salicylic acid induced adaptive response of …prr.hec.gov.pk/jspui/bitstream/123456789/7681/1/husn...

SALICYLIC ACID INDUCED ADAPTIVE RESPONSE OF

SUNFLOWER

(Helianthus annuus L.) TO DROUGHT STRESS

HUSN-E-SEHAR ZAIDI

07-arid-1273

Department of Botany

Faculty of Sciences

Pir Mehr Ali Shah

Arid Agriculture University Rawalpindi,

Pakistan

2015

2

SALICYLIC ACID INDUCED ADAPTIVE RESPONSE OF

SUNFLOWER

(Helianthus annuus L.) TO DROUGHT STRESS

by

HUSN-E-SEHAR ZAIDI

(07 -arid –1273)

A thesis submitted in partial fulfillment of the requirements for the degree of

DOCTOR OF PHILOSOPHY

in

BOTANY

Department of

Botany Faculty of

Sciences

Pir Mehr Ali Shah

Arid Agriculture University, Rawalpindi

Pakistan

3

2015

CERTIFICATION

I hereby undertake that this research is an original one and no part of this

thesis falls under plagiarism. If found otherwise, at any stage, I will be

responsible for the consequences.

Name: Husn-e-SeharZaidi Signature: ____________________________

Registration No: 07-arid-1273 Date:

_______________________________

It is certified that the contents and form of thesis entitled “Salicylic acid

induced

adaptive response of Sunflower (Helianthus annuus L.) to Drought stress ”by

Ms.

Husn-e-Sehar Zaidi has been found satisfactory for the requirement of the

degree.

Supervisor: ____________________________

(Prof. Dr. Abdul Waheed)

Co Supervisor: _________________________

(Dr. Jalal-ud-Din)

Member: ____________________________

(Prof. Dr. Muhammad Arshad)

Member: ____________________________

(Dr. Abdul Razzaq) Chairman: ________________________

4

Dean: ____________________________

Director Advanced Studies: __________________________

6

DEDICATED

TO

MY LOVING AMMI & DADDY

May Allah rest their souls in peace

And

MY BELOVED SISTER

Dr. Naveed-e-Sehar Zaidi

For her endless love, support and

encouragement

CONTENTS

Page

List of Tables 11

7

List of Figures 21

Abbreviations 29

Acknowledgements 30

Abstract 32

1 INTRODUCTION 1

2 REVIEW OF LITERATURE 8

2.1 SIGNIFICANCE OF SUNFLOWER 8

2.2 EFFECT OF DROUGHT ON CROP GROWTH AND 10

BIOMASS

2.3 EFFECT OF DROUGHT ON PHYSIOLOGICAL 12

ATTRIBUTES

2.4 EFFECT OF DROUGHT ON GAS EXCHANGE 13

CHARACTERISTICS

2.5 EFFECT OF DROUGHT ON BIOCHEMICAL ATTRIBUTES 14

2.6 EFFECT OF DROUGHT ON ANTIOXIDANT ACTIVITY 16

2.7 EFFECT OF DROUGHT ON YIELD ATTRIBUTES 17

2.8 STRATEGIES TO OVERCOME DROUGHT STRESS 19

2.8.1 Induction of Drought Resistance 19

2.9 EFFECT OF SALICYLIC ACID ON GROWTH AND BIOMASS 25

2.10 EFFECT OF SALICYLIC ACID ON PHYSIOLOGICAL 28

ATTRIBUTES

2.11 EFFECT OF SALICYLIC ACID ON BIOCHEMICAL 29

ATTRIBUTES

2.12 EFFECT OF SALICYLIC ACID ON ANTIOXIDANTS 31

ACTIVITY

2.13 EFFECT OF SALICYLIC ACID ON GRAIN YIELD 32

3 MATERIALS AND METHODS 35

3.1 EXPERIMENTAL SITE AND CONDITIONS 35

3.2 ACHENE MATERIAL 35

8

3.3 SCREENING OF SUNFLOWER HYBRIDS AT DIFFERENT

DROUGHT STRESS LEVELS FOR THEIR GROWTH

ATTRIBUTES

36

3.4 IMPROVING THE GROWTH OF DROUGHT-STRESSED

SUNFLOWER HYBRIDS BY SALICYLIC ACID APPLICATION

37

3.4.1 Effect Of Foliar Spray Of SA On Sunflower Hybrids At Seedling

Stage

39

3.4.2 Effect Of Seed Soaking In SA Of Sunflower Hybrids At Seedling

Stage

39

3.5 INFLUENCE OF FOLIAR APPLICATION OF SALICYLIC ACID

WITH DIFFERENT CONCENTRATIONS AT TWO GROWTH

STAGES OF SUNFLOWER HYBRIDS UNDER DROUGHT

STRESS

40

3.5.1 Seed Sowing 40

3.5.2 Development And Maintenance Of Water Stress Levels 40

3.6 PROCEDURES FOR DATA COLLECTION 42

3.6.1 Plant Growth And Development Attributes 42

3.6.2 Physiological Parameters 43

3.6.3 Biochemical Parameters 44

3.6.4 Anti-oxidative Enzymes 47

3.6.5 Determination Of Endogenous Level Of Salicylic Acid 49

3.6.6 Quality Parameters 50

3.6.7 Yield And Yield Related Traits 53

3.7 STATISTICAL ANALYSIS 54

4 RESULTS AND DISCUSSIONS 55



ATTRIBUTES

55

4.1.1 Seed Germination 55

4.1.2 Plant Height 58

4.1.3 Root Length 60

4.1.4 Plant Dry Matter 63



65

9


4.2.2 Root Length 71

4.2.3 Fresh Shoot Weight 77

4.2.4 Dry Shoot Weight 83

4.2.5 Fresh Root Weight 89

4.2.6 Dry Root Weight 93

4.2.7 Photosynthesis Rate 99

4.2.8 Stomatal Conductance 105

4.2.9 Relative Water Content 111

4.2.10 Water Potential 117

4.2.11 Osmotic Potential 123

4.2.12 Turgor Potential 128

4.2.13 Leaf Proline Content 134

4.2.14 Soluble Sugars 140

4.2.15 Leaf Protein 145

4.2.16 Free Amino Acid Content

152



158

4.3.1 Fresh Root Weight 158

4.3.2 Dry Root Weight 163

4.3.3 Fresh Shoot Weight 167

4.3.4 Dry Shoot Weight 171


4.3.6 Leaf Count 180

4.3.7 Leaf Area 183

10

4.3.8 Water Potential 188

4.3.9 Osmotic Potential 192

4.3.10 Turgor Potential 196

4.3.11 Relative Water Content 200

4.3.12 Photosynthesis Rate 205

4.3.13 Stomatal Conductance 209

4.3.14 Leaf Diffusive Resistance 214

4.3.15 Chlorophyll a Content 218

4.3.16 Chlorophyll b Content 220

4.3.17 Leaf Proline Contents 226

4.3.18 Soluble Sugar Contents 231

4.3.19 Leaf Protein Contents 235

4.3.20 Amino Acid Content 239

4.3.21 Superoxide Dismutase Activity 244

4.3.22 Peroxidase Activity 249

4.3.23 Catalase Activity 253

4.3.24 Endogenous Salicylic acid Level 258

4.3.25 Palmitic Acid Content 259

4.3.26 Stearic Acid Content 265

4.3.27 Oleic Acid Content 266

4.3.28 Linoleic Acid Content 272

4.3.29 Protein Content 277

4.3.30 Oil Content 282

4.3.31 Head Diameter 286

4.3.32 Number of Achenes 290

4.3.33 1000 Grain Weight 294

11

4.3.34 Achene Yield 299

4.3.35 Oil Yield 303

4.3.36 Biological Yield 307

SUMMARY 312

LITERATURE CITED 320

LIST OF TABLES

Table No Page

2.1.1 Plant height (cm) of sunflower hybrids under different

drought 67 stress levels as influenced by foliar

application of salicylic acid.

2.1.2 Plant height (cm) of sunflower hybrids under different

drought 68 stress levels as influenced by seed soaking in

salicylic acid.

12

2.2.1 Root length (cm) of sunflower hybrids under different

drought 73 stress levels as influenced by foliar


2.2.2 Root length (cm) of sunflower hybrids under different

drought 75 stress levels as influenced by seed soaking in

salicylic acid.

2.3.1 Fresh shoot weight (g/plant) of sunflower hybrids under

different 79 drought stress levels as influenced by foliar


2.3.2 Fresh shoot weight (g/plant) of sunflower hybrids under

different 80 drought stress levels as influenced by seed

soaking in salicylic acid.

2.4.1 Dry shoot weight (g/plant) of sunflower hybrids under



2.4.2 Dry shoot weight (g/plant) of sunflower hybrids under



2.5.1 Fresh root weight (g) of sunflower hybrids under different 90

drought stress levels as influenced by foliar application

of salicylic acid.

13

2.5.2 Fresh root weight (g) of sunflower hybrids under different

92 drought stress levels as influenced by seed soaking

in salicylic acid.

2.6.1 Dry root weight (g/plant) of sunflower hybrids under


application of

salicylic acid.

2.6.2 Dry root weight (g/plant) of sunflower hybrids under



2.7.1 Photosynthesis rate (µmol/m2/s) of sunflower hybrids under 100

different drought stress levels as influenced by foliar


2.7.2 Photosynthesis rate (µmol/m2/s) of sunflower hybrids under

102 different drought stress levels as influenced by seed


2.8.1 Stomatal conductance (mmol/m²/s) of sunflower hybrids

under 106 different drought stress levels as influenced

by foliar application of salicylic acid.

14

2.8.2 Stomatal conductance (mmol/m²/s) of sunflower hybrids

under 108 different drought stress levels as influenced

by seed soaking in salicylic acid.

2.9.1 Relative water content(%) of sunflower hybrids under



2.9.2 Relative water content(%) of sunflower hybrids under different 115

drought stress levels as influenced by seed soaking in salicylic

acid.

2.10.1 Water potential (-MPa) of sunflower hybrids under different 118

drought stress levels as influenced by foliar application of

salicylic acid.

2.10.2 Water potential (-MPa) of sunflower hybrids under different 119


acid.

2.11.1 Osmotic potential (-MPa) of sunflower hybrids under different 124


salicylic acid.

2.11.2 Osmotic potential (-MPa) of sunflower hybrids under different 126


acid.

15

2.12.1 Turgor potential(-MPa) of sunflower hybrids under different 129


salicylic acid.

2.12.2 Turgor potential(Mpa) of sunflower hybrids under different 131


acid.

Leaf proline concentration (µg/g) of sunflower hybrids under

2.13.1 different drought stress levels as influenced by foliar

application 136 of salicylic acid.

2.13.2 Leaf proline concentration (µg/g) of sunflower hybrids under



2.14.1 Leaf soluble sugar (mg/g) of sunflower hybrids under



2.14.2 Sugar content (mg/g) of sunflower hybrids under different

143 drought stress levels as influenced by seed soaking

in salicylic acid.

2.15.1 Leaf protein (µmol/g f wt) of sunflower hybrids under



16

2.15.2 Leaf protein (µmol/g f wt) of sunflower hybrids under



2.16.1 Amino acid content (µmol/g f wt) of sunflower hybrids under

153 different drought stress levels as influenced by

foliar application of salicylic acid.

2.16.2 Amino acid content (µmol/g f wt) of sunflower hybrids under



3.1 Root fresh weight (g/plant) of drought stressed sunflower

hybrids 159 under foliar applied various concentrations of

salicylic acid at two growth stages.

3.2 Root dry weight (g/plant) of drought stressed sunflower

hybrids 164 under foliar applied various concentrations

of salicylic acid at two growth stages.

3.3 Shoot fresh weight (g/plant) of drought stressed sunflower 169

hybrids under foliar applied various concentrations of


3.4 Shoot dry shoot weight (g/plant) of drought stressed

sunflower 174 hybrids under foliar applied various

concentrations of salicylic acid at two growth stages.

17

3.5 Plant height (cm) of drought stressed sunflower hybrids

under 177 foliar applied various concentrations of


3.6 Leaf count (#/plant) of drought stressed sunflower hybrids



3.7 Leaf area (cm2/plant) of drought stressed sunflower hybrids 186

under foliar applied various concentrations of salicylic

acid at two growth stages.

3.8 Water potential (-Mpa) of drought stressed sunflower



3.9 Osmotic (-Mpa) of drought stressed sunflower hybrids under 193

foliar applied various concentrations of salicylic acid at

two growth stages

3.10 Turgor potential (Mpa) of drought stressed sunflower



3.11 Relative water content (%) of drought stressed sunflower



18

3.12 Photosynthesis rate (µmol/m2/s) of drought stressed



3.13 Stomatal conductance (mmol/m²/s) of drouht stressed



3.14 Leaf diffusive resistance (cm/sec2) of drought stressed



3.15 Chlorophyll a content (mg/g FW) of drought stressed



3.16 Chlorophyll b content (mg/g FW) of drought stressed



3.17 Proline content (mg/g FW) of drought stressed sunflower



3.18 Sugar content (mg/g FW) of drought stressed sunflower



19

3.19 Protein content (mg/g FW) of drought stressed sunflower



3.20 Amino acid content (µmol/g) of drought stressed sunflower 241

hybrids under foliar applied various concentrations of


3.21 Superoxide dismutase activity (U/g FW) of drought stressed

245 sunflower hybrids under foliar applied various


3.22 Peroxidase activity (U/g FW) of drought stressed sunflower

250 hybrids under foliar applied various concentrations


3.23 Catalase activity (U/g FW) of drought stressed sunflower



3.24 Endogenous Salicylic acid level (ng/g) of drought stressed 260

sunflower hybrids under foliar applied various concentrations of


Palmitic acid content (%) of drought stressed sunflower

hybrids

3.25 under foliar applied various concentrations of salicylic acid at

263 two growth stages.

20

3.26 Stearic acid content (%) of drought stressed sunflower



3.27 Oleic acid content (%) of drought stressed sunflower hybrids

270 under foliar applied various concentrations of


3.28 Linoleic acid content (%) of drought stressed sunflower



3.29 Protein content (%) of drought stressed sunflower hybrids



3.30 Oil content (%) of drought stressed sunflower hybrids under

284 foliar applied various concentrations of salicylic


3.31 Head diameter (cm) of drought stressed sunflower hybrids



3.32 Number of achenes (# head-1) of drought stressed sunflower

292 hybrids under foliar applied various concentrations


21

3.33 1000 grain weight (g) drought stressed sunflower hybrids


salicylic acid at two growth stages

3.34 Achene yield (g/plant) of drought stressed sunflower hybrids



3.35 Oil yield (g/plant) of drought stressed sunflower hybrids under 305

foliar applied various concentrations of salicylic acid at two

growth stages.

Biological yield (g/plant) of drought stressed sunflower hybrids 308

3.36 under foliar applied various concentrations of salicylic


LIST OF FIGURES

Fig. No Page

1.1 Effect of different drought stress levels on seed germination

57 stress index (SGSI) of various sunflower hybrids.

1.2 Effect of different drought stress levels on plant height stress

59 index (PHSI) of various sunflower hybrids

1.3 Effect of different drought stress levels on root length stress

61 index (RLSI) of various sunflower hybrids

22

1.4 Effect of different drought stress levels on dry matter stress

64 index (DMSI) of various sunflower hybrids

2.1 Plant height of sunflower hybrids under various levels of 70

drought stress and salicylic acid application

2.2 Root length of sunflower hybrids under various levels of 76

drought stress and salicylic acid application

2.3 Fresh shoot weight of sunflower hybrids under various

levels 82 of drought stress and salicylic acid

application.

2.4 Dry shoot weight of sunflower hybrids under various levels

87 of drought stress and salicylic acid application

2.5 Fresh root weight of sunflower hybrids under various levels

94 of drought stress and salicylic acid application

2.6 Dry root weight of sunflower hybrids under various levels of

98 drought stress and salicylic acid application

2.7 Photosynthesis rate of sunflower hybrids under various


application

2.8 Stomatal conductance of sunflower hybrids under various 110

levels of drought stress and salicylic acid application.

23

2.9 Relative water content of sunflower hybrids under various 116

levels of drought stress and salicylic acid application.

2.10 Water potential of sunflower hybrids under various levels of

121 drought stress and salicylic acid application.

2.11 Osmotic potential of sunflower hybrids under various levels

127 of drought stress and salicylic acid application.

2.12 Turgor potential of sunflower hybrids under various levels of


2.13 Proline content of sunflower hybrids under various levels of


2.14 Sugar content of sunflower hybrids under various levels of


2.15 Protein content of sunflower hybrids under various levels of

151 drought stress and salicylic acid application

2.16 Amino acid content of sunflower hybrids under various


application

3.1 Fresh root weight of drought stressed sunflower hybrids 161



24

3.2 Dry root weight of drought stressed sunflower hybrids



3.3 Fresh shoot weight of drought stressed sunflower hybrids 170



3.4 Dry shoot weight of drought stressed sunflower hybrids 175



3.5 Plant height of drought stressed sunflower hybrids under 178


two growth stages.

3.6 Leaf count of drought stressed sunflower hybrids under

foliar 182 applied various concentrations of


3.7 Leaf area of drought stressed sunflower hybrids under foliar

187 applied various concentrations of salicylic acid at

two growth

stages

3.8 Water potential of drought stressed sunflower hybrids



25

3.9 Osmotic potential of drought stressed sunflower hybrids 195



3.10 Turgor potential of drought stressed sunflower hybrids



3.11 Relative water content of drought stressed sunflower

hybrids 203 under foliar applied various


3.12 Photosynthesis rate of drought stressed sunflower hybrids



3.13 Stomatal conductance of drought stressed sunflower



3.14 Leaf diffusive resistance of drought stressed sunflower 217

hybrids under foliar applied various concentrations


3.15 Chlorophyll a content of drought stressed sunflower



26

3.16 Chlorophyll b of drought stressed sunflower hybrids under



3.17 Proline content of drought stressed sunflower hybrids



3.18 Sugar content of drought stressed sunflower hybrids under



3.19 Leaf Protein of drought stressed sunflower hybrids under 238

foliar applied various concentrations of salicylic acid at two

growth stages.

3.20 Amino acid content of drought stressed sunflower hybrids 242



3.21 Superoxide dismutase activity of drought stressed



3.22 Peroxidase activity of drought stressed sunflower hybrids



27

3.23 Catalase activity of drought stressed sunflower hybrids



3.24 Endogenous Salicylic acid level of drought stressed 261

sunflower hybrids under foliar applied various


3.25 Palmitic acid content of drought stressed sunflower



3.26 Stearic acid content of drought stressed sunflower hybrids



3.27 Oleic acid content of drought stressed sunflower hybrids 271



3.28 Linoleic acid content of drought stressed sunflower



3.29 Protein content of drought stressed sunflower hybrids



28

3.30 Oil content of drought stressed sunflower hybrids under



3.31 Head diameter of drought stressed sunflower hybrids under


acid at two

growth stages.

3.32 Number of achenes of drought stressed sunflower hybrids 293



3.33 1000 grain weight of drought stressed sunflower hybrids 297



3.34 Achene yield of drought stressed sunflower hybrids under 302


two growth stages.

3.35 Oil yield of drought stressed sunflower hybrids under



3.36 Biological yield of drought stressed sunflower hybrids



29

LIST OF ABBREVIATIONS

Description

DS Drought Stress

SA Salicylic acid

H Hybrid

LA Leaf area Fig Figure m

Meter

m2 Meter square

m-2 Per meter square cm Centimeter mm

Millimeter

g Gram

kg Kilogram

RWC Relative water content

mM Milimolar

ppm Parts per million DAS Days after

sowing

C Control

PGR Plant growth regulator PEG Polyethylene

glycol

Abbreviation

30

ACKNOWLEDGEMENTS

In the beginning, I would like to express my deepest and sincere

gratitude to my supervisor Prof. Dr. Abdul Waheed, Director CIIT, Sahiwal

for his scientific advice, step to step guidance and moral encouragement. Deep

appreciation is extended to my

Co-supervisor, Dr. Jalal-ud-Din Principal Scientist, National Agricultural

Research Center, Islamabad who led me to the successful endings of my

research work by providing every possible facility.

This feat was possible only because of the unconditional support

provided by Dr. Sami-ullah-Khan with an amicable and positive disposition.

It is great privilege for me to record my heartiest and sincerest thanks to Dr.

Ayub, Dr. Buksh, Dr. Ghazala Kokab and Dr. Zahid Sharif who has

always made himself available to clarify my doubts despite their busy

schedules and I consider it as a great opportunity to learn from their research

expertise for their valuable suggestions and moral support.

I would like to express special thanks to M. Iftikhar (lab attendant NARC)

for his help and support.

I am also thankful to Higher Education Department, Govt. of Punjab for

giving me opportunity and facilitate me to achieve this degree.

31

Sincere thanks to all my friends especially Zahida Humayun,

Mamoona Illyas, Fakhra Jabeen , Kubra and others for their kindness and help.

Thanks for the friendship and memories.

I owe a lot to my parents, who encouraged and helped me at every

stage of my personal and academic life, and longed to see this achievement

come true. I deeply miss my mother, who took the lead to heaven before the

completion of this work. I am very much indebted to my family, my dear

husband Naeem who remains stood by me through the good times and bad.

My son, Aaliyan, who has suffered because of my work. I am greatly thankful

to my brother and his family and my sister, brother –in-law and nieces: Amna,

Ayesha and Maryam for their care and best wishes.

Finally I thank my Allah, for letting me through all the difficulties. I

have experienced Your guidance day by day. You are the one who let me

finish my degree. I will keep on trusting You for my future. Thank you.

HUSN-E-SEHAR

ZAIDI

32

ABSTRACT

Pakistan has very low production of edible oil, so spends huge amount

of foreign exchange on its import. Sunflower (Helianthus annuus L.) is the

third most widely cultivated crop for vegetable oil following soybean and

palm. However, drought stress causes severe adverse effect on growth and

yield of sunflower crop. To cope with this osmotic stress, plants respond by

production of hormones, accumulation of compatible solutes and acceleration

of antioxidant system. Salicylic acid (SA) is an endogenous plant regulator

that helps to sustain plant growth under stress conditions. It is not produced in

sufficient amount in plants to avert the negative effects of abiotic stresses

including drought. So, it has recently been suggested to apply SA exogenously

to the plants for enhancing stress tolerance. Growth stage of plant for SA

application, adequate dose, and its mechanism of action could vary among

species and genotypes.

This study was, therefore, performed to work out the optimum dose,

application time and probable physiological and biochemical response of SA

for ameliorating the negative consequences of drought stress in sunflower.

33

First, the effect of SA on six sunflower hybrids viz., Hyoleic-41, FH-

352, NX00989, Hysun-33, NX-19012 and Parsun-2 subjected to drought

stress at germination and seedling stage was assessed in a laboratory

experiment (25±3 °C). Four levels of water stress viz., 0.0 (control), -0.6, -

1.33, and -1.62 MPa were employed using polyethyleneglycol-8000 (PEG-

8000) under completely randomized design (CRD) with three replications..

Plant height and dry matter stress tolerance indices reduced with increasing

water stress in all the sunflower hybrids. Contrastingly, there was an increase

of RLSI in all sunflower hybrids except Hyoleic-41 and FH-352. Sunflower

hybrids

NX-00989 and NX-19012 performed better and were classified as drought tolerant.

Variation among hybrids for DMSI was found to be a reliable indicator of drought

tolerance in sunflower.

Second laboratory experiment was conducted to evaluate the mode of

SA application. It was divided in two sub-experiments, viz., foliar application

of SA, and seed soaking with SA, each with the following treatments: two

stress tolerant hybrids (NX-19012, NX-00989) and one sensitive hybrid (FH-

352), were subjected to three drought stress (PEG) levels (0, 10 and 20 %),

and further treated with four SA levels (0, 0.375, 0.75 and 1.50 mM) under

CRD with three replications in the lab. For the first set having foliar SA

application, Seeds of sunflower were grown in Hoagland’s nutrient solution.

Drought stress was applied to the 15 days old plants, and SA was sprayed on

the foliage of plants. For second set of experiment having seed treatment with

SA, seeds were soaked in SA solution for 10 hr, and grown in Hoagland’s

nutrient solution for 15 days. Drought stress was applied to the plants

afterwards. Results showed that fresh and dry biomass of sunflower was

34

reduced by the water stress but enhanced in both modes of SA application in

stressed and non-stressed plants. Plant water relations were improved with

foliar application of SA 0.75mM at all levels of stress, except in sensitive

genotype at 20% drought stress, more than pre treatment. It was concluded

that water deficit adversely affected all the growth, physiological and

biochemical parameters, which remained normal / restored in the presence of

salicylic acid in both stressed and non-stressed plants. Further, foliar SA

application enhanced the drought stress tolerance in plants better than pre-

treatment.

In order to study the interaction between drought stress and salicylic

acid on growth, physiological, water relations, biochemical, antioxidant, and

yield related attributes of sunflower hybrids, a factorial experiment with CRD

and three replications was undertaken. Treatments included four levels of

salicylic acid (0, 0.375, 0.75 and 1.5 mM) applied as foliar spray at vegetative

and flowering stages to water stressed plants under greenhouse conditions. To

one set of pots, water stress was given and sprayed with various concentrations

of SA at vegetative stage by withholding water till wilting to plants and then

re-watered. To second set of pots, water was withheld and foliar sprayed with

various concentrations of SA at flowering stage till wilting, then irrigated. The

control plants were irrigated normally till harvesting. Results indicated that

plant biomass was improved by SA application but non significantly,

however, leaf area, photosynthesis and stomatal conductance were

significantly increased by SA with 0.75 mM at vegetative stage. Water

relations parameters were also restored by SA at 0.75 significantly,

compatible solutes like proline and sugar contents at vegetative stage

enhanced more prominently in tolerant hybrids than in the sensitive ones

35

under stress and with SA applications. Activities of antioxidant enzymes

(superoxide dismutase, catalase and peroxidase) of all hybrids increased due

to moisture stress. Foliar applied SA caused a significant increase in leaf SOD

and POD activity at flowering stage. However, CAT activity and endogenous

SA level in leaves increased due to SA application under drought stress at both

vegetative and flowering stages in tolerant hybrids by 0.75 mM. Protein and

oil contents of seeds were improved more at vegetative stage by SA conc. of

0.75 and 1.5 mM. Application of water stress decreased the head diameter,

seed number per head, thousand seed weight, achene yield, oil yield,

biological yield and other related traits significantly. All the yield related

parameters were improved by the SA conc. 0.75 mM more than with 0.37 5

and 1.5 mM at vegetative stage in drought tolerant hybrids, except achene

number per head which was greater in sensitive FH-352. It is inferred that SA

is an environment friendly alternative that can be employed to improve the

growth and yield of drought-stressed sunflower plants at vegetative stage.

Chapter 1

INTRODUCTION

Pakistan is persistently low in the production of edible oil, and the situation

is getting worse day by day with increase in population growth (Asif et al., 2001).

Consumer‘s demand has increased from 0.32 to 2.76 million tonnes during the last

three decades with almost stagnant domestic production of 0.875 million tonnes.

Currently, indigenous oil seed production meets only 30% of the domestic need

36

(GOP, 2013).

Oilseed crops are categorized as traditional and non-traditional. Traditional

crops are rapeseed, mustard, linseed, sesame and safflower. As non-traditional crop,

sunflower can reduce the gap between production and import of edible oils (Khan et

al., 2003). Sunflower oil accounts for 80% of the value of sunflower crop and known

as premium oil due to high content of un-saturated fatty acids and lower linolenic

acid. The high percentage of polyunsaturated fatty acids 60%; including oleic acid

(16.2%) and linoleic acid (72.5%) that help to control blood cholesterol

(Satyabrata et al., 1998). The nutrients in sunflower seeds include 25-48% oil and

20-70% protein (Hatam and Abbasi, 1994), thiamine, vitamin A, D, E and K, iron,

phosphorous, potassium and calcium.

Water is a mandatory entity for the whole life cycle of sunflower crop, but

vegetative, heading and flowering stages are more susceptible to drought stress and

water insufficiency at these critical stages results in significant yield reduction. In

1

sunflower water stress at vegetative and reproductive growth stage may result in 61

and 40 % yield cutback, respectively (Iqbal, 2004). Further, it needs only 90-120

frost free days being grown twice as spring and autumn crop. The yield of sunflower

is destabilized by both biotic and abiotic stresses. Drought is an imperative climatic

anomaly that confines crop growth, biomass and yield.

Plants are always confronted with environmental constraints of both biotic

and abiotic nature and its response to both stresses can be additive, synergistic or

37

antagonistic and are affirmed by quantifying different quantitative and qualitative

traits (Ehdaie et al., 2008). Drought stress is characterized by water scarcity,

following unpredictable and substandard rainfall, high water demands or merging of

all these environmental constraints which twist productive land into barren terrain

annually (Ramakrishna et al., 2000). About one third of the earth‘s surface is

subjected to permanent drought due to which it is classified as arid and semiarid.

Most of the world food supply is produced in humid temperate regions, which are

often subjected to periods of severe drought (Rajaram et al., 1996; Van-Ginkel et al.,

1998).

Drought stress regardless of the growth stage decreases yield of the crops

(Jensen and Mogenson, 1984). Drought, the major abiotic stress, affects every aspect

of plant growth and is mainly responsible for limiting crop production (Golbashy et

al., 2010). Plants alleviate the harsh effect of drought stress by modifying different

physiological and biochemical adaptations (Hsieh et al., 2002).

Plants exhibit a wide range of response at molecular, cellular and whole plant

levels when exposed to osmotic stress (Zhu et al., 1997; Yeo, 1998; Hasegawa et al.,

2000). These changes include morphological and developmental changes (e.g life

cycle, inhibition of shoot and enhancement of root growth), adjustment in ion

transport (such as uptake, extrusion and sequestration of ions) and metabolic changes

(e.g carbon metabolism, the synthesis of compatible solutes). Drought resistance is

a complicated phenomenon in which different characteristics influence plant success

during vegetative period (Khan and Khan, 2010). It is accomplished by modulation

of gene expression and accumulation of specific protective proteins and metabolites

(Reddy et al., 2004; Zang and Komatsu, 2007). Almost all plants show water stress

tolerance, but its degree varies from species to species (Chaitanya et al., 2003).

38

Roots are supposed to be the primary sensors of water deficit, causing the

observed physiological and biochemical perturbations in the shoots and decline in

growth (Mujtaba and Alam, 2002).Water stress adversely affects the production of

fresh and dry biomass in crop plants (Ashraf and O‘Leary, 1996). Reduced biomass

production has been observed in almost all genotypes of sunflower resulting from

water stress (Tahir and Mehdi, 2001). Research on sunflower showed that water

stress significantly reduced, shoot length, fresh and dry weight, total leaf area,

chlorophyll a, b and total chlorophyll while increased the proline, free amino acids

and glycinebetaine contents (Manivannan et al., 2007).

Plant cells encounter the unfavorable environmental conditions by

accumulating a variety of small organic metabolites referred collectively as

compatible solutes such as proline, soluble sugars and proteins. Properties of

compatible solutes allow the maintenance of turgor pressure during water stress

(Sakamoto and Murata, 2002). This phenomenon of accumulation of compatible

solutes is called osmotic adjustment. Osmotic adjustment accepted as an effectual

way of drought resistance in many crops (Zhang et al., 2004).

Stomatal closure is the earliest response to drought and the dominant

limitations to photosynthesis at mild to moderate drought. However, in parallel,

progressive down-regulation or inhibition of metabolic processes leads to decreased

RuBP content, which becomes the dominant limitations at severe drought, and

thereby inhibits photosynthetic CO2 assimilation (Flexas and Medrano, 2002).

Concentrations of photosynthetic enzymes decrease, in response to drought, by

altering the hormonal balance. Non-stomatal limitations to photosynthesis under

39

water deficit conditions are linked to oxidative stress to macro- and micro-molecules.

Expression of reactive oxygen species (ROS) is severely enhanced in wheat,

sunflower and pea under drought stress because of photo-reduction of oxygen

(Sgherri and Navari-izzo, 1995; Sgherri, 1996).

Iqbal et al. (2005) working on two sunflower lines reported that there was a

marked adverse effect of water stress on 100-achene weight and achene oil contents

in both sunflower lines. Water stress reduced the head diameter, number of achenes,

1000-achene weight, achene‘s yield and oil yield in sunflower (Hussain

et al., 2008).

Biotechnology has provided considerable insights into mechanism of abiotic stress

tolerance in plants at molecular level (Zhu, 2001). For example, though stress

tolerance mechanisms may vary from species to species at different developmental

stages (Foolad and Lin, 2001), basic cellular responses to abiotic stresses are

conserved among most plant species (Zhu, 2002). Furthermore, different abiotic

stress factors may provoke osmotic stress, oxidative stress and protein denaturation

in plants, which lead to similar cellular adaptive responses such as accumulation of

compatible solutes, induction of stress proteins, and acceleration of reactive oxygen

species scavenging systems (Zhu, 2002; Khan et al., 2011 and 2012).

Salicylic acid (SA), an ubiquitous plant phenolic was recognized as an

endogenous growth regulator in plants after the finding that it is involved in many

plant physiological processes ,and regulator of plant metabolism, mainly involved in

biotic and abiotic stress (Yalpani et al., 1994; Szalai et al., 2000; Lian et al., 2000;

Aydin and Nalbantoglu 2011). In an extensive screening program using different

modern analytical technique, salicylic acid was detected in leaves and reproductive

40

organs of 34 argonomically important species (Raskin et al., 1990). SA is water-

soluble antioxidant compound that can regulate plant growth (Aberg, 1981).

Ameliorative effect of SA on growth of plants under abiotic stress conditions may

have been due to its role in nutrient uptake (Glass 1974), water relations (Barkosky

and Einhelling 1993), stomatal regulation (Arfan et al., 2007), photosynthesis and

growth (Khan et al., 2003; Arfan et al., 2007). Differnt modes of application as seed

treatment or foliar application of chemicals like glycinebetaine, kinetin, salicylic

acid (Gunes et al., 2007; Karlidag et al., 2009) may increase yield of different crops

due to reduction in stress induced inhibition of plant growth (Elwana and El-

Hamahmyb, 2009), enhanced photosynthetic rates, leaf area and plant dry matter

production (Khan et al., 2003).

Plant growth hormone salicylic acid produces protective effects on plants

under the activation of stress factors of different abiotic nature (Sakhabutdinova et al.,

2003). Thus, results have been obtained concerning the salicylic acid induced increase

in the resistance of wheat seedling to water deficit (Bezrukova et al., 2001), drought

resistance in wheat Singh and Usha (2003), low and high temp tolerance of tomatoe

and bean (Senaratna et al., 2000). Externally applied SA increased plant‘s tolerance to

several abiotic stresses, including osmotic stress (Wang et al., 2010), drought (Azooz

and Youssef, 2010), salinity (Gunes et al., 2007), and heavy metal stress (Moussa and

El-Gamel, 2010). SA appears to have the innate potentiality for enhancing antioxidants

and influencing antioxidant enzyme activity in plants subjected to oxidative stress

(Hayat et al., 2008; Kadioglu et al. 2011). In this respect salicylic acid was found to

improve the drought and salt stress (Hamada and Al-

Hakimi, 2001; Szalai et al., 2010; Shakeel and Mansoor, 2012).

41

Endogenous SA pool increased by the exogenous application of SA in the form

of o-glucosides (Raskin, 1992) that are more soluble and could be transported freely

inside the plant and generally enhance plant growth and final crop yield under stress

conditions. These reports clearly show that SA cannot induce abiotic stress tolerance

in all types of plants. However, numerous studies have demonstrated that the effect of

exogenous SA depends on various factors, including the species and developmental

stage, the mode of application and the concentration of SA (Vanacker et al., 2001;

Horvath et al., 2007). At low concentrations Brassinosteroids and

Salicylic acid promote growth in rice and gives resistance against abiotic stresses

(Farooq et al., 2009 a, b).

Although a lot of work has been done on different crops to mitigate the

adverse effects of water stress by exogenous application of different growth regulators,

little information is available regarding sunflower and exogenous application of

salicylic acid. Therefore, a series of experiments has been conducted to investigate the

effect of salicylic acid in mitigating the effects of moisture stress.

OBJECTIVES

To investigate the optimized concentration of salicylic acid effective in

drought stress.

To examine at what stage of growth salicylic acid application is significant.

To study the effect of salicylic acid on growth, yield and quality of sunflower

grown under drought stress.

Chapter 2

REVIEW OF LITERATURE

Sunflower (Helianthus annuus L.) is an annual plant native to North

America. It possesses a large inflorescence (flowering head). Sunflower got its name

from its huge, fiery blooms, whose shape and image is often used to depict the sun.

Sunflower has a rough, hairy stem, broad, coarsely toothed, rough leaves and circular

heads of flowers. The head consist of 1,000-2,000 individual flowers joined by a

receptacle base. Taxonomic description of sunflower is as follows:

Kingdom – Plantae (plants)

Subkingdom – Tracheobionta (vascular plants)

Superdivision – Spermatophyta (seed plants)

Division – Magnoliophyta (flowering plants)

Class – Magnoliopsida (dicotyledons)

Subclass – Asteridae

Order – Asterales

Family – Asteraceae (aster family)

Genus – Helianthus

Species – annuus (sunflower)

2.1 SIGNIFICANCE OF SUNFLOWER

Sunflower is one of the major and most important oilseed crops in the world due to

its excellent oil quality. Sunflower seed contains 25-48 % oil and 20-27 %

http://en.wikipedia.org/wiki/Annual_plant




http://en.wikipedia.org/wiki/Americas

http://en.wikipedia.org/wiki/Inflorescence

http://en.wikipedia.org/wiki/Inflorescence

http://plants.usda.gov/java/ClassificationServlet?source=display&classid=Magnoliopsida



http://plants.usda.gov/java/ClassificationServlet?source=display&classid=Asteridae



http://plants.usda.gov/java/ClassificationServlet?source=display&classid=Asterales



43

8

protein. Sunflower oil is composed of monounsaturated (C18:1) and

polyunsaturated (C18:2) fatty acids (60 %), accepted largely in diet to reduce

cholesterol in blood and prevents heart diseases (Rathore et al., 2001) and low

saturated fatty acids (C16:0 and C18:0). Sunflower cake is used as cattle feed

(Hussain et al., 2000). Sunflower is grown on 22 million hectares in the world;

producing 27 million tonnes total seed yield. It is primarily grown for oil, which is

mainly used for human consumption, but also as a raw material for the processing

industry, livestock feed and bee keeping. Sunflower breeders intend to achieve the

highest grain yield and oil content, through the best expression of heterosis

(Vrânceanu, 2000; Vrânceanu et al., 2005).

Imports provides the maximum portion of edible oil and only 30% of the

total domestic edible oil needs are met through local production. The share of

different crops in the domestic production of edible oil: Cottonseed 55.9%,

sunflower 29.1%, canola 7.60 %, and rapeseed / mustard 7.37% (GOP, 2008).

Among all these crops, sunflower is a high yielding crop. Okoko et al. (2008)

reported that sunflower is an important cash crop and a source of edible vegetable

oil of high quality.

Sunflower has been accepted as a high potential crop that can successfully

convene future oil requirements. In a wide range of climates of Pakistan sunflower

is being grown productively. It can endure a wide range of temperatures from 8 to

34°C, for better crop the optimum temperature is considered between 20 and 25°C

(Shah et al., 2005). Although sunflower is considered as moderately tolerant crop to

moisture stress yet drought greatly affects its area and production. The yield of

44

sunflower is destabilized by both biotic and abiotic stresses. Drought stress causes

severe adverse effect on growth, biomass and yield of the crop.

Drought stress is a major constraint for crop production in arid and semiarid

regions such as Pakistan. Drought severely impairs plant growth and development,

limits plant production and the performance of crop plants by affecting

photosynthesis,ion uptake, respiration, translocation and growth promoters, more

than any other enviryonmental factor (Shao et al., 2009; Jaleel et al., 2008 a-e).

Susceptibility of plants to drought stress varies due to independence of stress degree,

different accompanying stress factors, plant species, and their developmental stages

(Demirevska et al., 2009).

2.2 EFFECT OF DROUGHT ON CROP GROWTH AND BIOMASS

Plant growth is accomplished through cell division, cell enlargement and

differentiation, and it involves morphological, physiological, ecological and genetic

events and their complex interactions. The plant growth depends on the quality and

quantity of these events. The most drought-sensitive physiological process is cell

growth due to the reduction in turgor pressure (Taiz and Zeiger, 2006; Farooq et al.,

2010). In higher plants, under severe water loss, cell elongation can be inhibited by

interruption of water flow from the xylem to the surrounding elongating cells

(Nonami, 1998). Impaired mitosis, cell elongation and expansion result in reduced

plant height, leaf area and crop growth under drought (Hussain et al., 2008).Water

stress caused a visible increase in root length, soluble sugars and nitrogen, but a

considerable reduction in fresh and dry masses of root, growth vigor of shoot and

leaf area (Heshmat et al., 2012).

Meo (1999) studied the influence of nitrogen fertilizer and water stress on

leaf area of sunflower in pots. Urea as a nitrogen fertilizer was applied at the time

45

of sowing and sporadic stress was induced by a cycle of ten-day watering and tenday

stress period after 20, 30, 40 and 50 days of sowing. Results revealed that the

sporadic stress and urea fertilizer had a highly significant response, the leaf area

significantly decreased when either the stress period was increased or urea fertilizer

decreased. Drought stress affected plant height, dry matter, stem diameter, head size,

seed number/head, 100-seed weight and seed weight/ head, but leaf count was not

affected by either drought or defoliation (Nizami et al., 2008).

Sadeghipour and Aghaei (2012) showed that drought decreased plant height

and leaf area index (LAI). A general negative effect of water stress on crop plants is

the reduction in fresh and dry biomass production (Ashraf and O , Leary, 1996) and

reduced biomass production due to drought stress has been observed in almost all

genotypes of sunflower (Tahir and Mehdi, 2001). The predominant consequence of

drought is impaired germination and reduced stand establishment (Kaya et al.,

2006; Harris et al., 2002).

Water stress impaired the germination and early seedling growth of five

cultivar of pea (Okcu et al., 2005). Moreover, in alfalfa (Medicago sativa),

germination potential, hypocotyl length, and shoot and root fresh and dry weights

were decreased by induction of drought stress with polyethylene glycol, while it

increased the root length (Zeid and Shedeed, 2006). Water stress greatly reduced the

plant growth and development during the vegetative stage in rice (Tripathy et al.,

2000; Manikavelu et al., 2006). During water stress, water loss is decreased by

minimizing leaf area through reduced growth and shedding of older leaves. The

earliest response to drought is inhibition of leaf growth (Chaves et al., 2003). Leaf

expansion is severely inhibited at the onset of drought. Leaf cell expansion during

46

water stress is controlled by changes in the pH and inhibition is regulated by a rapid

decrease in extensibility of expansion in leaf cell walls (Hsiao and Xu, 2000).

Andrade et al. (2013) reported that water stress is likely the most important

factor that adversely affects plant growth and development. Two inbred lines of

sunflower with contrasting behavior to moisture deficit, the inbred lines B59,

sensitive, and B71, tolerant. Shoot and root relative fresh weight decreased in both

lines under water stress, although B71 showed a minor drop. Water stress affected

shoot dry weight in a greater proportion, than root dry weight in B59 and B71 line.

Saensee et al. (2012) tested seven sunflower genotypes and one commercial

hybrid, Pacific 77, at two water stress levels of -0.6 and -1.2 MPa, using

polyethylene glycol 6000 (PEG-6000). Dry matter stress index, plant height stress

index, root length stress index, relative water content stress index and germination

stress index were significantly decreased in all genotypes with increase in water

stress levels.

2.3 EFFECT OF DROUGHT ON PHYSIOLOGICAL ATTRIBUTES

Various forces acting through soil plant atmospheric continuum which allow

the uptake and loss of water constitute the water relations. Water potential, osmotic

potential, turgor potential and relative water content are the plant components. Plant

water relations are greatly influenced by relative water content, leaf water potential,

stomatal resistance, rate of transpiration, leaf temperature and canopy temperature.

During leaf growth relative water content of wheat was initially higher and reduced

as leaf matured and the dry matter accumulated (Siddique et al., 2001).

47

Generally, leaf osmotic and water potential decreases with water stress

intensity (Galle et al., 2002). Moisture stress significantly reduced the water

potential and relative water content, which had marked effect on rate of

photosynthesis (Siddique et al., 2000). Water stress significantly changed the

internal status of water in plants by lowering water potential and relative water

content of corn as a result inhibited photosynthetic rate and resulted in reduced final

yield (Atteya, 2003).

Ludlow and Muchow (1990) suggested that accumulation of solutes within

cell resulted in osmotic adjustment, which decreases osmotic potential and

maintains turgidity of plants in drought. Nayyar and Gupta (2006) reported that

drought stress decreased the relative water content (RWC).

2.4 EFFECT OF DROUGHT ON GAS EXCHANGE CHARACTERISTICS

A major effect of moisture stress is a reduction in photosynthesis, which

arises by a decrease in leaf expansion, impaired photosynthetic machinery,

premature leaf senescence and associated reduction in food production (Wahid and

Rasul, 2005). Moisture stress caused alterations in photosynthetic contents and

mechanism (Anjum et al., 2003), damage machinery of photosynthesis (Fu and

Huang, 2001) and cease Calvin cycle enzymes activities, the main causes of

reduction in yield of crop (Monakhova and Chernyadèv, 2002).

It was established that drought seriously inhibited the net rate of

photosynthesis (A) and rate of transpiration (E) in cow pea (Uzunova and Zlatev,

2013). Results had shown that stomatal or non-stomatal mechanisms were the cause

of low photosynthetic activity under moisture stress (Galle et al., 2010;

48

Samarah et al., 2009).

2.5 EFFECT OF DROUGHT ON BIOCHEMICAL ATTRIBUTES

During osmotic stress, plant cells accumulate solutes to prevent water loss

and to re-establish cell turgor. The solutes that accumulate during osmotic

adjustment include nitrogen containing compounds, such as praline and other amino

acids, polyamines and quaternary ammonium like glycine betain (Tamura et al.,

2003). These organic solutes are compatible with cellular processes and accumulate

in high level in cytosole with increasing drought. The accumulation and

mobilization of proline was found to be correlated with the levels of tolerance

towards drought stress in wheat (Nayyar and Walia, 2003). Drought stress

significantly reduced leaf RWC, chlorophyll a and b, carotenoids and soluble

proteins but enhanced the leaf proline (Ullah et al., 2012). Accumulation of proline

in wheat under water stress has been described (Sakhabutdinova et al., 2003).

Water stress also increased the free proline level (from 1.56 to 3.13 times). It is

concluded that proline may play a role in reducing the harm caused by water deficit

(Mohammadkhani and Heidari. 2008).

Proline possessive role in response to water stress was described by

Bandurska and Stroinski (2003). They observed that drought stress resulted in

proline addition in drought resistant wild accessions of Hordeuan Spontenum but no

accumulation in the sensitive H. oulgare. In rice high proline content was observed

in dry nursery as compared with plants in wet nursery (Zhao et al., 2001). Din et al.

(2011) showed that the chlorophyll (a, b) contents of all the Napus genotypes were

reduced under drought stress at both the growth stages. Tolerant genotype gave

slightest decline (12 %) during the flower initiation and pod filling stages in

49

chlorophyll content. Osmosis-regulating proline significantly enhanced under water

stress.

Sugars play an important role in Osmotic Adjustment (OA) as soluble sugars

concentration increased (from 1.18 to 1.90 times) in roots and shoots of both maize

varieties when subjected to drought stress. Appropriate exogenous application of

plant growth regulators may enhance plant growth and tolerance to moisture stress

(Arteca, 1996).

In drought sensitive (SQ1) and drought tolerant (CS) cultivars of wheat,

significant changes in physiological and biochemical attributes were reported when

SA (0.05 mM) or ABA (0.1 μM) was added to solutions containing PEG 6000

(−0.75 MPa). It has been shown that SA and ABA mitigated the adverse and after

effects of PEG, in drought resistant cultivar CS, by an increase in proline and

carbohydrate content as well as an increase in antioxidant activity, which help in the

better osmotic adjustment. Also, after the treatment of PEG about 98 % of osmotic

potential was decreased in wheat cultivars as compared to control

(Marcińska et al., 2013).

Parida et al., 2007 reported that the levels of biochemical components

(chlorophylls, carotenoids, proteins and starch contents) decreased in water stressed

plants compared to the control and increased during the recovery period. In the

sensitive genotype (Ca/H 631) the extent of decrease in chlorophylls, carotenoids

and protein contents under drought was higher as compared to the moderately

tolerant genotype (GM 090304). However, in drought stressed plants, proline, total

free amino acids, total sugars, reducing sugars and polyphenol contents were

increased and tended to decrease when the plants recovered from stress.

50

2.6 EFFECT OF DROUGHT ON ANTIOXIDANT ACTIVITY

When plants exposed to different environmental stresses it often generated

the ROS, including hydroxyl radicals (OH), hydrogen peroxide (H2O2), alkoxy

radicals (RO) and singlet oxygen (O1/2) (Munné-Bosch and Penuelas, 2003; Arora

et al. 2002). Production of ROS in excess can cause oxidative stress, which damages

plants by oxidizing photosynthetic pigments, membrane lipids, proteins and nucleic

acids (Yordanov et al., 2000). β-carotenes, ascorbic acid (AA), αtocopherol(α-toc),

reduced glutathione (GSH) and enzymes including: superoxide dismutase (SOD),

guaiacol peroxidase (POD), ascorbate peroxidase (APX), catalase (CAT),

polyphenol oxidase (PPO) and glutathione reductase (GR) are nonenzymatic

antioxidants (Xu et al., 2008).

Superoxide dismutases (SODs), a group of metalloenzymes, are considered

as the first defence against ROS. CAT, APX, POD are enzymes that catalyze the

conversion of H2O2 to water and O2 (Gratao et al., 2005). Balance between

production of ROS and activities of antioxidative enzymes determine whether

oxidative signalling and/or damage will occur (Moller et al., 2007). ROS may cause

oxidative damage and impairing the normal functions when react with proteins,

lipids and deoxyribonucleic acid (Foyer and Fletcher, 2001).

ROS production is linear with the water stress severity, which results in

enhanced peroxidation of membrane lipids and degradation of nucleic acids, and

both (structural and functional) proteins. First target of ROS created under water

stress is the cell organelles like chloroplasts, mitochondria and peroxisomes.

Drought stress enhanced preferentially the activities of superoxide dismutase (SOD)

and guaiacol peroxidase (POD) but decreased catalase (CAT) activity. Expression

51

of Mn-SOD and intensities of POD-4 and -5 were increased by water stress (Tayebe

and Hassan, 2010).

2.7 EFFECT OF DROUGHT ON YIELD ATTRIBUTES

In plants many yield-contributing physiological processes react to water stress.

Yield integrates in a complex way with many of these physiological processes.

Thus, to interpret how plants accumulate, combine and exhibit the everchanging and

indefinite physiological processes over the entire life cycle of crops is difficult. Iin

many field crops the reduction of yield due to water deficit at different growth stages

has been reported such as potato (Kawakami et al., 2006), common bean (Martinez

et al., 2007), rice (Lafitte et al., 2007). The severity, duration and timing of water

stress, as well as responses of plants and interaction between water stress and other

factors are particularly important, after removal of drought (Plaut, 2003).

Bajehbaj (2011) showed that four sunflower cultivars decreased plant height,

seed number per head, oil percentage, oil yield, biomass yield, seed yield, thousand

seed weight and other related traits significantly upon the application of water deficit

stress. Akhtar et al. (1993) observed that sunflower grown without water stress

shown maximum seed oil content (41.3%) than plants subjected to water stress at

seed setting, which gave the minimum seed oil content. MazaheryLaghab et al.

(2003) reported a 60% yield reduction in sunflower after drought application.

Nazariyan (2009) conducted an experiment with four levels of stress

(nowater stress, stress at head formation stage, stress at flowering stage and stress at

grain filling period) and the cultivars included Master and Lakomka and two hybrids

Euroflour and Azargol. Results depicted that Azargol had the highest yield under

no-stress conditions and Master had the lowest yield under stress at head formation

52

stage. Stress from head formation until the end of growing season had the highest

effect on components of yield, especially 1000-grain weight and head diameter as

the plants were exposed to drought stress for a longer time. Drought stress severely

decreased biological yield.

Dehkhoda (2013) evaluated the effect of water deficit on yield and yield

components of sunflower cultivars.Irrigation levels were: irrigation after 80 mm

evaporation from pan evaporation (control), irrigation after 130 mm evaporation

(ET130), irrigation of 180 mm evaporation (ET180). Results indicated that the

highest percentage of oil in ET130 treatment and lowest percentage of oil by ET180

treatment was obtained. The highest biological yield, harvest index, oil yield, seed

yield and seed weight was produced in control, and the lowest amount was pertained

to ET180.

2.8 STRATEGIES TO OVERCOME DROUGHT STRESS

Drought stress effects can be managed by production of the most suitable plant

genotypes. This is done to make sure that sensitive crop stages occurred at the time

when the possibility of drought is minimal. Various strategies of paramount

importance to achieve this objective may entail production of appropriate plant

varieties and improvement of the existing high-yielding varieties. Efforts have been

made to produce drought-tolerant genotypes using the knowledge of the responses

of plants to drought stress and mechanisms involved. The two most important

strategies may include: (a) selecting the desired materials as in traditional breeding

using molecular and biotechnological means and (b) inducing drought stress

tolerance in susceptible plants by priming and hormonal application. Only b is

discussed here.

53

2.8.1 Induction of Drought Resistance

Various strategies can be adopted to induce water stress resistance. Of these,

exogenous application of various growth regulating and other chemicals has

established in producing drought tolerance at various growth stages many plants.

An account of these strategies is given below:

2.8.1.1 Use of plant growth regulators

Exogenous application of plant growth regulators, both natural and

synthetic, has proved in improving growth against a variety of abiotic stresses.

Increase in hypocotyl length and fresh weight inhibited by drought stress, while

gibberelic acid (GA) inverted this effect. In this case, gibberelic acid partially

increased the water status of the seedlings and partially sustained protein synthesis

(Taiz and Zeiger, 2006). Exogenous application of GA increased the net

photosynthetic rate, stomatal conductance and transpiration rate in cotton (Kumar et

al., 2001), and stimulated pollen and seed cone production in Sitka spruce

(Piceasitchensis) under moisture stress (Philipson, 2003).

Under drought conditions, plant growth regulator treatments significantly

increased water potential, and improved chlorophyll content (Zhang et al., 2004).

Exogenous application of jasmonic acid induced drought resistance by enhancing

the betaine level in pear (Gao et al., 2004). Salicylic acid significantly enhances

plant growth under water stress conditions (Senaratna et al., 2000). In a study, when

salicylic acid applied exogenously under drought stress it improved the tolerance of

winter wheat (Horváth et al., 2007). Seed soaking or foliar spray of salicylic acid

and acetyl-salicylic acid (a derivative of salicylic acid), when applied at different

concentrations, protected muskmelon (Cucumis melo) seedlings prone to moisture

54

stress. However, the best results were obtained from pretreated seedlings with lower

concentrations of salicylic acid (Korkmaz et al., 2007).

Soaking of seeds with salicylic acid or acetylsalicylic acid confers stress

tolerance in plants is more consistent with signaling for gene expression rather than

their direct effects (Senaratna et al., 2000). The endogenous salicylic acid level was

enhanced in drought-stressed Phillyrea angustifolia (Munné-Bosch and Penuela,

2003), which suggest that salicylic acid might play role in the drought tolerance. In

wheat, abscisic acid content was shown to increase by the application of salicylic

acid, which results in the accumulation of proline (Shakirova et al., 2003).

Pretreatment with 0.5 mM SA for 1 day reduced the stress tolerance of 2-week-old

corn plants by enhancing their polyamine content (Németh et al., 2002).

Phytohormones are very much involved in directing the plant growth, in a

coordinated fashion in association with metabolism that provides energy and the

building blocks to develop the form that we recognize as plants. Out of the

recognized hormones, attention has largely been focused on auxins, gibberellins,

cytokinins, abscisic acid, ethylene and more recently to brassinosteroids. However,

a natural chemical- salicylic acid could be raised to the status of the above

phytohormones because it has significant impact on various aspects of the plant life.

Salicylic acid (SA) was first discovered as a major component in the extracts from

Salix (willow) whose bark from ancient time, was used as an antiinflammatory drug.

This acid (SA) is a phenol, ubiquitous in plants generating a significant impact on

plant growth and development, photosynthesis, transpiration, ion uptake and

transport and also induces specific changes in leaf anatomy and chloroplast

structure. The SA is recognized as an endogenous signal, mediating in plant defense,

against pathogens (Hayat, 2008).

55

2.8.1.2 Use of salicylic acid

Salicylic acid belongs to an extraordinary diverse group of plant phenolics

usually defined as substances that possess an aromatic ring bearing a hydroxyl group

or its functional derivative. Plant phenolics have often been referred to as secondary

metabolites. The term "secondary" implied that such compounds are only of minor

importance to the plant and could sometimes be equated with waste products. This

notion has been gradually replaced, however, by the view that many phenolic

compounds play an essential role in the regulation of plant growth, development,

and interaction with other organisms (Horborne, 1980).

Salicylic acid is an endogenous growth regulator of phenolic nature, which

participates in the regulation of physiological processes in plants and it is also

important in disease resistance (Raskin, 1992). Salicylic acid treatment reduced the

damaging action of salinity (Sakhabutdinova et al., 2003) water deficit (Singh and

Usha, 2003) and tomato on heat, chilling and drought stress (Senaratna et al., 2000)

on seedling growth and accelerated the restoration of growth processes. Salicylic

acid plays an important role in abiotic stress tolerance and considerable interests

have been focused on the ability of salicylic acid to induce a protective effect on

plants under stress.

Salicylic acid has been recently included in the category of phytohormones.

Using modern analytical techniques, it was found that salicylates are distributed in

many important agricultural plant species. In many plants such as rice, crabgrass,

barley, soybean the levels of SA have been found to be approximately 1µg g‾¹ fresh

weight.

The pathway for biosynthesis of salicylic acid include the formation of

benzoic acid and the most important mechanism for the formation of benzoic acid

56

in plants is the side chain degradation of cinnamic acids which are important

intermediates in the shikimic acid pathway. The conversion of cinnamic acid to SA

is likely to proceed via one of two pathways outlined in Figure a. These pathways

can operate independently in plants (Chadha and Brown, 1974). In plants, although

recent genetic finding indicates that over 90% of SA is synthesized from chorismate

(Chen et al., 2009).

The role of salicylic acid (SA) as a key molecule in the signal transduction

pathway of biotic stress responses has already been well described. Recent studies

indicate that it also participates in the signalling of abiotic stresses. The application

of exogenous SA could provide protection against several types of stresses such as

high or low temperature, drought, heavy metals, and so on. Although SA may also

cause oxidative stress to plants, partially through the accumulation of hydrogen

57

Fig. a. Pathway of salicylic acid biosynthesis in plants

peroxide, the results published so far show that the preliminary treatment of plants

with low concentrations of SA might have an acclimation-like effect, causing

enhanced tolerance toward most kinds of abiotic stresses primarily due to enhanced

antioxidative capacity.

The effect of exogenous SA depends on numerous factors such as the species

and developmental stage of the plant, the mode of application, and the concentration

of SA and its endogenous level in the given plant. Recent results show that not only

does exogenous SA application moderate stress effects, but abiotic stress factors

may also alter the endogenous SA levels in the plant cells. Salicylic acid is a potent

signalling molecule in plants and is involved in eliciting specific responses to biotic

and abiotic stresses (Krantev et al., 2006). SA is also involved in the activation of

the stress induced antioxidant system in plants, and is now considered to be a

hormonal substance that plays a key role in regulating plant growth and development

(Huang et al., 2008). A role for SA in plant growth and development, flowering

(Martin-Mex et al., 2005a; Martin-Mex et al., 2005b), ion uptake, stomatal

regulation and photosynthesis has been investigated by Uzunova and Popova

(2000). Intracellular SA concentration and SA signaling pathway(s) are associated

with the functions controlling cell growth, cell death and defense (Chen et al., 2001).

2.9 EFFECT OF SALICYLIC ACID ON GROWTH AND BIOMASS

Sayyari et al. (2013) observed the effect of salicylic acid on lettuce

(Lactuca sativa) with 3 levels of drought stress including stress-free conditions,

mild and severe stress and 3 concentrations of SA (0, 0.75 and 1.5 mM). Results

58

showed that drought stress imposed negative effects on plant growth and

productivity. In drought conditions, fresh and dry weight, leaf area, photosynthetic

pigments and RWC reduced, but EL, proline and MDA increased. Plant fresh and

dry weight, leaf area, carotenoids and proline significantly increased and MDA

accumulation and EL decreased by SA application.

Exogenous application of GB, SA or their interaction could counteract the

adverse effects of drought by improvement of growth vigor of root and shoot, leaf

area, retention of pigments content, increasing the concentration of organic solutes

(soluble sugars and soluble nitrogen) as osmoprotectants, keeping out the

polysaccharides concentration and/or stabilization of essential proteins in both

wheat cultivars.SA could improve the drought tolerance in wheat cultivars (Heshmat

et al., 2012). Salicylic acid is reported to cause an increase in root and shoot growth

(Khodary, 2004; El-Tayeb and Naglaa, 2010). However, it has also been observed

that foliar application of salicylic acid and acetyl salicylic acid to soybean and corn

plants @ 10 3 and 10 5 mol/L did not affect plant height and root length but showed

an increase in leaf area (Khan et al., 2003). Treatment of soybean seedlings with 0.1

mM salicylic acid increased the average leaf area (Lian et al., 2000).

Hamada and Al-Hakimi (2001) reported that soaking of wheat grain in

100ppm salicylic acid before sowing was generally effective in reducing the stress

effect of salinity and drought on leaf area. In potato, increasing concentration from

1-10mM acetyl salicylic acid significantly reduced total leaf area per plant (Haider

and Saffullah, 2001). SA applications (especially 0.5 mM) reduced adverse effects

of drought and increased plant height, leaf area index and protein yield in both

drought stress and normal conditions. Application of SA also reduced seed protein

content, results showed that exogenous application of this growth regulator under

59

water stress conditions can act as an effective tool in improving the growth and

production of common bean (Sadeghipour and Aghaei, 2012).

Singh and Usha (2003) and Sakhabutdinova et al. (2003) observed that

salicylic acid application increased the dry mass of wheat seedlings. Similarly,

though salicylic acid treatments have little effect on growth of barley seedling but

increased root and shoot fresh and dry weights (Metwally, 2003). Rice plants treated

with 100 ppm salicylic acid also showed greater biomass accumulation than

untreated plants (Maibangsa et al., 2000; Maibangsa et al., 2001). Sanna et al.

(2001) found that foliar spray of 2-5 mM of salicylic acid increased the fresh and

dry weight of Phaseolus oulgaris plants. Fariduddin et al. (2003) noted that when

sixty days old Brassica juncea plants sprayed with lowest used concentration

(105M) of salicylic acid possessed larger dry mass over the control. Agarwal et al.

(2005) demonstrated that reduction in biomass of seedling due to PEG induced water

stress is somewhat ameliorated by treatment with salicylic acid. An increase in fresh

and dry weights of root and shoot stressed and controlled plants were recorded with

the application of SA (El-Tayeb, 2005). An increase in the fresh weights of roots

and shoots and leaf area of the stressed maize plants treated with salicylic acid was

also obtained by Khodray (2004).

2.10 EFFECT OF SALICYLIC ACID ON PHYSIOLOGICAL ATTRIBUTES

Rao et al. (2012) suggested that foliar application of salicylic acid and

Ltryptophan can play a role to reduce the effect of drought in maize. Plants treated

with 100 ppm salicylic acid and 15 ppm L-tryptophan showed significantly higher

relative water content, chlorophyll and potassium content compared with other

treatments and control plants. Singh and Usha (2003) found that drought stressed

wheat seedlings showed a high moisture contents with salicylic acid than without

salicylic acid treatment. Exogenous application of 5 or 10 mg L-1 SA caused an

60

increase in stomatal conductance and transpiration rate in water stressed plants of

CV. S-24 whereas it was true for droughted plants of MH-97 only when 5 mg L -1

SA applied (Waseem et al., 2006).

Salicylic acid in combination with other phytohormones accelerated the rates

of photosynthesis and transpiration in soybean (Kumar et al., 2002). A similar

increase in photosynthetic rate seen at 42 hours after salicylic acid or acetyl salicylic

acid application at the rate of 10-3 or 10-5 Mol/L to corn and soybean plants under

stress, which was accompanied by increased or unchanged stomatal conductance

level and transpiration rate (Khan et al., 2003). Kabiri et al. (2014) examined the

effect of salicylic acid on black cumin (Nigella sativa) under drought stress in

hydroponic culture. Experimental treatments included salicylic acid at three levels

(0, 5, and 10μM) and drought stress (induced by PEG-6000) at four levels (0, –0.2,

–0.4, and –0.6 MPa). Salicylic acid could protect Nigella plant against drought stress

through increasing of photosynthetic pigments (chlorophyll a, b, total chlorophyll,

and carotenoids), relative water content, soluble sugar contents, and protein content,

and 10μM salicylic acid was the most effective level.

Salicylic acid also maintains almost the same photosynthetic rate and

stomatal conductance under water stress as those of water sufficient plants in the

wheat seedling. This shielding action of salicylic acid under water stress may be

associated with the reduction of transpiration rate and enhancement of

photosynthesis, which together enhanced water stress efficiency under drought

(Singh and Usha, 2003) salt stress( Poor et al., 2011). Al-Hakimi (2001) concluded

that treatment of wheat grains with sodium salicylate stimulated the growth via

enhancement of photosynthesis and transpiration rate.

Hamada and Al-Hakimi (2001) reported that soaking of wheat grain in 100

ppm salicylic acid was generally effective in reducing salinity and drought effect on

61

growth and transpiration rate and simultaneously increased the stimulatory effect of

drought on net photosynthesis. The exogenous SA application improved the growth

and photosynthetic rate in wheat (Hussein et al., 2007) under water stress. Salicylic

acid has also been observed to reverse the closure of stomata caused by abscisic acid

increased stomatal index and stomatal density on abaxial side, showing the opposite

on the adaxial side (Benavides-Mendoza et al., 2002).

2.11 EFFECT OF SALICYLIC ACID ON BIOCHEMICAL ATTRIBUTES

Salicylic acid is reported to induce more accumulation of praline contents in

NaCl and water stressed seedlings (Shakirova et al., 2003; Sakhabutdinova et al.,

2003; El-Tayeb, 2005). El-Tayeb and Naglaa (2010) observed an increase in proline

and protein and the results signify the role of SA in regulating the drought response

of plants suggested that SA could be used as a potential growth regulator, for

improving plant growth under water stress. Application of 100 ppm salicylic acid

enhanced the contents of total soluble proteins and grain proteins (Sivakumar et al.,

2001; Sivakumar et al., 2002). Sanna et al. (2001) showed that application of 5 mM

concentration of salicylic acid in Phaseolus vulgaris resulted in an increase of total

proteins in its leaves and fruits.

Sarangthem and Singh (2003) observed that under optimum dose of

treatment (0.1% V/V) salicylic acid, the levels of proteins in Phaseolus oulgaris

increased. Salicylic acid pre-treatment induces an increase in the contents of total

soluble proteins in roots and shoots while salinized plants showed significantly

decreased contents (El-Tayeb, 2005). It has also been observed that pre- treatment

of barley seedlings with salicylic acid before paraquat stress prevented the protein

loss (Popova et al., 2003).

62

Cag et al. (2009) reported that protein amount increased in all 0.001, 0.1,

and 10 μM concentrations except 1000 μM SA. Chlorophyll, carotenoid, protein

contents and POD activity increased in exogenic SA applications. It was also

documented that growth regulators, Salicylic acid (SA) and Putrescine (Put) were

highly effective on the canola in reducing the adverse effects of water stress.

Application of growth regulators under water stress protect canola plants by

maintaining the water budget, augmenting the accumulation of osmolyte, proline

and protecting photosynthetic pigments from its adverse effects (Ullah, 2012).

Maibangsa et al. (2000 and 2001) and Sivakumar et al. (2001 and 2002) showed

that plants treated with 100 ppm salicylic acid showed greater chlorophyll

accumulation.

2.12 EFFECT OF SALICYLIC ACID ON ANTIOXIDANT ACTIVITY

Salicylic acid (1mM) is the most effective concentration in increasing the

activities of superoxide dismutase (SOD), ascorbic peroxide (APOX) and catalase

(CAT) (Dat et al., 2000; Lindberg and Greger, 2002; Agarwal et al., 2005).

Salicylic acid enhanced the activities of antioxidant enzymes such as peroxidase

(POD), SOD and CAT, when sprayed exogenously to the drought stressed plants of

tomato (Hayat et al., 2008). Ananieva et al. (2004) reported that salicylic acid

treatment alone resulted in an increase in the activity of SOD, peroxidase and

catalase by 17, 25 and 20% respectively compared to the control plants. While

studying the changes in the activities of antioxidant enzymes in response to

treatment, it was noted that Phaseolus vulgaris treated with salicylic acid showed

elevated catalase and peroxidase activities while SOD activity remained the same in

water treated control (Clark et al., 2002).

63

Salicylic acid application increased peroxidase activity in different plant

species subjected to abiotic stresses (Kang and Saltvelt, 2002; Pal et al., 2005).

Singh and Usha (2003) recorded maximum SOD activity in wheat when sprayed

with 1 and 2 mM salicylic acid. Similarly, treatment of barely seedling with 500

µM salicylic acid caused an increase in SOD and catalase activities (Popova et al.,

2003). Contrastingly, salicylic acid treatment decreased the activities of catalase in

tomato (Senaratna et al., 2000).

In some studies, salicylic acid treatment did not cause any change in

superoxide dismutase activity, there was decrease in catalase activity and an increase

in peroxidase activity after one-day treatment of salicylic acid (Janda et al., 1999;

Kang et al., 2003). On the other hand, 0.25 mM salicylic acid application to the

shoot and soil enhanced SOD and catalase activity in Kentucky during heat stress

(He et al., 2005).

2.13 EFFECT OF SALICYLIC ACID ON GRAIN YIELD

Water stress both at vegetative and flowering stage badly affected the growth

and yield components of crop, but stress causes more damage at flowering stage.

Exogenous application of SA significantly ameliorated the negative effects of

moisture stress at both stages. (Buksh et al., 2009) observed that among different

treatments of exogenous application of SA, maximum number of achenes head -1

(986.11), achene yield (2601.00 kg ha-1), biological yield (10822.56 kg ha-1),

harvest index (25.43 %), and achene oil (40.72 %), and minimum achene protein

content (22.92 %) was recorded, when foliar application of SA (100 ppm) was done

at vegetative stage, but minimum values of above parameters were recorded at

flowering stage. Azimi et al. (2013) showed that water stress reduced all

characteristics of growth and yield, but amino acid and salicylic acid reduced

negative effect of water deficit on wheat.

64

Pre-sowing treatment of wheat seedling with salicylic acid was resulted in larger

ear size, higher mass of 100 seeds and higher grain yield (Shakirova et al.,

2003). De-Guang et al. (2001) noted that foliar application of 500mg/kg of acetyl

salicylic acid had obvious drought tolerance and yield increasing effects on maize.In

the same way, it was found that application of salicylic acid on the pearl millet plants

resulted in an increase of grain yield (Sivakumar et al., 2001,

Sivakumar et al., 2002).

Mainbangsa et al. (2000) demonstrated that salicylic acid treated rice plants

produced increase number of spikelets contributing to high yield. Zaghlool (2002)

described that the application of 20ppm salicylic acid (soaking+spraying) increased

yield in mung bean as indicated by pods per plant and 100 seed weight. Kumar et

al. (2002) observed that foliar application of salicylic acid alone and in combination

with any other phytohormones enhanced the grain yield of soybean. It was further

suggested that spraying 50 ppm salicylic acid increased the number of pods per

plant, number of seeds per pod, 100 seed weight, harvest index and yield of soybean

(Sharma and Kaur, 2003). It has been noted that 60 days old Brassica juncea plants

sprayed with 10-5 M salicylic acid increased number of pods and seed yield

enhanced by 13.7% and 8.4% respectively over the control (Fariduddin et al.,

2003).

Brinjal fruit yield was also increased with the application of 1000 ppm

salicylic acid (Karuppaiah et al., 2003). All treatments caused significant increase

in oil and protein % as those of the control. A marked decrease in total saturated

fatty acids accompanied by an increase in total unsaturated fatty acids was observed

in sunflower cultivars. Thus, SA treatments had a dogmatic effect on growth, seed

65

yield, total carbohydrate, phenolic content and the quality of the oil in favor of the

increase of unsaturated fatty acids of sunflower plant (Dawood et al.,

2012).

66

Chapter 3

MATERIALS AND METHODS

This study was conducted to assess the role of salicylic acid in alleviating

the water stress associated damages with autumn planted sunflower during 2009 and

2010 for increased productivity under water stress conditions.

3.1 EXPERIMENTAL SITE AND CONDITIONS

The study was conducted in the Stress Physiology Laboratory, National

Agriculture Research Centre (NARC) Islamabad, Pakistan. A series of experiments

were conducted under laboratory and greenhouse conditions. The laboratory

experiments were conducted in petri plates whereas the greenhouse experiment was

conducted in plastic pots of about 12 kg capacities.

Sandy clay soil was used for Greenhouse experiments. Soil composts used

has a composition of soil: sand: litter in a ratio of 2:1:1. Lab experiments were laid

out in completely randomized design in factorial arrangement with three

replications.

3.2 ACHENE MATERIAL

Six sunflower hybrids, Hyoleic-41, FH-352, NX-00989, Hysun-33, NX-

19102 and Parsun-2 were used for the present study. The achenes (seeds) of these

hybrids was obtained from the Oilseed Department, National Agriculture Research

Council (NARC),

67

35



ATTRIBUTES

The experiment was conducted under laboratory conditions (25±3 C) in order to

determine the response of different sunflower hybrids to water stress imposed

during germination and seedling stages. The morphological, biochemical and

physiological changes taking place in seedling stage by using salicylic acid under

water stress.

In this experiment, zero (control), -0.6, -1.33, and -1.62 MPa were developed

by dissolving 0,10, 15, and 20 g in PEG-8000 per 100mL Hoagland‘s solution, four

water stress levels of zero (control), 10% ,15% and 20% -MPa were applied to six

sunflower hybrids before germination using Polyethylene glycol (PEG-8000).

Sodium hypochlorite solution (10%) was used for five minutes to surface sterilize

the seeds and then washed three times with distilled water.Ten seeds of each six

sunflower hybrids (Hyoleic-41, FH-352, NX-00989, Hysun-33, NX-19012 and

Parsun-2) were sown in each petri plate containing filter papers. The experiment

was laid out in a completely randomized design with three replicates for each

experimental unit.

Ten millilitre of designated treatment solution was applied daily in each petri

plate after discarding out the previous solution. The number of seeds germinated

was counted daily and data was recorded for 14 days. A seed was considered

germinated when both plumule and radicle had emerged to 5 mm. Root and shoot

fresh and dry weights and their lengths were recorded after 14 days of the start of

experiment. Plant dry weights were recorded after drying at 70°C to a constant

68

weight. From these measurements the Promptness index (PI), Germination stress

tolerance index (GSI), Plant height stress tolerance index (PHSI), Root length stress

tolerance index (RLSl) and Dry matter stress tolerance index (DMS1) were

calculated using the following formulae given by Ashraf et al. (2006).

i.

Where, n is the number of seeds germinated on day d



In this experiment three water stress levels of zero (control), 10%, 20% -

MPa (PEG) were applied to three sunflower hybrids (selected from the previous

experiment) (NX-19012, NX-98900 (tolerant) and FH-352 (sensitive) after

germination. Ten seeds of each sunflower hybrid were sown in each petri plate

containing filter paper. The Hoagland‘s solution (10 mL) was added to these petri

plates and replaced daily after washing out the previous solution. The experiment

was laid out in a completely randomized design in factorial arrangement with three

replications. Seeds were allowed to germinate in 72 hours in the dark, then exposed

to light.

69

It is well documented now that exogenously applied compatible solutes are

effective in mitigating the bad effects caused by stress on plant growth depends on

concentration, stage of development at which applied, and mode of application

(Agboma et al., 1997; Ashraf and Foolad, 2007). Thus, the major objective to carry

out this experiment was to study the effect of exogenous application of SA during

different modes at the seedling stage of sunflower in counteracting the adverse

effects of water stress on sunflower differing in drought tolerance. This laboratory

experiment was further divided into two sub-experiments:

i) Effect of foliar spray of SA on sunflower hybrids at the seedling stage ii)

Effect of seed soaking in SA of sunflower hybrids at the seedling stage

These two sub-experiments were performed in laboratory conditions viz.,

10/14 light/dark period at 500-800 μmol m-2 s-1 PPFD, a temperature cycle of

29/15°C day/night and relative humidity of 65 ± 5%. Seeds of sunflower hybrids

were surface sterilized before experimentation in 5% sodium hypochlorite solution

for 5 minutes. Ten achenes of each hybrid were germinated for one week in each

petri plate, on moistened filter paper with distilled water. Seven days old sunflower

seedlings of same size of each hybrid were transplanted in plastic boxes having holes

in their covers containing 2 L of half strength Hoagland‘s solution. Sunflower

seedlings were further grown for another one week in hydroponics.

3.4.1 Effect of Foliar Spray of SA on Sunflower Hybrids at Seedling Stage

Two-week old plants were subjected to water stress levels 0, 10% and 20%

PEG (8000) in Hoagland‘s nutrient media. Sunflower plants were then sprayed with

different levels of SA concentrations (0, 0.375, 0.75 and 1.5 mM in 0.1% Tween-

20) ~5 ml per plant. The control (DS-0 + SA-0) plants were also sprayed with 0.1%

Tween-20 solution. For the maximum penetration and to avoid leaf injury pH of the

70

solution should be at 6.5. All the boxes were continuously aerated. At the end of the

experiment, plants were harvested, washed with distilled water, dry with blotting

paper and separated into shoots and roots, and fresh biomass were recorded. These

plants were then dried at 65oC for 72 h in an oven and dry biomass recorded. After

harvest, data for the shoot or root length were also recorded. However, relative water

content (RWC) water, osmotic and turgor potentials, leaf proline, protein, soluble

sugars and free amino acids and gas exchange parameters were measured, before

harvest.

3.4.2 Effect of Seed Soaking in SA of Sunflower Hybrids at Seedling Stage

Plant growth conditions were same for this experiment as in the experiment

(SA applied as a foliar spray), except that the sunflower seeds were pre-treated in

different SA concentrations (0, 0.375, 0.75 and1.5mM) for about 10 hours.. In this

sub-experiment, the sunflower plants were subjected to water stress 0, 10% and

20% PEG in Hoagland‘s nutrient solution after two weeks of germination. All the

treatment boxes were continuously aerated. After treatment, plants were harvested

when wilted, washed with distilled water, blotted dry and separated into shoots and

roots, as in the previous experiment, and fresh biomass were recorded. These plants

were then dried at 65 ºC for 72 h in an oven and dry biomass was recorded. After

harvest, data for the shoot or root length were also recorded. However, relative

water content (RWC) osmotic, water and turgor potentials, leaf proline, protein,

sugars and amino acid and gas exchange parameters were measured before harvest.

3.5 INFLUENCE OF FOLIAR APPLICATION OF SALICYLIC ACID WITH

DIFFERENT CONCENTRATIONS AT TWO GROWTH

71

STAGES OF SUNFLOWER HYBRIDS UNDER DROUGHT STRESS

3.5.1 Seed Sowing

For this greenhouse experiment, soil was initially sun dried, sieved and

mixed with IJ in order to avoid any plant residues. For greenhouse experiment 12

kg of this soil was filled carefully in each pot. In each pot five seeds were sown and

then watered with tap water. At the beginning all pots were kept at the field capacity

level for obtaining good germination and emergence. Later on the water was applied

according to the water stress level specified for the experiment. Before imposing

water stress the plants were thinned out and three healthy plants were kept in each

pot. Recommended amounts of P and K and half of N were applied in solution form

at the time of planting and the remaining half of the N was applied after 30 days of

planting. The plants were sprayed with insecticides to control insect attack.

3.5.2 Development and Maintenance of Water Stress Levels

This greenhouse experiment was performed to evaluate the genotypic

response of three sunflower hybrids towards four concentrations (0, 0.375, 0.75 and

1.5 mM) of salicylic acid at vegetative and flowering stages under water stress

conditions. Before experimentation, seeds of each sunflower hybrid were surface

sterilized for 5 minutes in 5% sodium hypochlorite solution. Seeds of each hybrid

were sown in plastic pots; each pot was filled with 12 kg thoroughly mixed sandy

clay soil. Plastic pots have holes for drainage covered with a muslin cloth piece at

the bottom. Five seeds were initially sown in pots and after one week of germination

thinned to three plants per pot of almost the same size and equidistantly placed. Each

pot was irrigated with 2 L of tap water. To one set of pots, water stress was given at

vegetative stage by withholding water till wilting and sprayed with varying

concentrations of SA (0, 0.375, 0.75 and 1.5mM in 0.1% solution of Tween20) to

plants. The control plants were irrigated normally and sprayed with 0.1%

72

Tween-20 solution.

The pH of the sprayed solution should be 6.5. As the signs of wilting

appeared the physiological, biochemical and gas exchange parameters were

recorded and rewatered, morphological and yield related parameters were recorded

after harvest. To second set of pots, water was withheld at flowering stage till wilting

and sprayed with varying concentrations of SA (0, 0.375, 0.75 and 1.5mM in 0.1%

solution of Tween-20) applied as a foliar spray to plants. The control plants were

irrigated normally and sprayed with 0.1% Tween-20 solution. The data were

measured for physiological, biochemical and gas exchange parameters and irrigated

normally till harvest, morphological and yield related parameters were recorded

after harvesting. Both sets of plants were harvested 90 DAS, heads were removed

and sun dried. Plants were washed with distilled water, dry and separated into shoots

and roots, and data for fresh biomass recorded. These plants were then dried at 65oC

for 72 h in an oven and dry biomass recorded.

3.6 PROCEDURES FOR DATA COLLECTION

3.6.1 Plant Growth and Development Attributes

3.6.1.1 Root fresh and dry weight

The plants were harvested, washed with distilled water, blotted dry and

separated into shoots and roots, and data for fresh biomass were recorded.

3.6.1.2 Shoot fresh and dry weight

The shoot and root of plants were then oven-dried at 65oC for 72 h and dry

biomass was recorded.

73

3.6.1.3 Plant height

Five plants were selected at random. Their heights were measured in cm with

measuring tape and then averaged

3.6.1.4 Leaf count

Total number of leaves per plant was counted by randomly selecting three

plants.

3.6.1.5 Leaf area

Leaf area meter (DT Area Meter, model MK2) was used for measurement.

Total leaf area was measured by randomly selecting three plants.

3.6.2 Physiological Parameters

3.6.2.1 Plant Water Relations

Parameters of plant water relations, including osmotic potential, water

potential, turgor potential and leaf water contents were taken after twenty days of

the water stress and foliar SA application, at both stages.

3.6.2.1.1 Leaf water potential

The fully expanded third leaf from the top of three plants of each treatment was

used to record the leaf water potential. The measurements were made in the morning

from 8.00 to 10.00 A.M. by using Scholander pressure chamber according to the

technique described by Scholander et al. (1965).

3.6.2.1.2 Leaf osmotic potential

The same leaf was used for osmotic potential determination which was used

for measurement of water potential. The leaf was frozen in 2.0 cm polypropylene

74

tubes for two weeks and then thawed, the sap was extracted by pressing it with a

glass rod. The sap so extracted was used directly for osmotic potential determination

in an osmometer, (Wescor, Model 5520, USA).

3.6.2.1.3 Leaf Turgor potential

Turgor potential was calculated as the difference between water potential

(Ψw) and osmotic potential (Ψs) values (Nobel, 1991).

(Ψp) = (Ψw) - (Ψs)

3.6.2.1.4 Leaf relative water contents

The third fully expanded leaf from the top was used. After excising at the base of

the lamina, fresh weight (FW) was determined. Then turgid weight (TW) was

obtained after placing leaves in distilled water for 16-18 h at room temperature for

re-hydration. Leaf turgid weight was measured after drying with tissue paper. Dry

weight (DW) was determined after oven drying the leaf samples at 70oC for 72

h. LRWC was calculated from Schonfeld et al., 1988 formula.

Where,

FW = fresh weight of leaf,

DW = dry weight of leaf,

TW = turgid weight of leaf.

3.6.2.2 Gas exchange parameters

Gas exchange parameters were observed using an open system LCA-4 ADC

portable infrared gas analyzer (IRGA). The measurements of photosynthesis (A) and

75

stomatal conductance (gs), were made for the youngest fully emerged leaf of each

plant. Leaf diffusive resistance was measured by porometer.

3.6.3 Biochemical Parameters

3.6.3.1 Proline content

The free proline was estimated according to the method of Bates et al.

(1973). Leaf material (1.0 g) was homogenized in 10 ml of 3% sulfosalicylic acid.

The homogenate was filtered through Whatman No. 2 filter paper. One ml of filtrate

was added in 1 ml of glacial acetic acid and 1 ml of ninhydrin solution (ninhydrin

solution was prepared by dissolving 1.25 g ninhydrin in 20 ml acetic acid and 20 ml

6M orthophosphoric acid). The filtrate was incubated at 100o C for 1 h and cooled

it in an ice bath afterwards. 4 ml of toluene was added and the mixture was vortexed

for 5 min. The chromophore containing toluene was aspirated from the aqueous

phase, warmed at room temperature and the absorbance was read at 520 nm using

toluene as a blank. The proline concentration was determined from a standard curve

and calculated on fresh weight basis as follows:

Proline (µmolg-1 fresh weight) = (µg proline mL-1 x mL of toluene/115.5) / (g of sample /10)

3.6.3.2 Total soluble sugars

Soluble sugars were estimated following Dubois (1951). 0.5g fresh leaf

material was taken in test tubes containing 10ml of 80% ethanol and heated at 80C

for one hour in water bath. 0.5 ml of this extract was taken in another set of test

tubes, then 1ml of 18% phenol was added and left for incubation for one hour at

room temperature followed by addition of 2.5ml of sulphuric acid, shaked and

absorbance was read at 490 nm.

76

3.6.3.3 Total soluble protein:

Fresh leaves (0.2 g) were ground in 4 mL of 50 mM cooled phosphate buffer

(pH 7.8). The homogenate was centrifuged at 6000 × g for 10 min at 4°C.

Protein concentration of the extract was measured as described by Bradford (1976).

3.6.3.4 Free amino acids

Free amino acids were determined according to Hamilton and Van Slyke

(1973). Fresh plant leaves (0.5 g) were cut into pieces and extracted with phosphate

buffer (0.2 M) having pH 7.0. 1 mL of the extract was poured in 25 mL test tube,

then 1 mL of pyridine (10 %.) and ImL of ninhydrin (2 %) solution was added in

each test tube. The test tubes with a sample mixture were placed in boiling water

bath for about 30 minutes; volume of each test tube was made up to 50 mL with

distilled water. Read the optical density of the colored solution at 570 nm using a

spectrophotometer. Developed a standard curve with Lueicine and determine free

amino acids by the formulae given below:

3.6.3.5 Chlorophyll contents

Leaf chlorophyll content was determined spectrophotometrically (Hiscox and

Israelstam 1979). Chlorophyll extraction was performed by using dimethyl

sulphoxide (DMSO). 0.05grams of leaf tissue in fractions was placed in a vial

containing 7 mL DMSO (B.D.H. Chemicals Ltd., Toronto) and chlorophyll were

extracted into the fluid without grinding at 65°C for 40 min in a water bath. After

that tubes were taken out and vortexed.The extract liquid was transferred to a

77

graduated tube and made up to a total volume of 10 mL with DMSO, assayed

immediately or transferred to vials and stored between 0-4°C until required for

analysis

Absorbance of the supernatant was read at 645 and 663 nm using a

spectrophotometer.The chlorophylls ‗a‘ and ‗b‘ were calculated by the following

formulae:

Chl. a = [12.7 (OD 663) -2.69 (OD 645)] Chl.

b = [22.9 (OD 645) -4.68 (OD 663)]

3.6.4 Antioxidant Enzymes

For the extraction of antioxidant enzymes, fresh leaves (0.5 g) were ground

using cooled pestle and mortar in 5 mL of 50 mM cooled phosphate buffer (pH

7.8). After filtration through cheese cloth, the homogenate was centrifuged at

15000 g for 20 min at 4°C and the supernatant was used for enzymes assays.

3.6.4.1 Superoxide dismutase (SOD)

The activity of SOD was assayed following the method of Giannopolitis and

Ries (1977) by monitoring the inhibition of photochemical reduction of nitroblue

tetrazolium (NBT) at 560 nm. The activity of SOD was determined by adding 50 μL

of the enzymatic extract to a solution containing (total reaction solution including

enzyme extract 3 mL) 50uM NBT (NBT dissolved in ethanol),

1.3 μM riboflavin, 13 mM methionine, 75 mM EDTA, 50 mM phosphate buffer

(pH 7.8), and 20 to 50 μl enzyme extract. The reaction solutions were kept in a

chamber under illumination of fluorescent lamps of 30 W.

78

The reaction was started by turning the fluorescent lamps on, and stopped 5

min later by turning them off. The blue formazane produced by NBT photo

reduction was measured as increase in absorbance at 560 nm. The reaction mixture

lacking leaf extract was taken as control and kept in light. However, blank solution

having the same complete reaction mixture (including enzyme extract) was kept in

the dark. The absorbance of the irradiated solution at 560 nm was read using a

UVvisible spectrophotometer (IRMECO U2020). One unit of SOD was defined as

the amount of enzyme required to cause 50% inhibition of the rate of NBT reduction

at

560 nm in comparison with tubes lacking the plant extract.

3.6.4.2 Catalase (CAT) and Peroxidase (POD)

Activities of CAT and peroxidase (POD) were assayed following Chance

and Maehly (1955) with some modification. The final volume of the reaction

mixture for CAT (3 mL) contained 50 mM phosphate buffer (pH 7.0), 5.9 mM H2O2,

and 0.1 mL enzyme extract. The reaction was initiated by adding 100 μL of the leaf

crude extract (enzyme extract) to the reaction mixture. Changes in absorbance of the

reaction solution due to decomposition of H2O2 at 240 nm were read every 20 s.

CAT activity was expressed as units (μmol of H2O2decomposed per min) per mg of

protein. One unit, CAT activity was defined as an absorbance change of 0.01 units

per min.

The activity of POD was determined by guaiacol oxidation method. The final

volume of the reaction mixture for POD (3 mL) contained 50 mM phosphate buffer

(pH 7.0), 20 mM guaiacol, 40 mM H2O2, and 0.1 mL enzyme extract.

79

Changes in absorbance of the reaction solution at 470 nm were determined every 20

s. One unit POD activity was defined as the change of 0.01 absorbance unit per min

per mg of protein.

3.6.5 Determination Endogenous Level of Salicylic Acid

Salicylic acid was extracted and purified following the procedures of

Enyedi et al. (1992) and Sarkar et al. (1998) with some modifications (Iqbal and

Ashraf, 2006). Fresh leaves already frozen and stored in 30 ml of 80% cold (-70o C)

aqueous methanol MeOH (4:1 v/v) supplemented with 20 mg L-1 butylated

hydroxytoluene (BHT) were ground in a mortar using aqueous 80% MeOH-BHT as

an extracting solvent. To check recoveries during extraction and purification, 300

ng naphthalence acetic acid NAA was added as standard before homogenization.

The homogenate was vortexed for 10 min filtered with suction through Whatman

no 42 filter paper. The residues in the flask and on the filter paper were rinsed three

times with 10 ml aliquots of MeOH-BHT and two times with 100% MeOH. The

extracts were combined mixed and used for SA

purification. The extract was concentrated to an aqueous residue by rotary flask

evaporation RFE at 40oC.

Sublimation of SA was prevented by the addition of 0.2 M sodium

hydroxide (Verberne et al., 2002). The aqueous fraction was transferred to 50 ml

polypropylene centrifuge tubes. The flask used for RFE was rinsed with 10 ml of

nhexane (pH-8) and then with n-hexane (pH-3) for SA clean up. The pH was then

adjusted to 2.8 and the samples were centrifuged for 15 minutes at 13000 g to

remove any precipitate. The supernatant fraction was decanted in to clean centrifuge

13000 g to remove any precipitate. The supernatant fraction was decanted in to the

clean centrifuge tube and partitioned thrice against 10 ml portions of 1ml

80

MHCH.The same 10 ml of 1m MHCH used for all the three 10 ml portions of cold

diethyl ether-BHT. All the three ether fractions were combined and evaporated by

RFE to dryness in vacuo. The residue as immediately dissolved in 500 ul of 80%

ice cold MeOH-BHT in a 1.5 ml eppendorf centrifuge tubes. These samples were

kept overnight in -70oC and then centrifuged (25000g) for 10 min at 10oC. Standards

were also prepared following the sample procedure. The supernatant was filtered

and subjected to HPLC (High performance liquid chromatography) analysis.

Analysis of SA was performed by HPLC Agilent Technologies USA

equipped with S-1121 dual piston solvent delivery system and S-3210 UV/VIS

diode array detector. The elution system consisted of 100 % methanol: 1% acetic

acid (52:48) v/v) as solvent; run isocratically with a flow rate of 1.10 ml min-1 twenty

microlitres of filtered extracts were injected in to Hypersil ODS reverse- phase (C-

18) column (4.6x250 mm. 5 um particale size: thermo hypersol GmbH, USA) fitted

with a C- 18 guard column. Detection of SA was performed at 280 nm by

choromatography with 2-hydroxbenzoic acid as standard.

3.6.6 Quality Parameters

3.6.6.1 Achene protein contents

Protein contents in the seeds were estimated according to Kjeldahl method

(Bremner, 1964). One gram of each sample was put in the Kjeldahl flask, a digestion

tablet was added to 5 ml of concentrated H2SO4 and the contents thoroughly mixed.

The flask was shifted on the digestion assembly and the heater and the exhaust fan

were turned on. The digestion was continued with occasional shaking of the flask.

When all the organic matter had been oxidized and the solution became clear, the

digestion was continued for another 30 minutes.

81

Cooled the digest and transferred it to a 100 ml volumetric flask and made

up the volume to 100 ml by washing the digest and mixed well. Pipette out 5 ml of

the digest and poured into a Markam Still Apparatus. 10 ml of NaOH (4% w/w) was

added slowly through the funnel stopper (did not remove the stopper, otherwise

ammonia may escape). Plugged the funnel and added distilled water a few ml. After

5 minute distillation, collected the droppings from the condenser for one minute in

a conical flask containing 5 ml of 2% boric acid. Washed the tip of the condenser

and titrated against standardized H2SO4. Percent crude protein was calculated using

the following formula:

Where V1 = Sample titration (in ml)

V2 = Blank titration (in ml)

N = Normality of standardized H2SO4

W = Sample weight (in g)

3.6.6.2 Achene oil content

Achene oil content was determined by the Soxhlet Fat Extraction method

(AOAC, 1990). Seeds were dried at 105oC in an oven for about 8 h. Seeds were

weighed before and after drying to measure moisture content. For analysis two gram

achenes per thimble were ground in a coffee mill. Thimbles were weighed and then

added ground seeds to record final weight. Afterwards, the thimbles with seeds were

placed in extractors. Solvent (petroleum ether) was added to six dry and clean round

bottom flasks (250 mL) already weighed, and then were connected to the extractors

and positioned on heating mantles, connected with condensers.

Flasks were heated and continued extraction for at least 6 hours. Then

extraction was stopped, thimbles were removed and then reheated the flasks, to

82

collect all the solvent in the Soxhlet extractors, the apparatus was allowed to cool

and flasks were dried at 105o C for 1 hour. After cooling, the flasks and oil were

weighed together. Percent oil content was calculated using the following equation.

3.6.6.3 Preparation of fatty acid methyl esters (fames) by official method

The IUPAC standard method was used for the preparation of FAMEs.

Fatty acid composition

Fatty acid methyl esters were analyzed by gas chromatography model Clarus

500 fitted with a cynopropyle polysiloxane polar capillary column RT- 2340 NB

(60m x 0.25mm I.D) with 0.20 μm film thickness and flame ionization detector.

Oxygen free nitrogen was used as a carrier gas at a flow rate of 67.4 Psi. Oven

temperature, 70oC with 2 min. hold; ramp rate, 3oC/min; final temperature,

190oC; injector temperature 200oC; detector temperature, 220oC. Fatty acid methyl

esters were identified by comparing their relative and absolute retention times to

those of authentic standards of fatty acid methyl esters. All the quantification was

done by using a CSW32 software program provided by SUPERMICR.

3.6.7 Yield and Yield Related Traits

3.6.7.1 Head diameter

The diameter was measured in cm with the help of a measuring tape and then

averaged of three randomly selected heads.

83

3.6.7.2 Number of achenes per head

Number of achenes were counted and then averaged in three randomly

selected heads from each treatment.

3.6.7.3 1000-achene weight

Three samples, each of 1000-achenes were randomly taken from the seed lot

of each to calculate the average weight of 1000-achenes.

3.6.7.4 Achene yield

After harvesting the plants, the heads were separated, sun dried, threshed

manually and the achene yield per head was recorded. The random achene samples

were taken from each treatment to determine the moisture contents. The achene

yield was adjusted to 10% moisture content in g head-1.

3.6.7.5 Oil yield

In order to calculate oil yield, the oil contents were determined by the

Soxhlet Fat Extraction method (AOAC, 1990) by taking random samples from each

treatment. After that, the achene yield was converted to oil yield.

3.6.7.6 Biological yield

Recorded the weight of air-dried plant (except achenes), then add it to

already calculated achene yield (g head-1) to find out biological yield.

3.7 STATISTICAL ANALYSIS

Data regarding all the plant parameters were collected using standard

procedures, and were statistically analyzed by using Statistix 8.1 software through

84

analysis of variance technique. The LSD test at 5% probability was used to compare

the differences among treatments, means (Steel et al., 1997).

Chapter 4

RESULTS AND DISCUSSIONS

This study includes four separate experiments on the effects of drought stress

on various plant growth and biochemical aspects of different sunflower hybrids, and

amelioration of drought stress by salicylic acid. Results indicated a negative impact

of various drought stress levels on both plant growth and biochemical parameters

differently. Also, treatment of stressed plants with salicylic acid helped them to

overcome the influence of drought stress to different degrees depending on the level

of stress and type of sunflower hybrid. Detailed description of the results from each

experiment is presented in the following paragraphs:


DROUGHT STRESS LEVELS FOR THEIR GROWTH ATTRIBUTES

This experiment was conducted to screen six sunflower hybrids in different

levels of drought stress produced through polyethylene glycol (PEG 8000) in

distilled water at 10, 15 and 20 % concentrations designated as DS-10, DS-15 and

DS-20, respectively. The action of different drought stress levels on various growth

parameters of sunflower hybrids are described separately in the followings:

4.1.1 Seed Germination

Effect of drought stress on the germination of sunflower hybrid seeds was

measured in terms of seed germination stress index (SGSI). Results indicated that

increased levels of drought stress (DS) decreased the SGSI significantly being the

86

55

lowest at DS-20 (Figure1.1). Various sunflower hybrids showed significant

difference for SGSI from each other; among them NX-19012 exhibited the highest

SGSI (80.3) and FH-352 had the lowest SGSI value of 39.7. While evaluating the

overall stress tolerance of all the hybrids at the highest stress level of DS-20, it was

found that hybrids NX-00989 and NX-19012 gave the best SGSI with non

significant difference with each other but differing significantly with rest of the

hybrids among which FH-352 performed the lowest. Interaction between stress

levels and sunflower hybrids was also statistically significant. Sunflower hybrid NX-

19012 at DS-10 gave the highest value of the seed germination stress index, whereas,

the lowest value was that of FH-352 under DS-20.

Although, earlier studies indicate the variable genetic response of various

crop cultivars to drought stress; however, there is no comprehensive work on the

sunflower hybrids compared in the present study. These hybrids were pre-screened

among from 100 sunflower hybrids / lines for drought tolerance in the germination

test (data not presented), where four hybrids (NX-19012, NX-00989, Hysun-33 and

Parsun-2) showed the highest drought tolerance and two hybrids (Hyoleic-41 and

FH-352) gave the lowest drought tolerance. Adverse effects of water deficit on

germination and seedling growth of sunflower were also reported by Mohammad et

al. (2002).

Seed germination is the most sensitive stage in the plant life cycle (Ashraf

and Mehmood, 1990) and unfavorable environmental conditions, e.g., water stress

could have a negative impact on the seed germination (Albuquerque and Carvalho,

87

Figure 1.1: Effect of different drought stress levels on seed germination stress index

(SGSI) of various sunflower hybrids

2003; Shao et al., 2008; Kusaka et al., 2005). According to Ahmad et al. (2009),

drought stress has an inhibitory effect on sunflower seed germination. Increasing

drought stress levels result in reduction of cell division and plant growth metabolism

which caused delay in seedling emergence. Sunflower is susceptible to water

shortage stress at the germination stage. Proper water level and osmotic pressure is

required during seed germination and for seedling establishment; nonetheless, due

to slight genetic difference, cultivars / hybrids of the same crop differ in drought

tolerance (Lenzi et al., 1995). Drought impairs seed germination and causes poor

stand; as it is the first and foremost effect of stress in crop establishment (Harris et

al., 2002). Sunflower seeds have been shown to decrease in percent germination with

increasing osmotic stress (Sajjan et al., 1999) while increase in germination time

with water deficit (El-Midaoui et al., 2001).

88

Moreover, Saensee et al. (2012) tested seven sunflower genotypes and one

commercial hybrid, Pacific 77, at two water stress levels of -0.6 and -1.2 MPa, using

PEG-6000.Dry matter stress index, plant height stress index, root length stress index,

relative water content stress index and germination stress index were significantly

decreased in all sunflower genotypes with increase in water stress levels.The results

of these previous findings are similar with that of present study.

4.1.2 Plant Height

Impact of drought stress on the plant growth of sunflower hybrid was

determined in terms of plant height stress index (PHSI) as shown in Figure1.2. At

higher levels of drought stress the PHSI was reduced significantly with the lowest

Figure 1.2: Effect of different drought stress levels on plant height stress index

(PHSI) of various sunflower hybrids

values at DS-20. Among the tested sunflower hybrids Hysun-33 followed non

significantly by NX-19012 gave the highest PHSI values with significant difference

from other hybrids; whereas, FH-352 exhibited the lowest PHSI value of 48.5. As

0.0

20.0

40.0

60.0

80.0

100.0

DS-10 DS-15 DS-20

Drought stress levels (% PEG)

Hyoleic-41

FH-352

NX-00989

Hysun-33

NX-19012

Parsun-2

89

far as evaluation of the hybrids at the highest stress level of DS-20 is concerned, four

hybrids NX-00989, Hysun-33, NX-19012 and Parsun-2 showed better tolerance in

terms of PHSI having non significant difference among themselves but significant

difference with other hybrids particularly FH-352 almost vanished after germination.

Interactions between stress levels and sunflower hybrids also showed a significant

difference, however, all the sunflower hybrids exhibited non significant difference

at DS-10 for the plant height stress index, whereas, under DS-20 the lowest value

(0.00) was that of FH-352 followed by

Hyoleic-41 (38.3).

Plants respond to drought stress and acclimatize through various

physiological and biochemical changes (Farooq et al., 2009).Water stress on

sunflower has been reported to reduce plant height and number of stomata

(PirjolSovulescu et al., 1974). Different cultivars of sunflower vary in their water

requirement, and drought stress impairs the seedling growth causing reduced PHSI

(Ahmad et al., 2009). Drought stress decreased the plant height and plant dry matter

(Nizami et al., 2008).

4.1.3 Root Length

Figure1.3 exhibits root length stress index (RLSI) of sunflower hybrids as

affected by drought stress. Significant reduction of RLSI was recorded at higher

90

Figure 1.3: Effect of different drought stress levels on root length stress index

(RLSI) of various sunflower hybrids

levels of drought stress with the lowest values at DS-20. The NX-00989 among from

other sunflower hybrids gave the highest RLSI value with significant difference from

the next highest value by NX-19012 and other hybrids. Whereas, Hyoleic-41 showed

the lowest RLSI. While evaluating the hybrids at the highest stress level of DS-20,

it was observed that NX-19012 got the best RLSI value followed by NX-00989 with

significant difference with each other and rest of the four hybrids. They showed

better tolerance in terms of RLSI, while two hybrids viz., FH-352 and Hyoleic-41did

not exist after germination. Interactions among stress levels and sunflower hybrids

also showed significant difference for the root length stress index being significantly

different at each stress level for almost all hybrids.

The results of present research are in complete agreement with that found by

Zeid and Shedeed (2006) that in alfalfa (Medicago sativa), germination potential was

decreased by polyethylene glycol-induced drought stress, while the root length was

increased. Water deficit stress, enhances the allocation of dry matter to the roots,

which causes increased water uptake by plants (Leport et al., 2006). Ahmad et al.

91

(2009) also reported that RLSI of sunflower hybrids increased with higher levels of

PEG induced drought stress. Moreover, Saensee et al. (2012) tested seven sunflower

genotypes and one commercial hybrid, Pacific 77, at two water stress levels of -0.6

and -1.2 MPa, using PEG-6000. Dry matter stress index, plant height stress index,

root length stress index, relative water content stress index and the germination stress

index were significantly decreased in all sunflower genotypes with increase in water

stress levels. Water stress caused a visible increase in root length and number of

adventitious roots, but a considerable reduction in fresh and dry masses of root,

growth vigor of shoot and leaf area

(Heshmat et al., 2012).

4.1.4 Plant Dry Matter

Data trend in Figure 1.4 indicates that higher concentrations of PEG (DS20)

significantly reduced the dry matter biomass weight of sunflower plants as shown by

the dry matter stress index (DMSI). Similarly, various sunflower hybrids differed

significantly for their tolerance to PEG-induced stress; NX-00989 exhibited the

highest DMSI value, followed by NX-19012 with statistical difference between the

both. The lowest DMSI value was recorded for FH-352 hybrid. Interaction between

stress levels and various sunflower hybrids was also significant. Water deficit stress

tolerance as checked by DMSI was greater in NX00989 at all levels of water stress

created by PEG-8000. The lowest values at all PEG concentrations were recorded

from FH-352 hybrid and they differed

significantly from all other treatments.

Drought, the major abiotic stress, affects every aspect of plant growth and is

mainly responsible for limiting crop production worldwide (Harris et al.,2002;

Golbashy et al., 2010). A general negative effect of water stress on crop plants is

92

, the reduction in

fresh and dry biomass production (Ashraf and O Leary 1996). Six sunflower hybrids

tested for stress tolerance in this study showed that the most tolerant (DMSI 70-77)

were Hysun-33, NX-19012 and NX-00989 in the ascending order, while the least

drought tolerant (DMSI 38-54) were FH-352 and Hyoleic-41.

Figure 1.4: Effect of different drought stress levels on dry matter stress index

(DMSI) of various sunflower hybrids

Hysun-33 hybrid exhibited medium tolerance to water stress in another study by

Ahmad et al. (2009). They also indicated that variation among hybrids for DMSI

was a reliable indicator of drought tolerance in sunflower. Saensee et al. (2012)

tested seven sunflower genotypes and one commercial hybrid, Pacific 77, at two

water stress levels of -0.6 and -1.2 MPa, PEG-6000. Dry matter stress index was

significantly decreased in all sunflower genotypes with increase in water stress

levels. Water stress caused a considerable reduction in fresh and dry masses of root

and shoot (Heshmat et al., 2012). Sayyari et al. (2013) studied the effect of salicylic

acid (SA) on lettuce (Lactuca sativa L.) under drought stress. They reported that

drought stress imposed negative effects on plant growth and productivity by

93

reducing the fresh and dry weight. Reduced biomass were recorded in several other

plant species, including soybean (Specht et al., 2001) maize and sunflower (Vanaja

et al., 2011).



This experiment was undertaken in the laboratory conditions using three

sunflower hybrids (NX-19012, NX-00989 and FH-352). Drought stress (DS) was

applied with PEG at three levels (0, 10 and 20 %) and treated to rehabilitate their

growth by applying four levels (0, 0.375, 0.75 and 1.50 mM) of salicylic acid (SA)

through either seed soaking or foliar spray on the seedlings. Results on various

growth, physiological, water relations and biochemical parameters of sunflower

seedlings are presented and discussed in the following paragraphs:

4.2.1 Plant Height

Under the first set of experiment with foliar application of salicylic acid, the

plant heights of three hybrids of sunflower were statistically similar to each other

(Table 2.1.1). Various concentrations of salicylic acid also did not have any

significant effect on plant height of sunflower. However, different levels of drought

stress (% PEG) caused a significant reduction in plant height, being the lowest with

DS-20 (20% PEG). The combined application of various drought stress levels and

salicylic acid doses had a significant interactive effect. The tallest plants (24.1 cm

height) were found under DS-0 (control) receiving 0.75 mM dose of salicylic acid,

while shortest plant height (16.9 cm) was recorded in treatment receiving 20% PEG

along with no SA concentration. Interaction among hybrids × drought levels ×

salicylic acid concentrations was not statistically significant. However, combination

of sunflower hybrids × DS levels had significant interaction with maximum plant

94

height (23.0 cm) of tolerant H-2 (NX-00989) genotype in control, while the smallest

shoot length (17.0 cm) was recorded for the sensitive H-3 (FH-

352) at DS-20.

Results of plant height under the second set of experiment on seed soaking

of sunflower hybrids with salicylic acid at different DS levels are shown in Table

2.1.2. Here also, the sunflower hybrids did not differ significantly; however, the SA-

0.75 concentration produced significantly longest plants (22.0 cm) as compared to

other SA levels. Different levels of drought stress also caused a significant reduction

in plant height, as the smallest height was with DS-20 (17.8). Interaction between

various levels of

Table 2.1.1: Plant height (cm) of sunflower hybrids under different drought

stress levels as influenced by foliar application of salicylic acid.

Treatments Salicylic acid (SA) levels (mM) Means

SA-0 SA-0.375 SA-0.75 SA-1.50

Hybrids (H) Interaction (H × SA) Means (H)

H-1 (NX-19012) 21.0NS 20.6 21.9 19.4 20.7 NS

H-2 (NX-00989) 20.4 19.1 21.7 19.6 20.2

H-3 (FH-352) 19.4 19.1 20.9 19.8 19.8

Drought stress (DS) Interaction (DS × SA) Means (DS)

DS-0 (Control) 22.3 abc 22.4 abc 24.1 a 22.1 a-d 23.0 A

DS-10 (10% PEG) 19.3 b-e 19.6 b-e 20.9 b-d 19.8 b-e 20.1 B

DS-20 (20% PEG) 16.9 e 17.0 e 18.6 de 17.9de 17.6 C

Hybrids × Drought Interaction (H × DS × SA) Means (H×DS)

H-1 × DS-0 23.0NS 22.8 23.7 20.9 22.6 AB

H-1 × DS-10 20.1 20.4 22.8 19.6 20.7 ABC

H-1 × DS-20 19.8 18.6 19.3 17.7 18.9 CD

H-2 × DS-0 23.4 21.7 24.2 22.8 23.0 A

95

H-2 × DS-10 20.1 19.7 22.7 20.0 20.6 ABC

H-2 × DS-20 17.5 16.0 18.2 16.0 16.9 D

H-3 × DS-0 23.4 22.8 24.5 22.7 23.4 A

H-3 × DS-10 18.7 17.9 20.1 19.6 19.1 BCD

H-3 × DS-20 16.3 16.6 18.2 17.1 17.0 D

Means (SA) 20.3NS 19.6 21.5 19.6

Means of each factor treatments in a column, row or interaction bearing same

letter(s) are statistically similar at P≤0.05. H=Hybrid, DS=Drought stress,

SA=Salicylic acid.

Table 2.1.2: Plant height (cm) of sunflower hybrids under different drought

stress levels as influenced by seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 21.0 abc 20.6 abc 21.9 ab 19.6 c 20.8NS

H-2 (NX-00989) 20.9 abc 19.8 c 22.6 a 20.0 bc 20.8

H-3 (FH-352) 20.3 bc 19.7 c 21.6 abc 20.4 bc 20.5


DS-0 (Control) 23.5 b 23.3 b 25.0 a 23.1 b 24.0 A

DS-10 (10% PEG) 19.7 c 19.5 c 21.2 bc 20.0 c 20.4 B

DS-20 (20% PEG) 17.1 d 17.2 d 18.9 cd 17.0 d 17.8 C


H-1 × DS-0 24.5 a 24.2 ab 24.5 a 22.4 a-g 23.9 A

H-1 × DS-10 19.1 f-k 19.8 e-i 22.5 a-f 19.2 f-k 20.2 BC

H-1 × DS-20 19.4 f-k 17.9 h-k 18.8 h-k 17.2 h-k 18.3 DE

H-2 × DS-0 24.1 ab 22.4 a-g 25.3 a 23.2 a-e 23.8 A

H-2 × DS-10 20.8 b-h 20.4 c-i 23.6 a-d 20.8 b-h 21.4 B

H-2 × DS-20 17.9 h-k 16.5 jk 18.8 g-k 16.1 k 17.3 E

H-3 × DS-0 25.0 a 23.5 a-d 25.2 a 23.6 abc 24.3 A

96

H-3 × DS-10 19.1 f-k 18.4 h-k 20.7 b-h 20.0 d-j 19.5 CD

H-3 × DS-20 16.9 ijk 17.3 h-k 19.0 f-k 17.6 h-k 17.7 E

Means (SA) 20.7 B 20.0 B 22.0 A 20.0 B


letter(s) are statistically similar at P≤0.05. H=Hybrid, SA=Salicylic acid,

DS=Drought stress

drought stress and salicylic acid concentrations were statistically significant. The

tallest plants (with 25.0 cm height) were found under DS-0 (control) receiving 0.75

mM dose of salicylic acid, statistically significant to other SA concentrations at all

DS levels. Whereas, the smallest plant height (17.0 cm) was recorded under DS-20

with 1.5 mM SA treatment, which had non-significant difference with other SA

levels at DS-20. Interaction among hybrids × drought levels × salicylic acid

concentrations was significant. All the three hybrids had statistically similar plant

height at all the SA concentrations with the best results in DS-0, while they gave the

smallest shoot length at DS-20 being statistically similar at all SA levels.

Combination of sunflower hybrids × DS levels had significant interaction with

maximum plant height of all genotypes in DS-0 being statistically similar; while the

smallest shoot lengths were recorded for all the genotypes at DS-20 with

nonsignificant difference among the hybrids.

Comparison of the SA application methods and the effect of DS levels on

different hybrids and interaction with various SA concentrations used are shown in

Figure 2.1. Increased levels of drought stress reduced the plant height of all the

sunflower hybrids, which differed non significantly at all DS levels under both foliar

spray and seed soaking treatments with SA, although H-1 (NX-19012) showed

slightly better height at DS-20 (Figure 2.1 a). Plant response to SA treatments was

better at SA-0.75, which produced taller plants at DS-10 under both foliar spray and

seed soaking treatments with SA (Figure 2.1 b). However, the

97

protective effect of

a. Drought stress × Hybrids

b. Drought stress × Salicylic acid

Figure 2.1: Plant height of sunflower hybrids under various levels of drought

stress and salicylic acid application.

salicylic acid on sunflower plants reduced with increasing DS levels, being the

lowest at the highest level of drought stress (DS-20).

15.0

17.0

19.0

21.0

23.0

25.0

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20

Foliar spray of S A Seed soaking in SA

Drought stress level (% PEG)

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

15.0

17.0

19.0

21.0

23.0

25.0

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20

Foliar spray of SA Seed soaking in SA


SA-0 SA-0.375 SA-0.75 SA-1.50

98

Table 2.1.1 and 2.1.2 showed that drought stress significantly reduces the

plant height under mild and severe PEG induced stress in both foliar and seed

soaking mode of applications. Under water stress, plant height reduces due to

impaired mitosis, cell elongation and expansion (Kaya et al., 2006; Hussain et al.,

2008). Results are in conformity with the findings that stem length was significantly

affected under water stress in Abelmoschus esculentus (Sankar et al., 2008); Vigna

unguiculata (Manivannan et al., 2007a); soybean (Zhang et al., 2004) and parsley

(Petroselinum crispum) (Petropoulos et al., 2008). Bajehbaj (2011) showed that four

sunflower cultivars decreased plant height and other growth traits significantly upon

the application of water deficit stress.

H-1 performed better than other hybrids and SA application 0.75mM

increased plant height in both modes of applications under stress (Fig 2.1a, b). The

SA application (especially 0.5 mM) diminished the drought damages and increased

plant height (Sadeghipour and Aghaei 2012). However, it has been observed that

foliar application of salicylic acid and acetyl salicylic acid to soybean and corn plants

@ 10 3 and 10 5 mol/L did not affect the plant height (Khan et al., 2003). These

results indicate that exogenous application of this phytohormone can act as an

effective tool in improving the growth and production of crops under water stress

conditions.

4.2.2 Root Length

The three sunflower hybrids showed significant differences in root length with the

foliar application of salicylic acid (Table 2.2.1). The greatest value of root length

(9.10 cm) was that of H-2 (NX-00989), while the smallest (6.46 cm) was of H-3

(FH-352). Application of salicylic acid at the concentration of 0.75 mM resulted in

a significant increase of root length compared to other rates. Higher levels of DS

caused a significant increase in root length being the highest at DS-20 except H-3

99

(which gives lower values at DS-20), while the lowest in control (DS-0) in both

tolerant hybrids. Therefore, increase in root length can be considered as the indicator

of drought tolerance. Interaction of salicylic acid × genotypes was significant with

the maximum root length (9.55 cm) observed in H-2 (NX-00989) with 0.75 mM SA

concentration, although having a non significant difference with SA-0 and SA-0.375.

The smallest root length values were recorded for H-3 (FH352) at all levels of

salicylic acid application. Levels of drought stress × salicylic acid had significant

interaction, showing higher values under DS-10 at all levels of SA, which differed

significantly with DS-0 and DS-20. Similarly, hybrids × drought stress levels

showed significant interaction. Lower values of root length were recorded in H-3

(FH-352) at DS-20 having significant difference with H-2 (NX-00989) at all DS

levels. Interaction among the sunflower hybrids × drought stress × salicylic acid

concentrations was also statistically significant. The smallest root length values were

recorded for H3 at DS-20, while the H-2 (NX-00989) rendered the highest values at

DS-20 followed by DS-10 treated with SA 0.75 mM.

The root length response of sunflower hybrids with salicylic acid seed

soaking was different from the foliar spray as expressed by data in Table 2.2.2.

Table 2.2.1: Root length (cm) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 8.09 cd 8.20 cd 8.60 bc 8.14 cd 8.26 B

H-2 (NX-00989) 8.82 ab 9.21 ab 9.55 a 8.81 bc 9.10 A

H-3 (FH-352) 6.08 g 6.48 f 6.93 e 6.35 f 6.46 C


DS-0 (Control) 7.44 e 7.57 de 7.92 cd 7.61 de 7.64 B

DS-10 (10% PEG) 8.48 ab 8.74 ab 9.04 a 8.46 bc 8.68 A

DS-20 (20% PEG) 7.07 f 7.58 de 8.11 cd 7.23 e 7.50 B

100


H-1 × DS-0 7.33 h-l 7.43 g-l 8.07 c-l 7.87 e-l 7.68 E

H-1 × DS-10 8.40 c-i 8.63 a-h 8.80 a-h 8.30c-j 8.53 C

H-1 × DS-20 8.53 a-h 8.53 a-h 8.93 a-f 8.27 c-j 8.57 BC

H-2 × DS-0 8.10 c-l 8.37 c-j 8.73 a-h 8.33 c-j 8.38 CD

H-2 × DS-10 8.84 a-d 9.37abc 9.90 ab 8.50b-h 9.15 AB

H-2 × DS-20 9.53 abc 9.90 ab 10.01 a 9.20 a-e

9.66 A

H-3 × DS-0 6.90 jkl 6.91 kl 6.97 i-l 6.63 l 6.85 F

H-3 × DS-10 8.20 c-j 8.23 c-j 8.43 c-i 8.19 e 8.26 D

H-3 × DS-20 3.13 o 4.30 n 5.40 m 4.23 n 4.27 G

Means (SA) 7.66 B 7.96 B 8.36 A 7.72 B



DS=Drought Stress

Here, the sunflower hybrids also differed significantly; the H-3 (FH-352) produced

shortest plant roots at all SA levels. Interaction between drought stress levels and

salicylic acid was statistically significant. The smallest plant roots were found under

DS-20 receiving SA-0. Whereas, longest plant root (7.19 cm) was observed under

DS-10 at SA-0.75 differing non significantly with all other treatments of same level

of stress. Interaction among hybrids × drought levels × salicylic acid concentrations

was also statistically significant; H-2 shows the highest root length (7.87 cm) at DS-

10 at SA-0.75 the smallest root length was of H-3 (FH-352) under DS-20.


longest roots of H-2 (NX-00989) (tolerant) genotype in DS-10 and DS-20 both being

statistically similar; while smallest root lengths were recorded for H-3 (sensitive)

genotype at DS-20 with all concentrations of SA.

101

Comparison of SA application methods for effect of DS levels on different

hybrids and interaction with various SA concentrations is in Figure 2.2. Foliar spray

of salicylic acid caused longer root lengths in all the treatments, indicating a better

stress protection effect as compared to that with seed soaking in SA. However, this

protective effect of SA on sunflower plants was reduced with increasing levels of

drought stress. High levels of drought stress, increased the root length of all the

sunflower hybrids, except H-3 at DS-20, under both foliar spray and seed soaking

treatments with SA (Figure 2.2 a). The H-2 (NX-00989) with foliar spray and seed

soaking showed significantly longer root lengths at all DS levels.

Table 2.2.1 and 2.2.2 showed that drought stress significantly increased the

root length under mild and severe PEG induced stress in both foliar and seed

Table 2.2.2: Root length (cm) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 7.01bc 7.08 bc 7.23 abc 7.02bc 7.08 B

H-2 (NX-00989) 7.25 ab 7.29 ab 7.47 a 7.28 ab 7.32 A

H-3 (FH-352) 5.18 e 5.61 d 6.02 d 5.51 d 5.58 C


DS-0 (Control) 6.39 d 6.54 bcd 6.72 bc 6.51 bcd 6.54 B

DS-10 (10% PEG) 6.91 ab 6.96 ab 7.19 a 6.94ab 7.00 A

DS-20 (20% PEG) 6.13 e 6.48 cd 6.82 bc 6.36 d 6.45 C

Hybrids×Drought Interaction (H × DS × SA) Means (H×DS)

H-1×DS-0 6.47f-j 6.57 d-j 6.74 c-j 6.48 f-j 6.56 CD

H-1×DS-10 7.23 d-j 7.33 a-f 7.52 a-g 7.29 d-j 7.34 AB

H-1×DS-20 7.31 a-f 7.34 a-h 7.41 a-h 7.30 c-j 7.34 AB

H-2×DS-0 6.77c-j 6.87 a-i 6.98 a-i 6.85 a-h 6.87 BC

H-2×DS-10 7.47 a-e 7.47 a-e 7.87 ab 7.47 a-e 7.57 A

102

H-2×DS-20 7.52 a-d 7.53 a-d 7.57a-e 7.47 b-i 7.52 A

H-3×DS-0 5.93 d-j 6.20 e-j 6.43 c-j 6.21 hij 6.19 D

H-3×DS-10 6.03 d-j 6.07 d-j 6.17 g-j 6.07 f-j 6.08 D

H-3×DS-20 3.57k 4.57d-j 5.47 d-j 4.24 d-j 4.46 E

Means (SA) 6.48 C 6.66 B 6.91A 6.60 B



DS=Drought Stress


3.00

5.00

7.00

9.00

11.00

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)


5.00

6.00

7.00

8.00

9.00

10.00

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

103

Figure 2.2: Root length of sunflower hybrids under various levels of drought


soaking mode of applications. The importance of root systems in acquiring water has

long been recognized.Water stress causes visible increase in root length. A prolific

root system can confer the advantage to support accelerated plant growth during the

early crop growth stage and extract water from shallow soil layers that is otherwise

easily lost by evaporation in legumes (Johansen et al., 1992). In alfalfa, the root

length increased by PEG-induced drought stress (Zeid and Shedeed, 2006). An

increased root growth due to water stress was reported in sunflower (Tahir et al.,

2002).

Tolerant sunflower hybrids performed better than sensitive and SA

application 0.75mM increases root length in both modes of applications under stress

especially in foliar spray (Fig 2.2 a, b). Salicylic acid is reported to cause an increase

in root and shoot growth (Khodary 2004; El-Tayeb and Naglaa, 2010). Exogenous

application of SA could counteract the adverse effects of drought by improving the

growth of root and shoot, retention of pigment content, increasing the concentration

of organic solutes (soluble sugars and soluble nitrogen) as osmoprotectants, keeping

out the polysaccharide concentration and/or stabilization of essential proteins


4.2.3 Fresh Shoot Weight

The fresh shoot weight of three sunflower hybrids differed significantly with

the foliar application of salicylic acid (Table 2.3.1). Genotype H-2 (NX00989) had

the highest fresh shoot weight while the lowest fresh shoot weight was obtained for

H-3 (FH-352). Application of different salicylic acid treatments significantly

104

increased the shoot fresh weight. Highest fresh shoot weight was recorded with

salicylic acid treatment of 0.75 mM, while lowest value was in control. Application

of drought stress levels significantly reduced the shoot fresh weight. There was

significant interaction between genotypes and salicylic acid concentrations.

Maximum fresh shoot weight (1.56 g) was found in genotype NX00989 receiving

0.75 mM salicylic acid, while minimum fresh shoot weight (0.98 g/plant) was

observed in genotype FH-352 without salicylic acid application. Interaction of

salicylic acid × DS and that of hybrids × salicylic acid × DS also showed significant

differences. Highest shoot fresh weight (2.0 g) was recorded for sunflower genotype

NX-00989 under DS-0 and with 0.75 mM salicylic acid and lowest (0.80 g/plant) in

H-3 (FH-352) at DS-20 without any application of salicylic acid.

Results of fresh shoot weight under seed soaking with salicylic acid of

sunflower hybrids at different DS levels are shown in Table 2.3.2. Here, the

sunflower hybrids showed similar responses to DS levels and SA concentrations as

that recorded under foliar spray of salicylic acid. Interaction between various levels

of drought stress and salicylic acid was statistically significant. Interaction among

hybrids × drought levels × salicylic acid levels also showed significant differences.


maximum fresh shoot weight of all genotypes in DS-0; while the lowest fresh shoot

weight was recorded for all the genotypes at DS-20 with significant difference

among the hybrids.


hybrids and interaction with various SA concentrations is expressed in Figure 2.3.

Drought stress (20% PEG) reduced the fresh shoot weight of all sunflower hybrids

Table 2.3.1: Fresh shoot weight (g/plant) of sunflower hybrids under different

drought stress levels as influenced by foliar application of salicylic acid.

105


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 1.13 c-f 1.21 b-f 1.38 ab 1.14 b-f 1.21 B

H-2 (NX-00989) 1.23 b-e 1.36 abc 1.56 a 1.29 bcd 1.36 A

H-3 (FH-352) 0.98 f 1.05 def 1.31 bc 1.01 ef 1.09 C


DS-0 (Control) 1.35 bcd 1.49 b 1.83 a 1.38 bc 1.51 A

DS-10 (10% PEG) 1.07 ef 1.12 def 1.29 b-e 1.09 ef 1.14 B

DS-20 (20% PEG) 0.92 f 1.01 f 1.13 c-f 0.97 f 1.01 C


H-1 × DS-0 1.30 b-j 1.41 b-g 1.80 ab 1.25 c-j 1.44 B

H-1 × DS-10 1.14 e-j 1.20 c-j 1.26 c-j 1.18 d-j 1.20 CD

H-1 × DS-20 0.95 g-j 1.01 f-j 1.09e-j 0.99 g-j 1.01 DEF

H-2 × DS-0 1.52 a-f 1.67 a-d 2.00 a 1.57 a-e 1.69 A

H-2 × DS-10 1.17 d-j 1.25 c-j 1.42 b-g 1.21 c-j 1.27 BC

H-2 × DS-20 1.01 g-j 1.17 d-j 1.26c-j 1.10 e-j 1.13 CDE

H-3 × DS-0 1.25 c-j 1.40 b-h 1.70 abc 1.32 b-i 1.41 B

H-3 × DS-10 0.89 hij 0.91g-j 1.18 d-j 0.89 hij 0.97 EF

H-3 × DS-20 0.80 j 0.84 ij 1.04 f-j 0.83 ij 0.88 F

Means (SA) 1.11 B 1.21 B 1.42 A 1.15 B



DS=Drought Stress

Table 2.3.2: Fresh shoot weight (g/plant) of sunflower hybrids under different

drought stress levels as influenced by seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 1.18 de 1.22 cd 1.35 b 1.15 def 1.22 B

106

H-2 (NX-00989) 1.26 bcd 1.34 b 1.48 a 1.30 bc 1.35 A

H-3 (FH-352) 1.00 g 1.03 fg 1.09 efg 1.00 g 1.03C

Drought stress (DS) Interaction s(DS × SA) Means (DS)

DS-0 (Control) 1.39 b 1.50 b 1.72 a 1.40 b 1.50 A

DS-10 (10% PEG) 1.09 cde 1.12 cd 1.19 c 1.10 cde 1.12 B

DS-20 (20% PEG) 0.95 f 0.97 ef 1.02 def 0.95 f 0.97 C


H-1 × DS-0 1.33 fg 1.44 def 1.80 b 1.31 fgh 1.47 B

H-1 × DS-10 1.22 g-k 1.23 g-k 1.23 f-j 1.19 g-k 1.22 CD

H-1 × DS-20 0.98 l-q 0.98 m-q 1.02 k-q 0.94 n-q 0.98 E

H-2 × DS-0 1.56 cde 1.70 bc 2.04 a 1.60 bcd 1.73 A

H-2 × DS-10 1.16 g-m 1.21 g-k 1.23 g-k 1.19 g-k 1.20 D

H-2 × DS-20 1.05 j-p 1.11 h-n 1.19 g-l 1.10 i-o 1.11 D

H-3 × DS-0 1.28 f-i 1.36 efg 1.33 fg 1.29 f-i 1.31 C

H-3 × DS-10 0.90 opq 0.91 n-q 1.10 h-o 0.90 opq 0.95 E

H-3 × DS-20 0.82 q 0.83 q 0.84 pq 0.81 q 0.83 F

Means (SA) 1.14 B 1.20 B 1.31 A 1.15 B



DS=Drought Stress

almost equally receiving either foliar or seed SA soaking treatments. Hybrid, H-2

(NX-00989) showed significantly higher fresh shoot weight than that of H-3

(FH352) genotype at all DS levels (Figure 2.3 a). SA application at 0.75 mM SA

produced greater plant fresh shoot weights at all DS levels especially under foliar

spray with SA (Figure 2.3 b). The curative effect of salicylic acid on sunflower plants

reduced to enhancing DS levels, being the minimum at the highest level of drought

stress.

107

Table 2.3.1 and 2.3.2 showed that drought stress significantly decreases the

shoot fresh weight under mild and severe PEG induced stress in both foliar and seed

soaking mode of applications. Moisture stress has adverse effects on the plant growth

and development. It causes a considerable reduction in growth vigor of shoot

(Andrade et al., 2013). Results are in line with the findings of other researchers,

biomass were reduced in moisture stressed soybean (Specht et al., 2001), Poncirus

trifoliatae seedlings (Wu et al., 2008), Shoot fresh and dry weights of alfalfa also

decreased by PEG-induced drought stress (Zeid and Shedeed, 2006). Severity,

duration and timing of water stress, and interaction between water stress and other

factors are particularly important (Plaut, 2003).

Sunflower hybrids H-1 and H-2 performed better than H-3(sensitive) and

SA application 0.75mM increased significantly shoot fresh weight in both modes of

application under stress especially giving higher values in foliar spray (Fig 2.3 a, b).

Similarly, salicylic acid is reported to neutralize the negative effects of drought by

improving the growth of root and shoot (El-Tayeb and Naglaa, 2010;

Heshmat et al., 2012). Salicylic acid treatments increased the shoot fresh


0.70

1.00

1.30

1.60

1.90

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

108

Figure 2.3: Fresh shoot weight of sunflower hybrids under various levels of

drought stress and salicylic acid application.

weight of barley seedlings (Metwally, 2003). Agarwal et al. (2005) demonstrated

that reduction in biomass of seedling due to PEG-induced water stress was slightly

recovered by salicylic acid treatment.

4.2.4 Shoot Dry Weight

Foliar application of salicylic acid, data showed that sunflower hybrids significantly

differed from each other regarding shoot dry weight (Table 2.4.1). Genotype NX-

00989 gained the highest weight while the lowest was observed for genotype FH-

352. Application of different salicylic acid treatments had significant effect on dry

shoot weight. The highest value of dry shoot weight was observed with salicylic acid

application at 0.75 mM, while the lowest was in control. Application of the drought

stress levels resulted in significant reduction of shoot dry weight. There was

significant interaction between genotypes and salicylic acid treatments. Maximum

weight (0.07 g) of dry shoots was gained by tolerant genotypes H-1 and H-2with

0.75 mM salicylic acid, although having a non significant difference with other SA


0.70

1.00

1.30

1.60

1.90

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

109

concentrations. The lowest weight of dry shoots (0.047 g) was observed in genotype

FH-352 without salicylic acid application. The interactions between SA treatments

× Drought (PEG) and among Hybrid × SA × Drought were also statistically

significant. The highest weight (0.09 g) of dry shoots was recorded for sunflower

genotype NX -19012 under 0.75 mM salicylic acid application without any drought

stress. The lowest weight (0.039 g) of dry shoots was observed in FH-352 genotype

at DS-20 without any application of salicylic acid.

Data for dry shoot weight obtained through seed soaking of sunflower

hybrids with salicylic acid at different DS levels are shown in Table 2.4.2.

Table 2.4.1: Dry Shoot weight (g) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.053 b 0.059 ab 0.070 a 0.058 ab 0.060 A

H-2 (NX-00989) 0.055 ab 0.062 ab 0.070 a 0.060 ab 0.062 A

H-3 (FH-352) 0.047 b 0.054 b 0.059 ab 0.051 b 0.053 B


DS-0 (Control) 0.062 bcd 0.072 ab 0.084 a 0.070 abc 0.072 A

DS-10 (10% PEG) 0.051 def 0.055 c-f 0.062 b-e 0.053 def 0.055 B

DS-20 (20% PEG) 0.041 abc 0.047 ef 0.053 def 0.046 f 0.047 C


H-1 × DS-0 0.062 a-f 0.071 a-f 0.090 a 0.070 a-f 0.073 A

H-1 × DS-10 0.053 a-f 0.057 b-f 0.064 a-f 0.055 b-f 0.057 BC

H-1 × DS-20 0.043 ef 0.048 c-f 0.057 b-f 0.049 c-f 0.049 CD

H-2 × DS-0 0.067 a-f 0.077 a-d 0.085 ab 0.074 a-e 0.076 A

H-2 × DS-10 0.055 b-f 0.058 a-f 0.066 a-f 0.056 b-f 0.059 BC

H-2 × DS-20 0.041 f 0.051 c-f 0.060 a-f 0.049 c-f 0.050 CD

H-3 × DS-0 0.057 b-f 0.069 a-f 0.078 abc 0.064 a-f 0.067 AB

H-3 × DS-10 0.045 def 0.051 c-f 0.056 b-f 0.049 c-f 0.051 CD

110

H-3 × DS-20 0.039 f 0.041f 0.043 ef 0.040 f 0.041 B

Means (SA) 0.051 B 0.058 B 0.066 A 0.056 B



DS=Drought Stress

Table 2.4.2: Shoot dry weight (g) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.056 b-e 0.059 b 0.066 a 0.062 ab 0.061 A

H-2 (NX-00989) 0.056 bcd 0.058 bc 0.058 bc 0.058 bc 0.057 A

H-3 (FH-352) 0.048 e 0.050 de 0.051 cde 0.052 cde 0.050 B


DS-0 (Control) 0.065 bc 0.067 b 0.076 a 0.069 b 0.069 A

DS-10 (10% PEG) 0.053 d 0.055 d 0.054 d 0.059 cd 0.055 B

DS-20 (20% PEG) 0.042 e 0.045 e 0.044 e 0.044 e 0.044 C


H-1 × DS-0 0.066 b-h 0.075 b 0.094 a 0.073 bc 0.077 A

H-1 × DS-10 0.056 h-m 0.057 g-m 0.057 f-l 0.068 b-g

0.060 B

H-1 × DS-20 0.045 mno 0.045 l-o 0.048 j-o 0.045 l-o

0.046 CD

H-2 × DS-0 0.070 b-e 0.069 b-f 0.074 b 0.072 bcd

0.071 A

H-2 × DS-10 0.057 g-m 0.058 f-k 0.056 g-m 0.056 h-m 0.057 B

H-2 × DS-20 0.041 no 0.047 j-o 0.043 no 0.045 l-o 0.044 CD

H-3 × DS-0 0.059 e-j 0.058 e-k 0.061 d-i 0.061 c-i 0.060 B

H-3 × DS-10 0.046 k-o 0.049 j-o 0.049 j-o 0.053 i-n 0.049 C

H-3 × DS-20 0.040 o 0.042 no 0.043 no 0.041 no 0.042 D

Means (SA) 0.053 B 0.056 AB 0.058 A 0.057 AB

111


letter(s) are statistically similar at P≤0.05 H=Hybrid, SA=Salicylic acid,

DS=Drought Stress

Sunflower hybrids differed significantly with lower dry shoot weight of H-3

(FH-352). SA-0.75 seed soaking increased significantly the dry shoot weight as

compared to control but statistically similar to that with other SA levels. Higher

drought stress levels reduced significantly the dry shoot weight, as the lowest weight

was with DS-20 (0.044). Interactions of genotypes and various levels of drought

stress with salicylic acid were statistically significant. Highest dry shoot weight of

plants (0.076 g) was found under DS-0 (Control) receiving 0.75 mM salicylic acid.

Whereas, the lowest weight (0.042 g) was recorded at DS-20 without SA application,

which had non significant difference with other SA levels at DS20. Interaction

among hybrids × drought levels × salicylic acid levels were also statistically

significant. Two hybrids (H-1 and H-2) had statistically similar dry shoot weight at

each SA level with the best results in DS-0, while H-3 (FH-352) gave the least dry

shoot weight at DS-20 being statistically similar at each SA level. Combination of

sunflower hybrids × DS levels had significant interaction with largest dry shoot

weight of all genotypes in DS-0 but lower in H-3; while the lowest dry shoot weights

were recorded for all the genotypes at DS-20 with non significant difference between

H-1 and H-2 hybrids.

Comparison of two methods of SA application (foliar and seed soaking) for

the effect of DS levels on different hybrids and interaction with various SA

concentrations is shown in Figure 2.4. Increased levels of drought stress reduced dry

shoot weight of all the sunflower hybrids, which differed non significantly for H-1

(NX-19012) and H-2 (NX-00989) at all DS levels under both foliar spray and seed

112

soaking treatments with SA, while H-3 (FH-352) showed significantly lower dry

shoot weight (Figure 2.4 a). Plant response to SA foliar spray was better at SA-


Figure 2.4: Dry shoot weight of sunflower hybrids under various levels of


0.75, which produced heavier plants at all PEG levels (Figure 2.4 b). Seed soaking

treatments with different SA concentrations gave non significant difference among

0.030

0.040

0.050

0.060

0.070

0.080

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)


0.040

0.050

0.060

0.070

0.080

0.090

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

113

them especially at DS-10 and DS-20. Positive effect of salicylic acid on sunflower

plants reduced with increasing DS levels, being the lowest at the highest level of

drought stress (20% PEG).

Table 2.4.1 and 2.4.2 showed that drought stress significantly reduces the shoot

dry weight under mild and severe stress induced by PEG in both foliar and seed

soaking mode of applications. Water stress has negative affects on the plant growth

and development.The common adverse effect of water deficit on plants is the

reduction in fresh and dry biomass production (Farooq et al., 2009). Plant

productivity is strongly related to the processes of dry matter partitioning and

temporal biomass distribution under water stress (Kage et al., 2004). Drought stress

greatly reduces the plant growth and development during the vegetative stage

(Tripathy et al., 2000; Manikavelu et al., 2006). In Medicago sativa, shoot weight

decreased by PEG-induced drought stress (Zeid and Shedeed, 2006). A moderate

stress tolerance was noticed in rice in expressions of shoot dry mass (Lafitte et al.,

2007). Drought stress affected shoot dry weight (DW) than root DW in a greater

proportion, in both sensitive and tolerant lines of sunflower Andrade et al. (2013).

Sunflower hybrids H-1 and H-2 performed better than H-3 ( sensitive) and

SA application 0.75mM increased significantly shoot dry weight in foliar application

but non significantly in pre treatment mode of application under stress (Fig 2.4 a, b).

Salicylic acid counteracts the negative effect of drought on plants by improving their

growth (El-Tayeb and Naglaa, 2010; Heshmat et al., 2012). Plant dry weight reduced

in drought conditions, but it increased significantly by SA application. Singh and

Usha (2003) and Sakhabutdinova et al. (2003) observed that salicylic acid

application increased the dry mass of wheat seedlings.

114

4.2.5 Fresh Root Weight

With the foliar application of salicylic acid there was significant difference

between sunflower genotypes in respect of fresh root weight (Table 2.5.1). Highest

fresh root weight (0.408 g/plant) was recorded for H-2 (NX-00989), while the lowest

(0.282 g/plant) in H-3 (FH-352). Application of salicylic acid at the concentration of

0.75 mM resulted in significant increase of fresh root weight. There was significant

decrease in fresh root weight with application of drought stress. The lowest fresh

root weight (0.267 g) was observed with DS-20, while highest (0.468 g) was

recorded in the control (DS-0). The salicylic acid × genotype interaction was

significant with maximum fresh root weight (0.498 g) was in NX00989 with 0.75

mM SA application. Minimum fresh root weight (0.266 g) was observed in FH-352

with 0.375 mM salicylic acid. The drought level × salicylic and the hybrid × drought

× salicylic acid interactions also had significant difference. The highest fresh root

weight (0.651 g/plant) was recorded for sunflower genotype NX-00989 without DS

through 0.75 mM SA application, while the lowest fresh root weight (0.117 g) was

found in H-3 (FH-352) genotype with DS-20 and 1.50 mM salicylic acid. Seed

soaking of sunflower hybrids with SA at different DS levels showed similar trend of

fresh root weight data as under foliar spray of SA (Table 2.5.2). Here also, the

sunflower hybrids differed significantly; and SA-0.75 produced significantly higher

fresh root weight as compared to other Table 2.5.1: Fresh root weight (g/plant) of

sunflower hybrids under different drought stress levels as influenced by foliar



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.344 cde 0.360 bc 0.422 ab 0.351 bcd 0.369 B

H-2 (NX-00989) 0.371 bc 0.387 bc 0.498 a 0.376 bc 0.408 A

115

H-3 (FH-352) 0.274 ef 0.266 f 0.313 c-f 0.277 def 0.282 C


DS-0 (Control) 0.428 bc 0.450 b 0.554 a 0.440 bc 0.468 A

DS-10 (10% PEG) 0.312 de 0.302 de 0.372 cd 0.311 de 0.324 B

DS-20 (20% PEG) 0.248 e 0.262 e 0.307 de 0.252 e 0.267 C


H-1 × DS-0 0.407 b-i 0.423 b-h 0.513 ab 0.413 b-i

0.439 B

H-1 × DS-10 0.331 d-j 0.341 c-j 0.380 b-j 0.330 d-j

0.346 C

H-1 × DS-20 0.293 g-k 0.317 d-j 0.373 b-j 0.310 f-j

0.323 CD

H-2 × DS-0 0.460 b-f 0.487 bcd 0.651 a 0.473 b-e

0.518 A

H-2 × DS-10 0.341 c-j 0.336 d-i 0.447 b-g 0.323 e-j

0.362 C

H-2 × DS-20 0.312 d-j 0.339 c-j 0.396 b-i 0.330 d-j

0.344 C

H-3 × DS-0 0.417 b-i 0.440 b-g 0.497 abc 0.433 b-h

0.447 B

H-3 × DS-10 0.265 i-m 0.230 j-m 0.290 g-k 0.280 h-l 0.266 D

H-3 × DS-20 0.140 klm 0.139 klm 0.151 klm 0.117 m

0.136 E

Means (SA) 0.330 B 0.339 B 0.411 A 0.335 B



DS=Drought Stress

SA concentrations. Different levels of drought stress also caused significant

reduction in fresh root weight, as the lowest was with DS-20. Interactions for both

sunflower hybrids and various levels of drought stress under different salicylic acid

rates had statistically significant difference. Interaction among hybrids × drought

stress levels × salicylic acid concentrations was also statistically significant.

Combination of sunflower hybrids × DS levels had significant interaction with the

highest fresh root weight of all genotypes under DS-0 but statistically different.

The lowest fresh root weights were recorded for the H-3 (FH-352) genotype at DS-

20 under all the SA levels.

116

Both SA application methods (foliar and seed soaking) showed similar

response with different DS levels on three hybrids and their interactions with various

SA treatments (Figure 2.5). Higher levels of drought stress reduced the fresh root

weight of all the sunflower hybrids under both foliar spray and seed soaking

treatments with SA, although H-1 (NX-19012) and H-2 (NX-00989) showed

significantly higher fresh root weight at DS-20 (Figure 2.5 a). Plant root response to

SA application was better at SA-0.75, which produced greater fresh root weight at

all DS levels under both foliar spray and seed soaking treatments with SA (Figure

2.4 b). However, protective effect of salicylic acid on sunflower plants reduced with

increasing drought stress levels, being the lowest at the highest level of drought stress

application.

Drought stress significantly reduces the root fresh weight. Current study

exhibits that root fresh weight was reduced in sunflower hybrids exposed to drought

stress in both methods of application (Table 2.5.1 and 2.5.2). The improvement in

root system increases the water uptake and maintains required Table 2.5.2: Fresh

Root weight (g/plant) of sunflower hybrids under different drought stress levels

as influenced by seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.361 de 0.393 cd 0.435 b 0.361 de 0.387 B

H-2 (NX-00989) 0.392 cd 0.416 bc 0.544 a 0.402 bc 0.439 A

H-3 (FH-352) 0.301 f 0.332 ef 0.381 cd 0.310 f 0.331 C


DS-0 (Control) 0.451 b 0.484 b 0.566 a 0.461 b 0.491A

DS-10 (10% PEG) 0.328 cd 0.362 c 0.456 b 0.337 c 0.370 B

DS-20 (20% PEG) 0.276 e 0.294 de 0.339 c 0.275 e 0.296 C


117

H-1 × DS-0 0.427 def 0.477 bcd 0.500 bc 0.430 de 0.458 B

H-1 × DS-10 0.338 hi 0.345 hi 0.423 d-g 0.333 hi

0.360 D

H-1 × DS-20 0.320 hi 0.357 h 0.381 e-h 0.319 hi

0.344 D

H-2 × DS-0 0.484 bcd 0.517 b 0.673 a 0.503 bc 0.544 A

H-2 × DS-10 0.363 fgh 0.380 e-h 0.520 b 0.350 h 0.403 C

H-2 × DS-20 0.330 hi 0.350 h 0.440 cde 0.353 h 0.368 B

H-3 × DS-0 0.443 cde 0.460 bcd 0.523 b 0.450 cd

0.469 B

H-3 × DS-10 0.283 i 0.360 gh 0.423 d-g 0.327 hi 0.348 D

H-3 × DS-20 0.177 j 0.177 j 0.195 j 0.153 j

0.175 E

Means (SA) 0.352 C 0.380 B 0.453 A 0.358 C



DS=Drought Stress

Osmotic pressure by accumulating proline in Phoenix dactylifera (Djibril et al.,

2005). Drought stress considerably affects the plant growth as indicated by reduced

fresh biomass of roots (Heshmat et al., 2012).

Sunflower hybrids H-2 and H-1 performed better than H-3 (sensitive) which dropped

to lower values under severe stress conditions and SA application 0.75mM increased

significantly shoot fresh weight in both modes of application (Fig 2.5 a, b). Salicylic

acid treatment increased the root fresh weight of barley seedlings (Metwally, 2003).

Agarwal et al. (2005) demonstrated that reduction in biomass of seedling due to

PEG-induced water stress is slightly ameliorated by treatment with salicylic acid.

Khodray (2004) reported an improved fresh root weight of stressed maize plants

treated with salicylic acid.

118

4.2.6 Root Dry Weight

In the first set of experiment for foliar application of salicylic acid,

sunflower genotypes differed significantly with respect to dry root weight (Table

2.6.1). The highest dry root weight (0.018 g) was recorded in both NX-19012 and

NX-00989 genotypes, while the lowest (0.015 g) was in FH-352. Application of

salicylic acid at the concentration of 0.75 mM resulted in significantly greater weight

of dry roots as compared to that with all other levels. There was significant decrease

in dry root weight with the application of 20% water stress (PEG) (0.015

g) while the highest dry root weight (0.018 g) was observed in control having non

significant difference with that of DS-10. Salicylic acid × genotypes interaction was

significant with maximum dry root weight (0.023 g) in NX-19012 and NX00989 at

0.75 mM salicylic acid application. Minimum dry root weight (0.014 g) -


0.100

0.200

0.300

0.400

0.500

0.600

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

119

Figure 2.5: Fresh root weight of sunflower hybrids under various levels of


Table 2.6.1: Dry Root weight (g/plant) of sunflower hybrids under different



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.016 c 0.016 c 0.023 a 0.016 c 0.018 A

H-2 (NX-00989) 0.016 c 0.018 bc 0.023 a 0.016 c 0.018 A

H-3 (FH-352) 0.014 c 0.015 c 0.015 c 0.014 c 0.015 B


DS-0 (Control) 0.016 bc 0.016 bc 0.023 a 0.016 c 0.018 A

DS-10 (10% PEG) 0.015 c 0.017 bc 0.021 ab 0.016 bc 0.017 A

DS-20 (20% PEG) 0.015 c 0.015 c 0.017 bc 0.014 c 0.015 B


H-1 × DS-0 0.016 bcd 0.016 bcd 0.022 bc 0.017 bcd 0.018 ABC

H-1 × DS-10 0.015 cd 0.016 bcd 0.025 b 0.017 bcd 0.018 ABC

H-1 × DS-20 0.015 cd 0.017 bcd 0.021 bcd 0.016 bcd 0.017 A-D

H-2 × DS-0 0.017 bcd 0.017 bcd 0.032 a 0.016 bcd 0.020 A


0.200

0.300

0.400

0.500

0.600

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

120

H-2 × DS-10 0.016 bcd 0.021 bcd 0.022 bc 0.018 bcd 0.019 AB

H-2 × DS-20 0.015 cd 0.016 bcd 0.016 bcd 0.015 cd 0.016 BCD

H-3 × DS-0 0.016 bcd 0.016 bcd 0.016 bcd 0.016 bcd 0.016 BCD

H-3 × DS-10 0.014 cd 0.014 cd 0.015 cd 0.014 cd 0.014 CD

H-3 × DS-20 0.013 cd 0.014 cd 0.014 cd 0.012 d 0.013 D

Means (SA) 0.015 B 0.016 B 0.020 A 0.016 B



DS=Drought Stress

was observed in FH 352 without salicylic acid application and 1.5 mM SA although

having a non significant difference with other SA levels. Drought stress × salicylic

acid and hybrids × drought × salicylic acid interactions were also statistically

significant. The highest dry root weight (0.032 g) was recorded for sunflower

genotype H-2 (NX-00989) at DS-0 with 0.75 mM salicylic acid, while the lowest

value (0.012 g) was observed in H-3 (FH-352) genotype at DS-20 and

1.50 mM salicylic acid application.

Dry root weight of sunflower hybrids obtained from seed soaking with

salicylic acid treatments at different DS levels is shown in Table 2.6.2. Sunflower

hybrids differed significantly with the lowest weight for H-3 (FH-352), and SA0.75

produced significantly higher dry root weight (0.021 g) as compared to other SA

treatments. Different levels of drought stress also caused significant reduction in dry

root weight, as the lowest dry root weight was in DS-20 (0.015 g). Interaction

between various levels of drought stress x salicylic acid was statistically significant.

The highest dry root weight (0.024 g) was found under DS-0 (Control) receiving

0.75 mM dose of salicylic acid, which was statistically different to other SA levels

in DS-0. The lowest dry root weight (0.014 g) was recorded in DS-20 combined with

121

0 and1.5 mM SA, which had non significant difference with SA0.375 concentration

at DS-20 level. Interaction among hybrids × drought stress levels × salicylic acid

concentrations was also statistically significant. All the three hybrids had statistically

similar dry root weight at all the SA levels with the best results in DS-0, while they

gave the smallest dry root weight at DS-20 being statistically similar at all SA

concentrations. Combination of sunflower hybrids × DS levels had significant

interaction with the maximum dry root weight of all Table 2.6.2: Dry Root weight

(g/plant) of sunflower hybrids under different drought stress levels as

influenced by seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.015 de 0.016 cd 0.022 b 0.016 cd 0.018 A

H-2 (NX-00989) 0.016 cd 0.016 cd 0.024 a 0.016 cd 0.018 A

H-3 (FH-352) 0.014 e 0.014 e 0.015 de 0.014 e 0.014 B


DS-0 (Control) 0.016 cde 0.017 cd 0.024 a 0.017 cd 0.019 A

DS-10 (10% PEG) 0.015 ef 0.015 ef 0.020 b 0.016 cde 0.016 B

DS-20 (20% PEG) 0.014 f 0.015 ef 0.017 cd 0.014 f 0.015 C


H-1 × DS-0 0.016 e-j 0.017 e-i 0.021 cd 0.016 e-j 0.017 BC

H-1 × DS-10 0.015 f-j 0.016 e-j 0.025 b 0.017 e-i 0.018 B

H-1 × DS-20 0.015 f-j 0.016 e-j 0.022 c 0.015 f-j 0.017 BC

H-2 × DS-0 0.018 de 0.018 de 0.036 a 0.017 e-i 0.022 A

H-2 × DS-10 0.015 f-j 0.016 e-j 0.021 cd 0.016 e-j 0.017 BC

H-2 × DS-20 0.015 f-j 0.015 f-j 0.016 e-j 0.015 f-j 0.015 D

H-3 × DS-0 0.015 f-j 0.016 e-j 0.016 e-j 0.016 e-j 0.016 CD

H-3 × DS-10 0.014 hij 0.014 hij 0.015 f-j 0.014 hij 0.014 DE

H-3 × DS-20 0.014 hij 0.013 ij 0.013 ij 0.013 ij 0.013 E

122

Means (SA) 0.015 B 0.016 B 0.021 A 0.015 B



DS=Drought Stress

genotypes in DS-0 being statistically similar; while the smallest dry root weight was

recorded for all the genotypes at DS-20 with non significant difference among the

hybrids.


hybrids and interaction with various SA treatments is shown in Figure 2.6. Increased

levels of drought stress reduced the dry root weight of all the sunflower hybrids,

which differed non significantly at all DS levels under both foliar spray and seed

soaking treatments with SA, although H-1 (NX-19012) showed slightly better dry

root weight at DS-20 (Figure 2.6 a). Plant response to SA application was better at

SA-0.75, which produced greater dry root weight at all DS levels under both foliar

spray and seed soaking treatments with SA (Figure 2.6 b).

Water stress significantly reduced the root dry weight. Present results

exhibits that root fresh weight was reduced in sunflower hybrids exposed to drought

stress in both modes of application (Table 2.6.1 and 2.6.2). Significant reduction in

dry mass of plant roots due to drought stress has also been reported previously for

different crops (Zeid and Shedeed, 2006; Nizami et al 2008). Andrade et al., 2013

compared two inbred lines of sunflower (sensitive B59, and tolerant B71) with

contrasting behavior to moisture déficit; he also reported decreased dry root weight

under water stress.

123

Tolerant sunflower hybrids performed better than H-3(sensitive) which

dropped to lower values under severe water stress conditions. SA application 0.75

mM increased significantly root dry weight in both modes of application (Fig 2.6 a,

b). Moreover, suitable exogenous application of PGR may enhance plant growth

and tolerance to water stress (Arteca, 1996). Salicylic acid is reported to counteract

the negative effects of drought by increasing the growth of root and shoot (ElTayeb

and Naglaa, 2010; Heshmat et al., 2012). El-Tayeb (2005) reported an increase in

dry root weight of drought stressed and controlled plants with the application of

salicylic acid.

4.2.7 Photosynthesis Rate

With foliar application of salicylic acid, the sunflower hybrids differed

significantly with respect to photosynthesis rate (Table 2.7.1). Genotype NX- 19012

(H-1) showed the highest photosynthesis rate while the lowest one was for genotype

H-3 (FH-352). Various concentrations of salicylic acid application exhibited

significant effect on photosynthesis rate, being the highest at 0.75 mM SA and the

lowest in control. Application of drought stress (PEG) caused significant reduction

in photosynthesis rate from 9.87 in control to 5.95 µmol/m2/s under DS-20. There

was significant interaction between genotypes and salicylic acid concentrations.

Higher photosynthesis rates were recorded in all genotypes receiving 0.75 and 1.50

mM doses of salicylic acid with non-significant difference. Relatively lower

photosynthesis rates were observed in genotype FH-352 without salicylic acid

application. The SA×DS levels interaction and H×SA×DS interaction were also

significant. The highest photosynthesis rate was recorded in sunflower genotype NX-

00989 without stress at 0.75 mM SA, while the lowest

photosynthesis rate (5.09) was observed for FH-352 genotype at 20% PEG without

any application of salicylic acid (SA-0).

124

Table 2.7.1: Photosynthesis rate (µmol/m2/s) of sunflower hybrids under

different drought stress levels as influenced by foliar application of salicylic

acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 7.72 abc 8.24 ab 8.95 a 8.36 ab 8.32 A

H-2 (NX-00989) 7.63 abc 7.03 bc 9.00 a 8.22 ab 7.97 AB

H-3 (FH-352) 6.68 c 7.45 bc 8.34 ab 7.81 abc 7.57 B


DS-0 (Control) 9.61 ab 9.94 ab 10.72 a 10.22 ab 10.12 A

DS-10 (10% PEG) 7.08 def 7.88 cde 9.03 bc 8.16 cd 8.04 B

DS-20 (20% PEG) 5.35 g 5.90 fg 6.54 efg 6.02 fg 5.95 C


H-1 × DS-0 9.81 a-e 9.99 a-d 10.48 ab 10.21 ab 10.12 A

H-1 × DS-10 7.69 b-j 8.82 a-h 10.04 abc 9.03 a-g 8.90 BC

H-1 × DS-20 5.67 ij 5.90 g-j 6.33 g-j 5.84 ij 5.94 D

H-2 × DS-0 9.92 a-e 7.17 d-j 10.92 a 10.21 abc 9.56 ABC

H-2 × DS-10 7.69 b-j 8.20 a-i 9.68 a-f 8.33 a-i 8.47 C

H-2 × DS-20 5.30 ij 5.71 ij 6.39 g-j 6.12 hij 5.88 D

H-3 × DS-0 9.09 a-g 9.66 a-f 10.76 a 10.23 ab 9.94 AB

H-3 × DS-10 5.86 ij 6.61 g-j 7.37 c-j 7.12 e-j 6.74 D

H-3 × DS-20 5.09 ij 6.08 hij 6.89 f-j 6.08 hij 6.04 D

Means (SA) 7.35 C 7.57 BC 8.76 A 8.13 B


letter(s) are statistically similar at P≤0.05.H=Hybrid, SA=Salicylic acid,

DS=Drought stress,

Same sunflower hybrids under another experiment through soaking of seeds

with salicylic acid at different DS levels exhibited significant difference for

photosynthesis rate being lower in FH-352 (Table 2.7.2). Medium concentration of

125

SA-0.75 produced significantly higher photosynthesis rate in plants (8.63µmol/m2/s)

as compared to other SA levels. Each increment in drought stress caused significant

reduction in plants photosynthesis rate, as the lowest one was with DS-20 (5.83

µmol/m2/s). Interactions of various salicylic acid concentrations with drought stress

levels and hybrids were statistically significant. The highest photosynthesis rate of

plants was found under DS-0 (Control) receiving 0.75 mM dose of salicylic acid,

being statistically different with other SA concentrations. Whereas, the lowest

photosynthesis rate (5.26 µmol/m2/s) was recorded in DS-20 without SA application,

which had significant difference with other SA levels at DS-20. Interaction among

hybrids × drought levels × salicylic acid concentrations was also statistically

significant. All the three hybrids had statistically similar photosynthesis rate at each

of the SA level with the best results in DS-0, while they gave the lowest

photosynthesis rate at DS-20 at all the SA concentrations. Combination of sunflower

hybrids × DS levels had significant interaction with maximum photosynthesis rate

of all genotypes in DS-0 being statistically similar. The lowest photosynthesis rates

were recorded for all the genotypes at DS-20 with non significant difference among

the hybrids.

Comparative results for modes of SA application (foliar spray vs. seed

soaking) indicated that increased levels of drought stress reduced the photosynthesis

rate of all the sunflower hybrids significantly and equally under both SA application

methods (Figure 7.1 a). Genotype H-3 (FH-352) differed Table 2.7.2:

Photosynthesis rate (µmol/m2/s) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


126

H-1 (NX-19012) 7.55 e 8.17 bc 8.84 a 8.21 b 8.20 A

H-2 (NX-00989) 7.57 e 7.94 cd 8.87 a 8.17 bc 8.14 A

H-3 (FH-352) 6.56 g 7.24 f 8.18 b 7.74 de 7.43 B


DS-0 (Control) 9.52 d 9.81 c 10.57 a 10.18 b 10.02 A

DS-10 (10% PEG) 6.91 g 7.79 f 8.94 e 8.01 f 7.91 B

DS-20 (20% PEG) 5.26 j 5.76 i 6.39 h 5.93 i 5.83 C


H-1 × DS-0 9.73 de 10.03 bcd 10.37 ab 10.13 bcd

10.07 A

H-1 × DS-10 7.39 jk 8.72 h 9.93 cde 8.77 h

8.70 C

H-1 × DS-20 5.53 pq 5.77 op 6.23 lmn 5.73 op

5.81 F

H-2 × DS-0 9.83 cde 10.09 bcd 10.68 a 10.20 bc

10.20 A

H-2 × DS-10 7.61 j 8.06 i 9.58 ef 8.26 i

8.38 D

H-2 × DS-20 5.28 qr 5.67 opq 6.36 lm 6.05 mno

5.84 F

H-3 × DS-0 8.99 gh 9.30 fg 10.67 a 10.20 bc

9.79 B

H-3 × DS-10 5.73 op 6.58 l 7.29 jk 7.02 k 6.66 E

H-3 × DS-20 4.96 r 5.83 nop 6.59 l 6.01 mno

5.85 F

Means (SA) 7.23 D 7.78 C 8.63 A 8.04 B



DS=Drought stress,

significantly from the other ones at DS-10 under both foliar spray and seed soaking

treatments with SA, although three genotypes showed non significant difference at

DS-0 and DS-20. Plant photosynthesis response to SA application was better at SA-

0.75 concentration, which caused better photosynthesis rate at all DS levels under

both foliar spray and seed soaking treatments with SA (Figure 2.7 b). Nonetheless,

photosynthesis rate of sunflower plants reduced with increasing DS levels, being the

lowest at the highest level of drought stress (DS-20) even with salicylic acid

treatments.

127

A major effect of moisture stress is reduction in photosynthesis, which arises

by a decrease in leaf expansion, impaired photosynthetic machinery, premature leaf

senescence and associated reduction in food production (Wahid and Rasul, 2005).

Moisture stress causes change in photosynthetic pigments and components (Anjum

et al., 2003), damaged photosynthetic apparatus (Fu and Huang, 2001), and

diminished activities of Calvin cycle enzymes, which are important causes of

reduced crop yield (Monakhova and Chernyadèv, 2002). Similarly, the present

investigation depicted that water stress significantly altered the photosynthetic

activity in sunflower hybrids exposed to drought stress in both modes of application

(Table 2.7.1 and 2.7.2). Uzunova and Zlatev (2013) reported that drought seriously

inhibited net photosynthetic rate (A) in cowpea. Studies have shown the decreased

photosynthetic activity under drought stress due to stomatal or non-stomatal

mechanisms (Del Blanco et al., 2000; Samarah et al., 2009). Drought significantly

lowers the plant internal water content that consequently reduces photosynthetic rate

(Atteya, 2003). Moisture stress significantly reduces

a. Drought stress× Hybrids

5.00

7.00

9.00

11.00

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

128

b. Drought stress× Salicylic acid

Figure 2.7: Photosynthesis rate of sunflower hybrids under various levels of


the leaf water potential and relative water content (RWC) which had marked effect

on photosynthesis rate (Siddique et al., 2000).

Sunflower hybrids H-1 and H-2 (tolerant) performed better than H-3

(sensitive) under severe water stress conditions and SA application 0.75mM

increased photosynthesis significantly in both modes of application (Fig 2.7 a, b).

Salicylic acid is involved in activation of the stress induced antioxidant system when

plants are exposed to stress, and is considered to be a hormonal substance that plays

a key role in regulating plant growth and development (Huang et al., 2008). Salicylic

acid maintains almost the same photosynthetic rate and stomatal conductance under

water stress as those of water sufficient plants. This shielding action of salicylic acid

under drought stress is associated with the reduction of transpiration rate and

enhancement of photosynthesis (Singh and Usha, 2003). Exogenous SA application

improved the growth and photosynthetic rate in wheat

5.00

7.00

9.00

11.00

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

129

under water stress (Hussein et al., 2007).

4.2.8 Stomatal Conductance

With foliar application of salicylic acid, there was significant difference

among three sunflower genotype for stomatal conductance (Table 2.8.1). The highest

value of stomatal conductance (240 mmol/m²/s) was found in H-2 (NX00989) while

the lowest (222 mmol/m²/s) in H-3 (FH-352). Foliar application of salicylic acid at

the concentration of 0.75 mM resulted significant increase in stomatal conductance.

There was significant decrease in stomatal conductance with enhanced drought stress

(PEG application). The lowest stomatal conductance

(217mmol/m²/s) was observed with application of drought stress (PEG)

Table 2.8.1: Stomatal conductance (mmol/m²/s) of sunflower hybrids under

different drought stress levels as influenced by foliar application of salicylic

acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 224 ef 233 d 240 b 228 e 231 B

H-2 (NX-00989) 234 cd 242 b 248 a 238 bc 240 A

H-3 (FH-352) 217 g 223 ef 227 e 217 g 222 C


DS-0 (Control) 243bc 248 b 254 a 245 b 248 A

DS-10 (10% PEG) 223 fg 229 e 237 d 226 ef 229 B

DS-20 (20% PEG) 209 h 221 g 225 ef 213 h 217 C


H-1×DS-0 241 d-f 245 c-f 251 abc 241 d-f 244 B

H-1×DS-10 224 nop 231 h-n 238 f-i 228 j-o 230 D

H-1×DS-20 206 rs 224 nop 232 h-n 214 qr 219 E

H-2×DS-0 249 b-e 256 ab 259 a 253 abc 254 A

H-2×DS-10 231 i-o 235 g-m 248 b-e 234 h-m 237 C

H-2×DS-20 221 opq 235 g-m 237 f-j 227 k-o 230 D

H-3×DS-0 240 e-i 244 c-g 250 a-d 241 d-f 243 B

H-3×DS-10 213 qr 222 n-q 225 l-p 217 pq 219 E

130

H-3×DS-20 199s 204 rs 206 rs 199 s 202 F

Means (SA) 225 C 233 B 239 A 228 C



DS=Drought stress,

at 20% concentration, while the highest value (248 mmol/m²/s) was in control.

Interactive effect of sunflower genotypes with salicylic acid concentrations (H ×

SA) was significant with maximum stomatal conductance (248 mmol/m²/s) in

NX00989 hybrid at 0.75 mM salicylic acid application. Minimum stomatal

conductance (217mmol/m²/s) was observed in H-3 (FH-352) without salicylic acid

application (control). The drought level × salicylic acid, and the hybrid × drought ×

salicylic acid interactions also had significant differences for various combinations.

Statistically higher stomatal conductance (259 mmol/m²/s) was recorded in

sunflower genotype NX-00989 without stress under 0.75 mM salicylic acid

concentration, while smallest value of stomatal conductance (199 mmol/m²/s) was

in FH-352 genotype at 20 % drought stress (PEG) without salicylic acid

application.

Data of stomatal conductance under the other set of experiment for seed

soaking of sunflower hybrids with salicylic acid at different DS levels is presented

in Table 2.8.2. Sunflower hybrid H-2 (NX-00989) performed better than others and

differed significantly with H-3 (FH-352). Higher levels of drought stress also

rendered significant reduction in stomatal conductance, and the smallest value was

with DS-20 (213 mmol/m²/s). The SA-0.75 caused significantly greater stomatal

conductance (237 mmol/m²/s) as compared to that with other SA levels. Interaction

between various levels of drought stress and salicylic acid was statistically

significant. The highest value of stomatal conductance (260 mmol/m²/s) was

131

recorded under DS-0 (Control) receiving 0.75 mM concentration of salicylic acid,

being statistically different from other SA levels in DS-0. Whereas, the smallest

value (204 mmol/m²/s) was recorded under DS-20 without SA or combined with

Table 2.8.2: Stomatal conductance (mmol/m²/s) of sunflower hybrids under

different drought stress levels as influenced by seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 220 de 230 c 237 ab 219 e 227 AB

H-2 (NX-00989) 223 d 230 c 240 a 219 e 228 A

H-3 (FH-352) 218 e 230 c 234 b 219 e 225 B


DS-0 (Control) 243 c 250 b 260 a 230 d 245 A

DS-10 (10% PEG) 215 g 224 e 228 d 220 f 221 B

DS-20 (20% PEG) 204 h 218 f 224 e 206 h 213 C


H-1×DS-0 243 de 248 cd 263 a 230 ghi 246 A

H-1×DS-10 215 nop 226 h-k 230 ghi 221 klm 223 C

H-1×DS-20 203 r 216 nop 220 lmn 206qr 211 F

H-2×DS-0 246 cd 250 bc 263 a 227 hij 246 A

H-2×DS-10 217 m-p 227 hij 232 gh 222 j-m 224 C

H-2×DS-20 206qr 213 op 225 i-l 207 qr 213 EF

H-3×DS-0 238 df 248 cd 254 b 234 fg 244 B

H-3×DS-10 212 pq 218 mno 222 klm 217 mno 217 D

H-3×DS-20 203 r 225 i-l 226 h-k 206qr 215 DE

Means (SA) 220 C 225 B 237 A 224 B



DS=Drought stress,

132

1.5 mM SA, and both had significant difference with other SA levels at DS-20.

Interaction among hybrids × drought stress × salicylic acid levels was also

statistically significant. Two hybrids (H-1 and H-2) had statistically similar stomatal

conductance at each SA level with the best results in DS-0, while they had the lowest

values at DS-20. Combination of sunflower hybrids × DS levels had significant

interaction with greater stomatal conductance of all genotypes in DS-0; while the

lowest one for all the genotypes at DS-20.

Figure 2.8 a shows the comparison between two methods of SA application

on three sunflower hybrids under different DS levels and interaction with various SA

levels. Higher levels of drought stress reduced the stomatal conductance of all the

sunflower hybrids, which differed non significantly at each DS level under seed

soaking treatment with SA but significantly under its foliar spray, where H-2

(NX00989) showed better performance (Figure 2.8 a). Plant response to SA

application in this regard was better at SA-0.75 mM, which caused greater stomatal

conductance at each DS level under both foliar spray and seed soaking treatments

with SA (Figure 2.8 b). However, stress-countering effect of salicylic acid in all the

sunflower hybrids diminished at increased DS levels, as the smallest value being at

the highest level (20% PEG) of drought stress.

The present findings revealed that drought stress decreased the stomatal

conductance (gs) in sunflower hybrids in both modes of application (Table 2.8.1,

2.8.2). Similar findings were reported by Mafakheri et al. (2010) that transpiration

and stomatal conductance decreased in all three varieties of chick pea when water

stress was imposed on them. Chaves and Oliviera (2004) concluded that stomatal

conductance only affects photosynthesis at severe drought stress.

133

Figure 2.8: Stomatal conductance of sunflower hybrids under various levels of


Salicylic acid applied under stress increased the stomatal conductance in

sunflower hybrids 0.75 being the best dose in ameliorating the negative effect of

stress (Fig 2.8 a, b). Results are in harmony with the previous studies that salicylic

acid maintains almost the same stomatal conductance under water stress as those of


200

220

240

260

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)


200

220

240

260

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

134

water sufficient plants (Singh and Usha, 2003). Application of salicylic acid at the

rate of 10-3 or 10-5 mol/L to corn and soybean plants under drought stress increased

or unchanged the stomatal conductance level and transpiration rate (Khan et al.,

2003). Hamada and Al-Hakimi (2001) reported that soaking of wheat grain in 100

ppm SA was generally effective in reducing the drought effects on growth and

transpiration rate.

4.2.9 Relative Water Content

Under the first set of experiment on foliar application of salicylic acid, it was

observed that each sunflower hybrid differed significantly with the other for its

relative water content (RWC) as evident from Table 2.9.1. Genotype H-1 (NX19012)

had the maximum RWC (79.5 %) while the lowest RWC (71.3%) was in H-3 (FH-

352). Employing different levels of drought stress resulted in statistically significant

reduction in RWC from 90 % in control to 64.3 % with 20% stress (PEG).

Application of various concentrations of salicylic acid also showed significant effect

on RWC. The highest RWC (77.1 %) was observed with salicylic acid application

at 0.75mM while the smallest value (74.0 %) was recorded in control. There was

significant interaction between genotype and salicylic acid concentrations.

Maximum RWC (81.2 %) was observed in H-1 (NX-19012) receiving 0.75 mM

salicylic acid, while minimum RWC (68.2 %) was observed in genotype FH-352

without salicylic acid application. The interactions between Table 2.9.1: Relative

water content (%) of sunflower hybrids under different drought stress levels as

influenced by foliar application of salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 78.5 abc 79.7 ab 81.2 a 78.7 abc 79.5 A

135

H-2 (NX-00989) 75.2 bcd 75.6 a-d 76.8 a-d 75.6 a-d 75.8 B

H-3 (FH-352) 68.2 e 72.2 de 73.2 cde 71.4 de 71.3 C


DS-0 (Control) 88.9 a 90.4 a 90.8 a 89.8 a 90.0 A

DS-10 (10% PEG) 71.0 bc 72.1 bc 74.1 b 71.9 bc 72.3 B

DS-20 (20% PEG) 62.1 d 65.0 d 66.3 cd 64.0 d 64.3 C


H-1×DS-0 90.9 ab 90.6 ab 90.2 ab 89.6 ab 90.3 A

H-1×DS-10 77.7 c-g 79.0 b-f 82.0 a-e 77.4 d-g 79.0 B

H-1×DS-20 67.0 f-j 69.6 f-i 71.5 e-h 69.0 f-i 69.3 CD

H-2×DS-0 91.1 a 90.3 ab 91.5 a 90.9 ab 90.9 A

H-2×DS-10 71.3 e-h 72.3 e-h 72.0 e-h 71.8 e-h 71.9 C

H-2×DS-20 63.0 hij 64.2 hij 66.9 g-j 64.2 hij 64.6 D

H-3×DS-0 84.6 a-d 90.3 ab 90.7 ab 89.1 a-d 88.7 A

H-3×DS-10 63.8 hij 65.0 hij 68.4 f-i 66.4 g-j 65.9 D

H-3×DS-20 56.2 j 61.2 hij 60.5 hij 58.9 ij 59.2 E

Means (SA) 74.0 B 75.8 AB 77.1 A 75.3 AB



DS=Drought stress,

salicylic acid × drought stress (PEG) and among Hybrid × salicylic acid × Drought

stress were also significant. The highest RWC (91.5 %) was recorded in sunflower

genotype H-2 (NX-00989) without water stress under 0.75 mM application, while

the lowest RWC (56.2 %) was observed in FH-352 genotype at 20% DS without any

application of salicylic acid. All genotypes showed statistically similar RWC at each

DS or SA level.

Relative water content in sunflower plants under second set of experiment

through seed soaking with salicylic acid at different DS levels are shown in Table

136

2.9.2. The sunflower hybrid H-1 (NX-19012) had statistically greater RWC than in

other hybrids, which differed non significantly with each other. The SA at 0.75 mM

produced significantly higher RWC (78.1 %) as compared to other SA levels.

Different levels of drought stress also caused significant reduction in RWC, as the

lowest with DS-20 (65.8 %). Interaction between various levels of drought stress and

salicylic acid was statistically significant. Plants with the highest RWC (90.8 %)

were found under DS-0 (Control) receiving 0.75 mM concentration of salicylic acid,

although statistically similar to that with other SA levels in DS-0. Whereas, the

lowest RWC (62.1 %) was recorded in DS-20 without SA. Interaction among hybrids

× drought levels × salicylic acid concentrations was also statistically significant. All

the three hybrids had statistically similar RWC at each SA concentration with the

best results in DS-0, while they gave the lowest RWC at DS-20 being. Interaction of

sunflower hybrids × DS levels was also statistically significant with maximum RWC

of all genotypes in DS-0 being statistically similar. The lowest RWC were recorded

for all the genotypes at DS-20 with significant difference among the hybrids.

In Figure 2.9 is given the comparison of SA application methods for effect

of DS levels on different hybrids and interaction with various SA levels with respect

to RWC. Increased levels of drought stress reduced the RWC in all the sunflower

hybrids, which differed significantly at higher DS levels under both foliar spray and

seed soaking treatments with SA (Figure 2.9 a). The H-1 (NX19012) showed

significantly higher RWC at DS-10 and DS-20. Response of sunflower hybrids to

DS levels was slightly better with SA-0.75 application under both foliar spray and

seed soaking treatments with SA (Figure 2.9 b). Influence of salicylic acid on RWC

in sunflower plants became more prominent with increasing DS levels, showing

wider difference among SA concentrations at the highest level of drought stress (DS-

20).

137

Various forces acting through soil plant atmospheric continuum, which allow

the uptake and loss of water, constitute the water relations. The present findings

revealed that drought stress decreased the relative water content in sunflower hybrids

in both modes of application (Table 2.9.1 and 2.9.2).

Generally, leaf water potential decreases with water stress intensity (Galle et

al., 2002). Plant water relations are greatly influenced by relative water content, leaf

water potential, stomatal resistance, rate of transpiration, leaf temperature and

canopy temperature (Siddique et al., 2001). Moisture stress significantly reduces the

leaf water potential and relative water content (Siddique et al., 2000). Drought Table

2.9.2: Relative water content (%) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 77.2 bcd 80.1 ab 82.0 a 79.9 abc 79.8 A

H-2 (NX-00989) 73.1 f 74.6 def 75.7 def 73.7 ef 74.3 B

H-3 (FH-352) 68.9 g 73.3 f 76.6 cde 73.5 ef 73.1 B


DS-0 (Control) 89.2 a 89.9 a 90.4 a 88.9 a 89.6 A

DS-10 (10% PEG) 68.6 d 70.5 cd 74.6 b 73.0 bc 71.7 B

DS-20 (20% PEG) 61.3 f 67.5 de 69.4 d 65.1 e 65.8 C


H-1×DS-0 88.5 ab 88.4 ab 89.4 a 88.2 ab 88.7 A

H-1×DS-10 76.9 def 79.0 cde 82.9 bc 80.4 cd 79.8 B

H-1×DS-20 66.2 h-l 73.0 fg 73.8 efg 71.1 fgh 71.0 C

H-2×DS-0 90.5 a 91.2 a 90.7 a 89.0 a 90.3 A

H-2×DS-10 66.1 h-l 67.9 g-k 70.8 ghi 69.2 g-j 68.5 CD

138

H-2×DS-20 62.6 kl 64.5 jkl 65.6 h-l 62.9 kl 63.9 EF

H-3×DS-0 88.7 ab 90.0 a 91.0 a 89.5 a 89.8 A

H-3×DS-10 62.8 kl 64.7 jkl 70.1 g-j 69.4 g-j 66.7 DE

H-3×DS-20 55.2 m 65.0 i-l 68.8 g-j 61.5 l 62.6 F

Means (SA) 73.1 C 76.0 B 78.1 A 75.7 B



DS=Drought stress,


55.0

65.0

75.0

85.0

95.0

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

139

Figure 2.9: Relative water content of sunflower hybrids under various levels of


significantly altered the plant internal water status by lowering water potential and

relative water content of corn (Atteya, 2003). Saensee et al. (2012) tested seven

sunflower genotypes and one commercial hybrid, using PEG-6000. Along with other

parameters, relative water content decreased significantly in all sunflower genotypes

with increase in water stress levels. Ullah et al. (2012) and Nayyar and Gupta (2006)

reported a decrease of relative water content in response to drought

stress.

SA increased the RWC more in tolerant hybrids with 0.75mM conc. (Fig 2.9

a, b) in both modes of application. It has been concluded that SA triggers some

metabolic processes in plants as well as affects plant water relations (Hayat et al,

2010). In this study, with that observed in wheat (Singh and Usha, 2003) and

Ctenanthe setosa plants grown under drought conditions (Kadioglu et al., 2011) and

in shallot plants Ahmad et al. (2014), the results showed that sunflower plants

sprayed with SA solution could maintain higher RWC compared with those of


55.0

65.0

75.0

85.0

95.0

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

140

drought stressed plants. It becomes evident that foliar treatment of SA and

LTryptophan had significant effect on RWC under osmotic stress enabling the plants

to maintain turgor, carry on photosynthetic activities (Rao et al., 2012).

4.2.10 WaterPotential

Results of the experiment on foliar application of various salicylic acid

concentrations under different drought stress levels indicated that three sunflower

hybrids differed significantly with respect to water potential (Table 2.10.1).

Genotype FH-352 had statistically lower water potential (0.60 -MPa) compared to

other two hybrids with high value (0.53-MPa) in H-1 non significantly different

effect on water potential. The maximum water potential (0.50 -MPa) was

Table 2.10.1: Water potential (-MPa) of sunflower hybrids under different



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.59 b 0.53 cd 0.48 de 0.53 cd 0.53 B

H-2 (NX-00989) 0.59 b 0.52 cd 0.47 e 0.52 cde 0.52 B

H-3 (FH-352) 0.65 a 0.60 ab 0.56 bc 0.60 ab 0.60 A


DS-0 (Control) 0.46 e 0.39 f 0.34 a 0.38 f 0.39 C

DS-10 (10% PEG) 0.65 bc 0.61 bc 0.55 d 0.60 cd 0.61 B

DS-20 (20% PEG) 0.72 f 0.65 bc 0.61 bc 0.66 b 0.66 A


H-1×DS-0 0.40 klm 0.33 lmn 0.27 n 0.32 lmn 0.33 E

H-1×DS-10 0.65 a-e 0.61 c-h 0.55 e-i 0.61 c-h 0.61 BC

H-1×DS-20 0.73 ab 0.65 a-e 0.62 c-h 0.67 a-d 0.67 A

H-2×DS-0 0.42 jkl 0.32 lmn 0.26 n 0.31 mn

0.33 E

H-2×DS-10 0.65 a-e 0.60 c-h 0.54 ghi 0.59 d-i 0.60 C

H-2×DS-20 0.70 abc 0.64 a-g 0.60 c-i 0.65 a-f 0.65 AB

H-3×DS-0 0.54 f-i 0.52 hij 0.49 ijk 0.52 hij 0.52 D

141

H-3×DS-10 0.66 a-e 0.62 b-h 0.56 e-i 0.61 c-h 0.61 BC

H-3×DS-20 0.74 a 0.66 a-e 0.62 b-h 0.67 a-d 0.67 A

Means (SA) 0.61 A 0.55 B 0.50 C 0.55 B



DS=Drought stress,

Table 2.10.2: Water potential (-MPa) of sunflower hybrids under different



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.49 b 0.42 d 0.37 d 0.42 d 0.42 C

H-2 (NX-00989) 0.53 a 0.46 bc 0.41 d 0.46 bc 0.47 AB

H-3 (FH-352) 0.55 a 0.49 b 0.44 cd 0.46 bc 0.48 A


DS-0 (Control) 0.42d 0.35 e 0.30 f 0.34 e 0.36 C

DS-10 (10% PEG) 0.53b 0.49 c 0.42 d 0.49 c 0.48 B

DS-20 (20% PEG) 0.61a 0.53b 0.50 c 0.51b 0.54 A


H-1 × DS-0 0.42 ij 0.31 nop 0.26 p 0.31 nop 0.33 D

H-1 × DS-10 0.49 e-h 0.45 gh 0.38kl 0.45 gh 0.44 C

H-1 × DS-20 0.55 bc 0.51 c-f 0.45 gh 0.51 c-f 0.51 B

H-2 × DS-0 0.41 ijk 0.33 lmn 0.27 op 0.32 mno 0.33 D

H-2 × DS-10 0.55 bc 0.50 c-f 0.43 hij 0.50 c-f 0.50 B

H-2 × DS-20 0.64 a 0.54 b-e 0.53 b-e 0.56 bc 0.57 A

H-3 × DS-0 0.44 ghi 0.41 ijk 0.37 klm 0.40jk 0.41 C

H-3 × DS-10 0.56 bc 0.51 c-f 0.44 ghi 0.51 c-f 0.50 B

H-3 × DS-20 0.64 a 0.55 bc 0.51 c-f 0.47 e-g 0.54 A

Means (SA) 0.52 A 0.46 B 0.41 C 0.45 B



142

DS=Drought stress,

recorded with 0.75 mM salicylic acid application while minimum water potential

(0.61 -MPa) was observed in control plants without SA application. Enhanced levels

of PEG resulted in significant decrease in water potential from 0.39 -MPa in control

to 0.66 -Mpa with 20% PEG. There was significant interaction between genotypes

and salicylic acid concentrations. Maximum water potential (0.47 -Mpa) was

observed in genotype H-2 (NX-00989) with salicylic acid application at the

concentration of 0.75 mM, while minimum water potential (0.65 -Mpa) was

observed in H-3 (FH-352) without any application of salicylic acid. The interactions

between salicylic acid × DS levels, and among hybrids × salicylic acid × PEG were

also significant. The highest water potential (0.26 -Mpa) was observed with NX-

00989 genotype without PEG under 0.75mM salicylic acid concentration. The

lowest value of water potential (0.74 -Mpa) was recorded with FH-352 sunflower

genotype at 20% PEG application rate without any application of

salicylic acid.

Water potential data obtained from another experiment on seed soaking of

sunflower hybrids with salicylic acid at different DS levels are expressed in Table

2.10.2. Three sunflower hybrids differed significantly from each other with the

highest (less negative) water potential in H-1 (NX-19012) and the more negative

value (0.48 -Mpa) in H-3. The SA-0.75 produced the plants with significantly higher

water potential (0.41 -Mpa) as compared to other SA concentrations. Different levels

of drought stress also caused significant decrease in water potential, as the highest

water potential was with DS-0 (0.36 -Mpa). Interaction between various levels of

drought stress and salicylic acid concentrations was statistically significant. The

lowest water potential (0.61 -Mpa) was found under

143

Figure 2.10: Water potential of sunflower hybrids under various levels of


DS-20 (control) without receiving salicylic acid, while 0.75mM SA reduced

the water potential significantly as compared to other SA concentrations at all DS

levels. So, the less negative water potential (0.30 Mpa) was recorded in DS-0

combined with 0.75 mM SA. Interaction among hybrids × drought levels × salicylic

acid concentrations was also statistically significant. Combination of sunflower


0.30

0.40

0.50

0.60

0.70

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)


0.30

0.40

0.50

0.60

0.70

0.80

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

144

hybrids × DS levels had significant interaction with minimum water potential of all

genotypes in DS-20 being statistically similar in H-2 and H-3.The highest values of

water potential were recorded for the H-1 (NX-19012) genotype at all DS-10 and

DS-20 levels having significant difference with other two hybrids.


hybrids and interaction with various SA levels is shown in Figure 2.10. Enhanced

levels of drought stress lowered the water potential in all the sunflower hybrids,

which differed non significantly at all higher DS levels under both methods of SA

application (Figure 2.10 a). However, H-1 (NX-19012) showed slightly higher water

potential through seed soaking treatments with SA. Plant response to SA application

was better at SA-0.75 mM, which produced plants with higher water potential at all

the DS levels under both foliar spray and seed soaking treatments of

SA (Figure 2.10 b).

The present findings revealed that drought stress decreased the water potential

in sunflower hybrids in both modes of application (Table 2.10.1 and 2.10.2). Leaf

water potential greatly influences the plant water relations (Siddique et al., 2001).

Generally, leaf water potential decreases with water stress intensity

(Galle et al., 2002). Moisture stress significantly reduces the leaf water potential

(Siddique et al., 2000). Similarly, drought significantly alters the plant internal water

status by lowering water potential (Atteya, 2003).

Exogenous application of SA on imposition of water stress maintained the

water potential level by increasing its value. SA application altered proline

metabolism significantly, leading to the maintenance of the turgor by accumulating

significant higher levels of proline content in sunflower, supporting its protection

145

from drought stress (Fig 2.10 a, b). Results are concurrent with previous findings,

exogenous application of plant growth regulators under drought conditions increase

the water potential, and improve chlorophyll content (Zhang et al., 2004). Under

drought stress exogenous application of SA, by accumulating significant higher

levels of proline content in shallot maintain turgor (Ahmad et al., 2014).

4.2.11 Osmotic Potential

Three sunflower genotypes differed significantly in respect of osmotic potential by

foliar application of salicylic acid (Table 2.11.1). The highest osmotic potential (less

negative) (1.41 -MPa) was recorded in H-3 (FH-352) and the lowest (more negative)

(1.53 -MPa) was recorded in H-2 (NX-98900).There was significant decrease in

osmotic potential with higher levels of drought stress. The lowest osmotic potential

(1.42 -MPa) was recorded in control, while the highest osmotic potential (1.50 -MPa)

was with DS-20. Salicylic acid ×genotype interaction was significant with lowest

(more negative) osmotic potential (1.54 -MPa) in H-2 (NX-

00989) treated with 0.75 mM salicylic acid concentration.

Table 2.11.1: Osmotic potential (-MPa) of sunflower hybrids under different



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 1.44 cd 1.46 bcd 1.49 bc 1.46 bcd 1.46 B

H-2 (NX-00989) 1.51 ab 1.53 a 1.54 a 1.52 ab 1.53 A

H-3 (FH-352) 1.40 e 1.41 de 1.43cd 1.40 ef 1.41 C


DS-0 (Control) 1.40 de 1.42 d 1.45 c 1.42 d 1.42 C

DS-10 (10% PEG) 1.46 c 1.47 bc 1.49 b 1.47 bc 1.47 B

DS-20 (20% PEG) 1.49 b 1.50 ab 1.52 a 1.50 ab 1.50 A

146


H-1×DS-0 1.37NS 1.40 1.45 1.41 1.41 D

H-1×DS-10 1.45 1.46 1.49 1.47 1.47 C

H-1×DS-20 1.50 1.51 1.54 1.51 1.52 B

H-2×DS-0 1.47 1.49 1.51 1.48 1.49 C

H-2×DS-10 1.50 1.52 1.53 1.52 1.52 B

H-2×DS-20 1.55 1.57 1.58 1.57 1.57 A

H-3×DS-0 1.36 1.38 1.40 1.37 1.38 DE

H-3×DS-10 1.42 1.43 1.44 1.42 1.43 CD

H-3×DS-20 1.42 1.42 1.44 1.42 1.43 CD

Means (SA) 1.45 C 1.46 B 1.49 A 1.46 B



DS=Drought stress

Highest (less negative) osmotic potential (1.40 -MPa) was observed in H-3 (FH352)

without salicylic acid application. Drought stress levels × salicylic acid

concentrations as well as the hybrids × drought × salicylic acid interactions were non

significant.

Results of osmotic potential under second set of experiment on seed soaking

of sunflower hybrids with salicylic acid at different DS levels are shown in Table

2.11.2. Sunflower hybrids differ significantly with each other, as H-2 (NX00989)

gives more negative values than other two hybrids. The SA-0.75 mM produced

plants with significantly lower osmotic potential (1.46 -MPa) as compared to other

SA concentrations. Higher levels of drought stress also caused significant decrease

in osmotic potential, as the lowest one was with DS-20 (1.46 MPa). Interaction

between various levels of drought stress and salicylic acid concentrations was

statistically significant. More negative osmotic potential (1.48 MPa) was found

147

under DS-20 receiving 0.75 mM concentration of salicylic acid, although statistically

similar to same SA concentration in DS-10. Whereas, less negative osmotic potential

(1.37 -MPa) was recorded in DS-0 + 1.5 SA non significantly different with DS-0

without SA. Interaction among hybrids × drought levels × salicylic acid

concentrations was statistically non significant. Combination of sunflower hybrids ×

DS levels had significant interaction with higher osmotic potential of two genotypes

(H-1 and H-2) in DS-20. Whereas, the maximum osmotic potentials in all the

genotypes were recorded at DS-0 with non significant difference between H-1 and

H-3 hybrids are shown in Figure 2.11. Enhanced levels of drought stress decreased

the osmotic potential of the sunflower hybrids in

Table 2.11.2: Osmotic potential (-Mpa) of sunflower hybrids under different



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 1.39 NS 1.41 1.43 1.37 1.40 C

H-2 (NX-00989) 1.50 1.51 1.54 1.51 1.52 A

H-3 (FH-352) 1.43 1.45 1.47 1.44 1.45 B


DS-0 (Control) 1.38 ij 1.41 gh 1.43 efg 1.37 j 1.40 C

DS-10 (10% PEG) 1.42fgh 1.44 def 1.47 abc 1.44 def 1.44 B

DS-20 (20% PEG) 1.44 def 1.46 bcd 1.48 a 1.45 cde 1.46 A


H-1×DS-0 1.35 NS 1.38 1.40 1.37 1.38 CD

H-1×DS-10 1.39 1.40 1.42 1.40 1.40 C

H-1×DS-20 1.42 1.44 1.46 1.43 1.41 C

H-2×DS-0 1.45 1.46 1.49 1.46 1.47 B

H-2×DS-10 1.51 1.53 1.56 1.52 1.53 A

H-2×DS-20 1.53 1.55 1.56 1.54 1.55 A

H-3×DS-0 1.34 1.36 1.38 1.35 1.36 D

148

H-3×DS-10 1.37 1.40 1.42 1.39 1.40 C

H-3×DS-20 1.38 1.40 1.42 1.39 1.40 C

Means (SA) 1.42 C 1.44 B 1.46 A 1.43 B



DS=Drought stress



Figure 2.11: Osmotic potential of sunflower hybrids under various levels of


1.30

1.35

1.40

1.45

1.50

1.55

1.60

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20


H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

1.30

1.35

1.40

1.45

1.50

1.55

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20


SA-0 SA-0.375 SA-0.75 SA-1.50

149

both foliar spray and seed soaking treatments with SA (Figure 2.11 a). Sunflower

genotype H-1 and H-2 exhibited progressively decreased osmotic potential with

enhanced DS levels, giving lower values at DS-20 compared to H-3 hybrid with

foliar spray. Genotypic response to SA application at SA-0.75 produced plants with

lower osmotic potential at all the DS levels under both foliar spray and seed soaking

treatments with SA (Figure 2.11 b) and the less negative osmotic potential values

were under SA-0 under each level of DS.

Uptake and loss of water influence the osmotic potential in plants (Galle et al.,

2002). The present findings revealed that drought stress decreased the osmotic

potential in sunflower hybrids in both modes of application (Table 2.11.1 and

2.11.2). Similar with the findings that salt stress significantly decreased (more

negative values) the leaf osmotic potential of all lines of safflower (Siddique and

Ashraf, 2008). Salinity and water shortage deduct the proline accumulation in

seedlings which regulate osmotic pressure (Inal, 2002).

SA decreased the osmotic potential more in tolerant hybrids with 0.75 mM

(Fig 2.11 a, b) in both modes of application. Osmotic potential in wheat cultivars

decreased by about 98% after PEG-6000 (0.75 −MPa) and treatment of SA (0.05)

mitigated the effects of stress (Marcińska et al., 2013).

4.2.12 Turgor Potential

Under the experiment on foliar application of salicylic acid, there was

significant difference between sunflower genotypes in respect of turgor potential

(Table 2.12.1). Higher turgor potential was observed in two genotypes (NX-00989

Table 2.12.1: Turgor potential (MPa) of sunflower hybrids under different


150


SA-0 SA0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.85 d 0.93 bc 1.01 ab 0.93 bc 0.93 A

H-2 (NX-00989) 0.95 bc 1.01 ab 1.07 a 1.00 ab 1.01 A

H-3 (FH-352) 0.75e 0.81 de 0.87 cd 0.80de 0.83 B


DS-0 (Control) 0.95 c 1.03 b 1.11 a 1.04 b 1.03A

DS-10 (10% PEG) 0.83 d 0.86 d 0.94 c 0.86 d 0.87 B

DS-20 (20% PEG) 0.78 f 0.85 d 0.91 c 0.84 d 0.84 B


H-1×DS-0 0.97 NS 1.07 1.18 1.09 1.08 B

H-1×DS-10 0.80 0.85 0.94 0.86 0.86 D

H-1×DS-20 0.77 0.86 0.92 0.84 0.85 D

H-2×DS-0 1.05 1.17 1.25 1.17 1.16 A

H-2×DS-10 0.92 0.92 0.99 0.92 0.94 C

H-2×DS-20 0.88 0.93 0.98 0.92 0.93 C

H-3×DS-0 0.82 0.86 0.91 0.85 0.86 D

H-3×DS-10 0.76 0.81 0.88 0.81 0.82 D

H-3×DS-20 0.68 0.76 0.82 0.75 0.75 E

Means (SA) 0.85 C 0.92 B 0.98 A 0.91 B



DS=Drought stress,

and NX-19012) while the lowest (0.83 Mpa) in FH-352. Application of salicylic acid

at the concentration of 0.75 mM resulted in significant increase in turgor potential

over the control and other treatments. There was significant decrease in turgor

potential with the application of PEG at 10%, which was non significantly different

with PEG application at the concentration of 20%. The lowest turgors potential (0.84

Mpa) was observed in DS-20 while the highest turgor potential (1.03 MPa) was

151

observed with in control. Salicylic acid × genotype and Drought level × salicylic

acid interaction were significant with maximum turgor potential

(1.07 Mpa) observed in NX-00989 with 0.75 mM salicylic acid application.

Minimum turgor potential (0.75 Mpa) was observed in FH-352 without salicylic acid

application and the hybrid × drought × salicylic acid interaction was non significant.

The highest turgor potential (1.25 Mpa) was recorded in sunflower genotype NX-

00989 without PEG under the application of 0.75mM salicylic acid concentration.

While the lowest turgor potential (0.68 Mpa) was observed in FH-

352 genotype at 20% PEG without salicylic acid application.

Results of plant turgor potential under second set of experiment on seed

soaking of sunflower hybrids with salicylic acid at different DS levels are shown in

Table 2.12.2. Here, three sunflower hybrids differed significantly from each other

exhibiting the highest value in H-2 (NX-00989) and the lowest for H-3 (FH-352).

The SA-0.75mM concentration produced plants with significantly higher turgor

potential (1.05 Mpa) as compared to other SA levels. Enhanced levels of drought

stress caused significant decrease in turgor potential, as the lowest value (0.92 Mpa)

was with DS-20 level. Interaction between various levels of drought stress Table

2.12.2: Turgor potential (Mpa) of sunflower hybrids under different drought



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 0.90 de 0.98 cd 1.06 abc 0.98 cd 0.98 B

H-2 (NX-00989) 0.96 d 1.06 abc 1.13 a 1.05 bcd 1.05 A

H-3 (FH-352) 0.82 f 0.90de 0.97 cd 0.89 ef 0.90 C


DS-0 (Control) 0.96 cd 1.05 b 1.12 a 1.05 b 1.05 A

152

DS-10 (10% PEG) 0.89 f 0.96 cd 1.05 b 0.95 d 0.96 B

DS-20 (20% PEG) 0.83 g 0.93 de 0.98 c 0.92 e 0.92 BC


H-1×DS-0 0.93 NS 1.07 1.14 1.06 1.05 B

H-1×DS-10 0.90 0.95 1.04 0.95 0.96 E

H-1×DS-20 0.87 0.93 1.01 0.92 0.93 A

H-2×DS-0 1.04 1.13 1.22 1.14 1.13 A

H-2×DS-10 0.96 1.03 1.13 1.02 1.04 C

H-2×DS-20 0.89 1.01 1.03 1.00 0.98 D

H-3×DS-0 0.90 0.95 1.01 0.95 0.95 F

H-3×DS-10 0.81 0.89 0.98 0.88 0.89 G

H-3×DS-20 0.74 0.85 0.91 0.85 0.84 H

Means (SA) 0.89 C 0.98 B 1.05 A 0.97 B



DS=Drought stress,

and salicylic acid was statistically significant. The greatest turgor potential (1.12

Mpa) was found under DS-0 (control) receiving 0.75 mM concentration of salicylic

acid whereas, the smallest turgor potential (0.83 Mpa) was recorded under DS-20

without SA, which had significant difference with other SA levels. Interaction

among hybrids × drought levels × salicylic acid levels was statistically non

significant. All the three hybrids had statistically different turgor potential at various

SA levels with lower values mostly under DS- 20, while they got the highest turgor

potential at DS-0. Combination of sunflower hybrids × DS levels had significant

interaction with maximum turgor potential in H-1 and H-2 genotypes under DS-0;

while in H-3 the lowest turgor potential was recorded at

DS-20.

153


hybrids and interaction with various SA levels is shown in Figure 2.12. Enhanced

levels of drought stress decreased the turgor potential of sunflower hybrids H-1, H2

and H-3 especially at DS-20 showing lower values for both foliar spray and seed

soaking treatments with SA. Plant response to SA application was better at SA0.75,

which produced plants with significantly higher turgor potential at all DS levels

under both foliar spray and seed soaking treatments with SA (Figure 2.12 b). Foliar

spray caused a higher turgor potential as compared to that with seed soaking

treatments of SA. However, remedial effect of salicylic acid on sunflower plants

reduced with increasing DS levels, being lower at the highest level of drought stress

(DS-20) under seed soaking of SA.


0.40

0.60

0.80

1.00

1.20

1.40

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

154

Figure 2.12: Turgor potential of sunflower hybrids under various levels of


Plant water relations are greatly influenced by relative water content, leaf


canopy temperature (Siddique et al., 2001). Water potential, osmotic potential,

turgor potential and relative water contents are the components of plant. The results

of present study showed that water stress in both modes of application significantly

reduced the leaf turgor potential (Tables 2.12.1, 2.12.2). Generally, leaf water

potential decreases with water stress intensity (Galle et al., 2002). Osmotic

adjustment resulted from the accumulation of solutes within cell, lowers the osmotic

potential and helps maintain turgor of plants experiencing water stress (Ludlow and

Muchow, 1990). A significant reduction in leaf turgor potential was observed in all

lines due to salt stress in Safflower (Siddique and Ashraf, 2008).

Maintenance of turgor potential plays an important role in drought tolerance

of plants which may be due to its involvement in stomatal regulation and hence

photosynthesis (Ludlow, 1985). Salicylic acid applied under stress increased the


0.40

0.60

0.80

1.00

1.20

1.40

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

155

turgor potential significantly among hybrids 0.75 being the best dose in reducing the

negative effect of stress (Fig 2.12 a, b). Drought stress decreased leaf relative water

contents, water potential, osmotic potential, turgor pressure, by applying

glycinebetaine and salicylic acid exogenously improves water relations of sunflower

under water stress conditions (Hussain et al., 2009). Exogenous application of SA,

by accumulating significant higher levels of proline content in shallot maintains

turgor under water deficit conditions (Ahmad et al., 2014).

4.2.13 Leaf Proline Content

The leaf proline of three sunflower hybrids differed significantly from each other

when plants of sunflower were sprayed with salicylic acid (Table 2.13.1).

Genotype NX-00989 (H-2) had the highest proline (545 µg/g) while genotype H-3

(FH-352) had lowest (410 µg/g). Application of different concentrations of salicylic

acid had significant effect on proline accumulation. Highest proline (535

µg/g) was recorded with 0.75 mM SA and smallest (465 µg/g) in the control.

Application of drought stress (PEG) levels significantly increased proline from173

µg/g in control to753 µg/g in 20% stress (PEG). There was significant interaction

between genotype and salicylic acid treatments. Maximum proline (609 µg/g) was

recorded in the genotype NX-00989 receiving 0.75 mM SA while minimum (391

µg/g) was found in genotype FH-352 without salicylic acid application. The

interactions salicylic acid × Drought stress (PEG) and Hybrid × salicylic acid ×

Drought stress were also statistically significant. Highest proline (966 µg/g) was

recorded in the sunflower hybrid; NX-00989 with 20% drought stress and 0.75 mM

salicylic acid. While the lowest (124 µg/g) in FH-352 without any application of

stress and salicylic acid.

156

Proline concentrations in sunflower hybrids plants with SA seed soaking

under DS levels are shown in Table 2.13.2. The hybrids differ significantly with each

other; 0.75 mM SA caused significantly greater proline accumulation in leaf

(453µg/g) as compared to that of other SA levels. There was significant rise in

proline accumulation with increasing levels of drought stress, as the highest (678

µg/g) with DS-20. Interaction between drought levels and salicylic acid was

statistically significant. The greatest proline contents were found under DS-20

receiving 0.75 mM SA, and statistically significant to other SA levels in any of the

Table 2.13.1: Leaf proline concentration (µg/g of fresh weight) of sunflower

hybrids under different drought stress levels as influenced by foliar



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 496 d 507 d 554 b 506 d 516 B

H-2 (NX-00989) 506 d 521 c 609 a 544 b 545 A

H-3 (FH-352) 391 g 400 fg 443 e 407 f 410 C


DS-0 (Control) 133 h 159 g 237 f 164 g 173 C

DS-10 (10% PEG) 535 e 540 e 567 d 539 e 545 B

DS-20 (20% PEG) 726 c 729 c 802 a 755 b 753 A


H-1 × DS-0 143 opq 164 no 230 m 165 no 176 F

H-1 × DS-10 562 i 568 hi 583 ghi 562 i 569 D

H-1 × DS-20 783 e 787 e 848 c 791 e 802 B

H-2 × DS-0 133 pq 171 n 253 l 172 n 182 F

H-2 × DS-10 568 hi 573 ghi 608 f 572 ghi 580 C

H-2 × DS-20 818 d 820 d 966 a 889 b 873 A

H-3 × DS-0 124 q 141 pq 227 m 155 nop 161 G

H-3 × DS-10 475 k 479 k 509 j 482 k 486 E

157

H-3 × DS-20 576 ghi 580 ghi 593 fg 585 gh 584 C

Means (SA) 465 D 476 C 535 A 486 B



DS=Drought Stress

Table 2.13.2: Leaf proline concentration (µg/g of fresh weight) of sunflower

hybrids under different drought stress levels as influenced by seed soaking in

salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 448 cd 454 bcd 492 ab 448 cd 461 B

H-2 (NX-00989) 455 bcd 460 bc 503 a 464 b 470 A

H-3 (FH-352) 329 f 331 f 363 e 334 f 340 C


DS-0 (Control) 137 g 128 g 203 f 136 g 151 C

DS-10 (10% PEG) 435 e 438 e 457 d 435 e 441 B

DS-20 (20% PEG) 660 c 679 b 698 a 675 b 678 A


H-1 × DS-0 146 l 140 lm 209 k 137 lm 158 G

H-1 × DS-10 457 hi 452 i 479 fg 451 i 460 E

H-1 × DS-20 741 d 770 bc 790 ab 757 cd 764 B

H-2 × DS-0 138 lm 124 m 209 k 138 lm 152 GH

H-2 × DS-10 466 ghi 473 gh 495 ef 466 ghi 475 D

H-2 × DS-20 760 cd 783 b 805 a 787 ab 784 A

H-3 × DS-0 125 m 122 m 191 k 133 lm 143 H

H-3 × DS-10 383 j 388 j 398 j 388 j 389 F

H-3 × DS-20 480 fg 485 efg 500 e 482 efg 487 C

Means (SA) 411 B 415 B 453 A 415 B



DS=Drought Stress

158

DS levels. Whereas, the smallest proline content was recorded in DS-0 combined

with 0.375 mM SA, which had non significant difference with other SA levels at

DS-0 except that with SA-0.75. Interaction among hybrids × drought levels ×

salicylic acid was also statistically significant. All the three hybrids had statistically

similar proline content at all the SA levels under DS-0, while they gave the higher

proline content at DS-20 but statistically dissimilar at various SA concentrations.


maximum proline content of all genotypes in DS-20 being statistically different;

while the lowest proline contents were recorded for all the genotypes at DS-0 with

significant difference between the hybrids H-1 and H-3.

Figure 2.13 exhibits comparison of SA application methods for the effect of

DS levels on sunflower hybrids and interaction with various SA levels. Increased

levels of drought enhanced the proline content in all the sunflower hybrids, which

differed significantly at higher DS levels under both SA foliar spray and seed

soaking. H-2 (NX-00989) showed significantly higher proline content at DS-20

(Figure 2.13 a). Plant response to at SA-0.75 was better, which caused greater proline

accumulation at all DS levels both under SA foliar and seed soaking treatments

(Figure 2.13 b).

Osmosis-regulating substance proline significantly increases under water

stress (Din et al., 2011). The present findings also revealed that drought stress

augmented the leaf proline in sunflower hybrids in both modes of application (Table

2.13.1, 2.13.2). Accumulation and mobilization of proline correlated with

the levels

159


Figure 2.13: Proline content of sunflower hybrids under various levels of


of tolerance towards drought stress in wheat (Nayyar and Walia, 2003).

Accumulation of proline in drought stress has been described in wheat

(Sakhabutdinova et al., 2003) and in rice (Chou et al., 1991). The free proline level

also increased (from 1.56 to 3.13 times) in response to water stress. It seems proline

may play a role in minimizing the damage caused by water deficit (Mohammadkhani

.


100

300

500

700

900

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

100

300

500

700

900

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

160

and Heidari. 2008). Bandurska and Stroinski (2003) found that water stress resulted

in proline accumulation in drought tolerant wild accessions of

Hordeuan spontaneum but not in the sensitive.

Salicylic acid application further enhanced the leaf proline more in tolerant

hybrids with 0.75mM (Fig 2.13 a, b) in both modes of application. In wheat, salicylic

acid was shown to increase the abscisic acid content, leading to the accumulation of

proline (Shakirova et al., 2003). Sayyari et al. (2013) studied the effect of SA on

lettuce (Lactuca sativa L.) in drought conditions, proline significantly increased by

SA application. El-Tayeb and Naglaa (2010) observed an increase in proline

signifying the role of SA in regulating drought response of plants.

4.2.14 Soluble Sugars

Under the first set of experiment on foliar application of salicylic acid,

sunflower genotypes differed significantly in respect of sugar content (Table 2.14.1).

Highest level of sugar (15.6 mg/g) was observed in NX-00989 while lowest (12.1

mg/g) in FH-352. Salicylic acid application enhanced significantly sugar contents

from 13.0 mg/g under control to 14.9 mg/g at the concentration of Table 2.14.1:

Soluble sugars (mg/g of fresh weight) of sunflower hybrids under different

drought stress levels as influenced by foliar application of salicylic acid


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 13.1 d 13.5 cd 14.3bc 13.2 d 13.5 B

H-2 (NX-00989) 14.5 bc 15.4 b 17.2 a 15.4 b 15.6 A

H-3 (FH-352) 11.5 e 11.9 e 13.3 cd 11.9 e 12.1 C


DS-0 (Control) 10.6 d 10.9 d 11.5 d 10.9 d 11.0 C

161

DS-10 (10% PEG) 13.0 c 14.0 c 15.7 b 13.4 c 14.1 B

DS-20 (20% PEG) 15.4 b 16.1 b 17.6 a 15.9 b 16.5 A


H-1 × DS-0 10.1 pq 10.4 m-q 10.6 pq 10.3 opq 10.3 G

H-1 × DS-10 13.8 f-k 14.4 f-j 15.2 efg 14.1 f-j 14.4 CD

H-1 × DS-20 15.3 efg 15.7 def 17.1 cde 15.1 e-h 15.8 B

H-2 × DS-0 11.4 l-q 12.0 j-q 13.0 g-n 12.6 i-o 12.3 F

H-2 × DS-10 13.7 f-l 14.9 e-i 18.0 bcd 13.9 f-j 15.1 BC

H-2 × DS-20 18.3 bc 19.3 bc 20.7 a 19.6 b 19.5 A

H-3 × DS-0 10.3 opq 10.4 m-q 10.8 m-q 10.4 m-q 10.3 G

H-3 × DS-10 11.5 k-q 12.7 h-m 14.1 f-j 12.3 j-p 12.7 EF

H-3 × DS-20 12.6 i-n 13.1 g-m 15.0 e-h 12.9 g-m 13.4 DE

Means (SA) 13.0 C 13.7 B 14.9 A 13.4 BC



DS=Drought Stress

0.75 mM. Elevated drought stress also increased significantly the sugar level.The

lowest sugar content (11.0 mg/g) was recorded in control while the highest content

(16.5 mg/g) was observed with 20% PEG (drought stress) treatment. Salicylic acid

× Hybrid interaction was significant with maximum sugar content (17.2 mg/g) in

NX-00989 with 0.75 mM salicylic acid. The smallest content of sugar (11.5 mg/g)

was observed in hybrid FH-352 without salicylic acid application. Interactions for

drought levels × salicylic acid, and sunflower hybrids × drought levels × salicylic

acid treatments were also significant. Highest sugar (20.7 mg/g) was recorded in

sunflower hybrid NX-00989 receiving DS-20 (20% PEG) and 0.75 mM salicylic

acid, while the lowest one (10.3 mg/g) in FH-352 genotype without drought stress

and SA-0.

162

Sugar contents in sunflower hybrids receiving different levels of salicylic

acid seed soaking and drought stress are shown in Table 2.14.2. Here also, the sugar

contents of sunflower hybrids differed significantly with a greater value in H2 (NX-

00989). Further, SA-0.75 treated plants accumulated higher sugar (14.2 mg/g) as

compared to other SA levels. Increased levels of drought also caused significant rise

in sugar content, as the highest value (14.0 mg/g) was under DS-20. Interaction

between various levels of drought stress and salicylic acid was statistically

significant. The highest sugar content in plants (15.6 mg/g) was found under DS-20

receiving 0.75 mM of salicylic acid, which was statistically different from other SA

concentrations at any DS level. Whereas, the smallest value of sugar content (9.6

mg/g) was recorded in DS-0 without SA, which had non significant difference with

that from SA-1.50 at DS-0. Interaction among hybrids × drought levels × salicylic

acid concentrations was also statistically significant. All the three Table 2.14.2:

Soluble Sugars (mg/g of fresh weight) of sunflower hybrids under different



SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 11.6 e 12.3 e 13.1 bcd 12.1 e 12.3 B

H-2 (NX-00989) 12.3 de 13.2 bc 15.7 a 12.5 cde 13.4 A

H-3 (FH-352) 10.6 f 12.0 e 13.9 b 11.9 e 12.1 B


DS-0 (Control) 9.6 h 11.0 g 12.5 ef 9.8 h 10.7 C

DS-10 (10% PEG) 12.0 f 12.8de 14.7 b 12.8 de 13.0 B

DS-20 (20% PEG) 12.9 de 13.6 cd 15.6 a 13.9 bc 14.0 A


H-1 × DS-0 9.5 n 10.2 lmn 10.5 lmn 9.8 n 10.0 C

H-1 × DS-10 12.2 ijk 12.8 g-k 13.7 d-h 12.5 f-k 12.8 B

H-1 × DS-20 13.1 e-j 13.8 d-h 15.2 a-d 13.9 c-h 14.0 A

163

H-2 × DS-0 11.5 klm 12.6 f-k 15.0 a-d 10.2 mn 12.3 B

H-2 × DS-10 12.3 h-k 13.2 e-j 15.8 ab 13.1 e-j 13.6 A

H-2 × DS-20 13.1 e-j 13.9 c-g 16.2 a 14.1 c-f 14.3 A

H-3 × DS-0 7.9 o 10.3 mn 12.0 jkl 9.3 no 9.9 C

H-3 × DS-10 11.4 klm 12.4 g-k 14.4 b-e 12.7 f-k 12.7 B

H-3 × DS-20 12.5 f-k 13.2 e-j 15.4 abc 13.7 d-i 13.7 A

Means (SA) 11.5 C 12.5 B 14.2 A 12.1 B



DS=Drought Stress

hybrids had statistically different sugar content at each SA level with the best results

in DS-20, while they gave the smallest values at DS-0. Combination of sunflower

hybrids × DS levels had significant interaction with maximum plant sugar content

of all genotypes in DS-20 being statistically similar; while the smallest sugar content

were recorded for all the genotypes at DS-0 with non significant difference between

H-1 and H-3 hybrids.

Difference between two methods of SA application under different DS levels

on three hybrids and interaction with various SA levels is presented in Figure 2.14.

Increased levels of drought stress improved the sugar contents in the plants of all the

sunflower hybrids, which differed significantly under foliar spray but not for seed

soaking treatments with SA, and H-2 (NX-00989) showed slightly higher sugar

content (Figure 2.14 a). Sugar contents response in plants by SA application was

better at SA-0.75, being higher at high DS levels under both SA foliar and seed

soaking treatments (Figure 2.14 b). Higher concentration of salicylic acid treatment

(SA-1.50) did not increase sugar contents in the sunflower plants even at higher

DS-20 (20% PEG).

164

The present results also revealed that drought stress augmented the soluble

sugars in sunflower hybrids in both modes of application (Tables2.14.1, 2.14.2).The

increase in sugar concentration may be a result from the degradation of starch Fischer

and Höll (1991). Starch may play an important role in accumulation of soluble sugars

in cells. Starch depletion in grapevine leaves was noted by

Patakas and Noitsakis (2001) in response to drought stress.

The present investigation found that soluble sugars augmented under water

stressed conditions under foliar and seed soaking treatments of SA (Fig. 2.14).

Results corroborated with the findings of Kabiri et al. (2014) who examined the

effect of salicylic acid at three levels (0, 5, and 10μM) on black cumin (Nigella sativa

L.) under drought stress in hydroponic culture. Salicylic acid protected the Nigella

plant against drought stress through increasing of photosynthetic pigments and

soluble sugar contents, while 10μM SA was the most effective level. Exogenous

application of SA to drought stressed shallot plants showed more profound results as

compared to other groups. This may result from increased starch hydrolysis,

synthesis by other pathways or decreased conversion to other products (Ahmad et

al., 2014). Soluble sugar may act as a typical osmoprotectant, stabilizing cellular

membranes and maintaining turgor pressure. Both ABA and SA seed soaking

triggered the drought tolerance mechanism in Wafaq-2001 through enhanced sugar

accretion (Khan et al., 2012).

4.2.15 Leaf Protein

With the foliar application of salicylic acid, there was significant difference

among sunflower genotypes for protein contents in plants (Table 2.15.1). The highest

protein content (1154 µg/g) was recorded in H-2 (NX-00989), while the lowest one

(1071 µg/g) was in H-3 (FH-352) hybrid. Application of salicylic acid at 0.75 mM

significantly increased leaf protein as compared to the other concentrations. Drought

stress at 10% and 20% PEG significantly decreased protein content as compared to

165

that in control. Lowest protein (1017 µg/g) was found under 20% PEG application

and highest (1213 µg/g) in control (no-stress).

Figure 2.14: Soluble Sugars of sunflower hybrids under various levels of


Table 2.15.1: Leaf protein (µg/g f wt) of sunflower hybrids under different



9.0

12.0

15.0

18.0

21.0

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)


9.0

12.0

15.0

18.0

21.0

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

166


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 1069 gh 1126 d 1159 bc 1111g 1116 B

H-2 (NX-00989) 1092 ef 1144 c 1214 a 1166 b 1154 A

H-3 (FH-352) 1014 i 1069 gh 1124 d 1078 fg 1071 C


DS-0 (Control) 1166 d 1226 b 1258 a 1201bc 1213 A

DS-10 (10% PEG) 1062 g 1110 ef 1154 d 1121 e 1112 B

DS-20 (20% PEG) 947 j 1003 i 1085 g 1033 h 1017 C


H-1 × DS-0 1172 de 1281 a 1283 a 1244 b 1245 A

H-1 × DS-10 1077 j 1097 ij 1134 fg 1091ij 1100 C

H-1 × DS-20 959 q 1001 n 1059 k 997 o 1004 F

H-2 × DS-0 1217 bc 1236 b 1286 a 1234 b 1243 A

H-2 × DS-10 1094 ij 1155 ef 1198 d 1171 de 1154 B

H-2 × DS-20 965 n 1040 kl 1159 ef 1093 ij 1064 DE

H-3 × DS-0 1110 ghi 1160 ef 1203 c 1124 gh 1149 B

H-3 × DS-10 1016 lm 1079 j 1131 fg 1101 hij 1082 D

H-3 × DS-20 916 r 968 pq 1036 kl 1010 m 983 G

Means (SA) 1059 C 1113 B 1166 A 1118 B



DS=Drought Stress

The interaction of salicylic acid × genotypes was significant with maximum protein

content (1214 µg/g) in NX-00989 with 0.75 mM salicylic acid application.

Minimum protein content (1014 µg/g) was observed in FH-352 hybrid without

salicylic acid application. Similarly, drought levels × salicylic acid, and sunflower

hybrids × drought levels × salicylic acid interaction were statistically significant.

The highest protein content (1286 µg/g) was recorded in NX-00989 sunflower

167

genotype without drought stress with 0.75 mM salicylic acid application. Whereas,

the smallest value of protein content (916 µg/g) was observed in the plants of FH-

352 genotype at 20 % PEG without salicylic acid application.

Data of leaf protein content under seed soaking of sunflower hybrids with

salicylic acid at different DS levels have been depicted in Table 2.15.2. Sunflower

hybrids had significant difference for their protein contents, exhibiting the highest in

H-2 (NX-00989) and the lowest in H-3 (FH-352) genotype. On the other hand, plants

treated with SA-0.75 produced significantly greater protein (1042 µg/g) as compared

to other SA treatments. Different levels of drought stress caused significant reduction

in protein content, as the smallest protein content was with DS-20 (888 µg/g).

Interaction between various levels of drought stress and salicylic acid concentrations

was statistically significant. The highest protein content (1141 µg/g) in sunflower

plants was found under DS-0 receiving 0.75 mM concentration of salicylic acid,

which had statistically significant difference with other SA levels under each DS

level. Whereas, the lowest protein content (800µg/g) was recorded in DS-20 without

SA, which had significant difference with other SA levels at each

DS level. Interaction among hybrids × drought levels × salicylic acid Table 2.15.2:

Leaf protein (µg/g f wt) of sunflower hybrids under different drought stress

levels as influenced by seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 931 i 997 e 1065 b 1012 d 1001 B

H-2 (NX-00989) 954 g 1005 d 1076 a 1028 c 1016 A

H-3 (FH-352) 876 j 931 i 985 f 940 h 933 C


DS-0 (Control) 1053 d 1087 b 1141 a 1076 c 1089 A

DS-10 (10% PEG) 907 j 972 g 1025 e 988 f 973 B

DS-20 (20% PEG) 800 l 874 k 960 h 916 i 888 C

168


H-1 × DS-0 1109 c 1140 b 1208 a 1147 b 1151 A

H-1 × DS-10 889 mn 960 ij 1024 fg 970 i 961 D

H-1 × DS-20 795 q 892 m 963 ij 920 l 893 G

H-2 × DS-0 1079 d 1098 c 1148 b 1096 c 1105 B

H-2 × DS-10 956 j 1017 g 1059 e 1033 f 1016 C

H-2 × DS-20 827 p 902 m 1020 fg 955 j 926 F

H-3 × DS-0 972 i 1022 fg 1065 e 986 h 1011 C

H-3 × DS-10 878 no 941 k 993 h 963 ij 944 E

H-3 × DS-20 778 r 830 p 898 m 872 o 844 H

Means (SA) 920 D 978 C 1042 A 993 B



DS=Drought Stress

concentrations were also statistically significant. All the three hybrids were

statistically different for protein content at all SA levels with the best results in DS0,

while they gave the smallest protein content at DS-20 being statistically dissimilar

at all SA levels. Combination of sunflower hybrids × DS levels had significant

interaction with maximum protein content in of all genotypes in DS-0 being

statistically significant; while the smallest protein content were recorded for all the

genotypes at DS-20 with significant difference among the hybrids.


hybrids and interaction with various SA levels has been drawn in Figure 2.15. DS20

reduced the protein contents in all the sunflower hybrids, which differed at all DS

levels both with SA foliar and seed soaking treatments. H-2 (NX-00989) showed

slightly better protein content at DS-20 (Figure 2.15 a). Plant response to SA

application was better at SA-0.75, which produced higher protein contents in plants

at all DS levels under both foliar spray and seed soaking (Figure 2.15 b). However,

169

protein content of sunflower plants reduced with increasing DS levels, being the

lowest at the highest level of drought stress (DS-20).

The present investigation found that leaf protein decreased under water

stressed conditions under foliar and seed soaking treatments (Table 2.15.1 and

2.15.2). A stress occurrence which inhibits cell division and expansion, and thus leaf

expansion, will also apprehend protein synthesis. For instance, in Avena coleoptiles

water deficit caused a significant decrease in rate of protein synthesis


800

900

1000

1100

1200

1300

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

170

Figure 2.15: Leaf Protein of sunflower hybrids under various levels of drought


Dhindsa and Cleland (1975). Levels of biochemical components such as

chlorophylls, carotenoids, protein and starch decreased in drought stressed plants,

and increased when the plants recovered from stress (Haider and Saiffullah, 2001).

Lazcano and Lowatt (1999) found variable protein level in different cultivars of

Phaseolus under water stress.

Exogenous application of SA enhanced leaf protein content in all hybrids of

sunflower with 0.75 mM conc. (Fig.2.15 a, b). The results of present study are in

agreement with those of Singh and Usha (2003) who reported that exogenously

applied SA enhanced soluble protein contents in wheat seedlings under water stress

conditions. Application of salicylic acid caused significant increase in protein

content of brinjal (Karuppaiah et al., 2003). Sarangthem and Singh (2003) observed

that under optimum dose of treatment (0.1% V/V) salicylic acid, the levels of

proteins in Phaseolus oulgaris increased. It has also been observed that pre-

treatment of barely seedlings with salicylic acid before paraquat stress prevented the

protein loss (Popova et al., 2003).


800

900

1000

1100

1200

1300

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA0.375 SA-0.75 SA-1.50

171

4.2.16 Free Amino Acid Content

Foliar application of salicylic acid increased significantly amino acid content

in the three sunflower hybrids (Table 2.16.1). The highest amino acid content (6.82

µmol/g) was recorded with salicylic acid concentration of 0.75 mM while the lowest

(5.56 µmol/g) was found in control. Drought levels also increased amino acid content

from 3.65µmol/g in control to 8.22 µmol/g in 20% PEG. There was significant

interaction between genotype and salicylic acid concentrations.

Maximum amino acid (6.97µmol/g) was observed in genotype NX-00989 receiving

SA-0.75 while minimum (5.28 µmol/g) was observed in genotype FH-352 without

Table 2.16.1: Free amino acid content (µmol/g f wt) of sunflower hybrids under

different drought stress levels as influenced by foliar application of

salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 5.60 d 5.76 cd 6.78 a 6.19 b 6.08 AB

H-2 (NX-00989) 5.81 cd 6.08 bc 6.97 a 6.26 b 6.28 A

H-3 (FH-352) 5.28 e 5.51 de 6.70 a 5.69 d 5.79 C


DS-0 (Control) 3.30 h 3.25 h 4.38 f 3.65 g 3.65 C

DS-10 (10% PEG) 5.41 e 6.05 d 7.39 c 6.29 d 6.29 B

DS-20 (20% PEG) 7.97 b 8.05 b 8.68 a 8.20 b 8.22 A


H-1 × DS-0 3.20 no 3.00 o 4.38 m 3.77 mn 3.59 D

H-1 × DS-10 5.20 l 5.95 ijk 6.92 gh 6.35 hi 6.10 C

H-1 × DS-20 8.40 bcd 8.33 cd 9.04 ab 8.46 a-d 8.56 A

H-2 × DS-0 3.65 no 3.57 no 4.35 m 3.60 no 3.79 D

H-2 × DS-10 5.43 kl 6.20 ij 7.45 efg 6.43 hi 6.38 C

H-2 × DS-20 8.35 cd 8.47 a-d 9.10 a 8.75 abc 8.67 A

172

H-3 × DS-0 3.07 o 3.19 no 4.40 m 3.60 no 3.56 D

H-3 × DS-10 5.60 jkl 6.01 ijk 7.82 def 6.08 ijk 6.38 C

H-3 × DS-20 7.16 fg 7.34 efg 7.89 de 7.38 efg 7.44 B

Means (SA) 5.56 D 5.78 C 6.82 A 6.05 B



DS=Drought Stress.

salicylic acid application. The salicylic acid concentrations × drought stress (PEG)

levels interaction and sunflower hybrids × salicylic acid × drought stress interaction

were also significant. The highest content of amino acid (9.10 µmol/g) was recorded

in NX-00989 sunflower genotype with DS- 20 and SA-0.75, and lowest (3.07

µmol/g) in FH-352 without any application of PEG or salicylic acid.

Amino acid contents under experiment on seed soaking of sunflower hybrids

with salicylic acid at different DS levels are shown in Table 2.16.2. Here also, the

sunflower hybrids differed significantly; and the SA-0.75 produced significantly

higher amino acid contents in plants (6.51µmol/g) as compared to other SA levels.

Different levels of drought stress also caused significant increase in amino acid

content, as the highest value was with DS-20 (7.73 µmol/g). Interaction between

various levels of drought stress × salicylic acid treatments was statistically

significant. Highest amino acid content (8.44 µmol/g) was found under

DS-20 receiving SA-0.75, which was statistically different from other SA levels in

DS-20. Whereas, the smallest amino acid content (3.13 µmol/g) was recorded in

0.375 mM SA, which had non significant difference with that in control and SA1.50

under DS-0. Interaction among sunflower hybrids × drought levels × salicylic acid

concentrations was also statistically significant. All the three hybrids had statistically

different amino acid contents at each SA concentrations with the highest results in

173

DS-20, while they gave the smallest amino acid contents at DS-0. Combination of

sunflower hybrids × DS levels had significant interaction with maximum amino acid

content of all genotypes under DS-20 but statistically different; while the smallest

amino acid contents were recorded for all the genotypes at DS-0 with significant

difference among the hybrids. However, at Table 2.16.2: Free amino acid (µmol/g

f wt) of sunflower hybrids under different drought stress levels as influenced by

seed soaking in salicylic acid.


SA-0 SA-0.375 SA-0.75 SA-1.50


H-1 (NX-19012) 5.21 e 5.23 e 6.42 b 5.75 d 5.65 B

H-2 (NX-00989) 5.70 d 6.03 c 6.95 a 6.09 c 6.19 A

H-3 (FH-352) 4.94 f 5.15 ef 6.17 bc 5.33 e 5.40 C


DS-0 (Control) 3.26 i 3.13 i 4.11 h 3.23 i 3.43 C

DS-10 (10% PEG) 5.32 g 5.78 f 6.99 d 6.25 e 6.08 B

DS-20 (20% PEG) 7.28 c 7.50 bc 8.44 a 7.69 b 7.73 A


H-1×DS-0 3.02 s 2.75 s 3.75 op 2.95 s 3.12 G

H-1 × DS-10 5.18 m 5.53 klm 6.94 efg 6.82 fg 6.12 D

H-1 × DS-20 7.43 d 7.40 de 8.58 b 7.47 d 7.72 B

H-2 × DS-0 3.64 pq 3.51 pqr 4.38 n 3.54 pqr 3.77 E

H-2 × DS-10 5.39 lm 6.05 ij 7.22def 5.98 ijk 6.16 D

H-2 × DS-20 8.08 c 8.53 bc 9.27 a 8.75 b 8.66 A

H-3 × DS-0 3.13 rs 3.14 rs 4.20 no 3.22 qrs 3.42 F

H-3 × DS-10 5.38 lm 5.74 jkl 6.83 fg 5.94 ijk 5.97 D

H-3 × DS-20 6.32 hi 6.58 gh 7.47 d 6.84 fg 6.80 C

Means (SA) 5.28 D 5.47 C 6.51 A 5.72 B



174

DS=Drought Stress.

DS-10 all the three hybrids showed statistically similar contents of amino acid in the

plants.


hybrids and interaction with various SA levels is presented in Figure 2.16. Higher

levels of drought stress increased the amino acid contents of all the sunflower

hybrids, which differed non significantly at DS-0 and DS-10 under both foliar spray

and seed soaking treatments with SA, however, H-1 (NX-19012) and H-2

(NX-00989) showed significantly higher amino acid content at DS-20 than that in

H-3 (Figure 2.16 a). Plant response to SA application was better at SA-0.75, which

caused higher amino acid content in plants at all DS levels under both foliar spray

and seed soaking treatments (Figure 2.16 b). Effect of salicylic acid on sunflower

plants amino acid content with 0.75 mM concentration remained constant with

increasing DS levels and significantly different with that under SA-0 at all levels of

drought stress.

The present investigation revealed that leaf amino acid decreased under water

stressed conditions under foliar and seed soaking treatments (Table 2.16.1 and

2.16.2). During osmotic stress, plant cells accumulate solutes to prevent water loss

and to re-establish cell turgor. The solutes that accumulate during osmotic

adjustment include nitrogen-containing compounds, such as proline and other amino

acids (Tamura et al., 2003). The accumulation of ROS during drought stress, along

with increasing H2O2, often enhances protein oxidation in plant species (Jiang and

Nhunag, 2001). The other reason for accumulation of amino acid may be taking place

in response to the change in osmotic adjustment of their cellular

175


Figure 2.16: Free amino acids of sunflower hybrids under various levels of


contents (Shao et al., 2007). Total free amino acids contents are augmented in

drought stressed plants, and tend to decrease during the period of recovery (Parida

et al., 2007). A common response of plants to environmental stress is an

accumulation of amino acids (Aspinall and Paleg, 1981).


3.00

5.00

7.00

9.00

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

3.00

5.00

7.00

9.00

DS-0 DS-10 DS-20 DS-0 DS-10 DS-20



SA-0 SA-0.375 SA-0.75 SA-1.50

176

Exogenous application of SA further increased amino acid content in all

sunflower hybrids with 0.75mM conc. (Fig.2.16 a, b). Similar results were given by

El-Tayeb (2005) that all amino acids increased with salicylic acid in maize plants.

Azimi et al. (2013) showed that water stress reduced all attributes of growth and

yield, but amino acid and salicylic acid reduced negative effects of water deficit on

wheat. The free amino acid accumulation under water stressed shallot plants treated

with SA was found higher as compared to control plants (Ahmad et al., 2014).



STAGES OF SUNFLOWER HYBRIDS UNDER DROUGHT STRESS

4.3.1 Fresh Root Weight

Difference between foliar application of salicylic acid at two growth stages

of sunflower plants was non significant for fresh root weight (Table 3.1). Drought

stressed three sunflower hybrids also differed non significantly with each other for

fresh root weight under foliar application of salicylic acid in different concentrations

Table 3.1: Root fresh weight (g/plant) of drought stressed sunflower hybrids

under foliar applied various concentrations of salicylic acid at two growth

stages.

Treatments Growth Stages (GS) Means

Vegetative Flowering

Hybrids (H) Interaction (H × GS) Means (H)

H-1 (NX-19012) 3.36 NS 3.31 3.33 NS

H-2 (NX-00989) 3.52 3.40 3.46

H-3 (FH-352) 3.24 3.18 3.21

SA concentrations mM Interaction (SA × GS) Means (SA)

Control (DS-0 + SA-0) 3.89 NS 3.80 3.85 A

DS + SA-0 2.89 2.94 2.92 C

DS + SA-0.375 3.27 3.10 3.18 B

DS + SA-0.75 3.52 3.40 3.46 AB

DS + SA-1.50 3.31 3.22 3.27 B

Hybrids × Salicylic acid Interaction (H × SA × GS) Means (H × SA)

177

H-1 × DS-0 + SA-0 3.73NS 3.70 3.72 NS

H-1 × DS + SA-0 2.87 2.93 2.90

H-1 × DS + SA-0.375 3.27 3.03 3.15

H-1 × DS + SA-0.75 3.63 3.57 3.60

H-1 × DS + SA-1.50 3.30 3.30 3.30

H-2 × DS-0 + SA-0 4.10 3.97 4.03

H-2 × DS + SA-0 2.89 3.07 2.98

H-2 × DS + SA-0.375 3.50 3.30 3.40

H-2 × DS + SA-0.75 3.63 3.40 3.52

H-2 × DS + SA-1.50 3.50 3.27 3.38

H-3 × DS-0 + SA-0 3.83 3.77 3.80

H-3 × DS + SA-0 2.90 2.83 2.87

H-3 × DS + SA-0.375 3.03 2.97 3.00

H-3 × DS + SA-0.75 3.30 3.23 3.27

H-3 × DS + SA-1.50 3.13 3.10 3.12

Means (GS) 3.37 NS 3.30



DS=Drought Stress

at two growth stages. However, there was significant difference among different

concentrations of salicylic acid application. The highest fresh root weight (3.85

g/plant) was recorded in control (DS-0+SA-0) without receiving both drought stress

and salicylic acid, while the lowest (2.92 g/plant) was withDS+SA-0 (drought

stressed but receiving no salicylic acid). Application of salicylic acid at the

concentration of 0.75 mM resulted the highest increase of fresh root weight of

drought stressed plants although non significantly. There was significant decrease in

fresh root weight with the employment of drought stress, which was slightly

addressed by salicylic acid application. Interactions were statistically non significant

for: sunflower hybrids (H) × growth stages (GS); concentrations of salicylic acid

(SA) × GS; H×SA; and H × SA × GS.

Interactive effect of three sunflower hybrids with two growth stages of

sunflower at which SA foliar application was made, has been drawn in Figure 3.1 a.

178

It indicates that sunflower hybrids differed more if SA was applied at vegetative

growth stage even all giving slightly greater fresh root weight as compared to that at

flowering stage. H-2 (NX-00989) showed slightly better results while H-3 (FH352)

performed the least although the difference was statistically non significant.

Similarly, differential response of drought stressed sunflower hybrids to foliar

application of SA at two growth stages was non significant (Figure 3.1 b). However,

SA application in 0.75 mM concentration at both stages produced significantly

higher fresh root weight of drought stressed plants as compared to those receiving

no SA. Whereas, control treatment (DS-0 + SA-0) rendered the highest value for

fresh root weight of non stressed plants, which was significantly

a. Hybrids× Growth stages

2.00

2.50

3.00

3.50

4.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


179

b. Salicylic acid concentrations × Growth stages

Figure 3.1: Fresh root weight of drought stressed sunflower hybrids under

foliar applied various concentrations of salicylic acid at two growth stages.

superior to all the treatments in drought stressed sunflower hybrids plants. Within

all the treatments, interaction of DS with various SA concentrations showed better

response of sunflower hybrids to SA foliar application at vegetative growth stage.

Further, protective effect of salicylic acid on drought-stressed sunflower plants

enhanced with increasing SA concentration, being highest at the 0.75 mM, and lesser

at next higher concentration of 1.50 mM.

Drought stress significantly reduced the root fresh weight. Current study

exhibits that root fresh weight was reduced in sunflower plant exposed to drought

stress at both stages whereas, root fresh weight markedly reduced at flowering stage

(Table 3.I). The development of root system increases the water uptake and

maintains requisite osmotic pressure through higher proline levels in Phoenix

dactylifera (Djibril et al., 2005). An increased root growth due to water stress was

reported in sunflower (Tahir et al., 2002) and Catharanthus roseus (Jaleel et al.,

2.50

3.00

3.50

4.00

4.50

DS-0 + SA-0 DS + SA-0 DS + SA-0.375 DS + SA-0.75 DS + SA-1.50

Salicylic acid treatments (mM ) to droughted plants


180

2008 b, c). However, drought stress significantly reduced the root growth by resisting

the root penetration into the dry soil and reducing the root respiratory efficiency

(Borrell and Hammer, 2000; Thomas and Howarth, 2000). Drought stress

considerably affects the plant growth as indicated by reduced fresh biomass of roots


Root fresh weight increased by the application of SA, tolerant hybrids

showed better response than sensitive one (Fig. 3.1 a, b). Similarly, under water

stress, root fresh weight decreased significantly in sensitive inbred line of sunflower

(B59), while the tolerant one (B71) showed a minor drop (Andrade et al., 2013). An

increase in fresh root weight of drought-stressed plants was recorded with SA

application (El-Tayeb, 2005). Khodray (2004) reported an improved fresh root

weight of stressed maize plants treated with salicylic acid. Application of acetyl

salicylic acid (ASA) at 20 ppm on pea plants enhanced plant growth as indicated by

plant height, number of leaves, fresh and dry weights in both seasons

(El-Shraiy and Hegazi, 2009).

4.3.2 Dry Root Weight

There was non significant difference between foliar applications of salicylic acid at

two growth stages of sunflower plants for dry root weight (Table 3.2). Three

sunflower hybrids grown under drought stress, however, differed significantly with

each other for dry root weight under foliar application of salicylic acid. H-2

(NX00989) exhibited greater dry root weight as compared to other two hybrids of

sunflower, both of which had non significant difference between them. Further, there

was non significant difference among different concentrations of salicylic acid foliar

application to drought-stressed plants. Whereas, sunflower plants in control (DS-0 +

SA-0) showed significantly higher dry root weight as compared to drought-stressed

181

plants even treated with salicylic acid. The highest dry root weight (0.61g/plant) was

recorded in control plants without receiving drought stress as well as salicylic acid,

while the lowest weight (0.42 g/plant) was underDS+SA-0 treatment. Salicylic acid

application at 0.75 mM concentration caused better recovery of dry root weight of

drought stressed plants although non significantly with other SA treatments. There

was significant decrease in the dry root weight of plants under drought stress. All the

treatment interactions, viz. H × GS, concentrations of SA × GS, H×SA, and H × SA

× GS were statistically non

significant for dry root weight of sunflower hybrids.

Table 3.2: Root dry weight (g/plant) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 0.45 NS 0.43 0.44 B

H-2 (NX-00989) 0.62 0.59 0.61 A

H-3 (FH-352) 0.43 0.41 0.42 B

SA concentrations (mM) Interaction (SA × GS) Means (SA)

Control (DS-0 + SA-0) 0.63 NS 0.60 0.61 A

DS + SA-0 0.44 0.39 0.42 C

DS + SA-0.375 0.47 0.43 0.45 BC

DS + SA-0.75 0.53 0.52 0.53 B

DS + SA-1.50 0.48 0.47 0.47 BC

Hybrids × Salicylic acid Interaction (H × SA × GS) Means (H×SA)

H-1 × DS-0 + SA-0 0.63 NS 0.59 0.61NS

H-1 × DS + SA-0 0.36 0.33 0.35

H-1 × DS + SA-0.375 0.38 0.36 0.37

H-1 × DS + SA-0.75 0.50 0.47 0.49

H-1 × DS + SA-1.50 0.39 0.38 0.39

H-2 × DS-0 + SA-0 0.66 0.65 0.65

H-2 × DS + SA-0 0.56 0.51 0.54

H-2 × DS + SA-0.375 0.62 0.54 0.58

H-2 × DS + SA-0.75 0.65 0.67 0.66

H-2 × DS + SA-1.50 0.62 0.62 0.62

H-3 × DS-0 + SA-0 0.51 0.47 0.49

H-3 × DS + SA-0 0.39 0.33 0.36

H-3 × DS + SA-0.375 0.41 0.39 0.40

182

H-3 × DS + SA-0.75 0.43 0.43 0.43

H-3 × DS + SA-1.50 0.42 0.41 0.41

Means (GS) 0.50 NS 0.48



DS=Drought Stress.

Interaction between sunflower hybrids and growth stages for SA application

as shown in Figure 3.2 a. Sunflower genotypes differed equally with SA applied at

vegetative growth as well as flowering stage, although they produced higher dry root

weight through foliar application of SA at vegetative stage as compared to that at

flowering stage. H-2 (NX-00989) showed significantly better results while H-3 (FH-

352) performed the least. Response of drought stressed sunflower hybrids to foliar

application of SA at two growth stages was non significant (Figure 3.2 b). SA

application in 0.75 mM concentration at both stages produced significantly higher

dry root weight of drought stressed sunflower plants as compared to those receiving

no SA. Under the control treatment, the highest dry root weight of sunflower plants

was recorded, which was significantly superior to all other treatments in drought

stressed plants. Within all the treatments, interaction of DS with various SA

concentrations showed slightly better response of sunflower hybrids to SA foliar

application at vegetative growth stage. Protective effect of salicylic acid on drought-

stressed sunflower plants enhanced with increasing concentration of SA up to 0.75

mM, after that it was lesser at 1.50 mM concentration.

In the present investigation water stress reduced the root dry weight in

sunflower hybrids especially at flowering stage (Table 3.2). Present findings are in

hormony as reported that the root dry weight was decreased under mild and severe

water stress in Populus species (Wullschleger et al., 2005). A common adverse effect

of water stress on crop plants is the reduction in fresh and dry biomass production

(Farooq et al., 2009). Diminished biomass due to water stress was

183



Figure 3.2 Dry root weight of drought stressed sunflower hybrids under foliar applied

various concentrations of salicylic acid at two growth stages.

observed in almost all genotypes of sunflower (Tahir and Mehdi, 2001). Reduced

biomass was seen in water stressed common bean and green gram (Webber et al.,

2006) and Petroselinum crispum (Petropoulos et al., 2008). Heshmat et al. (2012)

also found decreased dry weight of root in different lines of sunflower.

0.30

0.40

0.50

0.60

0.70

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


0.30

0.40

0.50

0.60

0.70


Salicylic acid treatments (mM) to droughted plants


184

Salicylic acid application improved root dry weight under water deficit stress

(Fig. 3.2 a, b). These results are in line with the findings that foliar spray with

salicylic acid increased the fresh and dry weight of plant, pod setting and total

proteins of leaves and fruits of broad bean (Liu Xini et al., 2000 and Sanaa et al.,

2001). Salicylic acid is reported to counteract the adverse effects of drought by

improving the growth of root and shoot (El-Tayeb and Naglaa, 2010). Salicylic acid

treatments increased root fresh and dry weights of barley seedlings (Metwally,

2003).

4.3.3 Shoot Fresh Weight

Foliar application of salicylic acid at vegetative growth stage of sunflower

plants produced significantly higher fresh shoot weight as compared to that under

SA application at flowering stage (Table 3.3). Among three water stressed sunflower

hybrids, H-2 (NX-00989) gained significantly greater fresh shoot weight as

compared to others under SA foliar application. While, H-3 (FH-352) could fetch

the lowest fresh shoot weight. There was significant difference among different

treatments of salicylic acid application. The highest fresh shoot weight (21.2 g/plant)

was recorded in control followed by DS + SA-0.75 (18.6 g/plant), while the lowest

(16.0 g/plant) was with DS+SA-0 (water stressed but no salicylic acid spray).

Application of salicylic acid resulted in an increase of fresh shoot weight in water

stressed plants. There was significant decrease in fresh shoot weight with the

employment of water stress, which was significantly addressed by salicylic acid

application. Interactions among all sort of factors / treatments (H ×

GS, SA × GS, H×SA, and H × SA × GS) were statistically non significant.

185


sunflower at which SA foliar application was made, has been shown in Figure 3.3 a.

Sunflower hybrids H1 and H2 differed more for SA applied at two growth stages,

and they had greater fresh shoot weight if SA was applied at vegetative stage. H-1

(NX-19012) and H-2 (NX-00989) showed better results while H-3 (FH352)

performed the least. Similarly, response of water stressed sunflower hybrids to foliar

application of SA at two growth stages was significant (Figure 3.3 b). The SA

application in 0.75 mM treatment at both stages produced significantly higher fresh

shoot weight of water stressed plants as compared to those receiving no SA.

Within all the treatments, interaction of DS with various SA concentrations showed

better response of sunflower hybrids to SA foliar application at vegetative growth

stage. Recovering effect of salicylic acid on drought-stressed sunflower plants

enhanced with increasing SA concentration, being maximum at 0.75 mM, and

significantly lesser at further increased concentration.

The present study revealed that water stress reduces plant shoot fresh weight

in all sunflower hybrids at both stages of growth (Table 3.3). This reduction in the

shoot fresh weight with the increase in drought stress may be due to the adaptation

of sunflower plants to drought stress. Results are in line with the various treatments

at

Table 3.3: Shoot fresh weight (g/plant) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 18.9 NS 17.2 18.0AB

H-2 (NX-00989) 18.9 18.2 18.6A

H-3 (FH-352) 18.3 17.3 17.8B


Control (DS-0 + SA-0) 21.5 NS 20.9 21.2 A

DS + SA-0 16.8 15.2 16.0 C

186

DS + SA-0.375 17.6 16.6 17.1 BC

DS + SA-0.75 19.1 18.1 18.6 B

DS + SA-1.50 18.5 17.1 17.8BC


H-1 × DS-0 + SA-0 21.8 NS 20.5 21.1 NS

H-1 × DS + SA-0 16.9 15.1 16.0

H-1 × DS + SA-0.375 17.5 16.2 16.9

H-1 × DS + SA-0.75 19.2 18.1 18.7

H-1 × DS + SA-1.50 18.8 16.3 17.6

H-2 × DS-0 + SA-0 21.2 20.6 20.9

H-2 × DS + SA-0 17.6 16.1 16.9

H-2 × DS + SA-0.375 18.0 17.1 17.6

H-2 × DS + SA-0.75 19.5 19.0 19.3

H-2 × DS + SA-1.50 18.3 18.4 18.4

H-3 × DS-0 + SA-0 21.6 21.7 21.6

H-3 × DS + SA-0 15.8 14.4 15.1

H-3 × DS + SA-0.375 17.3 16.5 16.9

H-3 × DS + SA-0.75 18.6 17.1 17.8

H-3 × DS + SA-1.50 18.4 16.7 17.5

Means (GS) 18.7 A 17.6 B



DS=Drought Stress


12.0

15.0

18.0

21.0

24.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


187


Figure 3.3: Fresh Shoot weight of drought stressed sunflower hybrids under


previous findings of many researchers as described that with the onset of drought

plants tend to reduce the shoot growth by reducing the leaf area, number of leaves

and plant height and increase root growth (Schuppler et al., 1998). Shoot fresh and

dry weights in maize and soybean plants decreased when exposed to drought due to

reduced shoot growth, increased senescence and switching over of the plant growth

from shoot growth towards root growth (Sharp et al., 1988; Hamayun et al., 2010).

Reduced biomass was seen in water stressed soybean (Specht et al., 2001). Andrade

et al. (2013) compared two inbred lines of sunflower (sensitive B59, and tolerant

B71) with contrasting behavior to moisture deficit. Their fresh shoot weight

decreased under water stress.

Results showed that shoot fresh weight increased in all hybrids when SA

was applied especially at vegetative growth stage in tolerant hybrids H-1 and H-2

(Fig 3.3 a, b). These results are in conformity with the observations that salicylic

12.0

15.0

18.0

21.0

24.0




188

acid treatments increased the shoot fresh weight of barley seedlings (Metwally,

2003). Foliar application of salicylic acid 150 ppm gave the highest increment in

number of branches, fresh weight and dry weight and total protein of snap bean.

(Kmal et al., 2006). Spraying potato plants with salicylic acid at 100 ppm had

beneficial effects on vegetative growth characters, total tuber yield and chemical

contents of potato tubers (Awad and Mansour, 2007).

4.3.4 Shoot Dry Weight

There was non significant difference in dry shoot weight for foliar application

of salicylic acid at two growth stages of sunflower (Table 3.4).

However, water stressed three sunflower hybrids treated with salicylic acid in two

growth stages showed significant difference among them. The highest weight of dry

shoots was recorded for H-2 (NX-00989) followed by H-1 (NX-19012) and the

lowest value was found in H-3 (FH-352). Similarly, there was significant difference

among different treatments of salicylic acid foliar application. Significantly greater

dry shoot weight (7.85 g/plant) was in control given no water stress as well as no SA

application, while the lowest (4.94 g/plant) was in water stress. Foliar spray of

salicylic acid at 0.75 mM treatment caused the highest increase of dry shoot weight.

There was significant recovery in dry shoot weight of water stressed plants with SA

application. Interactions between all three treatment factors were statistically non

significant for dry shoot weight.

Interactive effect of sunflower hybrids with two growth stages has been

shown in Figure 3.4 a, which reflects greater dry shoot weight in all sunflower

hybrids if SA was applied at vegetative growth stage as compared to that at flowering

stage. Comparative response of water stressed sunflower hybrids to foliar application

of SA at two growth stages was non significant (Figure 3.4 b). However, SA

189

application with 0.75 mM treatment at both stages produced significantly greater dry

shoot weight of water stressed plants as compared to those receiving no SA.

Whereas, control treatment (DS-0 + SA-0) rendered the highest value for dry shoot

weight, which was significantly superior to all the treatments in water stressed plants.

Generally, within all the treatments, interaction of DS with different SA treatments

showed slightly better response of sunflower hybrids to SA foliar application at

vegetative growth stage. The remedial effect of SA on waterstressed plants increased

with higher SA treatment, being best at 0.75 mM.

It was concluded from the results water stress negatively affects shoot dry

weight at both stages (Table 3.4). These findings are in line with Nizami et al.,

2008 who found that drought stress badly affects the sunflower plant dry matter.

Water stress affected in a greater proportion shoot dry weight (DW) than root DW

in sensitive and tolerant lines of sunflower Andrade et al. (2013). A common adverse

effect of water stress on crop plants is the reduction in fresh and dry biomass

production (Farooq et al., 2009). Mild water stress affected the shoot dry weight,

while shoot dry weight was greater than root dry weight loss under severe stress in

sugar beet genotypes (Mohammadian et al., 2005).

Foliar application of salicylic acid at two growth stages resulted in increased

shoot dry weight in all hybrids especially when applied at vegetative growth stage

(Fig. 3.4 a, b). Salicylic acid counteracts the negative effects of drought on plants by

improving their growth (Heshmat et al., 2012). Sayyari et al. (2013) showed that

plant dry weight of lettuce (Lactuca sativa L.) reduced in drought conditions, but it

increased significantly by SA application. Also spraying tomato plants with salicylic

acid at 100 ppm increased vegetative growth, dry weight, yield and its components

and NPK content as well as total protein (Ali et al., 2009). 1.5 mM concentration of

190

salicylic acid had a stimulating effect on the growth, dry weight and protein of pepper

as compared with other concentrations 5 and 10 mM (Canakci, 2011).

4.3.5 Plant Height

Foliar application of salicylic acid at two growth stages had significant

difference for the height of sunflower plants (Table 3.5). Three sunflower hybrids

Table 3.4: Shoot dry weight (g/plant) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 6.51 NS 5.38 5.95B

H-2 (NX-00989) 7.02 6.10 6.56 A

H-3 (FH-352) 6.21 5.62 5.92 B


Control (DS-0 + SA-0) 7.92 NS 7.78 7.85 A

DS + SA-0 5.51 4.37 4.94 D

DS + SA-0.375 6.23 5.14 5.68 C

DS + SA-0.75 6.94 5.77 6.36 B

DS + SA-1.50 6.30 5.46 5.88 BC


H-1 × DS-0 + SA-0 7.83 NS 7.60 7.72 NS

H-1 × DS + SA-0 5.33 4.08 4.71

H-1 × DS + SA-0.375 6.07 4.81 5.44

H-1 × DS + SA-0.75 6.85 5.29 6.07

H-1 × DS + SA-1.50 6.49 5.13 5.81

H-2 × DS-0 + SA-0 8.23 8.07 8.15

H-2 × DS + SA-0 6.00 4.93 5.47

H-2 × DS + SA-0.375 6.83 5.50 6.17

H-2 × DS + SA-0.75 7.33 6.21 6.77

H-2 × DS + SA-1.50 6.70 5.81 6.26

H-3 × DS-0 + SA-0 7.70 7.67 7.68

H-3 × DS + SA-0 5.20 4.10 4.65

H-3 × DS + SA-0.375 5.80 5.10 5.45

H-3 × DS + SA-0.75 6.63 5.82 6.23

191

H-3 × DS + SA-1.50 5.70 5.43 5.57

Means (GS) 6.58 NS 5.70



DS=Drought Stress.



Figure 3.4: Dry Shoot weight of drought stressed sunflower hybrids under foliar

applied various concentrations of salicylic acid at two growth stages. differed

4.00

5.00

6.00

7.00

8.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


4.00

5.00

6.00

7.00

8.00

9.00




192

significantly for plant height under foliar application of salicylic acid in different

treatments at two growth stages H-1(NX-19012) gave highest value (50.4 cm).

Further, there was significant difference among three treatments of salicylic acid

application to water stressed sunflower plants. The best plant height (52.8cm) was

recorded in control without receiving both water stress and salicylic acid at flowering

stage, while the smallest height (44.8 cm) was with DS+SA-0 at vegetative stage.

Application of salicylic acid at the treatment of 0.75 mM caused the highest and

significant increase of plant height of water stressed plants. There was significant

decrease in plant height with the employment of water stress; however, the negative

impact of water stress was reduced by salicylic acid spray. Interactions were

statistically non significant for treatments of SA × GS and H ×

SA × GS.

Interactive effect of three sunflower hybrids with SA foliar application at two

growth stages of sunflower indicated that plant height of H-1 (NX-19012) was

greater if SA was applied at flowering growth stage (Figure 3.5 a). However, H2

and H3 did not show any statistical difference between two stages of SA application.

Difference of water stressed sunflower hybrids for foliar application of SA at two

growth stages was non significant for most of the treatments (Figure 3.5 b).

However, SA application in 0.75 mM treatment at flowering growth stage produced

significantly greater plant height of water stressed plants as compared to those

receiving SA spray at vegetative stage. Control (DS-0 + SA-0) rendered the highest

value for plant height, which was significantly superior to all the treatments in water

stressed sunflower plants. Within all the treatments, interaction of DS with Table

3.5: Plant height (cm) of drought stressed sunflower hybrids under foliar

applied various concentrations of salicylic acid at two growth stages.


193



H-1 (NX-19012) 49.0NS 51.5 50.3NS

H-2 (NX-00989) 48.9 49.3 49.1

H-3 (FH-352) 47.7 48.1 47.9


Control (DS-0 + SA-0) 52.5 NS 52.8 52.6 A

DS + SA-0 44.8 45.9 45.3 D

DS + SA-0.375 47.3 48.4 47.8C

DS + SA-0.75 49.6 51.2 50.1 B

DS + SA-1.50 48.7 49.7 49.2 BC


H-1 × DS-0 + SA-0 51.9NS 52.8 52.4B

H-1 × DS + SA-0 44.2 47.6 45.9 EF

H-1 × DS + SA-0.375 49.5 52.2 50.9 CD

H-1 × DS + SA-0.75 50.0 52.5 51.3 C

H-1 × DS + SA-1.50 49.6 52.3 51.0 C

H-2 × DS-0 + SA-0 53.6 53.8 53.7 A

H-2 × DS + SA-0 45.1 45.2 45.2 F

H-2 × DS + SA-0.375 46.1 46.2 46.2 EF

H-2 × DS + SA-0.75 50.1 51.3 50.7 CD

H-2 × DS + SA-1.50 49.8 49.9 49.9 D

H-3 × DS-0 + SA-0 51.9 51.9 51.9 BC

H-3 × DS + SA-0 45.0 45.0 45.0 F

H-3 × DS + SA-0.375 46.3 46.8 46.6 E

H-3 × DS + SA-0.75 48.8 49.9 49.4 D

H-3 × DS + SA-1.50 46.7 46.9 46.8 E

Means (GS) 48.6 B 49.6 A



DS=Drought Stress.

194



Figure 3.5: Plant height of drought stressed sunflower hybrids under foliar


various SA treatments showed better response of sunflower hybrids to SA foliar

application at flowering growth stage. Further, protective effect of salicylic acid on

water-stressed sunflower plants enhanced with increasing SA treatment, being

highest at 0.75 mM, and lesser at the next higher treatment of 1.50 mM.

42.0

46.0

50.0

54.0

58.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


42.0

46.0

50.0

54.0

58.0




195

Plant height of sunflower hybrids decreases non significantly at both stages

under the water deficit conditions (Table 3.5) compared with control. Under water stress

the reduction in plant height might be due to disturbed plant water relations specifically

turgor potential, which then severely affects the cell elongation and cell division (Arnon,

1972). Bakhsh et al. (1999) also reported reduced plant height in response to decreasing

irrigation frequencies. Due to impaired mitosis, cell elongation and expansion, plant

height reduces under drought stress (Kaya et al., 2006; Hussain et al., 2008). Nizami et

al. (2008) reported that drought stress affected the plant height, and other growth / yield

attributes in sunflower. Bajehbaj (2011) showed that four sunflower cultivars decreased

plant height and other growth traits significantly upon the application of water deficit

stress.

SA application especially (0.75 mM) improved the plant height significantly than

other treatments (Fig.3.5 b). SA application especially (0.5 mM) diminished the drought

damages and increased plant height (Sadeghipour and Aghaei, 2012). However, it has

also been observed that foliar application of salicylic acid to soybean and corn plants @

10 3 and 10 5 mol/L did not affect the plant height (Khan et al.,

2003).

4.3.6 Leaf count

Number of leaves per plant of sunflower hybrids was non significantly

different with foliar application of salicylic acid at flowering stage to that with SA

spray at vegetative growth stage (Table 3.6). Water stressed three sunflower hybrids

not differed significantly with each other for per plant leaf count under foliar

application of salicylic acid in different treatments at two growth stages. There is no

pronounced difference of leaf number among the three sunflower hybrids. However,

there was non significant difference among three applied treatments of salicylic acid

196

application. The highest number of leaves per plant (21.6) was recorded in control

without receiving both water stress and salicylic acid, while the lowest (20.4) was in

DS+SA-0. There was non significant decrease in leaf count with the employment of

water stress, which was slightly addressed by SA application. Salicylic acid spray at

the treatment of 0.75 mM resulted in the highest increase of leaf count of water

stressed plants. Most of the interactions were statistically non significant.

Figure 3.6 a showing the interactive effect of three sunflower hybrids with

two growth stages of sunflower for SA foliar application indicates that they

performed equally showing statistically non significant difference. Differential

response of water stressed sunflower hybrids to foliar application of SA at two

growth stages was also non-significant (Figure 3.6 b). Among all water stress

treatments, SA application in all treatments at both stages produced similar leaf count

as compared to those receiving no SA. Leaf number shows no effect of water stress

neither at both stages or the application of SA.

Table 3.6: Leaf count (Number/plant) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 21.8NS 22.0 21.9 NS

H-2 (NX-00989) 21.8 22.1 22.0

H-3 (FH-352) 20.2 20.3 20.2


Control (DS-0 + SA-0) 21.4 NS 21.7 21.6NS

DS + SA-0 20.0 20.9 20.4

DS + SA-0.375 21.3 21.3 21.3

DS + SA-0.75 21.4 21.8 21.6

DS + SA-1.50 20.9 21.2 21.0


H-1 × DS-0 + SA-0 20.0 NS 23.0 21.5 NS

H-1 × DS + SA-0 19.3 21.0 20.2

H-1 × DS + SA-0.375 21.0 21.0 21.0

197

H-1 × DS + SA-0.75 22.3 23.0 22.7

H-1 × DS + SA-1.50 21.0 22.0 21.5

H-2 × DS-0 + SA-0 23.0 21.0 22.0

H-2 × DS + SA-0 20.3 21.3 20.8

H-2 × DS + SA-0.375 21.7 22.0 21.8

H-2 × DS + SA-0.75 22.3 23.0 22.7

H-2 × DS + SA-1.50 21.7 22.0 21.8

H-3 × DS-0 + SA-0 21.3 21.0 21.2

H-3 × DS + SA-0 20.3 20.3 20.3

H-3 × DS + SA-0.375 21.1 21.0 21.0

H-3 × DS + SA-0.75 19.7 19.3 19.5

H-3 × DS + SA-1.50 19.9 19.7 19.8

Means (GS) 21.0NS 21.4



DS=Drought Stress.


12.0

15.0

18.0

21.0

24.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


198


Figure 3.6: Leaf count of drought stressed sunflower hybrids under foliar


Leaf score was not significantly affected by water stress at both stages (Table

3.6). Drought stress affected plant height, plant dry matter, stem diameter, head size,

seed number/head, 100-seed weight and seed weight/ head but Leaf number was not

affected by either drought or defoliation ( Nizami et al., 2008).

Leaf score was not decreased /increased by water stress and exogenous

application of SA in all sunflower hybrids at both growth stages (Fig 3.6 a, b) Earlier,

Senaranta et al. (2000) reported no observable differences in leaf number between

SA treated and untreated bean and tomato plants under heat, chilling and drought

stresses.

4.3.7 Leaf Area

Difference between two growth stages of SA application to sunflower for

their effect on leaf area of plants was statistically significant (Table 3.7). Application

of SA at vegetative stage caused significantly greater leaf area as compared to that

12.0

15.0

18.0

21.0

24.0




199

with SA application at flowering growth stage. Three sunflower hybrids also differed

significantly with each other for leaf area as H-1 (NX-19012) showed the highest

value and H-3 (FH-352) with least one. There was significant difference among

different treatmentsof salicylic acid application. The greatest leaf area value (152

cm2/plant) was recorded in control without water stress and SA application, while

the lowest (105 cm2/plant) was in water stressed plants without receiving SA.

Application of SA at the treatment of 0.75 mM caused the highest and significant

increase in leaf area of water stressed plants. There was significant decrease in leaf

area with the employment of water stress, which was significantly addressed by SA

application. Interactions among various treatment factors were also statistically

significant for leaf area of sunflower hybrids under foliar application of salicylic acid

in different treatments at two growth stages.


sunflower, at which SA foliar application was made, has been drawn in Figure 3.7 a.

Sunflower hybrid plants differed more if SA was applied at vegetative stage even all

giving greater leaf area as compared to that with SA applied at flowering growth

stage. H-1 (NX-19012) showed significantly better results while H-3 (FH-352)

performed the least, and the difference of both was statistically significant with H-2

(NX-00989)) giving intermediate results. Similarly, response of drought stressed

sunflower hybrids to SA application at two growth stages was significant (Figure 3.7

b). The SA application with 0.75 mM concentration at both stages produced

significantly greater leaf area of drought stressed plants as compared to those

receiving no SA. In control (DS-0 + SA-0) having non stressed plants, the value for

leaf area was highest, and it was significantly superior to all the treatments in drought

stressed sunflower hybrids plants. Within all the treatments, interaction of

200

DS with various SA concentrations showed better response of sunflower hybrids to

SA foliar application at flowering stage. Remedial effect of salicylic acid on drought-

stressed sunflower plants increased by elevating the SA concentration to the level of

0.75 mM, after which it reduced at further higher 1.50 mM concentration.

Leaf area (LA) indicates the size of the assimilatory system of the crop. Water

stress at both the stages resulted in decreased LA (Table 3.7). Leaf area reduction

occurred only due to decreased leaf expansion rate (leaf size) because leaf score

remained unaffected in this study (Table 3.6). Reduction in leaf expansion rate might

be the result of decreased turgor potential which may have resulted in the reduced

cell elongation and thus leaves could not attain the full size. Similarly, Sadras et al.

(1993) also reported reduction in leaf expansion rate due to decreased water potential

under water stress in sunflower. Similarly during water stress, total leaf area

decreased significantly in sorghum (Yadave et al., 2005).

Photosynthesis and dry matter yield depends on the optimal development of

leaf area. Drought stress mostly decreases leaf growth and in turns the leaf areas in

many plant species (Jaleel et al., 2009, Lazaridou et al., 2003). Leaf turgor,

temperature and assimilating supply for growth are important for leaf area

expansion. Drought-induced reduction in leaf area is ascribed to suppression of leaf

expansion through reduction in photosynthesis (Anjum et al., 2011a). Reduction of

leaf area in bean under water stress conditions has been mentioned in many studies

(Emam et al., 2010; Nielsen and Nelson, 1998).

Under drought conditions SA treatment improves leaf area more in tolerant

hybrids (Fig. 3.7 a, b). SA treatment under water stress causes maintenance of

relative water contents and photosynthesis so improves leaf area. Similarly,

increasing of leaf area under treatment with SA has been reported in pearl millet

201

(Mathur and Vyas, 2007) and wheat (Hayat et al., 2005). Khan et al. (2003) reported

increased leaf areas in corn and soybean through enhancing photosynthesis by foliar

application of SA. Treatment of 0.1 mM salicylic acid increased the average leaf area

in soybean seedlings (Lian et al., 2000).

Table 3.7: Leaf area (cm2/plant) of drought stressed sunflower hybrids under





H-1 (NX-19012) 137 a 128b 133A

H-2 (NX-00989) 130 b 124c 127B

H-3 (FH-352) 125c 121d 123C


Control (DS-0 + SA-0) 153 a 151 a 152 A

DS + SA-0 110f 101g 105 D

DS + SA-0.375 127d 120e 118 C

DS + SA-0.75 136b 130 c 129 B

DS + SA-1.50 127d 122e 118 C


H-1 × DS-0 + SA-0 165 a 160 b 163 A

H-1 × DS + SA-0 112 nop 100 r 106 I

H-1 × DS + SA-0.375 132 ef 125 ij 129 E

H-1 × DS + SA-0.75 141 d 134 ef 138 C

H-1 × DS + SA-1.50 133 k 124 j 128 E

H-2 × DS-0 + SA-0 147 c 147 c 147 B

H-2 × DS + SA-0 109 p 100 r 100 J

H-2 × DS + SA-0.375 128 hi 122 jk 122 F

H-2 × DS + SA-0.75 136 e 131 fgh 131 D

H-2 × DS + SA-1.50 129 gh 122 jk 122 F

H-3 × DS-0 + SA-0 146 c 147 c 147 B

H-3 × DS + SA-0 110 op 104 q 110 H

H-3 × DS + SA-0.375 119 kl 114 no 119 G

H-3 × DS + SA-0.75 131 fgh 125 bc 131 D

H-3 × DS + SA-1.50 118 lm 115 mn 118 DE

Means (GS) 131A 125B

202



DS=Drought Stress

a. Hybrids × Growth stages


Figure 3.7: Leaf area of drought stressed sunflower hybrids under foliar applied


75

90

105

120

135

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids

Vegetative Flowering Leaf

area

( cm 2 /plant)

60

75

90

105

120

135

150

165




203

4.3.8 Water Potential

Regarding water potential of water stressed sunflower hybrids there was

significant difference between two growth stages of sunflower plants (Table 3.8).

Foliar spray of SA at vegetative growth stage resulted in higher (less negative) water

potential as compared to that with flowering stage (more negative). Water stressed

three sunflower hybrids differed non significantly with each other for their water

potential. However, there was significant difference among different concentrations

of salicylic acid application. The highest water potential (0.71 Mpa) was recorded in

control (DS-0+SA-0) without receiving both water stress and salicylic acid treatment

followed by that under DS+SA-0.75 (0.88 -Mpa).The lowest water potential (0.99 -

Mpa) was with DS+SA-0 treatment (water stress without SA application). There was

significant decrease of water potential with water stress, which was significantly

improved by salicylic acid applications.

Interactions of all the treatment factors were statistically non significant.

Comparison of three sunflower hybrids with two growth stages of sunflower

at which DS was employed along with SA foliar application is shown in Figure 3.8

a. Sunflower hybrids differed slightly with higher values of water potential if SA

was applied at vegetative growth stage as compared to that at flowering stage. The

H-1 and H-2 showed slightly lower value than H-3 although the difference was

statistically non significant. However, differential response of water stressed

sunflower hybrids to foliar application of SA at two growth stages was significant

(Figure 3.8 b). The SA application in 0.75 mM concentration at both stages caused

significantly higher water potential of water stressed plants as Table 3.8: Water

potential (-Mpa) of drought stressed sunflower hybrids under foliar applied



204



H-1 (NX-19012) 0.85 NS 0.90 0.87B

H-2 (NX-00989) 0.87 0.90 0.88 A

H-3 (FH-352) 0.82 0.91 0.86 C


Control (DS-0 + SA-0) 0.71g 0.72 gf 0.71D

DS + SA-0 0.90 c 1.09 a 0.99 A

DS + SA-0.375 0.88 d 0.92 b 0.90 B

DS + SA-0.75 0.87 e 0.89 cd 0.88C

DS + SA-1.50 0.87 e 0.91bc 0.89C


H-1 × DS-0 + SA-0 0.71 NS 0.72 0.72 NS

H-1 × DS + SA-0 0.90 1.07 0.99

H-1 × DS + SA-0.375 0.89 0.92 0.91

H-1 × DS + SA-0.75 0.88 0.89 0.89

H-1 × DS + SA-1.50 0.88 0.91 0.90

H-2 × DS-0 + SA-0 0.72 0.71 0.72

H-2 × DS + SA-0 0.93 1.09 1.01

H-2 × DS + SA-0.375 0.92 0.93 0.93

H-2 × DS + SA-0.75 0.90 0.89 0.90

H-2 × DS + SA-1.50 0.89 0.90 0.90

H-3 × DS-0 + SA-0 0.69 0.73 0.71

H-3 × DS + SA-0 0.86 1.10 0.98

H-3 × DS + SA-0.375 0.84 0.92 0.88

H-3 × DS + SA-0.75 0.84 0.90 0.87

H-3 × DS + SA-1.50 0.85 0.91 0.88

Means (GS) 0.85 B 0.91 A



DS=Drought Stress

205



Figure 3.8: Water potential of drought stressed sunflower hybrids under foliar


compared to those receiving no concentration of SA. Whereas, DS-0+SA-0 got the

maximum value for water potential, which was significantly higher from all other

treatments in water stressed sunflower hybrids. Within all the treatments, interaction

of DS with various SA concentrations showed better response of sunflower hybrids

to SA foliar application at vegetative growth stage. Thus, remedial effect of salicylic

0.60

0.80

1.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


0.00

0.20

0.40

0.60

0.80

1.00

1.20




206

acid on water-stressed sunflower plants was significantly enhanced with increasing

SA concentration, being highest at 0.75

mM.

Results showed that the leaf water potential became more negative by

imposing water stress at vegetative and flowering stage, flowering being the most

sensitive stage (Table 3.8). These findings are in line with those of Lopez et al.

(2002) who found decrease in water potential upon the imposition of water stress.

The reduction in water potential due to moisture stress has been reported previously

in sunflower (Sgherri and Navari-izzo, 1995) and in other crops such as cotton (Meek

et al., 2003) and maize (Atteya, 2003). Salt stress significantly lowered the leaf water

potential (more negative values) of all 10 lines of Safflower (Siddique and Ashraf,

2008). Plant water relations are greatly influenced by relative water content, leaf


canopy temperature (Siddique et al., 2001).

Fig. (3.8 a, b) showed that SA application increased the water potential under

stress conditions. Mamenko and Iaroshenko (2009) determined that the treatment of

plants by salicylic acid contributes to a decrease of water loss and intensity of lipid

peroxidation in the winter wheat leaves under drought conditions.

Exogenous application of plant growth regulators under drought conditions increase

the water potential, and improve chlorophyll content (Zhang et al., 2004).

4.3.9 Osmotic Potential

Water stress and foliar application of salicylic acid at two growth stages of

sunflower plants had significant difference in osmotic potential with more negative

values for DS and SA treatments at flowering stage (Table 3.9). However, water

207

stressed three sunflower hybrids differed non significantly with each other for

osmotic potential under foliar application of salicylic acid in different concentrations

at two growth stages. Water stress caused a significant decrease in osmotic potential.

The highest reading of osmotic potential (1.28 -Mpa) was recorded for plants without

water stress and no salicylic acid treatment (DS-0+SA0), while the lowest (1.42 -

Mpa) was in treatment (DS-0+SA-0.75). It was observed that among various

concentrations of salicylic acid application there was no difference for addressing

the negative effect of water stress. Interactions among various treatment factors were

statistically non significant. Interactive effect of three sunflower hybrids with two

growth stages of sunflower at which SA foliar application was made, has been drawn

in Figure 3.9 a. It indicates that sunflower hybrids differed more if SA was applied

at vegetative growth stage even all giving higher (less negative) osmotic potential as

compared to that at flowering stage. Genotype H-1 and H-2 showed slightly different

osmotic potential at both stages, different osmotic potential at both stages, while H-

3 (FH-352) gave less negative value when stress was given at vegetative growth

stage than flowering stage

Table 3.9: Osmotic potential (-Mpa) of drought stressed sunflower hybrids under





H-1 (NX-19012) 1.35 NS 1.36 1.36 NS

H-2 (NX-00989) 1.41 1.42 1.42

H-3 (FH-352) 1.32 1.37 1.34


Control (DS-0 + SA-0) 1.26 NS 1.29 1.28 C

DS + SA-0 1.36 1.39 1.37B

DS + SA-0.375 1.38 1.40 1.39 AB

DS + SA-0.75 1.41 1.43 1.42A

DS + SA-1.50 1.40 1.41 1.41A


H-1 × DS-0 + SA-0 1.27 NS 1.29 1.28 NS

H-1 × DS + SA-0 1.36 1.37 1.37

208

H-1 × DS + SA-0.375 1.37 1.38 1.38

H-1 × DS + SA-0.75 1.40 1.39 1.40

H-1 × DS + SA-1.50 1.37 1.39 1.38

H-2 × DS-0 + SA-0 1.29 1.32 1.31

H-2 × DS + SA-0 1.41 1.42 1.42

H-2 × DS + SA-0.375 1.43 1.43 1.43

H-2 × DS + SA-0.75 1.46 1.47 1.47

H-2 × DS + SA-1.50 1.45 1.44 1.45

H-3 × DS-0 + SA-0 1.23 1.27 1.25

H-3 × DS + SA-0 1.31 1.37 1.34

H-3 × DS + SA-0.375 1.33 1.39 1.36

H-3 × DS + SA-0.75 1.37 1.42 1.40

H-3 × DS + SA-1.50 1.37 1.41 1.39

Means (GS) 1.36 B 1.38 A



DS=Drought Stress

However, response of water stressed sunflower hybrids to foliar application of SA

at two growth stages was statistically significant (Figure 3.9 b). SA application in

0.75 mM concentration at both stages lowered the osmotic potential of water stressed

plants significantly as compared to those receiving no SA.Whereas control treatment

(DS-0+SA-0) rendered the lowest value. Within all the treatments, interaction of DS

with various SA concentrations showed better response of sunflower hybrids to SA

foliar application at flowering growth stage. Further, protective effect of salicylic

acid on water-stressed sunflower plants enhanced with increasing SA concentration,

being the best at 0.75 mM.

Active lowering of osmotic potential is generally considered as an adjustment

in maintaining turgor under water deficit conditions (Ludlow and Muchow, 1990)

and this process is accomplished by the addition of solutes in a process named as

osmotic adjustment (Morgan, 1984). The results clearly shows drop in leaf osmotic

potential in response of drought stress espicially at flowering stage (Table 3.9).

209

Lower leaf water contents directly decrease the leaf osmotic potential (Table 3.11),

low leaf water potential (Table 3.8) and more accumulation of solutes like proline

(Table 3.17). These results are in line with the findings of Atteya, (2003) who found

reduction in leaf osmotic potential, lowering of water potential and RWC of corn by

imposing water stress at vegetative and tasselling stage in maize that resulted in

reduced final yield. It was observed that salinity significantly decreased (more

negative values) the leaf osmotic potential of all safflower lines of (Siddique and

Ashraf, 2008). Moisture stress significantly reduces the leaf water potential

(Siddique et al., 2000). Accumulation of solutes


1.28

1.30

1.32

1.34

1.36

1.38

1.40

1.42

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


1.15

1.20

1.25

1.30

1.35

1.40

1.45




210


Figure 3.9: Osmotic potential of drought stressed sunflower hybrids under


within cell resulted in osmotic adjustment, which lowers the osmotic potential and

helps maintain turgor of plants under water stress (Ludlow and Muchow, 1990).

Exogenous application of SA further lowered the leaf osmotic potential in

sunflower hybrids at flowering stage (Fig. 3.9 a, b). It might be due to the

accumulation of compatible solutes like proline (Table 3.17) and hence showing

osmotic adjustment. Addition of SA (0.05 mM) ameliorated the harmful effects of

osmotic stress, especially in drought resistant cultivar (Marcińska et al., 2013).

4.3.10 Turgor Potential

Turgor potential of sunflower plants had significant difference for water

stress along with foliar SA application at two growth stages of sunflower hybrids

(Table 3.10). Significantly higher turgor potential was recorded with SA application

at vegetative growth stage as compared to that at flowering stage. Water stressed

three sunflower hybrids differed non significantly with each other for turgor potential

under foliar application of SA. However, there was significant difference among

different concentrations of salicylic acid application. The highest turgor potential

(0.57 Mpa) was recorded with DS-0+SA-0 (no stress +no salicylic acid), while the

lowest (0.38 Mpa) was under DS+SA-0 treatment. Application of salicylic acid at

the concentration of 0.75 mM resulted the highest (0.53 MPa) and significant

increase of turgor potential in water stressed plants. There was significant decrease

in turgor potential with the employment of water stress, which was highly increased

211

by SA application. Interactions were statistically non significant for H×GS, SA×GS,

H×SA, and H×SA×GS.

Table 3.10: Turgor potential (Mpa) of drought stressed sunflower

hybrids under foliar applied various concentrations of salicylic acid at two

growth stages.




H-1 (NX-19012) 0.50 NS 0.46 0.48B

H-2 (NX-00989) 0.54 0.51 0.53 A

H-3 (FH-352) 0.51 0.46 0.49 B


Control (DS-0 + SA-0) 0.56b 0.57a 0.57 A

DS + SA-0 0.46e 0.30f 0.38 E

DS + SA-0.375 0.49d 0.48 0.49 D

DS + SA-0.75 0.54c 0.53 c 0.53 BC

DS + SA-1.50 0.52c 0.51cd 0.52 C


H-1 × DS-0 + SA-0 0.56 NS 0.57 0.57B

H-1 × DS + SA-0 0.45 0.30 0.38 K

H-1 × DS + SA-0.375 0.48 0.46 0.47 I

H-1 × DS + SA-0.75 0.52 0.50 0.51 F

H-1 × DS + SA-1.50 0.49 0.48 0.49 GH

H-2 × DS-0 + SA-0 0.57 0.61 0.59 A

H-2 × DS + SA-0 0.48 0.33 0.41 J

H-2 × DS + SA-0.375 0.51 0.50 0.51 F

H-2 × DS + SA-0.75 0.56 0.56 0.56 B

H-2 × DS + SA-1.50 0.56 0.54 0.55 BC

H-3 × DS-0 + SA-0 0.54 0.54 0.54 CD

H-3 × DS + SA-0 0.45 0.27 0.36 L

H-3 × DS + SA-0.375 0.49 0.47 0.48 HI

H-3 × DS + SA-0.75 0.53 0.52 0.53 DE

H-3 × DS + SA-1.50 0.52 0.50 0.51 F

Means (GS) 0.52A 0.48B



DS=Drought Stress

Graphical presentation of turgor potential for the interactive effect of three

sunflower hybrids with two growth stages of sunflower is given in Figure 3.10 a.

212

Sunflower hybrids differed more for turgor potential if SA was applied at flowering

growth stage, while there was no difference among them when DS / SA treatments

were given at vegetative stage. The H-2 (NX-98900) performed better than the other

two hybrids at both growth stages. However, the response of water stressed

sunflower hybrids to SA application at two growth stages was statistically significant

(Figure 3.10 b). Application of SA with 0.75 mM concentration at vegetative stage

rendered significantly higher turgor potential in water stressed plants as compared to

those receiving no or lower dose of SA. After that, DS+SA-0 rendered the lowest

value for turgor potential of stressed plants at flowering stage. Interaction of DS with

various SA concentrations showed better response of sunflower hybrids to SA foliar

application at vegetative growth stage. Further, protective effect of salicylic acid on

water-stressed sunflower plants enhanced with increasing SA concentration, being

highest at the 0.75 mM, and lesser at next higher concentration.

Turgor maintenance plays an important role in drought tolerance of plants

which may be due to its involvement in stomatal regulation and hence photosynthesis

(Ludlow et al., 1985). The results of present study showed that water stress at both

the stages significantly reduced the leaf turgor potential. Minimum leaf turgor

potential was recorded when crop faced water stress at flowering stage (Table 3.10).

Plant water relations are greatly influenced by relative water content, leaf water

potential, stomatal resistance, rate of transpiration, leaf temperature and canopy

temperature. Water potential, osmotic potential, turgor potential and relative water

content are the plant components. Leaf

213



Figure 3.10: Turgor potential of drought stressed sunflower hybrids under


turgor potential significantly reduced in all lines due to salt stress in Safflower

(Siddique and Ashraf, 2008).

Generally, leaf water potential decreases with water stress intensity (Galle et

al., 2002). In most plants, osmoregulation through the accumulation of solutes has

the function of reducing the osmotic potential of the cell in order to maintain cell

turgor and growth (Mafakheri et al., 2010).

0.42

0.44

0.46

0.48

0.50

0.52

0.54

0.56

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


0.00

0.10

0.20

0.30

0.40

0.50

0.60

0.70




214

4.3.11 Relative Water Content

As Table 3.11 for relative water content (RWC) indicates, there was

significant difference between two growth stages of sunflower plants at which water

stress (DS) along with foliar SA were employed. The RWC in the plants was greater

if SA was applied at vegetative growth stage. Water stressed three sunflower hybrids

also differed significantly with each other for RWC under DS and SA application.

Genotype H-2 (NX-00989) showed statistically greater RWC (77.0 %) than that of

other two and the lowest in H-3 (FH-352). Similarly, there was significant difference

among different concentrations of salicylic acid application to water stressed

sunflower hybrids. The highest RWC (82.3%) was recorded in control receiving no

water stress or salicylic acid. The lowest (73.3%) was with DS+SA-0 (water stressed

but receiving no salicylic acid). Application of SA with the concentration of 0.75

mM resulted in the highest and significant increase of RWC in water stressed plants.

There was significant decrease in RWC with the employment of water stress, which

was significantly addressed by SA application. Interactions were statistically non

significant for H×GS, and SA×GS; while significant for H×SA, and H×SA×GS.

Table 3.11: Relative water content (%) of water stressed sunflower hybrids


stages.




H-1 (NX-19012) 76.8 NS 76.1 76.5 B

H-2 (NX-00989) 77.5 76.4 77.0 A

H-3 (FH-352) 75.7 75.0 75.4 C


Control (DS-0 + SA-0) 82.8 NS 81.9 82.3 A

DS + SA-0 73.8 72.8 73.3 D

DS + SA-0.38 74.4 73.9 74.1 C

DS + SA-0.75 77.7 77.1 77.4 B

DS + SA-1.50 74.7 73.5 74.1 C

215


H-1 × DS-0 + SA-0 82.3 b 81.9 b 82.1 B

H-1 × DS + SA-0 75.5 def 73.2 ghi 74.3 E

H-1 × DS + SA-0.38 74.1 fg 74.2 fg 74.1 E

H-1 × DS + SA-0.75 77.1 cd 76.5 cde 76.8 D

H-1 × DS + SA-1.50 75.2 d-g 74.9 efg 75.0 E

H-2 × DS-0 + SA-0 84.6 a 82.9 ab 83.7 A

H-2 × DS + SA-0 74.1 fg 73.5 f-i 73.8 EF

H-2 × DS + SA-0.38 75.1 d-g 74.0 fg 74.6 E

H-2 × DS + SA-0.75 78.5 c 77.9 c 78.2 C

H-2 × DS + SA-1.50 75.4 def 73.9 fgh 74.7 E

H-3 × DS-0 + SA-0 81.5 b 80.9 b 81.2 B

H-3 × DS + SA-0 71.8 i 71.8 i 71.8 G

H-3 × DS + SA-0.38 74.1 fg 73.4 ghi 73.7 EF

H-3 × DS + SA-0.75 77.7 c 76.9 cde 77.3 CD

H-3 × DS + SA-1.50 73.6 f-i 71.9 hi 72.7 FG

Means (GS) 76.7 A 75.8 B



DS=Water Stress

Regarding RWC, Figure (3.11 a) shows the interactive effect of three

sunflower hybrids with two growth stages of sunflower at which SA foliar

application was made. It indicates that sunflower hybrids differed more if SA was

applied at vegetative growth stage even all giving greater RWC as compared to that

at flowering stage. H-2 (NX-00989) showed slightly better results while H-3

(FH352) performed the least although the difference was statistically non significant.

Similarly, response of water stressed sunflower hybrids to foliar application of SA

at two growth stages was non significant (Figure 3.11 b). However, SA application

in 0.75 mM concentration at both stages produced significantly higher RWC of water

stressed plants as compared to those receiving no SA. Whereas, control (DS0+SA-

0) showed the highest value for RWC of non stressed plants, which was significantly

superior to all the treatments in water stressed sunflower plants.

216

Among all the treatments, interaction of DS with various SA concentrations

showed better response of sunflower hybrids to SA application at vegetative growth

stage. Positive effect of salicylic acid on RWC in water-stressed sunflower plants

was improved by increasing the SA concentration, as it was highest at 0.75 mM, but

slightly reduced with further higher (1.50 mM) concentration.

RWC indicate the internal water status of plant tissue and is a convenient

method for following changes in tissue water content without errors caused by

continually changing tissue dry weight (Erickson et al., 1991). During this study,

RWC decreased significantly by imposing moisture stress and minimum RWC were

observed at flowering stage (Table 3.11). These results are in line with the findings

of Atteya (2003), who reported that plants suffering from water stress at


74.0

75.0

76.0

77.0

78.0

79.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


217


Figure 3.11: Relative water content of drought stressed sunflower hybrids


stages.

tasselling stage had decreased RWC values compared with the plants suffering water

stress at vegetative stage and well irrigated plants in maize. Plant and cell water

balance is measured by the difference of water absorbed from the soil and

transpirational water loss to the atmosphere. RWC tend to decline when transpiration

exceeds water absorption under drought condition (Tas and Tas,

2007) leading to decrease in cell turgor Hussain et al., 2009. Maintenance of high

RWC under drought due to relatively more growth of the roots than shoots and/or

abscisic acid induced reduction in stomatal opening (Makoto et al., 1990) tends to

maintain cell turgidity, chlorophyll content (Keyvan, 2010) and photosynthesis.

Saensee et al. (2012) also reported that relative water content decreased significantly

in all sunflower genotypes with increase in water stress levels. Ullah et al. (2012)

observed a decrease of RWC on imposition of drought stress.

68.0

72.0

76.0

80.0

84.0

88.0




218

SA increased the RWC more in tolerant hybrids with 0.75mM conc. (Fig.

3.11 a, b). It has been proved that SA triggers some metabolic processes in plants as

well as affects plant water relations (Hayat et al., 2010). In this study, with that

observed in wheat (Singh and Usha, 2003) and Ctenanthes etosa (Kadioglu et al.,

2011 ) and the shallot (Ahmad et al., 2014) plants grown under drought conditions

sprayed with SA solution could maintain higher RWC compared with those of

drought stressed plants. It becomes evident that foliar application of SA and

LTryptophan had pronounced effect on RWC under water stress conditions they may

regulate stomatal openings and reduce transpirational water loss under drought

conditions enabling the plants to maintain turgor, carry on photosynthesis and be

productive under moisture deficit Rao et al. (2012). He et al., (2005) and

Sakhabutdinova et al., (2003) postulated that salicylic acid increased the production

of photosynthetic apparatus that produced more photosynthates. This enhanced

photosynthetic activity increased sap production in the leaf lamella which resulted

in maintenance of RWC in leaf and better growth.

.

4.3.12 Photosynthesis Rate

Difference between two growth stages of sunflower plants at which water

stress (DS) and foliar spray of SA were applied, was significant for photosynthesis

rate (Table 3.12). The rate of photosynthesis was greater if DS along with SA was

given at vegetative stage of sunflower plants. Water stressed three sunflower hybrids

also differed significantly with each other for photosynthesis rate, H-3 (FH-352) had

the lowest value (6.21 µmol/m2/s), while other two genotypes (H1 and H2) were

superior and statistically similar to each other for photosynthesis rate under foliar

application of salicylic acid. However, there was significant difference among

different concentrations of salicylic acid application at two growth stages. The

highest photosynthesis rate (9.66 µmol/m2/s) was recorded in control, while the

219

lowest (4.26 µmol/m2/s) was with DS+SA-0. There was significant decrease in

photosynthesis rate with the employment of water stress, which was slightly

addressed by SA spray. Application of salicylic acid at of 0.75 mM caused

significantly higher increase of photosynthesis rate of water stressed plants over

other concentrations. All interactions were statistically significant except for

sunflower hybrids (H) × growth stages (GS).

Sunflower hybrids had positive but non significant interactive

relationshioship with two growth stages of sunflower at which DS / SA foliar Table

3.12: Photosynthesis rate (µmol/m2/s) of drought stressed sunflower hybrids

under foliar applied various concentrations of salicylic acid at two growth stages.




H-1 (NX-19012) 7.47 NS 6.50 6.98 A

H-2 (NX-00989) 7.05 6.46 6.76 A

H-3 (FH-352) 6.64 5.78 6.21 B


Control (DS-0 + SA-0) 9.85a 9.53a 9.66 A

DS + SA-0 4.27 e 4.26 e 4.26 D

DS + SA-0.375 5.84cd 5.33 d 5.59 C

DS + SA-0.75 7.74b 6.54c 7.14 B

DS + SA-1.50 6.01cd 5.56 d 5.79 C


H-1 × DS-0 + SA-0 10.91ab 9.96 bc 10.43 AB

H-1 × DS + SA-0 4.52 j 4.31j 4.41 FG

H-1 × DS + SA-0.375 5.28 ij 5.61 hij 5.44 EFG

H-1 × DS + SA-0.75 8.92 cde 6.73 f-i 7.83 C

H-1 × DS + SA-1.50 7.71 def 5.90 f-j 6.80 CD

H-2 × DS-0 + SA-0 10.07 ab 9.43 bcd 10.75 A

H-2 × DS + SA-0 4.19 j 4.26 j 4.23 G

H-2 × DS + SA-0.375 6.07 f-j 5.87 f-j 5.96 DE

H-2 × DS + SA-0.75 8.86 cde 7.01 e-i 7.94 C

H-2 × DS + SA-1.50 6.05 f-j 5.72 g-j 5.88 DE

H-3 × DS-0 + SA-0 9.36 bcd 9.20 bcd 9.28 B

H-3 × DS + SA-0 4.31 j 4.20j 4.26 G

H-3 × DS + SA-0.375 5.84 f-j 4.51 j 5.18 EFG

220

H-3 × DS + SA-0.75 7.62d-g 5.86 f-j 6.74 CD

H-3 × DS + SA-1.50 6.08 f-j 5.06ij 5.57 DEF

Means (GS) 7.05 A 6.24 B



DS=Drought Stress

application was made (Figure 3.12 a). These hybrids differed more if SA was applied

at vegetative growth stage even all giving higher photosynthesis rate as compared to

that at flowering stage. Sunflower hybrids H-1 (NX-19012) and H-2 (NX-00989)

showed better response as compared with H-3 (FH-352) although the difference was

statistically non significant. Whereas, interactive effect of water stressed sunflower

hybrids with foliar application of SA at two growth stages was statistically

significant (Figure 3.12 b). SA application in 0.75 mM concentration at both stages

produced significantly higher photosynthesis rate of water stressed plants as

compared to those receiving no SA. Control (DS-0+SA-0) showed the highest value

for photosynthesis rate, which was significantly superior to all the treatments in

water stressed plants. Among all the treatment combinations, interaction of DS with

various SA concentrations showed better response of sunflower hybrids to SA

application at vegetative growth stage. Positive effect of salicylic acid on water-

stressed sunflower plants was improved by increasing SA concentration up to 0.75

mM, but further higher concentration of 1.50 mM caused comparatively lower rate

of photosynthesis.

The present study shows that water deficit significantly inhibited the

photosynthesis process in all sunflower hybrids (Table 3.12). Similarly, decline in

photosynthetic rate under drought stress was also documented by Bacelar et al.

(2007) and Ben Ahmed et al. (2009). It was reported that decreased photosynthetic

activity under drought stress might be due to stomatal or non-stomatal mechanisms

221

(Samarah et al., 2009) also explained on reduced leaf structure and reduction of

RuBP activity (Reddy et al., 2004). Moisture stress causes reduction in

photosynthesis, which results in decreased leaf expansion, impaired photosynthetic



Figure 3.12: Photosynthesis rate of drought stressed sunflower hybrids under


machinery, premature leaf senescence and associated reduction in food production

(Wahid and Rasul, 2005). Uzunova and Zlatev (2013) reported that drought seriously

4.00

5.00

6.00

7.00

8.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


2.00

4.00

6.00

8.00

10.00

12.00


Salicylic acid treatments (mM)to droughted plants


222

inhibited net photosynthetic rate (A) in cowpea. Drought significantly lowers the

plant internal water content that consequently reduces photosynthetic rate (Atteya,

2003). For screening sunflower germplasm photosynthetic ability could be used as

selection criteria for water stress tolerance, in sunflower genotypes a positive

relationship between photosynthesis and water stress tolerance was observed (Kiani

et al., 2007).

Reduction in photosynthesis was well addressed by salicylic concentrations

especially at vegetative growth stage in all hybrids (Fig. 3.12 a, b). Salicylic acid

maintains almost the same photosynthetic rate and stomatal conductance under water

stress as those of water sufficient plants. This shielding action of salicylic acid under

drought stress is associated with the reduction of transpiration rate and enhancement

of photosynthesis (Singh and Usha, 2003). Results are concomitant with previous

findings that exogenous SA application improved the growth and photosynthetic rate

in wheat under water stress (Hussein et al., 2007).Increased photosynthetic rate at

42 hours after salicylic acid or acetyl salicylic acid application to corn and soybean

plants under drought stress was accompanied by increased stomatal conductance and

transpiration rate (Khan et al., 2003).

4.3.13 Stomatal Conductance

Difference between foliar applications of salicylic acid at two growth stages of

sunflower plants was statistically significant for stomatal conductance (Table 3.13).

There was greater stomatal conductance when along with SA was sprayed at

vegatative stage of water stressed plants. Three sunflower hybrids also differed

significantly with each other for stomatal conductance under foliar application of

salicylic acid in different concentrations at two growth stages. The H-2 showed the

highest stomatal conductance (395 mmol/m²/s) while H-3 had the lowest value (340

223

mmol/m²/s). Further, there was significant difference among different concentrations

of salicylic acid applied to water stressed plants. The highest stomatal conductance

(423mmol/m²/s) was recorded in control (DS-0+SA-0), and the lowest (280

mmol/m²/s) was under DS+SA-0 treatment. Application of SA at 0.75 mM

concentration resulted in statistically best increase of stomatal conductance of water

stressed plants compared to other SA concentrations. There was significant decrease

in stomatal conductance with the employment of water stress, which was

significantly improved by salicylic acid application. Interactions were statistically

non significant for all combinations of treatment factors except the H×SA.


sunflower at which SA foliar application was made, has been drawn in Figure 3.13

a. It reflects that sunflower hybrids differed equally in response to SA application at

both growth stages, although all showing greater stomatal conductance values at

vegetative as compared to that at flowering stage. The H-2 (NX-00989) showed

significantly better results than the other two hybrids, while H-3 (FH-352) performed

the least. Similarly, differential response of water stressed sunflower hybrids to foliar

application of SA at two growth stages was statistically significant (Figure 3.13 b).

The SA application in any concentration at both stages caused significantly higher

values of stomatal conductance as compared to those receiving Table 3.13:

Stomatal conductance (mmol/m²/s) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 382 NS 375 379 B

H-2 (NX-00989) 398 391 395 A

H-3 (FH-352) 343 336 340C

224


Control (DS-0 + SA-0) 423NS 423 423 A

DS + SA-0 285 276 280 E

DS + SA-0.375 356 356 356 D

DS + SA-0.75 413 400 408 B

DS + SA-1.50 394 383 388 C


H-1 × DS-0 + SA-0 423 NS 421 422B

H-1 × DS + SA-0 272 262 267H

H-1 × DS + SA-0.375 369 376 372 D

H-1 × DS + SA-0.75 424 412 418BC

H-1 × DS + SA-1.50 420 402 411 BC

H-2 × DS-0 + SA-0 448 446 447 A

H-2 × DS + SA-0 294 284 289G

H-2 × DS + SA-0.375 381 375 378 D

H-2 × DS + SA-0.75 448 439 444 AB

H-2 × DS + SA-1.50 420 411 415BC

H-3 × DS-0 + SA-0 398 402 400C

H-3 × DS + SA-0 288 280 284 GH

H-3 × DS + SA-0.375 320 317 318F

H-3 × DS + SA-0.75 368 348 358 DE

H-3 × DS + SA-1.50 344 334 339E

Means (GS) 374A 367B



DS=Drought Stress

no SA. Whereas, control treatment (DS-0 + SA-0) rendered the highest value for

stomatal conductance, which was significantly superior to all the treatments in water

stressed sunflower hybrids. Within all the treatments, interaction of DS with various

SA concentrations showed better response of sunflower hybrids to SA foliar


water-stressed sunflower plants was enhanced with increasing SA concentration,

being the highest at 0.75 mM and followed by 1.5 mM.

One of the first responses of plants to drought is stomatal closure, restricting gas

exchange between the atmosphere and the inside of the leaf. The present findings

revealed that drought stress decreased the stomatal conductance

225

(gs) in sunflower hybrids (Table 3.13). Similar findings were reported by Mafakheri

et al. (2010) that transpiration and stomatal conductance reduced in all varieties of

chick pea when water stress was imposed on them. Chaves and Oliviera (2004)

concluded that gs only affect A at severe drought stress. Reduction in photosynthesis

underwater stressed plants can be ascribed both to stomatal (stomatal closure) and

non-stomatal (impairments of metabolic processes) factors. Most researchers

concurred that the stomatal closure ( Abbate et al., 2004) and the resulting CO2

deficit in chloroplast is the main cause of reduced photosynthesis under mild and

moderate stresses (Flexas and Medrano, 2002).

Salicylic acid applied under stress increased the stomatal conductance but

non significantly among hybrids and growth stages (Fig. 3.13 a, b). Salicylic acid

maintains almost the same stomatal conductance in drought stress plants as those of

water sufficient plants (Singh and Usha, 2003).


300

350

400

450

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


226


Figure 3.13: Stomatal conductance of drought stressed sunflower hybrids under


Salicylic acid application at the rate of 10-3 or 10-5 mol/L to corn and soybean plants

under drought stress increased or unchanged the stomatal conductance level and

transpiration rate (Khan et al., 2003). Hamada and AlHakimi (2001) reported that

soaking of wheat grain in 100 ppm SA was generally effective in reducing the

drought effects on growth and transpiration rate.

4.3.14 Leaf Diffusive Resistance

Difference between two growth stages of sunflower plants for SA application

was significant with respect to leaf diffusive rate (Table 3.14). Water stress and SA

spray at vegetative growth stage produced significantly higher diffusive rate (2.07

sec/cm) in sunflower hybrids. Water stressed three sunflower hybrids also differed

significantly with each other for leaf diffusive rate under foliar application of

salicylic acid. H-1 (NX-19012) and H-2 (NX-00989) had the similar higher rates

(2.01 sec/cm), while lower (1.96 sec/cm) in H-3 (FH-352) genotype. Further, there

was significant difference between control and water stressed sunflower plants. The

lowest value (1.48 sec/cm) was detected in control (without receiving both water

200

250

300

350

400

450




227

stress and salicylic acid), while the highest (2.13 sec/cm) was with DS+SA-0.75non

significantly different with other treatments. There was significant increase in

diffusive rate with the employment of water stress. Interactions were statistically

significant for: sunflower hybrids (H) × growth stages (GS) and SA ×GS while non

significant in H×SA and H×SA×GS.

Interaction of three sunflower hybrids with SA application at two growth

stages of sunflower was significant (Figure 3.14 a). Generally, all the genotypes had

lower leaf diffusive rate if water stressed / sprayed with salicylic acid at flowering

stage as compared to vegetative stage. The differential response of water Table

3.14: Leaf diffusive resistance of drought stressed sunflower hybrids under





H-1 (NX-19012) 2.09 b 1.92d 2.01A

H-2 (NX-00989) 2.11a 1.92d 2.01 A

H-3 (FH-352) 2.01 c 1.91d 1.96 B


Control (DS-0 + SA-0) 1.52 c 1.44 d 1.48C

DS + SA-0 2.20 a 2.03 b 2.11B

DS + SA-0.375 2.20 a 2.04 b 2.12 B

DS + SA-0.75 2.21 a 2.05 b 2.13 A

DS + SA-1.50 2.21a 2.04 b 2.12AB


H-1 × DS-0 + SA-0 1.54 NS 1.44 1.495 NS

H-1 × DS + SA-0 2.23 2.04 2.135

H-1 × DS + SA-0.375 2.23 2.03 2.135

H-1 × DS + SA-0.75 2.23 2.05 2.145

H-1 × DS + SA-1.50 2.22 2.04 2.141

H-2 × DS-0 + SA-0 1.55 1.44 1.495

H-2 × DS + SA-0 2.24 2.03 2.140

H-2 × DS + SA-0.375 2.24 2.04 2.143

H-2 × DS + SA-0.75 2.25 2.04 2.148

H-2 × DS + SA-1.50 2.25 2.04 2.148

H-3 × DS-0 + SA-0 1.47 1.45 1.460

228

H-3 × DS + SA-0 2.13 2.02 2.080

H-3 × DS + SA-0.375 2.14 2.03 2.085

H-3 × DS + SA-0.75 2.16 2.03 2.101

H-3 × DS + SA-1.50 2.14 2.03 2.088

Means (GS) 2.07 A 1.91 B



DS=Drought Stress

stressed sunflower hybrids to foliar application of SA at two growth stages was

significant only between control and stressed plants SA concentration showed no

prominent effect in increasing resistance (Figure 3.14 b). Salicylic acid spray on

water-stressed sunflower plants slightly improved with increasing SA

concentration, being best at 0.75 mM but similar to other treatments.

The present study shows that water deficit significantly enhanced the leaf

diffusive resistance in all sunflower hybrids (Table 3.12). Results are in agreement

with Khalilvand and Yarnia (2007) who showed that in sunflower stomatal resistance

increased under water stress condition as a result of relative closing of stomata,

therefore it increases the total resistance of the sunflower against its H2O movement

in comparison CO2.

Salicylic acid act as a regulator increased/ decreased the stomatal activity.

Salicylic acid applied under stress showed higher stomatal resistance in drought

stress conditions in comparison with control and significantly among hybrids and

growth stages (Fig. 3.13 a, b).

This is in accordance with our knowledge about plants reaction in water stressed

conditions. There is a close relationship between stomatal behavior and plants

survival ability under drought conditions. Stomatal closure significantly decreases

transpiration rate and results in maintaining positive turgor of the cells (Saei et al.,

229

2006). Similar results were reported by (Souza et al., 2004) in cowpea and Farzane

et al. (2014) who reported high stomatal resistance in drought stress conditions and

salicylic acid as compared to control in 10 varieties of barley.



Figure 3.14: Leaf diffusive resistance of drought stressed sunflower hybrids


stages.

s

1.8

1.8

1.9

1.9

2.0

2.0

2.1

2.1

2.2

2.2

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)


0

0.5

1

1.5

2

2.5




230

4.3.15 Chlorophyll a Content

Difference between foliar applications of salicylic acid at two growth stages

of sunflower plants was non significant for chlorophyll a content (Table 3.15). Water

stressed three sunflower hybrids differed significantly with each other for

chlorophyll a content under foliar application of salicylic acid in different

concentrations at two growth stages. However, there was significant difference

among different concentrations of salicylic acid application. The highest chlorophyll

a content (1.53 mg/g FW) was recorded in control (DS-0+SA-0) without receiving

both water stress and salicylic acid, while the lowest (1.16 mg/g

FW) was with DS+SA-0 (water stressed but receiving no salicylic acid). Application

of salicylic acid at the concentrations of 0.75 mM and 1.5 mM resulted the highest

increase of chlorophyll a content of water stressed plants although non significantly.

There was significant decrease in chlorophyll a content with the employment of

water stress, which was slightly addressed by salicylic acid application. Interactions

were statistically significant for: sunflower hybrids (H) × growth stages (GS); H ×

SA.



a. It indicates that sunflower hybrids differed more if SA was applied at vegetative

growth stage even all giving greater chlorophyll a content as compared to that at

flowering stage. H-2 (NX-00989) showed slightly better results while H-1

(NX19012) and H-3 (FH-352) performed somewhat similar although the difference

was statistically non significant. Similarly differential response of water stressed

Table 3.15: Chlorophyll a content (mg/g FW) of drought stressed sunflower


growth stages.



231


H-1 (NX-19012) 1.31 BC 1.30C 1.30 B

H-2 (NX-00989) 1.34 A 1.33 AB 1.34 A

H-3 (FH-352) 1.29 C 1.28 C 1.29 B


Control (DS-0 + SA-0) 1.53 NS 1.53 1.53 A

DS + SA-0 1.15 1.16 1.16 D

DS + SA-0.375 1.22 1.20 1.21 C

DS + SA-0.75 1.34 1.32 1.33 B

DS + SA-1.50 1.32 1.31 1.31 B


H-1 × DS-0 + SA-0 1.60 NS 1.60 1.60 A

H-1 × DS + SA-0 1.14 1.16 1.15 G

H-1 × DS + SA-0.375 1.19 1.17 1.18 FG

H-1 × DS + SA-0.75 1.31 1.29 1.30CD

H-1 × DS + SA-1.50 1.30 1.27 1.29 CD

H-2 × DS-0 + SA-0 1.63 1.62 1.62 A

H-2 × DS + SA-0 1.17 1.16 1.17 G

H-2 × DS + SA-0.375 1.24 1.22 1.23 EF

H-2 × DS + SA-0.75 1.36 1.33 1.34 BC

H-2 × DS + SA-1.50 1.32 1.31 1.33 BC

H-3 × DS-0 + SA-0 1.37 1.37 1.37 B

H-3 × DS + SA-0 1.15 1.16 1.15 G

H-3 × DS + SA-0.375 1.24 1.22 1.23 EF

H-3 × DS + SA-0.75 1.35 1.33 1.33 BC

H-3 × DS + SA-1.50 1.33 1.34 1.33 BC

Means (GS) 1.31 NS 1.30



DS=Drought Stress

sunflower hybrids to foliar application of SA at two growth stages was non

significant (Figure 3.15 b). However, SA application in 0.75 mM concentration at

both stages produced significantly higher chlorophyll a content of water stressed

plants as compared to those receiving no SA. Whereas, control treatment (DS-0 +

SA-0) rendered the highest value for chlorophyll a content of non stressed plants,

which was significantly superior to all the treatments in water stressed sunflower

232

hybrids plants. Within all the treatments, interaction of DS with various SA

concentrations showed better response of sunflower hybrids to SA foliar


water-stressed sunflower plants enhanced with increasing SA concentration, being

highest at the 0.75 mM, and is similar to next higher concentration of 1.50 mM.

4.3.16 Chlorophyll b Content

Chlorophyll b contents had significant difference for SA applications at two

growth stages of sunflower hybrids (Table 3.16). Foliar applications of salicylic acid

to drought stressed plants at vegetative growth stage yielded significantly greater

amount of chlorophyll b than that with SA spray at flowering stage. Similarly, three

sunflower hybrids differed significantly with lesser chlorophyll b content (0.51 mg/g

FW) in H-3 (FH-352) as compared to other two genotypes, which had no statistical

difference. Further, different concentrations of salicylic acid application to drought

stressed plants showed significant difference. There was significant decrease in

chlorophyll b content with the employment of drought stress, which was highly

improved by SA. The highest contents of chlorophyll b (0.80 mg/g FW) were found

in control (without receiving both drought stress and


1.24

1.27

1.30

1.33

1.36

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


233


Figure 3.15: Chlorophyll a content of drought stressed sunflower hybrids under


Table 3.16: Chlorophyll b content (mg/g) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 0.80 NS 0.79 0.79 A

H-2 (NX-00989) 0.80 0.79 0.79 A

H-3 (FH-352) 0.53 0.50 0.51 B


Control (DS-0 + SA-0) 0.80 NS 0.80 0.80 A

DS + SA-0 0.64 0.63 0.64 C

DS + SA-0.38 0.66 0.64 0.65 C

DS + SA-0.75 0.72 0.70 0.71 B

DS + SA-1.50 0.72 0.69 0.70 B


H-1 × DS-0 + SA-0 0.87 NS 0.85 0.86 A

H-1 × DS + SA-0 0.74 0.75 0.75 C

H-1 × DS + SA-0.38 0.76 0.77 0.76 BC

H-1 × DS + SA-0.75 0.81 0.80 0.80 B

H-1 × DS + SA-1.50 0.81 0.79 0.80 B

H-2 × DS-0 + SA-0 0.88 0.87 0.88 A

H-2 × DS + SA-0 0.74 0.74 0.74 C

1.00

1.10

1.20

1.30

1.40

1.50

1.60




234

H-2 × DS + SA-0.38 0.75 0.74 0.75 C

H-2 × DS + SA-0.75 0.81 0.80 0.80 B

H-2 × DS + SA-1.50 0.82 0.79 0.80 B

H-3 × DS-0 + SA-0 0.64 0.69 0.67 D

H-3 × DS + SA-0 0.44 0.41 0.42 F

H-3 × DS + SA-0.38 0.47 0.42 0.44 F

H-3 × DS + SA-0.75 0.55 0.51 0.53 E

H-3 × DS + SA-1.50 0.53 0.48 0.51 E

Means (GS) 0.71 A 0.69 B



DS=Drought Stress

salicylic acid), while the lowest (0.64 mg/g FW) was in plants under drought stress

but without SA application. Salicylic acid at 0.75 mM concentration caused the

highest and significant increase of chlorophyll b content in drought stressed plants.

All interactions of treatment factors were statistically non significant except for

H×SA.

Three sunflower hybrids had non significant interactive effect with two

growth stages of sunflower at which SA foliar application was made (Figure 3.16

a). Sunflower hybrids did not differ much either SA was applied at vegetative growth

stage or at flowering stage. Similarly, differential response of drought stressed


significant (Figure 3.16 b). The SA application in all concentrations at both stages

produced statistically similar chlorophyll b contents of drought stressed plants as

compared to those receiving no SA. Control treatment although showed the highest

values for chlorophyll b content of non stressed plants, however, it did not differ

significantly with any other treatment in drought stressed sunflower plants.

Nevertheless, remedial effect of salicylic acid on drought-stressed sunflower plants

was enhanced by increasing the concentration of salicylic acid.

235

The first target of ROS produced under water stress is the cell organelles like

chloroplasts, mitochondria and peroxisomes (Tayebe and Hassan, 2010). Water

stress significantly minimize the leaf chl a, chl b, and carotenoids (Ullah et al.,

2012).Table 3.15 and 3.16 revealed that chlorophyll a,b content decreased under

water stress, are similar as found by Kauser et al. (2006). The reduction in

chlorophyll under water deficit conditions might be due to the result of decreased

synthesis of the main chlorophyll pigment complexes encoded by the cab gene



0.40

0.50

0.60

0.70

0.80

0.90

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


0.40

0.50

0.60

0.70

0.80

0.90




236

Figure 3.16: Chlorophyll b content of drought stressed sunflower hybrids under


family (Allakhverdiev, et al. 2000) or destruction of chiral macro-aggregates of light

harvesting chlorophyll ‗a‘ or ‗b‘ pigment protein complexes (CHCIIs) which

protect the photosynthetic apparatus (Gussakovsky et al. 2002) or due to oxidative

damage of chloroplast lipids, pigments and proteins (Tambussi, et al. 2000).

Decrease in chlorophyll content under moisture deficit is a commonly observable

fact (Chaves, et al. 2003, Reynolds, et al. 2005). Singh, et al. (2003) found a

significant reduction under water stress in chlorophyll contents of the leaves of

mustard plant.

Foliar application of SA enhanced both chl a, b at both stages in all hybrids

of sunflower particularly in tolerant ones (Fig 3.15 a, b and 3.16 a, b). Similarly,

foliar application 100 ppm of SA followed by 15 ppm L-TRP significantly increased

chlorophyll contents over control under stress conditions Rao et al. (2012). Similar

results were also reported by Arfan et al., (2007) and Yildirim et al., (2008). Foliar

spray of SA is also involved in stomatal regulation thus controlling photosynthetic

process (Khan et al., 2003). However, the beneficial effect of SA application

depends on genotype (Bezrukova et al., 2004). SA application, usually, induced

noticeable increase in pigments content (chl ‗a‘ and chl ‗b‘) in leaves of drought

stressed shallot plants Ahmad et al. (2014). The stimulating effect of SA may be due

to the fact that SA led to increase leaf longevity on drought stressed plants by

retaining their pigment content, therefore inhibit their senescence. It was also noticed

that treatment with SA increased pigment contents in soybean (Zhao et al., 1995),

maize (Khodary, 2004), and wheat (Singh and Usha, 2003; Arfan et al., 2007))

grown under non stress or stress conditions. Salicylic acid could protect the plants

against drought stress through increasing of photosynthetic pigments (chlorophyll a,

237

b, total chlorophyll, and carotenoids), and 10 μM salicylic acid was the most

effective level (Kabiri et al., 2014). Cag et al. (2009) reported that chlorophyll and

carotenoid contents increased in canola with exogenic SA applications (0.001-10

μM). Sivakumar et al. (2002) showed that plants treated with 100 ppm salicylic acid

showed greater chlorophyll accumulation. The degree of decrease in chlorophylls

and carotenoids contents under stress is higher in the sensitive genotypes as

compared to the moderately tolerant genotypes (Parida et al., 2007). Under drought

conditions, treatment with plant growth regulators significantly improves the

chlorophyll content (Zhang et al., 2004). Din et al. (2011) observed decrease in

chlorophyll a and b content of all the Napus genotypes at both the growth stages

under drought stress. Tolerant genotype gave least decline in chlorophyll content

during the flower initiation and pod filling stage.

4.3.17 Leaf Proline Contents

Difference between two growth stages of sunflower plants for SA application

was significant with respect to proline content (Table 3.17). Drought stress and SA

spray at vegetative growth stage produced significantly higher proline content

(0.49mg/g FW) in sunflower hybrids. Drought stressed three sunflower hybrids also

differed significantly with each other for proline content under foliar application of

salicylic acid. The H-1 (NX-19012) contained the least amount of proline content

(0.39 mg/g FW), while H-2 (NX-00989) had the highest content (0.52 mg/g FW),

both differing significantly from H-3 (FH-352) genotype. Further, there was

significant difference among various concentrations of salicylic acid application to

drought stressed sunflower plants. The lowest proline content Table 3.17: Leaf

proline contents (mg/g FW) of drought stressed sunflower hybrids under foliar




238


H-1 (NX-19012) 0.44 NS 0.34 0.39 C

H-2 (NX-00989) 0.57 0.48 0.52 A

H-3 (FH-352) 0.46 0.38 0.42 B


Control (DS-0 + SA-0) 0.16 d 0.15 d 0.16 C

DS + SA-0 0.54 b 0.43 c 0.48 B

DS + SA-0.375 0.52 b 0.40 c 0.46 B

DS + SA-0.75 0.63 a 0.51 b 0.57 A

DS + SA-1.50 0.62 a 0.50 b 0.56 A


H-1 × DS-0 + SA-0 0.16 NS 0.16 0.16 H

H-1 × DS + SA-0 0.50 0.375 0.44 F

H-1 × DS + SA-0.375 0.39 0.27 0.33 G

H-1 × DS + SA-0.75 0.59 0.46 0.52 D

H-1 × DS + SA-1.50 0.58 0.45 0.52 D

H-2 × DS-0 + SA-0 0.14 0.14 0.14 H

H-2 × DS + SA-0 0.61 0.50 0.55 CD

H-2 × DS + SA-0.375 0.65 0.54 0.60 BC

H-2 × DS + SA-0.75 0.72 0.61 0.67 A

H-2 × DS + SA-1.50 0.71 0.60 0.65 AB

H-3 × DS-0 + SA-0 0.17 0.17 0.17 H

H-3 × DS + SA-0 0.51 0.41 0.46 EF

H-3 × DS + SA-0.375 0.50 0.40 0.45 F

H-3 × DS + SA-0.75 0.57 0.47 0.52 D

H-3 × DS + SA-1.50 0.56 0.46 0.51 DE

Means (GS) 0.49 A 0.40 B



DS=Drought Stress

(0.16 mg/g FW) was detected in control (without receiving both drought stress and

salicylic acid), while the highest (0.57 mg/g FW) was with DS+SA-0.75. There was

significant increase in proline content with the employment of drought stress.

Application of salicylic acid at the concentration of 0.75 mM resulted the highest

increase of proline content of drought stressed plants although nonsignificantly with

1.50 mM. Interactions were statistically non significant for: sunflower hybrids

239

(H) × growth stages (GS); and H×SA×GS.

Interaction of three sunflower hybrids with SA application two growth stages

of sunflower was non significant (Figure 3.17 a). Generally, all the genotypes had

lower proline contents if drought stressed / sprayed with salicylic acid at flowering

stage. All the sunflower hybrids responded equally to SA application at two growth

stages. However, differential response of drought stressed sunflower hybrids to foliar

application of SA at two growth stages was significant (Figure 3.17 b). The SA

application at concentrations higher than 0.375 mM caused a significant increase of

proline content sprayed at both stages of drought stress, whereas, control treatment

(DS-0+SA-0) rendered the lowest value for proline content. Within all the

treatments, interaction of DS with various SA concentrations showed better response

of sunflower hybrids to SA foliar application at vegetative growth stage. The

shielding effect of salicylic acid on drought-stressed sunflower plants improved with

increasing SA concentration, being best at 0.75 mM.

Proline accumulation increased under the water stress in all sunflower

hybrids (Table 3.17). Osmotic adjustment is the main part of physiological

a. Hybrids× Growt h stages

0.20

0.30

0.40

0.50

0.60

0.70

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


240


Figure 3.17: Leaf proline content of drought stressed sunflower hybrids under


processes through which plants react to the drought stress (Zhu, 2003; Zhao, 2008)

and proline plays important role in this adjustment. Proline accumulation in plants

in response to water deficit is generally accepted (Sakhabutindova et al., 2003).

Compatible solutes act as osmolytes as the changes in osmolity occur in external

environment and produce higher concentrations intracellular without changing the

normal internal cell metabolism (Ramanjulu and Bartels, 2002). Proline may

perform roles other than osmotic adjustments like scavenging hydroxyl ions,

stabilizing membrane and protein structure, and act as a sink for carbon and nitrogen

for stress recovery, and buffering cellular redox potential, Moreover, high levels of

proline lowers the water potentials. By lowering water potentials, the accumulation

of compatible osmolytes, involved in osmoregulation allows additional water to be

taken up from the environment, thus buffering the immediate effect of water

shortages within the organism (Kumar et al., 2001).

0.10

0.20

0.30

0.40

0.50

0.60

0.70




241

The present investigation revealed that proline accumulation enhanced by the

foliar application of SA concentrations at both stages significantly in all hybrids (Fig.

3.17 a, b).The accumulation of proline, mainly in the cytosol, often occurs in plants

under stress with a correlation between stress tolerance and proline accumulation

(Desnigh and Kanagaraj, 2007), but the relationship is not universal and may be

species dependent (Ashraf and Foolad, 2007). Proline accumulation increases by

salicylic acid treatment in wheat, oat, bean and tomato, under oxidative stresses,

(Tasgin et al., 2006). Proline can thus be considered as an important component in

the spectra of SA-induced ABA-mediated protective reactions of wheat plants in

response to water deficit, reducing the injurious effects of drought and an acceleration

of the repairing processes following stress, evidencing the protective action of SA

on wheat plants (Shakirova, 2003). The SA ameliorates the harmful effects of

osmotic stress through increased proline and carbohydrate content (Marcińska et al.,

2013). Sayyari et al. (2013) studied the effect of SA on lettuce (Lactuca sativa L.)

in drought conditions; proline increased significantly by SA application. El-Tayeb

and Naglaa (2010) suggested using SA as a potential growth regulator for improving

plant growth under water stress. SA was highly effective in mitigating the negative

effects of drought stress in canola cultivars through enhanced accumulation of

proline.

4.3.18 Soluble Sugar Contents

Total content sugars differed significantly for salicylic acid application at two

growth stages of sunflower plants (Table 3.18). Foliar application of salicylic acid at

vegetative growth stage produced significantly greater sugar content as compared to

that with SA applied at flowering stage. Three sunflower hybrids also differed

significantly as H-3 (FH-352) had statistically lower sugar content compared to other

two genotypes having non significant difference between them. Foliar application of

http://www.ncbi.nlm.nih.gov/pubmed/?term=Marci%26%23x00144%3Bska%20I%5Bauth%5D

http://www.ncbi.nlm.nih.gov/pubmed/?term=Marci%26%23x00144%3Bska%20I%5Bauth%5D

242

salicylic acid at two growth stages of drought stressed plants also showed significant

difference among different SA concentrations. The lowest sugar content (40.7 mg/g

FW) was recorded in control (DS-0+SA-0), which was followed by DS+SA-0

(drought stressed but receiving no salicylic acid) but with significant difference.

Application of salicylic acid at the concentration of 0.75 mM caused the highest and

significant increase of sugar content in drought stressed plants. There was significant

decrease in sugar content with the employment of drought stress, which was

significantly addressed by salicylic acid spray.

Table 3.18: Soluble sugar contents (mg/g FW) of drought stressed sunflower


growth stages.




H-1 (NX-19012) 71.0 NS 68.8 69.9 A

H-2 (NX-00989) 71.9 70.6 71.2 A

H-3 (FH-352) 63.7 62.1 62.9 B


Control (DS-0 + SA-0) 40.0 E 41.5 E 40.7 D

DS + SA-0 69.2 D 69.4 D 69.3 C

DS + SA-0.375 71.6 CD 70.4 D 71.0 C

DS + SA-0.75 83.9 A 78.9 B 81.4 A

DS + SA-1.50 79.6 AB 75.7 BC 77.7 B


H-1 × DS-0 + SA-0 40.2 NS 42.6 41.4 F

H-1 × DS + SA-0 70.3 71.0 70.7 CD

H-1 × DS + SA-0.375 75.6 71.6 73.6C

H-1 × DS + SA-0.75 86.9 80.8 83.9 A

H-1 × DS + SA-1.50 82.1 78.2 80.1 AB

H-2 × DS-0 + SA-0 41.6 44.3 42.9 F

H-2 × DS + SA-0 72.4 72.7 72.6 C

H-2 × DS + SA-0.375 71.7 73.6 72.7 C

H-2 × DS + SA-0.75 87.5 82.5 85.0 A

H-2 × DS + SA-1.50 86.2 79.9 83.0 A

H-3 × DS-0 + SA-0 38.1 37.5 37.8 F

H-3 × DS + SA-0 65.0 64.3 64.7 E

H-3 × DS + SA-0.375 67.6 65.9 66.7 DE

H-3 × DS + SA-0.75 77.2 73.4 75.3 BC

243

H-3 × DS + SA-1.50 70.6 69.1 69.9 CDE

Means (GS) 68.9 A 67.2 B



DS=Drought Stress

Interactions of the treatment factorts: sunflower hybrids (H) × growth stages

(GS) and H×SA×GS were statistically non significant. Interactive relation of three

sunflower hybrids with two growth stages of sunflower for SA application was

although non significant, however, it was obvious that sunflower hybrids differed

more if SA was applied at flowering stage (Figure 3.18 a). The H-3 (FH-352)

contained the lowest sugars, while H-1 (NX-19012) and H-2 (NX-00989) had

statistically higher contents although both differed non significant. Response of

drought stressed sunflower hybrids to foliar application of SA at two growth stages

was significant (Figure 3.18 b). The SA application in 0.75 mM concentration at both

stages produced significantly higher sugar content of drought stressed plants as

compared to those receiving no SA. Whereas, control treatment (DS-0+SA-0)

rendered the lowest value for sugar content, and it was significantly inferior to all

the treatments in drought stressed plants. Except in control treatment, interaction of

DS with various SA concentrations showed slightly better response of sunflower

hybrids to SA foliar application at vegetative growth stage. Positive effect of

salicylic acid on sugar content of sunflower plants under drought stress, enhanced

by increasing SA concentration being highest at the 0.75 mM, but lesser at further

higher concentration.

The present research indicates that accumulation of soluble sugar increased

under water deficit conditions in all hybrids significantly in tolerant ones (Table

3.18). This may result from increased starch degradation, synthesis by other

244

processes or decreased conversion to other products. Several researches reported an

increase in amylase activity in water stressed leaves (Keller and Ludlow, 1993).


Figure 3.18: Soluble sugar contents of drought stressed sunflower hybrids under


The enhancement of soluble sugars is strongly correlated to the attainment of drought

tolerance in plants (Hoekstra and Buitink, 2000). Soluble sugars act as

osmoprotectants as they stabilize the cellular membranes and maintain the turgor


60.0

62.0

64.0

66.0

68.0

70.0

72.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


30.0

40.0

50.0

60.0

70.0

80.0

90.0

100.0




245

potential. Xu et al. (2008) found that water stressed wheat leaves caused a decrease

in photosynthesis and required high concentrations of osmolyte synthesis especially

total soluble sugars.

Exogenous application of SA to water stressed sunflower plants presented high

levels of sugar in tolerant hybrids (Fig. 3.18 a, b). Results agreed with the findings

of El-Tayeb (2005) found an additional increase in Na, soluble proteins and soluble

sugars in salt-stressed barley grains with application of SA. With salicylic acid, the

leaves fill up more soluble sugar and proline (Szepesi, 2006). Kabiri et al. (2014)

reported that salicylic acid protected the Nigella plant against drought stress through

increase of photosynthetic pigments and soluble sugar contents, while 10 μM SA

was the most effective level.

4.3.19 Leaf Protein Contents

Difference between foliar application of salicylic acid at two growth stages

of sunflower plants was non significant for protein content (Table 3.19). However,

three sunflower genotypes differed significantly with each other for protein content

under foliar application of salicylic acid. H-2 (NX-00989) showed the highest

protein content (6.83 mg/g FW), while H-3 (FH-352) had the lowest one (5.94 mg/g

FW).Similarly, there was statistically significant difference among different

concentrations of salicylic acid application at two growth stages. The highest protein

content (7.58 mg/g FW) was recorded in control (without receiving both Table 3.19:

Protein content (mg/g FW) of drought stressed sunflower hybrids under foliar





H-1 (NX-19012) 6.60 NS 6.68 6.64 B

246

H-2 (NX-00989) 6.80 6.86 6.83 A

H-3 (FH-352) 5.94 5.94 5.94 C


Control (DS-0 + SA-0) 7.56 A 7.60 A 7.58 A

DS + SA-0 5.44 D 6.15 BC 5.80 C

DS + SA-0.375 5.78 CD 6.23 BC 6.01 BC

DS + SA-0.75 6.84 B 6.35 B 6.60 B

DS + SA-1.50 6.61 B 6.16 BC 6.30 BC


H-1 × DS-0 + SA-0 8.02 NS 8.06 8.04 NS

H-1 × DS + SA-0 5.28 6.25 5.77

H-1 × DS + SA-0.375 5.51 6.29 5.90

H-1 × DS + SA-0.75 7.23 6.51 6.87

H-1 × DS + SA-1.50 6.98 6.31 6.64

H-2 × DS-0 + SA-0 8.14 8.15 8.14

H-2 × DS + SA-0 5.89 6.49 6.19

H-2 × DS + SA-0.375 5.90 6.61 6.26

H-2 × DS + SA-0.75 7.20 6.66 6.93

H-2 × DS + SA-1.50 6.89 6.39 6.64

H-3 × DS-0 + SA-0 6.52 6.58 6.55

H-3 × DS + SA-0 5.15 5.71 5.43

H-3 × DS + SA-0.375 5.94 5.78 5.86

H-3 × DS + SA-0.75 6.11 5.87 5.99

H-3 × DS + SA-1.50 5.97 5.80 5.87

Means (GS) 6.45 NS 6.50



DS=Drought Stress

drought stress and salicylic acid), while the lowest (5.80 mg/g FW) was with

DS+SA-0. Application of salicylic acid at the concentration of 0.75 mM resulted the

highest increase of protein content of drought stressed plants although non

significantly differing with DS+SA-1.50.

There was significant decrease in protein content with the imposition of

drought stress, which was significantly improved by SA application. Treatment

247

interactions were statistically non significant except for concentrations of salicylic

acid (SA) × sunflower growth stages (GS).

As shown in Figure 3.19 a, interactive effect of three sunflower hybrids with

two sunflower growth stages of SA spray was statistically non significant. Sunflower

hybrids showed no observable difference when SA was applied at vegetative and

flowering growth stages. The tolerant hybrids showed significantly better results,

while H-3 (FH-352) performed the least. Similarly, differential response of drought

stressed sunflower hybrids to foliar application of SA at two growth stages was


both stages produced significantly higher protein content of drought stressed plants

as compared to those receiving no SA. Whereas, control treatment without drought

stress (DS-0+SA-0) rendered the highest value for protein content of sunflower

plants. Within all the treatments, interaction of DS with various SA concentrations

showed better response of sunflower hybrids to SA foliar application at vegetative

growth stage except DS+0.375 which showed no response at flowering stage.

Salicylic acid improved the recovering impact on drought-stressed sunflower plants,

being maximum with 0.75 mM concentration


4.00

5.00

6.00

7.00

8.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


248


Figure 3.19: Protein content of drought stressed sunflower hybrids under foliar

applied various concentrations of salicylic acid at two growth stages. beyond

which there was no further increase in protein contents.

The present investigation showed that leaf protein decreased under drought

stress (Table 3.19). Increased /decreased levels of amino acids and protein caused by

water stress have been mentioned in many reports which stated that it depends on

the stress level and type of plant. A stress event which inhibits cell division and

expansion, and thus leaf expansion, will also arrest protein synthesis. For instance,

in Avena coleoptiles water deficit caused a significant decrease in rate of protein

synthesis (Dhindsa and Cleland, 1975).

Results also indicated that SA foliar application augmented the leaf protein

and reduced the effect of water stress and degree of decrease in protein contents

under drought was higher in the sensitive genotype as compared to the tolerant one.

(Fig. 3.19 a, b). Exogenous application of SA protects the plant against drought

4.00

5.00

6.00

7.00

8.00




249

stress by increasing the protein content, and 10 μM is the best level (Kabiri et al.,

2014). Water stress both at vegetative and flowering stage badly affected the growth

and yield components of sunflower crop but it caused more damage at flowering

stage (Ahmad et al., 2009). While, exogenous SA application significantly

ameliorated the negative effects of moisture stress at both stages. Application of 100

ppm SA enhanced the contents of total soluble proteins and grain proteins

(Sivakumar et al., 2002).

4.3.20 Amino Acid Content

Foliar application of salicylic acid to drought stressed plants at two growth

stages of sunflower showed non significant difference for amino acid contents (Table

3.20). However, three sunflower hybrids differed significantly among themselves for

amino acid content under SA application in various concentrations at two growth

stages. The H-2 (NX-00989) had significantly higher amino acid contents as

compared to other two sunflower hybrids. Also, there was significant difference

among various concentrations of salicylic acid sprayed. The lowest contents of

amino acids (8.1 µmol/g) were recorded in control treatment having no drought

stress and without SA application (DS-0+SA-0), whereas the highest content (17.0

µmol/g) was with DS+SA-0.75. Application of salicylic acid at the concentration of

0.75 mM caused the maximum and significant increase in amino acid content of

drought stressed plants. All the treatment combinations (H×GS; SA×GS; H×SA; and

H×SA×GS) showed statistically significant interactions among themselves.

Figure 3.20a illustrates the interaction between three sunflower hybrids and

two growth stages of sunflower at which DS and SA were employed. It is obvious

that sunflower hybrids differed more if SA was applied at flowering growth stage

although they had lower amino acid content as compared to that for vegetative stage.

250

H-2 (NX-00989) showed significantly better results, while H-1 (NX-19012) and H-

3 (FH-352) had statistically non significant difference. All the genotypes showed

statistically greater amino acid contents for DS and SA application at vegetative

stage as compared to that for flowering stage. Similarly, comparative response of

drought stressed sunflower hybrids to foliar application of SA in various

concentrations at two growth stages was statistically significant (Figure

3.20 b).

Table 3.20: Amino acid content (µmol/g) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 14.5 c 14.0 e 14.2 B

H-2 (NX-00989) 15.0 a 14.8 b 14.9 A

H-3 (FH-352) 14.4 c 14.1 d 14.2 B


Control (DS-0 + SA-0) 8.2 h 7.9 i 8.1 E

DS + SA-0 15.6 e 15.2 g 15.4 D

DS + SA-0.375 15.9 d 15.4 f 15.7 C

DS + SA-0.75 17.2 a 16.9 b 17.0 A

DS + SA-1.50 16.2 c 15.9 d 16.1 B


H-1 × DS-0 + SA-0 8.3 n 8.0 o 8.2 H

H-1 × DS + SA-0 15.3 jk 14.8 m 15.0 G

H-1 × DS + SA-0.375 15.9 gh 15.1 kl 15.5 EF

H-1 × DS + SA-0.75 16.9 cd 16.6 ef 16.8 B

H-1 × DS + SA-1.50 16.0 gh 15.3 jk 15.7 E

H-2 × DS-0 + SA-0 8.3 n 8.2 no 8.3 H

H-2 × DS + SA-0 16.4 f 15.8 gh 16.1 C

H-2 × DS + SA-0.375 16.4 f 16.0 gh 16.2 C

H-2 × DS + SA-0.75 17.5 a 17.1 bc 17.3 A

H-2 × DS + SA-1.50 16.6 ef 16.7 de 16.7 B

H-3 × DS-0 + SA-0 8.0 o 7.6 p 7.80 I

H-3 × DS + SA-0 15.1 kl 15.0 lm 15.1 G

H-3 × DS + SA-0.375 15.4 ij 15.2 jkl 15.3 F

H-3 × DS + SA-0.75 17.3 ab 16.8 cde 17.1 A

H-3 × DS + SA-1.50 16.1 g 15.7 hi 15.9 D

251

Means (GS) 14.6 NS 14.3



DS=Drought Stress


Figure 3.20: Amino acid content of drought stressed sunflower hybrids under


All the DS+SA treatments differed significantly with each other, and DS+SA

employment at vegetative stage caused higher amino acid contents as compared to


13.5

14.0

14.5

15.0

15.5

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


6.0

9.0

12.0

15.0

18.0




252

that for flowering stage. The SA application in 0.75 mM concentration at both stages

produced significantly higher amino acid content of drought stressed plants as

compared to those receiving no SA or its lower / higher concentration. Whereas,

control treatment (DS-0+SA-0) rendered the lowest value for amino acid content of

non stressed plants, which was significantly inferior to all the treatments in drought

stressed sunflower hybrids. Among all the treatments, interaction of DS with various


application at vegetative growth stage. Positive effect of salicylic acid applicationon

drought-stressed sunflower plants for improving amino acid contents increased by

enhancing the SA concentration up to 0.75 mM, and further increase in SA

concentration has significantly lesser impact.

Changes of amino acids and protein have been documented in many reports

which have stated that different responses caused by water stress depend on the level

of stress and plant type. The free amino acid accumulation other than proline in

leaves of drought stressed sunflower plants was found higher at both stages as

compared to control plants (Table 3.20). The increased levels of the amino acid

might be due to the breakdown of proteins by the action of reactive oxygen species

(ROS), produced at water stress conditions, enhance the decomposition of proteins

(Yazdanpanah et al., 2011). These osmolytes play a major role in osmotic adjustment

and protect the cell by scavenging ROS (Pinhero et al., 2001). Total free amino acids

contents are augmented in drought stressed plants, and tend to decrease during the

period of rehydration (Parida et al., 2007). Treshow (1970) concluded that water

stress inhibited amino acid utilisation and protein synthesis. While amino acid

synthesis was not impaired, the cellular protein levels decreased and since utilisation

of amino acids was blocked, amino acids accumulated, giving a 10- to 100-fold

253

accumulation of free asparagine. Valine levels increased, and glutamic acid and

alanine levels decreased.

Foliar spray of SA significantly enhanced the amino acid concentration (Fig.

3.20 b). Similar results were given by El-Tayeb (2005) that all amino acids increased

with salicylic acid in maize plants. Azimi et al. (2013) showed that water stress

reduced all attributes of growth and yield, but amino acid and salicylic acid reduced

negative effects of water deficit on wheat.

4.3.21 Superoxide Dismutase Activity

Table 3.21 shows the difference between two growth stages of sunflower

plants for foliar application of salicylic acid to water stressed sunflower plants

regarding the activity of superoxide dismutase (SOD) in them. Salicylic acid spray

at flowering stage resulted in significantly higher SOD activity as compared to

vegetative growth stage. Three sunflower hybrids also differed significantly with

each other for SOD activity under foliar application of salicylic acid at two growth

stages. The H-2 (NX-00989) genotype had the highest SOD activity (21.1 U/g FW),

while H-3 (FH-352) had the lowest one (17.5 U/g FW). Further, different

concentrations of salicylic acid application showed significant difference among

them. The lowest SOD activity (14.1 U/g FW) was detected in control (DS-0+SA0),

while the highest (22.5 U/g FW) was underDS+SA-0.75 treatment. Application of

salicylic acid at the concentration of 0.75 mM resulted in the highest and

Table 3.21: Superoxide dismutase activity (U/g FW) of drought stressed

sunflower hybrids under foliar applied various concentrations of salicylic acid

at two growth stages.




H-1 (NX-19012) 16.3 D 22.2 B 19.2 B

H-2 (NX-00989) 16.4 D 25.4 A 20.9 A

254

H-3 (FH-352) 15.4 D 19.4 C 17.4 C


Control (DS-0 + SA-0) 14.0 f 14.3f 14.1 C

DS + SA-0 16.5 de 22.3 c 19.4 B

DS + SA-0.375 16.1 e 23.5 bc 19.8 B

DS + SA-0.75 18.0 d 27.1 a 22.5 A

DS + SA-1.50 15.8 e 24.4 b 20.1 B


H-1 × DS-0 + SA-0 14.5 hi 14.2 hi 14.3 IJ

H-1 × DS + SA-0 17.2 fgh 20.9 cde 19.0 EFG

H-1 × DS + SA-0.375 16.7 fgh 21.1 cde 18.9 EFG

H-1 × DS + SA-0.75 18.2 efg 28.0 ab 23.1 AB

H-1 × DS + SA-1.50 14.9 hi 26.9 b 20.9 CDE

H-2 × DS-0 + SA-0 14.9 hi 15.3 ghi 15.1 HI

H-2 × DS + SA-0 16.9 fgh 26.7 b 21.8 BC

H-2 × DS + SA-0.375 15.8 gh 27.2 b 21.5 BCD

H-2 × DS + SA-0.75 18.9 ef 30.5 a 24.7 A

H-2 × DS + SA-1.50 15.7 gh 27.3 b 21.5 BCD

H-3 × DS-0 + SA-0 12.5 j 13.4 i 13.0 J

H-3 × DS + SA-0 15.5 ghi 19.5 def 17.5 GH

H-3 × DS + SA-0.375 15.7 gh 22.1 cd 18.9 EFG

H-3 × DS + SA-0.75 16.8 fgh 22.8 c 19.8 DEF

H-3 × DS + SA-1.50 16.7 fgh 19.1 def 17.9 FGH

Means (GS) 16.1 B 22.3 A



DS=Drought Stress

significant increase of SOD activity in water stressed plants. Also, there was

significant increase of SOD activity with the employment of water stress, which was

further increased by salicylic acid application. Interactions among all the treatment

combinations (H×GS; SA×GS; H×SA; and H×SA×GS) had statistically

significant difference.

Interaction between three sunflower hybrids and two growth stages of SA

foliar application has been shown in Figure 3.21 a. Sunflower hybrids differed

255

significantly if SA was sprayed at flowering growth stage, and all gave higher SOD

activity as compared to that for vegetative growth stage. The H-2 (NX-00989) had

statistically higher SOD activity, while H-3 (FH-352) rendered significantly the

lowest results. Response of water stressed sunflower hybrids to foliar application of

SA in different concentrations at two growth stages was also significant for SOD

activity (Figure 3.21 b). The SA application in 0.75 mM concentration at both stages

produced significantly higher SOD activity of water stressed plants as compared to

those receiving other SA concentrations. Whereas, control treatment (DS-0+SA-0)

rendered the lowest value for SOD activity, which was significantly inferior to all

the treatments in water stressed sunflower hybrids. For all the treatments, interaction

of DS with various SA concentrations showed better response of sunflower hybrids

to SA foliar application at flowering stage. Protection of water-stressed sunflower

plants improved through enhanced SOD activity by SA application up to 0.75 mM

concentration. Plant drought stress tolerance needs the activation of complex

metabolic including anti-oxidative pathways, especially reactive oxygen species

scavenging systems within the cells


10.0

15.0

20.0

25.0

30.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


256


Figure 3.21: Superoxide dismutase activity of drought stressed sunflower


growth stages.

which can provide to continue growth under moisture stress (Ezzat-Ollah et al.,

2007), SOD, CAT and PODs are main antioxidants involved in detoxification

ofsuperoxide and hydrogen peroxide respectively (Noctor and Foyer, 1998).ROS

homeostasis act as a regulator in relationships between the soil water threshold range

of chemical signals and drought tolerance (Wang et al., 2008).

Superoxide dismutase activity increased with the induction of drought stress

(Table 3.21). For keeping the levels of active oxygen species under control, plants

have non-enzymatic and enzymatic antioxidant systems to protect cells from

oxidative damage (Mittler, 2002). Superoxide dismutases (SODs), a group of

metalloenzymes, are the first defence against ROS (Gratao et al., 2005).Drought

stress enhances the SOD and peroxidase (POD) but decreases catalase (CAT)

activity (Tayebe and Hassan, 2010).

10.0

15.0

20.0

25.0

30.0




257

As results showed that salicylic acid concentration 0.75 mM gives highest

activity of SOD (Fig. 3.21 b). Similar results were obtained by Lindberg and Greger,

2002 reported that salicylic acid (1mM) is the most effective concentration in

increasing SOD, ascorbic peroxide (APOX) and CAT activities. Salicylic acid

enhanced the activities of POD, SOD and CAT, when sprayed on drought-stressed

tomato plants (Hayat et al., 2008). Ananieva et al. (2004) reported that SA treatment

alone resulted in an increase of SOD, POD and CAT activities by 17, 25 and 20%,

respectively, while higher concentrations (150-200 mg/L) restrained their activities.

Singh and Usha (2003) recorded maximum SOD activity in wheat if sprayed with 1

and 2 mM salicylic acid.

4.3.22 Peroxidase Activity

For the activity of peroxidase (POD) in sunflower plants, difference between

two growth stages of sunflower with respect to foliar spray of salicylic acid was

statistically significant (Table 3.22). The POD activity was greater (15.1 U/g FW) if

SA was applied at vegetative rather at flowering growth stage (13.7 U/g FW).

Drought stressed three sunflower hybrids also differed significantly with each other

for POD activity under application of salicylic acid. The H-1 (NX-19012) rendered

the highest POD activity (15.0 U/g FW) having non significant difference with H-2

(NX-00989) while H-3 (FH-352) showed statistically lower value (13.6 U/g FW).

Similarly, there was significant difference among different concentrations of

salicylic acid application. The lowest POD activity (6.2 U/g FW) was observed in

control (not receiving drought stress or salicylic acid), while the highest (19.6 U/g

FW) was in drought stressed plants receiving salicylic acid in

0.75 mM concentration. DS+SA-0.375 differing non significantly with that under

DS+SA-1.50. There was significant increase in POD activity with the employment

of drought stress, which was further increased by salicylic acid application.

258

Interactions were statistically significant for: sunflower hybrids (H) × growth stages

(GS); concentrations of salicylic acid (SA) × GS; H×SA; and H × SA × GS.

Interaction of three sunflower hybrids with two growth stages of sunflower

for SA application has been shown in Figure 3.22 a. It reflects that sunflower hybrids

differed significantly if SA was applied at vegetative stage, and they all got Table

3.22: Peroxidase activity (U/g FW) of drought stressed sunflower hybrids under





H-1 (NX-19012) 15.8 a 14.1c 14.9 A

H-2 (NX-00989) 15.5b 13.9d 14.7B

H-3 (FH-352) 14.1c 13.1e 13.6 C


Control (DS-0 + SA-0) 6.8g 5.7h 6.2 D

DS + SA-0 13.3 e 12.1f 12.7 C

DS + SA-0.375 17.1c 16.5 d 16.8 B

DS + SA-0.75 21.3a 17.9b 19.6 A

DS + SA-1.50 17.1 c 16.4d 16.8B


H-1 × DS-0 + SA-0 6.8i 5.4j 6.1G

H-1 × DS + SA-0 13.2 g 12.2 h 12.7E

H-1 × DS + SA-0.375 18.0 d 17.2e 17.6 B

H-1 × DS + SA-0.75 22.9 a 18.8 b 20.9 A

H-1 × DS + SA-1.50 18.1 cd 17.1 de 17.6B

H-2 × DS-0 + SA-0 6.9i 6.4i 6.7 F

H-2 × DS + SA-0 13.4g 12.2 h 12.8E

H-2 × DS + SA-0.375 17.2 e 16.2f 16.7C

H-2 × DS + SA-0.75 22.6 a 18.6bc 20.6 A

H-2 × DS + SA-1.50 17.2 e 16.0f 16.7 C

H-3 × DS-0 + SA-0 6.7 i 5.2j 6.0 G

H-3 × DS + SA-0 13.2 g 12.0 h 12.6E

H-3 × DS + SA-0.375 16.2 e 16.0 f 16.1 D

H-3 × DS + SA-0.75 18.5 bcd 16.2f 17.4 B

H-3 × DS + SA-1.50 16.1f 15.9f 16.0 D

Means (GS) 15.1 A 13.7 B

259



DS=Drought Stress

greater POD activity as compared to that for flowering growth stage. Genotypes H1

(NX-19012) and H-2 (NX-00989) showed significantly better results, while H-3

(FH-352) had the lowest value of POD activity. Similarly, differential response of

drought stressed sunflower hybrids to foliar application of SA at two growth stages

was significant (Figure 3.22 b). The SA application in 0.75 mM concentration at

vegetative stage produced significantly higher POD activity of drought stressed

plants as compared to those receiving no or other SA concentration. Whereas, control

treatment (DS-0+SA-0) rendered the lowest value for POD activity of non stressed

plants, which was significantly inferior to all the treatments in drought stressed

sunflower plants. Among all the treatments, interaction of DS with various SA

concentrations showed better response of sunflower hybrids to SA foliar application

at vegetative stage of plants showing POD activity decreases as the plant grows.

Salicylic acid spray on drought-stressed sunflower plants enhanced the POD activity

with increasing SA concentration, which declined at the highest concentration of

1.50 mM.

There is a defensive system in plants, that is to say, plants have an internal

protective enzyme-catalyzed clean up system, which is fine and elaborate enough to

avoid injuries of active oxygen, thus guaranteeing normal cellular function (Horváth

et al., 2007). The results depicted that POD activity enhanced under water deficit

conditions but gradually decreased as the plant grows (Table 3.22). Antioxidant

enzymes (CAT, APX, and POD) catalyze the conversion of H2O2 to water and O2

(Gratao et al., 2005). Balance between production of ROS and activities of

antioxidant enzymes determine whether oxidative signalling and/or damage will

occur (Moller et al., 2007). Drought stress enhanced the POD activity,

260



Figure 3.22: Peroxidase activity of drought stressed sunflower hybrids under


and intensities of POD-4 and -5 (Tayebe and Hassan, 2010).

10.0

12.0

14.0

16.0

18.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


0.0

5.0

10.0

15.0

20.0

25.0




261

Salicylic acid foliar application 0.75mM increased the activity of POD more

at vegetative stage and in tolerant hybrids of sunflower (Fig 3.22 a, b). These results

are in line with the observations of Ananieva et al. (2004) who reported that

SA treatment alone resulted in an increase of SOD, POD and CAT activities by 17,

25 and 20%, respectively. Chuanjie et al. (2003) found that low SA concentrations

(25-100 mg/L) induced higher SOD and POD activities‘ in Vanilla planifolin, while

higher concentrations (150-200 mg/L) restrained their activities. Cag et al. (2009)

reported that POD activity was stimulated with all concentrations (0.001-

1000 μM) of exogenic SA applications to canola, and SA was highly effective in

ameliorating the adverse effects of drought stress also suggested that that POD

activity decreases during growth period.

4.3.23 Catalase Activity

Drought stress to hybrid sunflower genotypes along with foliar application

of salicylic acid at two growth stages indicated that catalase activity was significantly

greater if applied at vegetative stage (Table 3.23). Among three drought-stressed

sunflower hybrids, the H-2 (NX-00989) showed the highest catalase activity (20.0

U/g FW) and differed significantly with others. The H-3 (FH-352) had the lowest

catalase activity (18.6 U/g FW) under foliar application of

salicylic acid in different

Table 3.23: Catalase activity (U/g FW) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 19.3 bc 19.6 abc 19.5 B

H-2 (NX-00989) 20.1 a 19.8 ab 20.0 A

H-3 (FH-352) 19.0 c 18.2 d 18.6 C


Control (DS-0 + SA-0) 14.9 NS 14.5 14.7 D

DS + SA-0 18.9 18.6 18.8 C

262

DS + SA-0.375 20.3 20.0 20.2 B

DS + SA-0.75 23.4 22.8 23.1 A

DS + SA-1.50 20.0 20.0 20.0 B


H-1 × DS-0 + SA-0 14.7 NS 15.0 14.9 E

H-1 × DS + SA-0 18.9 19.2 19.0 C

H-1 × DS + SA-0.375 20.4 20.5 20.5 B

H-1 × DS + SA-0.75 23.1 23.1 23.1 A

H-1 × DS + SA-1.50 19.7 20.1 19.9 BC

H-2 × DS-0 + SA-0 15.8 15.9 15.8 E

H-2 × DS + SA-0 19.6 19.8 19.7 BC

H-2 × DS + SA-0.375 20.9 20.1 20.5 B

H-2 × DS + SA-0.75 23.5 23.0 23.3 A

H-2 × DS + SA-1.50 20.8 20.3 20.6 B

H-3 × DS-0 + SA-0 14.3 12.7 13.5 F

H-3 × DS + SA-0 18.2 16.8 17.5 D

H-3 × DS + SA-0.375 19.7 19.4 19.6 BC

H-3 × DS + SA-0.75 23.5 22.3 22.9 A

H-3 × DS + SA-1.50 19.4 19.6 19.5 BC

Means (GS) 19.5 A 19.2 B



DS=Drought Stress

concentrations at two growth stages. Similarly, there was significant difference

among different concentrations of salicylic acid application. The lowest catalase

activity (14.7 U/g FW) was in control (DS-0+SA-0) without receiving both drought

stress and salicylic acid. While, application of salicylic acid at the concentration of

0.75 mM resulted the highest and significant increase in catalase activity of drought

stressed plants. There was significant increase in catalase activity with the

employment of drought stress, which was further increased by SA application.

Interactions among treatment factors were statistically non significant for:

concentrations of salicylic acid (SA) × growth stages (GS); and H × SA × GS.

263

Interaction between three sunflower hybrids and two growth stages of SA

application to sunflower for their effect on catalase activity has been presented in

Figure 3.23 a. All sunflower hybrids had greater catalase activity if SA was applied

at vegetative growth stage as compared to that at flowering stage. The H-3 (FH352)

showed significantly lower results, while having statistically significant difference

between two stages of SA application. Interactive response of drought stressed



both stages caused significantly higher catalase activity in drought stressed plants as

compared to those receiving lower or higher concentrations of SA. Whereas, control

treatment (DS-0 + SA-0) rendered the smallest value of catalase activity, which was

significantly lower than with all the SA treatments to drought stressed sunflower

plants. Generally, the catalase activity in drought-stressed sunflower


17.5

18.0

18.5

19.0

19.5

20.0

20.5

21.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


264


Figure 3.23: Catalase activity of drought stressed sunflower hybrids under


plants enhanced with increasing concentration of salicylic acid up to 0.75 mM, and

further increase in SA concentration (1.50 mM) reduced the catalase activity

significantly.

Drought stress increases the catalase (CAT) activity in plants (Nazarli et al.,

2011). (Table 3.23) also showed that catalase activity enhanced as drought was

imposed on sunflower hybrids. The increase in CAT activities in leaves is probably

a response to the enhanced production of ROS, and particularly H2O2, under water

stress (Smirnoff, 1993). It is suggested that the higher concentrations of catalase and

ascorbate peroxidase might have removed the O2 radicals and its products such as

H2O2.The increase of this antioxidant may be triggered by excess production of

reactive oxygen species in the photosynthetic apparatus under water stress

conditions.

10.00

15.00

20.00

25.00




265

Salicylic acid further enhanced the activities of CAT when applied under water

stress (Fig. 3.24 b).These results are in line with the findings as Salicylic acid (1

mM) is the most effective concentration in increasing CAT activities (Lindberg and

Greger, 2002). Salicylic acid enhanced the activities of POD, SOD and CAT, when

sprayed on drought-stressed tomato plants (Hayat et al., 2008). Ananieva et al.

(2004) reported that SA treatment alone resulted an increase of SOD, POD and CAT

activities by 17, 25 and 20%, respectively. Similarly, treatment of barely seedlings

with 500 µM SA caused an increase in CAT activity (Popova et al.,

2003). Contrastingly, SA treatment decreased the activities of catalase in tomato

(Senaratha et al., 2000), and other plants after one day treatment of SA (Janda et al.,

1999; Kang et al., 2003).

4.3.24 Endogenous Salicylic acid Level

Endogenous level of SA had significant difference for exogenous SA

applications at two growth stages of sunflower hybrids (Table 3.24). Foliar

applications of salicylic acid to water stressed plants at vegetative growth stage

yielded significantly higher values (1169 ng/g) as compared with SA spray at

flowering stage. Similarly, three sunflower hybrids differed significantly with higher

SA content (1108 ng/g) in H-2 (NX-00989) while H-3 (FH-352) had lesser value

(907.9 ng/g) which had significant statistical difference. Further, different

concentrations of salicylic acid application to water stressed plants also showed

significant difference. There was significant increase in SA content with the

employment of water stress, which was further enhanced by SA applications. The

highest content of SA (1364.5 ng/g) was found in 0.75 mM salicylic acid

concentration while, the lowest (481.9 ng/g) in control. All interactions of treatment

factors were statistically significant.

266

Three sunflower hybrids had significant interactive effect with two growth

stages of sunflower at which SA foliar application was made (Figure 3.24 a).

Sunflower hybrids differ significantly when SA was applied at vegetative growth

stage. Similarly, differential response of water stressed sunflower hybrids to foliar

application of SA at two growth stages was also significant (Figure 3.24 b). The SA

application in all concentrations at both stages produced statistically different SA

contents of water stressed plants. Control treatments showed the lowest values for

SA content, and differed significantly with other treatments in water stressed

sunflower plants. The remedial effect of salicylic acid on water-stressed sunflower

plants was enhanced by increasing the concentration of salicylic acid maximum at

0.75 mM.

Endogenous level of SA increased with the imposition of drought stress at

both the stages of growth in all sunflower hybrids (Table 3.24). Tolerant hybrids

exhibited more accumulation of SA than sensitive one. Hayat (2013) reported

drought stress increased the endogenous level of SA. The results are similar with the

findings of Munne-Bosch and Penuelas (2003) who reported accumulation of five

folds endogenous salicylic acid in Phyllyrea angustifolia when grown in water

stressed field.

Foliar application of SA enhanced the endogenous level of salicylic acid

under water stressed conditions (Fig. 3.24 b). Exogenous application of SA increased

the endogenous content of salicylic acid in leaves of Phlox setacea (Talieva and

Kondrat, 2002). The role of salicylic acid in extenuating the adverse affects of water

stress might be due to its association with the enhancing effects on antioxidant

enzymes which increased membrane stability and protects the membrane damage

(Singh and Usha, 2003). Salicylic acid application may also contribute in the process

of anti stress reactions by the accumulation of soluble proteins (Sakhabutindova et

267

al., 2003). Exogenous application of SA enhanced the levels of endogenous SA

contents (Hayat, 2013).

4.3.25 Palmitic Acid Content

Difference between two growth stages of sunflower plants for foliar application of

salicylic acid was non significant with respect to palmitic acid content (Table 3.25).

Whereas, drought stressed three sunflower hybrids differed significantly with each

other for palmitic acid content under foliar application of SA in different

Table 3.24: Endogenous salicylic acid level of drought stressed sunflower hybrids

under foliar applied various concentrations of salicylic acid at two growth stages.




H-1 (NX-19012) 1178.5 b 970.4 e 1074.4 B

H-2 (NX-00989) 1216.2 b 1000.8 d 1108.5 A

H-3 (FH-352) 1112.2 c 703.6 f 907.9 C


Control (DS-0 + SA-0) 537.4 i 426.4 j 481.9 E

DS + SA-0 1190.5 e 858.7 h 1024.6 D

DS + SA-0.375 1335.0 b 1008.7 f 1171.9 B

DS + SA-0.75 1512.0 a 1216.9 d 1364.5 A

DS + SA-1.50 1270.0 c 947.3 g 1108.6 C


H-1 × DS-0 + SA-0 536.3 w 436.5 y 486.4 M

H-1 × DS + SA-0 1210.4 k 960.5 r 1085.5 H

H-1 × DS + SA-0.375 1348.8 e 1107.8m 1228.3 D

H-1 × DS + SA-0.75 1545.2 b 1305.4 g 1425.3 B

H-1 × DS + SA-1.50 1251.5 i 1041.8 o 1146.7 F

H-2 × DS-0 + SA-0 544.9 v 458.3 x 501.6 L

H-2 × DS + SA-0 1257.3 i 984.5 q 1120.9 G

H-2 × DS + SA-0.375 1368.4 d 1129.9 l 1249.1 C

H-2 × DS + SA-0.75 1589.2 a 1349.6 e 1469.4 A

H-2 × DS + SA-1.50 1321.3 f 1081.6 n 1201.5 E

H-3 × DS-0 + SA-0 531.0 w 384.3 z 457.6 N

268

H-3 × DS + SA-0 1103.9 m 631.0 u 867.5 K

H-3 × DS + SA-0.375 1287.7 h 788.5 s 1038.1 I

H-3 × DS + SA-0.75 1401.5 c 995.8 p 1198.6 E

H-3 × DS + SA-1.50 1237.0 j 718.4 t 977.7 J

Means (GS) 1169.0 A 891.6 B



DS=Drought Stress



200.0

400.0

600.0

800.0

1000.0

1200.0

1400.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


200

400

600

800

1000

1200

1400

1600

1800




269

Figure 3.24: Endogenous salicylic acid level of drought stressed sunflower


growth stages.

concentrations at both growth stages. H-1 (NX-19012) had the highest palmitic acid

content (6.65 %) while H-3 (FH-352) contained the lowest amount. Further, there

was significant increase by drought stress and salicylic acid application in different

concentrations. The lowest palmitic acid content (5.92%) was recorded in control

(without both drought stress and salicylic acid), while the highest (6.47%) was with

DS+SA-0 (drought stressed plants receiving no salicylic acid). There was significant

increase in palmitic acid content with the employment of drought stress, which was

slightly addressed by salicylic acid application. Application of salicylic acid at the

concentration of 0.75 mM lowered down the palmitic acid content of drought

stressed plants although non significantly. Interactions among various types of

treatment factors were statistically significant only for sunflower hybrids (H) ×

growth stages(GS); whereas there was non significant interactions for various

concentrations of salicylic acid (SA) × GS; H×SA; and H × SA × GS.

As expressed in Figure 3.25 a, the interactive effect of three sunflower

hybrids with two growth stages of sunflower for SA application was statistically

significant. Sunflower hybrids differed more if SA was applied at vegetative growth

stage giving higher palmitic acid content (except in H-2 genotype) as compared to

that at flowering stage. H-3 (FH-352) showed the lowest contents, while H-1 (NX-

19012) got the highest value for SA application at vegetative growth stage and the

difference was statistically significant. Nonetheless, differential response of drought

stressed sunflower hybrids to foliar application of SA at two growth stages was non

significant (Figure 3.25 b). Whereas, SA application 0.75 mM concentration at both

stages produced slightly lower palmitic acid content giving better response in

drought stressed plants as compared to those receiving lower / higher SA

270

concentrations. Whereas, control treatment (DS-0 + Table 3.25: Palmitic acid

content (%) of drought stressed sunflower hybrids under foliar applied various





H-1 (NX-19012) 6.80 a 6.49 b 6.65 A

H-2 (NX-00989) 6.14 c 6.72 a 6.43 B

H-3 (FH-352) 5.91 d 5.85 d 5.88 C


Control (DS-0 + SA-0) 5.98 NS 5.86 5.92 B

DS + SA-0 6.43 6.52 6.47 A

DS + SA-0.375 6.35 6.48 6.42 A

DS + SA-0.75 6.29 6.43 6.36 A

DS + SA-1.50 6.36 6.48 6.42 A


H-1 × DS-0 + SA-0 6.36 NS 6.03 6.20 NS

H-1 × DS + SA-0 7.07 6.66 6.86

H-1 × DS + SA-0.375 6.89 6.61 6.75

H-1 × DS + SA-0.75 6.78 6.57 6.68

H-1 × DS + SA-1.50 6.90 6.60 6.75

H-2 × DS-0 + SA-0 6.12 6.32 6.22

H-2 × DS + SA-0 6.15 6.83 6.49

H-2 × DS + SA-0.375 6.14 6.84 6.49

H-2 × DS + SA-0.75 6.13 6.78 6.46

H-2 × DS + SA-1.50 6.14 6.83 6.49

H-3 × DS-0 + SA-0 5.47 5.23 5.35

H-3 × DS + SA-0 6.06 6.06 6.06

H-3 × DS + SA-0.375 6.02 6.00 6.01

H-3 × DS + SA-0.75 5.96 5.93 5.95

H-3 × DS + SA-1.50 6.04 6.01 6.03

Means (GS) 6.3 NS 6.4



DS=Drought Stress

271


Figure 3.25: Palmitic acid content of drought stressed sunflower hybrids under


SA-0) rendered the lowest values for palmitic acid content, which was

significantly different from all the treatments in drought stressed sunflower plants.

Under all the treatments (except control), interaction of DS with various SA

concentrations showed that sunflower hybrids had relatively lower content of

palmitic acid with SA application at vegetative growth stage. It is obvious from the


5.60

5.80

6.00

6.20

6.40

6.60

6.80

7.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


5.60

5.80

6.00

6.20

6.40

6.60




272

results that salicylic acid application on drought-stressed sunflower plants has

remedial effect by lowering palmitic acid content in drought-stressed plants, and the

most appropriate SA concentration for this purpose is 0.75 mM.

4.3.26 Stearic Acid Content

Data indicated that statistical difference between two growth stages of sunflower

plants for SA application was significant regarding the stearic acid contents (Table

3.26). Foliar application of salicylic acid at vegetative growth stage rendered

significantly higher content of stearic acid (4.71 %) as compared to that with SA

application at flowering stage of sunflower hybrids (3.93 %). Drought stressed three

sunflower hybrids also differed significantly with each other for stearic acid content

under foliar application of salicylic acid in different concentrations at two growth

stages. H-1 (NX-19012) contained greater amount of stearic acid (5.08 %) whereas

H-2 (NX-00989) had the lowest stearic acid contents (3.46 %). Similarly, there was

significant difference among different concentrations of salicylic acid application.

The highest stearic acid content (4.88 %) was recorded in control (DS0+SA-0), while

the lowest (4.13 %) was withDS+SA-0 (drought stressed but not receiving salicylic

acid). There was significant decrease in stearic acid content with the employment of

drought stress, which was significantly improved by SA application. Salicylic acid

spray at the concentration of 0.75 mM caused the highest and significant increase of

stearic acid content in drought stressed plants. All the interactions as for: sunflower

hybrids (H) × growth stages (GS) of salicylic acid foliar application; concentrations

of salicylic acid (SA) × GS;H×SA; and H × SA × GS were statistically significant.

Interaction between sunflower hybrids and their growth stages for SA

application indicated that only H-1 (NX-19012) differed significantly by having

greater stearic acid content if SA was applied at vegetative growth stage as compared

to that at flowering stage (Figure 3.26 a). H-2 (NX-00989) and H-3 (FH352) showed

statistically similar results at both stages. Comparative response of drought stressed

273

sunflower hybrids to foliar application of SA at two growth stages was significantly

different (Figure 3.26 b). In all the treatment combinations, application of salicylic

acid at vegetative growth stage produced significantly higher content of stearic acid

as compared to that with SA application at flowering stage of sunflower hybrids. The


higher stearic acid content of drought stressed plants. However, control treatment

rendered the highest value for stearic acid content, which was significantly superior

to all the treatments in drought stressed sunflower hybrids plants. Within all the

treatments, interaction of DS with various SA concentrations showed better response

of sunflower hybrids to SA foliar application at vegetative growth stage. Positive

effect of salicylic acid on droughtstressed sunflower plants in terms of stearic acid

content enhanced non significantly by increasing SA concentration, being slightly

higher with 0.75 mM.

4.3.27 Oleic Acid Content

Difference between two growth stages of sunflower regarding foliar

application of salicylic acid was statistically significant for oleic acid content

(Table 3.27). Data reflected that oleic acid contents were higher with SA applied at

Table 3.26: Stearic acid content (%) of drought stressed sunflower hybrids


stages.




H-1 (NX-19012) 6.21 a 3.96 c 5.08 A

H-2 (NX-00989) 3.47 d 3.44 d 3.46 C

H-3 (FH-352) 4.45 b 4.40 b 4.43 B


Control (DS-0 + SA-0) 5.31 a 4.45 c 4.88 A

DS + SA-0 4.53 bc 3.74 e 4.13 C

DS + SA-0.375 4.57 b 3.82 de 4.19 BC

DS + SA-0.75 4.58 b 3.84 d 4.21 B

DS + SA-1.50 4.57 b 3.82 de 4.20 BC

274


H-1 × DS-0 + SA-0 6.34 a 4.15 ef 5.25 A

H-1 × DS + SA-0 6.10 b 3.85 h 4.98 B

H-1 × DS + SA-0.375 6.19 ab 3.91 h 5.05 B

H-1 × DS + SA-0.75 6.20 ab 3.95 fgh 5.08 B

H-1 × DS + SA-1.50 6.21 ab 3.92 gh 5.07 B

H-2 × DS-0 + SA-0 4.53 d 4.23 e 4.38 C

H-2 × DS + SA-0 3.19 i 3.23 i 3.21 E

H-2 × DS + SA-0.375 3.21 i 3.25 i 3.23 E

H-2 × DS + SA-0.75 3.22 i 3.26 i 3.24 E

H-2 × DS + SA-1.50 3.20 i 3.24 i 3.22 E

H-3 × DS-0 + SA-0 5.07 c 4.98 c 5.03 B

H-3 × DS + SA-0 4.29 e 4.13 efg 4.21 E

H-3 × DS + SA-0.375 4.30 e 4.29 e 4.30 CD

H-3 × DS + SA-0.75 4.31 e 4.32 de 4.32 CD

H-3 × DS + SA-1.50 4.30 e 4.30 e 4.30 CD

Means (GS) 4.71 A 3.93 B



DS=Drought Stress


2.00

3.00

4.00

5.00

6.00

7.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


275


Figure 3.26: Stearic acid content of drought stressed sunflower hybrids under


vegetative growth stage (28.8 %) as compared to that at flowering stage (26.3 %).


for oleic acid content under foliar application of salicylic acid in different

concentrations. H-2 (NX-00989) had the highest oleic acid content (29.4 %), while

H-3 (FH-352) contained the lowest amount (26.1 %). Further, there was significant

difference among various concentrations of salicylic acid application at two growth

stages. The highest oleic acid content (33.7 %) was recorded in control (DS-0+SA0)

without receiving both drought stress and salicylic acid, while the lowest (24.9 %)

was with DS+SA-0 (drought stressed but receiving no salicylic acid). There was

significant decrease in oleic acid content with the employment of drought stress,

which was highly restored by salicylic acid. Application of salicylic acid at the

concentration of 0.75 mM caused the highest and significant increase of oleic acid

content (26.9 %) in drought stressed plants. All the interactions were statistically

significant for three treatment factors as: H × GS; SA × GS; H×SA; and H × SA ×

2.00

3.00

4.00

5.00

6.00




276

GS. Interaction of three sunflower hybrids with two growth stages of sunflower for

SA application has been presented in (Figure 3.27 a). Apparently, sunflower hybrids

differed more if SA was applied at flowering stage even all giving lesser oleic acid

content as compared to that at vegetative growth stage. The H-2 (NX00989) showed

significantly better results, while H-3 (FH-352) performed the least. Similarly,

differential response of drought stressed sunflower hybrids to foliar application of

SA at two growth stages was statistically significant (Figure 3.27 b). The SA

application in 0.75 mM concentration at both stages produced significantly higher

oleic acid content of drought stressed plants as compared to Table 3.27: Oleic acid

content % of drought stressed sunflower hybrids under foliar applied various

concentrations of salicylic acid at two growth stages




H-1 (NX-19012) 27.3 c 26.9 d 27.1 B

H-2 (NX-00989) 29.9 a 29.0 b 29.4 A

H-3 (FH-352) 29.1 b 23.1 e 26.1 C


Control (DS-0 + SA-0) 35.2 a 32.3 b 33.7 A

DS + SA-0 26.4 e 23.4 g 24.9 E

DS + SA-0.375 27.0 d 24.7 f 25.8 D

DS + SA-0.75 27.8 c 26.1 e 26.9 B

DS + SA-1.50 27.5 cd 25.1 f 26.3 C


H-1 × DS-0 + SA-0 38.1 a 34.0 c 36.0 A

H-1 × DS + SA-0 23.8 j 23.6 j 23.7 J

H-1 × DS + SA-0.375 24.1 j 25.4 hi 24.7 I

H-1 × DS + SA-0.75 25.2 i 25.9 hi 25.6 H

H-1 × DS + SA-1.50 25.2 i 25.5 hi 25.3 HI

H-2 × DS-0 + SA-0 36.0 b 34.8 c 35.4 B

H-2 × DS + SA-0 28.0 fg 26.3 h 27.1 F

H-2 × DS + SA-0.375 28.3 efg 27.4 g 27.8 E

H-2 × DS + SA-0.75 29.1 e 28.8 ef 28.9 D

277

H-2 × DS + SA-1.50 28.3 efg 27.7 g 28.0 E

H-3 × DS-0 + SA-0 31.5 d 28.2 efg 29.8 C

H-3 × DS + SA-0 27.4 g 20.3 l 23.8 J

H-3 × DS + SA-0.375 28.7 ef 21.4 k 25.0 HI

H-3 × DS + SA-0.75 29.1 e 23.6 j 26.4 G

H-3 × DS + SA-1.50 28.8 ef 22.0 k 25.4 H

Means (GS) 28.8 A 26.3 B



DS=Drought Stress



15.0

20.0

25.0

30.0

35.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


15.0

20.0

25.0

30.0

35.0

40.0




278

Figure 3.27: Oleic acid content of drought stressed sunflower hybrids under

foliar applied various concentrations of salicylic acid at two growth stages. those

receiving no or other SA concentrations. Whereas, control treatment (DS-0 + SA-0)

rendered the highest value for oleic acid content of non stressed plants, which was

significantly superior to all the treatments in drought stressed sunflower plants.

Within all the treatments, interaction of DS with various SA concentrations showed

better response of sunflower hybrids to SA foliar application at vegetative growth

stage. Restoring effect of salicylic acid on drought-stressed sunflower plants

enhanced by increasing the SA concentration up to 0.75 mM; and above this

concentration, the oleic acid content remained lower.

4.3.28 Linoleic Acid Content

Difference between applications of salicylic acid at two growth stages of

sunflower plants was significant for linoleic acid content (Table 3.28). Foliar SA

application at flowering stage resulted in higher content of linoleic acid (60.5 %)

with a significant difference from that if sprayed at vegetative stage (58.0 %).


for linoleic acid content under foliar application of salicylic acid. H-2 (NX-00989)

contained the highest amount of linoleic acid (61.7 %) while H-3 (FH-352) had the

lowest one (56.9 %) Further, there was significant difference among various

concentrations of salicylic acid application to drought-stressed sunflower plants. The

lowest linoleic acid content (52.9 %) was found in control (without receiving both

drought stress and salicylic acid), whereas the highest (61.7 %) was in drought-

stressed plants without receiving SA. Application of salicylic acid at all

concentrations caused significant reduction in linoleic acid content of drought

stressed plants having non significant difference among them.

Table 3.28: Linoleic acid % of drought stressed sunflower hybrids under foliar


279




H-1 (NX-19012) 57.0 d 61.2 b 59.1 B

H-2 (NX-00989) 59.9 c 63.6 a 61.7 A

H-3 (FH-352) 57.1 d 56.7 e 56.9 C


Control (DS-0 + SA-0) 51.5 i 54.3 h 52.9 C

DS + SA-0 60.4 d 63.1 a 61.7 A

DS + SA-0.375 59.6 e 61.6 c 60.6 B

DS + SA-0.75 59.2 g 61.9 b 60.5 B

DS + SA-1.50 59.4 f 61.7 bc 60.6 B


H-1 × DS-0 + SA-0 46.5 r 54.5 o 50.5 L

H-1 × DS + SA-0 60.5 fg 64.1 c 62.3 C

H-1 × DS + SA-0.375 59.3 hi 62.3 d 60.8 E

H-1 × DS + SA-0.75 59.5 h 62.6 d 61.1 D

H-1 × DS + SA-1.50 59.3 hi 62.5 d 60.9 DE

H-2 × DS-0 + SA-0 56.3 n 57.7 m 57.0 J

H-2 × DS + SA-0 61.5 e 66.4 a 63.9 A

H-2 × DS + SA-0.375 60.8 f 64.3 c 62.5 B

H-2 × DS + SA-0.75 60.2 g 65.1 b 62.7 B

H-2 × DS + SA-1.50 60.6 f 64.4 c 62.5 B

H-3 × DS-0 + SA-0 51.7 p 50.6 q 51.2 K

H-3 × DS + SA-0 59.0 i 58.7 j 58.9 F

H-3 × DS + SA-0.375 58.7 j 58.2 kl 58.4 G

H-3 × DS + SA-0.75 57.9 lm 57.9 lm 57.9 I

H-3 × DS + SA-1.50 58.2 kl 58.3 k 58.2 H

Means (GS) 58.0 B 60.5 A



DS=Drought Stress

280


Figure 3.28: Linoleic acid content of drought stressed sunflower hybrids under


There was significant increase in linoleic acid content with the employment of

drought stress, which was partly restricted by SA application. Interactions were

statistically significant for all the combinations of treatment factors as: H × GS; SA

× GS; H×SA; and H × SA × GS.

Interaction between sunflower hybrids and growth stages indicated that H-3


50.0

54.0

58.0

62.0

66.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


50.0

54.0

58.0

62.0

66.0




281

(FH-352) had lesser linoleic acid content with smaller difference for growth stages

(Figure 3.28 a). Other two sunflower hybrids differed equally for linoleic acid

content with SA application whether at vegetative or flowering stage. Similarly,

differential response of drought stressed sunflower hybrids to foliar application of

SA at two growth stages was statistically significant (Figure 3.28 b). The SA

application in all the concentrations at both stages produced significantly lower

linoleic acid content of drought stressed plants as compared to those receiving no

SA. Whereas, control treatment (DS-0 + SA-0) rendered the lowest value for linoleic

acid content, which was significantly lower in all the treatments of drought- stressed

sunflower hybrid plants. Within all the treatments, interaction of DS with various

SA concentrations showed lower contents of linoleic acid in sunflower hybrids to

SA foliar application at vegetative growth stage. Generally, there was positive effect

of salicylic acid on drought-stressed sunflower plants for reducing the linoleic acid

content being statistically similar at all concentrations.

Results of fatty acids comprise palmitic acid, stearic acid, oleic acid and

linoleic acid showed that water stress affected the composition of different fatty

acids. The saturated fatty acid (palmitic and stearic acid) contents were affected by

water stress. The palmitic acid concentration increased under drought conditions and

stearic acid decreased, there was a significant negative effect of drought on the oleic

acid concentration in all studied sunflower hybrids. On the contrary, the linoleic acid

concentration increases in sunflower seeds provided by drought stress as compared

to control. (Tables 3.25-3.28). It was reported previously that beside the genotype

which is the most important factor that defines the fatty acid composition, water

stress also affects the fatty acid composition of sunflower oil

Stefanoudaki et al., 2009. The results are concurrent with previous findings of Javed

et al., 2013 who reported saturated fatty acid (palmitic and stearic acid) contents

were insignificantly affected by water stress. Palmitic acid concentration increases

282

under drought conditions (with 0.39 to 0.74%) and stearic acid concentration

decreases under the same conditions (up to 1.33) in safflower. It was established that

drought stress in sunflower plants was linked to free radical processes in membrane

components leading to alterations in membrane stability and increasing their

permeability. The peroxidation of unsaturated lipids in biological membranes is the

most prominent symptom of oxidative stress in animals and plants (Cho and Seo,

2005). In present research differential effect upon fatty acid synthesis was observed

by different hybrids. The linoleic and oleic are the fatty acids, which affect the

quality of oil. Drought modified fatty acids composition and eventually the food

quality and it is considered to be very important in stress tolerance of plants Azachi

et al., 2002. Moreover extent of unsaturation of fatty acids is correlated with

potential of photosynthetic machinery to tolerate stress. Generally abiotic stress

induces inactivation of PSII and PSI Allakhverdiev et al., 2000 and unsaturation of

fatty acids in membrane lipids shelter PSII and PSI as one of effective protective

strategy.

SA treatments increased the oleic acid and stearic acid but decreased the

palmitic and linoleic acid in drought stressed sunflower hybrids especially in H-1

and H-2 (Fig. 3.25a, b -3.28a, b). Similarly, Noreen and Ashraf (2010) mentioned

that high doses of salicylic acid caused a marked increase in sunflower achene oil

content as well as some key fatty acids and significant decrease in stearic acid. The

research work of Baldini et al. (2000) revealed that water stress causes a significant

reduction of about 15% in the concentration of oleic acid in standard hybrid. He had

also established that from the 8th days after flowering, with the increase in the

biosynthesis of the –9 desaturase started to be active. This

enzyme has been proposed as being responsible for the accumulation of oleic acid

(18:1) by desaturing stearic acid (18:0), (Mckeon and Stumpf, 1982). Another

enzyme leading to the oleic acid accumu –12 desaturase, which catalyses

283

the second desaturation of oleic acid in linoleic acid (Stymme and Appelqvist, 1980).

Abdel Rahim et al., (2000) reported that the percentage of unsaturated fatty acids

proved the efficiency of desaturation in oil. Contrastingly, ASA treatments were

found to increase saturated fatty acids such as palmitic acid, but decreased stearic

acid content. Even though, saturated fatty acids of ground nut seeds increased due to

treatments of ASA, increased levels of oleic acid with ASA treatments may maintain

the quality and stability of oil thereby increasing shelf life of oil as indicated by

Izquierdo et al. (2002).

4.3.29 Protein Content

Contents of protein in hybrid sunflower seeds differed non significantly for

foliar application of salicylic acid at two growth stages of drought-stressed sunflower

plants (Table 3.29). The SA spray at flowering stage caused greater protein content

(23.0%) as compared to that for vegetative growth stage (22.9%). Three sunflower

hybrids differed significantly with each other for protein content under foliar

application of salicylic acid in different concentrations at two growth stages. The H-

3 (FH-352) accumulated statistically lower protein content (21.1 %) as compared to

other two hybrids which had non significant difference. Similarly, there was

significant difference among different concentrations of salicylic acid application

drought-stressed sunflower hybrids. The highest protein content (24.2 %) was

recorded in DS+SA-0 (drought stressed plants without receiving salicylic acid), and

the lowest amount was with DS-0 + SA-0. Application of salicylic acid at the

concentration of 1.50 mM gave higher protein content of drought-stressed plants

than with lower SA concentration or in control, although there was non significant

difference. There was significant increase in protein content with the employment of

drought stress as compared to that in control. Most of the interactions among various

284

treatment factors were statistically non significant except for: concentrations of

salicylic acid (SA) × growth stages (GS).


sunflower at which SA was sprayed, has been drawn in Figure 3.29 a. It indicates

that sunflower hybrids differed more if SA was applied at vegetative growth stage

giving lesser protein content as compared to that at flowering stage in H-3 sensitive

while other hybrids have no difference among the growth stages. The H-2

(NX00989), showed slightly higher contents of protein than in H-1 (NX-19012),

while H-3 (FH-352) had the least and the difference was statistically significant.

Differential response of drought stressed sunflower hybrids to foliar application of

SA at two growth stages was non significant (Figure 3.29 b). The SA application in

1.50 mM concentration at both stages gives higher protein content of droughtstressed

plants showing least affect. Within all the treatments except (DS + SA-0), interaction

of DS with various SA concentrations showed equal response of sunflower hybrids

to SA foliar application at both stages except in H-3. Generally, SA concentrations

(0.375 and 0.75 mM) on drought-stressed sunflower plants give better response to

the protein contents, being least at the 1.50 mM concentration.

Achene protein % increases under the drought stress conditions (Table 3.29)

as Reddy et al. (2004) pointed out that imposition of drought stress at flowering

drastically reduced achene yield but improved achene protein content. This increase

in protein content was due to negative relationship between oil and protein content

(Debaeke et al., 1998) and accumulation of LEA (late embryogenesis abundant)

protein like dehydrin in sunflower (Natali et al., 2003). These proteins protected

plant cells from damage under drought stress. Degree of decrease in protein contents

under drought was higher in the sensitive genotype as compared to moderately

tolerant one (Parida et al., 2007). These results confirm our findings (Fig. 3.29 a).

By imposition of water stress especially at flowering stage, the grains got shrunken

285

and 1000-achene weight decreased drastically resulting in yield reduction but achene

protein contents were increased.

In present study SA application reduced the protein content (Fig. 3.29 b) as

SA improves the oil content (Fig. 3.30 b). Exogenous applications of SA increased

the achene weight and achene yield but achene protein contents were reduced

(Hussain, 2008). Protein and lipid synthesis in seeds are negatively correlated with

each other. Photo-assimilation is the only source for all kind of seed reserves

(starches, fats, proteins), so increase in any of these constituents should be followed

by the corresponding decrease in the others (or at least one). Under stress conditions,

each plant sp. has its preferred form of food reserves, so the most of

Table 3.28: Protein content (%) of drought stressed sunflower hybrids under foliar





H-1 (NX-19012) 23.9 NS 23.6 23.8 A

H-2 (NX-00989) 24.0 24.1 24.0 A

H-3 (FH-352) 20.8 21.3 21.1 B


Control (DS-0 + SA-0) 22.3 bcd 22.3bcd 22.4 BC

DS + SA-0 23.8 ab 24.6 a 24.2 A

DS + SA-0.375 22.7abc 22.6 abc 22.5BC

DS + SA-0.75 22.6 abc 22.4 bc 22.5 BC

DS + SA-1.50 23.0 abc 22.9 abc 22.9 B


H-1 × DS-0 + SA-0 23.3 NS 23.1 23.2 NS

H-1 × DS + SA-0 24.9 25.5 25.2

H-1 × DS + SA-0.375 23.5 23.3 23.4

H-1 × DS + SA-0.75 23.5 23.2 23.4

H-1 × DS + SA-1.50 24.1 23.4 23.8

H-2 × DS-0 + SA-0 23.2 23.1 23.1

H-2 × DS + SA-0 25.5 26.2 25.9

H-2 × DS + SA-0.375 23.8 23.2 23.5

H-2 × DS + SA-0.75 23.7 23.1 23.4

286

H-2 × DS + SA-1.50 23.9 23.8 23.8

H-3 × DS-0 + SA-0 20.3 20.6 20.5

H-3 × DS + SA-0 21.1 22.2 21.6

H-3 × DS + SA-0.375 20.9 21.3 21.1

H-3 × DS + SA-0.75 20.7 21.0 20.8

H-3 × DS + SA-1.50 21.0 21.4 21.2

Means (GS) 22.9 NS 23.0



DS=Drought Stress



19.0

20.0

21.0

22.0

23.0

24.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


18.0

20.0

22.0

24.0

26.0

DS-0+SA-0 DS+SA-0 DS+SA-0.38 DS+SA-0.75 DS+SA-1.50

Salicylic acid treatments (mM L – 1 ) to drought stressed plants


287

Figure 3.29: Protein content of drought stressed sunflower hybrids under foliar


the reserves would be stored in that form and consequently, the amount of other

constituents of food reserves would be decreased. Contradictory to our findings SA

application (0.5 mM) increased protein yield in both water stress and optimum

conditions (Sadeghipour and Aghaei, 2012). Kabiri et al. (2014) found that SA at 10

μM enhanced the protein content to the maximum in black cumin (Nigella sativa)

under drought stress.

4.3.30 Oil Content

Applications of salicylic acid at two growth stages of sunflower plants

differed significantly for percentage of oil content (Table 3.30). The SA spray at

vegetative growth stage caused higher oil contents in sunflower hybrids significantly

than that at flowering stage. Drought stressed three sunflower hybrids also differed

significantly with each other for oil content due to SA application; H2 (NX-00989)

gave higher oil contents (33.1 %) as compared to other two hybrids. Further, there

was significant difference among different concentrations of salicylic acid

application. The highest oil content (35.4%) was recorded in control (DS0+SA-0),

whereas the lowest (27.0%) was with DS+SA-0 (drought stressed but receiving no

salicylic acid). Salicylic acid application at 0.75 mM concentration caused the

maximum increase of oil content in drought stressed plants although having non

significant difference with other SA concentrations. There was significant decrease

in oil content with the employment of drought stress, which was significantly

countered by the application of SA. However, interactions among all the treatment

factors viz. sunflower hybrids (H), growth stages (GS), and concentrations of

salicylic acid (SA) were statistically non significant.


sunflower, at which SA foliar application was made, has been drawn in Fig 3.30 a.

288

It indicates that sunflower hybrids differed almost equally whether SA was applied

at vegetative growth stage or flowering stage. The H-2 (NX-00989) showed higher

oil contents than in H1 and H3; whereas H-1 (NX-19012) showed slightly better

results while H-3 (FH-352) performed the least although the difference was

statistically non significant. Differential response of drought stressed sunflower

hybrids to foliar application of SA at two growth stages showed non significant

difference between two growth stages (Figure 3.30 b). However, SA application in

0.75 mM concentration at both stages produced significantly higher oil content in

drought stressed plants as compared to those receiving no SA. Whereas, control

treatment (DS-0 + SA-0) rendered the highest value for oil content in non-stressed

plants, which was significantly superior to all the treatments in drought stressed

sunflower hybrids plants. Within all the treatments, interaction of DS with various


application at vegetative growth stage. Generally, the protective effect of salicylic

acid on drought-stressed sunflower plants enhanced with increasing SA

concentration up to 0.75 mM, but reduced at further higher concentration.

Significant reduction was produced in oil percentage when stress was

imposed at both stages particularly at flowering stage (Table 3.30). Similarly,

Bajehbaj (2011) showed that four sunflower cultivars decreased oil percentage and

oil yield significantly upon the application of water deficit stress. Akhtar et al. (1993)

observed that sunflower grown without water stress had maximum seed oil content

(41.3%) than plants subjected to water stress at seed-setting which gave the

minimum seed oil content.

Application of SA concentrations improved the oil % better at vegetative

stage (Fig. 3.30 b). These results are in line with the findings that exogenous foliar

Table 3.30: Oil content (%) of drought stressed sunflower hybrids under foliar


289




H-1 (NX-19012) 31.0 NS 29.8 30.4 B

H-2 (NX-00989) 33.7 32.5 33.1 A

H-3 (FH-352) 30.0 28.8 29.4 B


Control (DS-0 + SA-0) 36.0 NS 34.8 35.4 A

DS + SA-0 27.6 26.4 27.0 C

DS + SA-0.375 30.8 29.6 30.2 B

DS + SA-0.75 31.8 30.6 31.2 B

DS + SA-1.50 31.6 30.4 31.0 B


H-1 × DS-0 + SA-0 35.8 NS 34.6 35.2 NS

H-1 × DS + SA-0 27.9 26.7 27.3

H-1 × DS + SA-0.375 29.0 27.8 28.4

H-1 × DS + SA-0.75 31.4 30.2 30.8

H-1 × DS + SA-1.50 30.8 29.6 30.2

H-2 × DS-0 + SA-0 38.8 37.6 38.2

H-2 × DS + SA-0 28.7 27.5 28.1

H-2 × DS + SA-0.375 34.4 33.2 33.8

H-2 × DS + SA-0.75 33.1 31.9 32.5

H-2 × DS + SA-1.50 33.6 32.4 33.0

H-3 × DS-0 + SA-0 33.5 32.3 32.9

H-3 × DS + SA-0 26.1 24.9 25.5

H-3 × DS + SA-0.375 29.1 27.9 28.5

H-3 × DS + SA-0.75 31.0 29.8 30.4

H-3 × DS + SA-1.50 30.3 29.1 29.7

Means (GS) 31.6 A 30.4 B



DS=Drought Stress

290


Figure 3.30: Oil content of drought stressed sunflower hybrids under foliar

applied various concentrations of salicylic acid at two growth stages. application

of SA (100 ppm) to sunflower at vegetative stage produced maximum achene oil

(40.7 %), but lower values with SA treatment at flowering stage (Ahmad et al.,

2009). Salicylic acid treatment caused significant increase in oil and protein content

in sunflower (Dawood et al., 2012). Contrastingly, Gharib (2006) stated that


27.0

29.0

31.0

33.0

35.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


20.0

25.0

30.0

35.0

40.0




291

exogenously applied SA did not increase oil yield by increasing photosynthesis and

nutrient up take.

4.3.31 Head Diameter

Difference between two growth stages of sunflower plants for the application

of salicylic acid was statistically significant for head diameter (Table 3.31). The SA

applied to drought stressed plants at vegetative growth stage produced greater head

diameter as compared to that sprayed at flowering stage. Three sunflower hybrids

differed significantly with each other for head diameter under foliar application of

salicylic acid in different concentrations at two growth stages. The H-3 (FH-352)

plants had greater head diameter than that of other two genotypes, and H-1 (NX-

19012) had the smallest one. Further, there was significant difference among

different concentrations of salicylic acid application to water-stressed plants. The

greatest head diameter (13.7 cm) was in control plants without receiving both

drought stress and salicylic acid, while the smallest one (8.9 cm) was under DS+SA-

0 treatment (drought stressed but receiving no salicylic acid). Application of salicylic

acid at the concentration of 0.75 mM resulted in the highest and significant increase

of head diameter of drought stressed plants. There was significant decrease in head

diameter due to drought stress, which was significantly addressed by salicylic acid

application. All the interactions among different treatment factors (H, GS and SA)

were statistically significant.


sunflower at which SA was applied, has been shown in Figure 3.31 a. It indicates

that sunflower hybrids differed significantly for SA application; head diameter of all

genotypes was greater if SA was sprayed at vegetative growth stage as compared to

that at flowering stage. The H-3 (FH-352) gave significantly better results, while H-

1 (NX-19012) performed the least as the difference was statistically significant.

292

Similarly, drought-stressed sunflower hybrids showed significant differential

response to foliar application of SA at two growth stages (Figure 3.31 b). However,


greater head diameter of drought stressed plants as compared to those receiving no

SA. Whereas, control treatment (DS-0 + SA-0) rendered the highest value for head

diameter of non stressed plants, which was significantly superior to all the treatments

in drought stressed sunflower plants. Within all the treatments, interaction of DS

with various SA concentrations showed better response of sunflower hybrids to SA

foliar application at vegetative growth stage. Further, shielding effect of salicylic

acid on drought-stressed sunflower plants enhanced with increasing SA

concentration, being highest at the 0.75 mM, and lesser at next higher concentration

of 1.50 mM.

Head diameter reduced under water stress especially at flowering stage, SA

application well ameliorated the negative effects of drought (Table 3.30). The

partitioning of dry matter to the head is critical in the process of yield determination

in water stressed Parsley (Petropoulos et al., 2008). There was a Table 3.31: Head

diameter (cm) of drought stressed sunflower hybrids under foliar applied





H-1 (NX-19012) 10.7 d 9.4 f 10.1 C

H-2 (NX-00989) 11.4 b 10.3 e 10.9 B

H-3 (FH-352) 11.8 a 11.1 c 11.5 A


Control (DS-0 + SA-0) 13.8 a 13.7 a 13.7 A

DS + SA-0 9.4 e 8.4 f 8.9 E

DS + SA-0.375 10.8 c 9.4 e 10.1 C

DS + SA-0.75 12.2 b 10.6 cd 11.4 B

DS + SA-1.50 10.4 d 9.2 e 9.8 D


293

H-1 × DS-0 + SA-0 12.9 b 12.9 b 12.9 B

H-1 × DS + SA-0 8.3 q 7.4 r 7.8 I

H-1 × DS + SA-0.375 10.5 hij 8.5 pq 9.5 GH

H-1 × DS + SA-0.75 11.9 de 9.4 lm 10.7 EF

H-1 × DS + SA-1.50 10.0 jk 8.6 opq 9.3 H

H-2 × DS-0 + SA-0 14.1 a 14.0 a 14.1 A

H-2 × DS + SA-0 9.8 kl 8.8 nop 9.3 H

H-2 × DS + SA-0.375 10.6 hi 9.2 mn 9.9 G

H-2 × DS + SA-0.75 12.4 cd 10.6 hi 11.5 D

H-2 × DS + SA-1.50 10.3 hijk 8.9 mnop 9.6 GH

H-3 × DS-0 + SA-0 14.3 a 14.2 a 14.2 A

H-3 × DS + SA-0 10.3 hijk 9.1 mno 9.7 GH

H-3 × DS + SA-0.375 11.4 fg 10.4 hij 10.9 E

H-3 × DS + SA-0.75 12.3 de 11.7 ef 12.0 C

H-3 × DS + SA-1.50 10.8 gh 10.1 ijk 10.5 F

Means (GS) 11.3 B 10.3 B



DS=Drought Stress


6.00

8.00

10.00

12.00

14.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


294


Figure 3.31: Head diameter of drought stressed sunflower hybrids under foliar

applied various concentrations of salicylic acid at two growth stages. negative

correlation of head diameter with fresh root and shoot weight, while a positive one

between dry shoot weight and achene yield per plant under water stress (Tahir and

Mehdi, 2001). Drought stress affects head size in sunflower (Nizami et al., 2008).

The stress from head formation until the end of growing season had the highest effect

on components of yield, especially 1000-grain weight and head diameter (Nazariyan,

2009).

4.3.32 Number of Achenes

Number of sunflower achenes per head of a plant was significantly greater when

salicylic acid was applied at vegetative growth stage (473) as compared to that for

foliar application of SA at flowering stage (453) of plants (Table 3.32). Drought

stressed three sunflower hybrids also had significant difference with each other for

number of achenes under foliar application of salicylic acid at two growth stages.

The H-3 (FH-352) produced significantly higher number of achenes (523) as

compared to other two hybrids, which had no statistical difference. Likewise, there

6.0

8.0

10.0

12.0

14.0

16.0




295

was significant difference among different concentrations of salicylic acid

application to drought-stressed sunflower genotypes. Although the highest number

of achenes (520) was recorded in control, but the lowest number (426) was in

drought stressed plants without salicylic acid (DS+SA-0). Reduction in the number

of achenes due to drought stress was significantly countered by SA application

salicylic acid at the concentration of 0.75 mM caused maximum and significant

increase in the number of achenes in drought-stressed plants as compared to other

SA concentrations. All the treatment interactions viz.H×GS, SA×GS,H×SA, and

H×SA×GS were statistically significant for mean values regarding number of

achenes in sunflower genotypes.

Interaction of three sunflower hybrids with two growth stages of sunflower

for SA application showed that the number of achenes in all sunflower hybrids

differed equally by SA applied at vegetative growth stage or that at flowering stage

(Figure 3.32 a). The H-3 (FH-352) showed statistically better results, while H-1 (NX-

19012) and H-2 (NX-00989) performed lesser as the difference between them was

also non significant. Similarly, differential response of drought stressed sunflower

hybrids to foliar application of SA at two growth stages was significant (Figure 3.32

b). The SA application in 0.75 mM concentration at both stages produced

significantly higher number of achenes in drought-stressed plants as compared to

those receiving no SA or lower / higher SA concentrations. Whereas, control

treatment (DS-0 + SA-0) rendered the highest values for number of achenes, which

was significantly superior to all the treatments in drought-stressed sunflower hybrid

plants. Among all the treatments, interaction of DS with various SA concentrations

showed better response of sunflower hybrids to SA application at vegetative growth

stage especially for 0.75 mM concentration. These results reflected that positive

effect of salicylic acid on drought-stressed sunflower plants increased with elevated

296

SA concentration only up to 0.75 mM concentration, beyond which its impact

declined significantly.

Water stress both at vegetative and flowering stage badly affects the yield

components of crop but stress causes more harm at flowering stage (Table 3.313.36).

Results are in line with the findings of Petcu et al., (2003), who stated that irrigation

stopped at flowering stage significantly reduced number of achenes/ head. Water

provides an important media for the biochemical and physiological reactions the

deficiency of which badly affected the final yield. Reduced number Table 3.32:

Number of achenes/headof drought stressed sunflower hybrids under foliar





H-1 (NX-19012) 439 c 425 d 432 B

H-2 (NX-00989) 442 c 424 d 433 B

H-3 (FH-352) 537 a 510 b 523 A


Control (DS-0 + SA-0) 521 a 520 a 520 A

DS + SA-0 436 e 415 f 426 D

DS + SA-0.375 461 c 437 e 449 C

DS + SA-0.75 490 b 453 d 471 B

DS + SA-1.50 456 d 441 e 449 C


H-1 × DS-0 + SA-0 479 fg 479 fg 479 F

H-1 × DS + SA-0 403 k 387 l 395 K

H-1 × DS + SA-0.375 429 i 418 j 423 H

H-1 × DS + SA-0.75 463 h 422 ij 443 G

H-1 × DS + SA-1.50 423 ij 421 ij 422 HI

H-2 × DS-0 + SA-0 502 de 499 e 501 D

H-2 × DS + SA-0 404 k 386 l 395 K

H-2 × DS + SA-0.375 428 i 406 k 417 IJ

H-2 × DS + SA-0.75 456 h 427 i 441 G

H-2 × DS + SA-1.50 422 ij 405 k 413 J

H-3 × DS-0 + SA-0 581 a 582 a 582 A

297

H-3 × DS + SA-0 502 de 472 g 487 E

H-3 × DS + SA-0.375 527 c 487 f 507 C

H-3 × DS + SA-0.75 550 b 510 d 530 B

H-3 × DS + SA-1.50 523 c 497 e 510 C

Means (GS) 473 A 453 B



DS=Drought Stress


Figure 3.32: Number of achene of drought stressed sunflower hybrids under



350

400

450

500

550

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


350

400

450

500

550




298

of achenes might be related to the decreased turgor potential which affects the

elongation and expansion of cells leads to inhibition of leaf area which resulted in

reduced photosynthesis.

Exogenous application of SA could significantly ameliorate the negative

effects of moisture stress at both stages especially at vegetative stage (Fig. 3.32 b).

Similar results were obtained by Ahmad et al. (2009) that among different treatments

of exogenous application of SA, maximum number of achenes head-1 (986.11) was

recorded, when foliar application of SA (100 ppm) was done at vegetative stage, but

minimum values at flowering stage. Results are also in agreement with the findings

of Sharma and Kaur (2003) who concluded that in soybean foliar application of SA

increased the number of grains pod-1.

4.3.33 1000 Grain Weight

Difference between two growth stages of plants for SA application was significant

regarding 1000 grain weight of sunflower, being higher (55.1) for vegetative growth

stage (Table 3.33). Among three drought-stressed sunflower hybrids, H-3 (FH-352)

had significantly lower 1000 grain weight (42.3 g) as compared to other two

genotypes which differed non significantly with each other. There was significant

difference among different concentrations of salicylic acid applied to drought-

stressed sunflower plants. The highest 1000 grain weight (64.6g) was recorded under

control treatment (DS-0+SA-0), while the lowest (46.0 g) was that from drought

stressed plants receiving no salicylic acid (DS+SA-0). Spray with salicylic acid at

0.75 mM concentration resulted in the maximum and significant counter of drought

stress for 1000 grain weight. There was significant decrease in

1000 grain weight with the employment of drought stress, which was significantly

addressed by salicylic acid application. Treatment interactions were statistically

significant only for concentrations of salicylic acid (SA) × growth stages (GS).

299

Three sunflower hybrids showed non significant interaction with two growth

stages for SA foliar application with respect to 1000 grain weight (Figure 3.33 a). It

was obvious that sunflower hybrids differed similarly with SA applied at vegetative

growth stage and flowering stage. The H-3 (FH-352) showed significantly lower

results, while H-1 (NX-19012) and H-2 (NX-00989) performed better, and the

difference between them was statistically non significant. However, differential

response of drought-stressed sunflower genotypes to SA application at two growth

stages was significant (Figure 3.33 b). Further, SA application in 0.75 mM

concentration at both stages produced significantly greater 1000 grain weight of

drought stressed plants as compared to those receiving lower or higher SA

concentrations. Whereas, control treatment rendered the highest value for 1000 grain

weight, which was significantly superior to all the treatments in droughtstressed

sunflower plants. For all the drought-stress treatments, interaction of DS with various

SA concentrations showed better response of sunflower hybrids to SA application at

vegetative growth stage. Results indicated that salicylic acid application to drought-

stressed sunflower plants countered the stress effect better with increasing SA

concentration up to 0.75 mM, and further higher concentration had lower impact.

The seed weight is an important and efficient component of plant yield. 1000

achene weight was reduced under water deficit conditions in all sunflower hybrids

at both stages, particularly at flowering stage (Table 3.33). The factor which causes

changes in thousand seed weight is the potential number of flowers Table 3.33:

1000 grain weight (g) drought stressed sunflower hybrids under foliar applied





H-1 (NX-19012) 60.7 NS 56.8 58.8 A

H-2 (NX-00989) 60.5 56.1 58.3 A

300

H-3 (FH-352) 44.1 40.5 42.3 B


Control (DS-0 + SA-0) 64.6 a 64.6 a 64.6 A

DS + SA-0 47.6 e 44.3 f 46.0 D

DS + SA-0.375 53.2 c 47.7 e 50.5 C

DS + SA-0.75 58.4 b 50.8 d 54.6 B

DS + SA-1.50 51.7 cd 48.1 e 49.9 C


H-1 × DS-0 + SA-0 70.6 NS 70.7 70.6 NS

H-1 × DS + SA-0 51.7 49.2 50.5

H-1 × DS + SA-0.375 59.1 53.4 56.2

H-1 × DS + SA-0.75 65.1 56.0 60.6

H-1 × DS + SA-1.50 57.1 54.8 55.9

H-2 × DS-0 + SA-0 71.0 69.3 70.1

H-2 × DS + SA-0 54.1 50.7 52.4

H-2 × DS + SA-0.375 58.3 52.7 55.5

H-2 × DS + SA-0.75 62.9 56.0 59.4

H-2 × DS + SA-1.50 56.4 51.6 54.0

H-3 × DS-0 + SA-0 52.1 53.8 53.0

H-3 × DS + SA-0 37.1 33.1 35.1

H-3 × DS + SA-0.375 42.2 37.0 39.6

H-3 × DS + SA-0.75 47.3 40.3 43.8

H-3 × DS + SA-1.50 41.6 38.1 39.9

Means (GS) 55.1 A 51.1 B



DS=Drought Stress

301


Figure 3.33: 1000 grain weight of drought stressed sunflower hybrids under


which is determined during the plant growth period, particularly by leaf distribution

(Jonson, 2003; Lio et al., 2004). Physiological and biochemical features of plant

changed under drought (Keyvan, 2010). Reduction in head diameter, 1000-achene

weight and number of achenes per head due to water stress can be related with

reduced photosynthetic area as is evident from lower leaf area, which then resulted


30.0

40.0

50.0

60.0

70.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


30.0

40.0

50.0

60.0

70.0




302

in reduced crop growth. Also reduced turgor can affect the cell elongation, thus

reducing the leaf area (Sadras et al. 1993). As water is also one of the important input

for p/s, medium for reactions and dry matter accumulation, so reduced water

availability sternly affected the allometry and ultimately the final yield. Drought

stress at flowering stage was found to be more damaging. It might be due to the fact

that water stress at flowering stage increases the chances of pollen sterility, abortion,

pollen germination and fertilization in-compatibility, which directly reduces the

number of achenes and achene yield. Drought stress also increases the ROS

production (Alscher and Hess, 1993), which attack all the biological membranes,

DNA, lipids and proteins; also cause stomatal closure and decreases the RuBP

content (Flexas and Medrano, 2002). Similar findings were reported by Nazariyan

(2009) as stress from head formation until the end of growing season had the highest

effect on components of yield, especially 1000grain weight and head diameter.

SA increased the achene weight especially in drought tolerant hybrids at

vegetative stage (3.33 a, b). SA application improved the head diameter, 1000achene

weight and number of achenes per head under stress conditions (Hussain et al.,

2009). SA might act as signaling molecule to protect the photosynthetic machinery

from ROS and also by increasing RuBP (Singh and Usha, 2003; Yang et

al., 2004).

4.3.34 Achene Yield

Comparison for foliar spray of salicylic acid at two growth stages of drought-

stressed sunflower plants showed significantly higher achene yield (25.9 g/plant)

with SA applied at vegetative stage (Table 3.34). Drought stressed three sunflower

hybrids also differed significantly with higher achene yields of H-1 and H-2, as H-3

(FH-352) had the lowest yield (22.3 g/plant). Further, there was significant

difference among different concentrations of salicylic acid applied to drought-

stressed sunflower plants. The highest achene yield (33.3 g/plant) was obtained from

control treatment (DS-0+SA-0), while the lowest (19.3 g/plant) was that ofdrought

stressed plants receiving no salicylic acid (DS+SA-0). Application of salicylic acid

303

at the concentration of 0.75 mM resulted inthe maximum achene yield in drought-

stressed plants being significantly higher than that with all other lower or higher SA

concentrations. There was significant decrease in achene yield with the induction of

drought stress, which was highly addressed by salicylic acid application. Interactions

among treatments were statistically non significant except for salicylicacid (SA) ×

growth stages (GS).



a. It indicates that sunflower hybrids did not differ statistically for SA applied

whether at vegetative growth stage or that at flowering stage. H-3 (FH-352) showed

significantly lower results, while H-1 (NX-19012) and H-2 (NX-00989) performed

similarly, and the difference between them was statistically non significant.

Differential response of drought stressed sunflower hybrids to foliar application of

different SA concentrations at two growth stages was significant (Figure 3.34 b).

The SA application in 0.75 mM concentration at both stages produced significantly

higher achene yield of drought stressed plants as compared to those receiving no SA

or its lower / higher concentrations whereas, control treatment (DS-0 + SA-0)

rendered the highest value for achene yield, which was significantly superior to all

the treatments in drought-stressed sunflower hybrid

plants. Within all the DS treatments, interaction of DS with various SA

concentrations showed better response of sunflower hybrids to SA foliar

application at vegetative growth stage. Ameliorative effect of salicylic acid on

drought-stressed sunflower plants significantly improved with increasing SA

concentration, being highest at the 0.75 mM, but lower at next higher concentration

of 1.50 mM.

Drought stress causes great reduction in achene yield in all the hybrids of

sunflower at both stages but damage was more pronounced at flowering stage Table

3.34). These results are in agreement with the findings of Laghrab et al. (2003) and

304

Demir et al. (2006) also reported that drought condition occurred at flowering stage

greatly reduced achene yield. Flowering stage was supposed to be more damaging

to drought stress. It might be the chances of pollen infertility, abortion, pollen

germination and fertilization in-compatibility increases due to moisture stress at

flowering stage, which straightly reduces the number of achenes per head and achene

yield. Water stress also enhanced the production of ROS (Alscher and Hess, 1993),

which attack all the biological membranes, DNA, lipids and proteins; also cause

stomatal closure and lowers the RuBP content (Flexas and Medrano, 2002). Similar

results were observed by Demir et al. (2006), who reported that maximum achene

yield was produced when sunflower was irrigated at three critical stages i.e. heading,

flowering and milking. The limitation of irrigation

at the flowering period should be avoided. Results are corroborated with the findings

of Reddy et al. (2004), who observed decrease in seed yield in sunflower by

imposing moisture stress.

Table 3.34: Achene yield (g/plant) of drought stressed sunflower hybrids under

foliar applied various concentrations of salicylic acid at two growth

stages.




H-1 (NX-19012) 26.9 NS 24.4 25.6 A

H-2 (NX-00989) 27.0 24.1 25.5 A

H-3 (FH-352) 23.8 20.9 22.3 B


Control (DS-0 + SA-0) 33.3 a 33.2 a 33.3 A

DS + SA-0 20.4 e 18.1 f 19.3 D

DS + SA-0.375 24.2 c 20.6 e 22.4 C

DS + SA-0.75 28.3 b 22.7 d 25.5 B

DS + SA-1.50 23.3 cd 21.0 e 22.1 C


H-1 × DS-0 + SA-0 33.8 NS 33.8 33.8 NS

H-1 × DS + SA-0 20.9 19.0 20.0

305

H-1 × DS + SA-0.375 25.3 22.3 23.8

H-1 × DS + SA-0.75 30.2 23.6 26.9

H-1 × DS + SA-1.50 24.1 23.1 23.6

H-2 × DS-0 + SA-0 35.7 34.6 35.1

H-2 × DS + SA-0 21.9 19.6 20.7

H-2 × DS + SA-0.375 25.0 21.4 23.2

H-2 × DS + SA-0.75 28.6 23.9 26.3

H-2 × DS + SA-1.50 23.8 20.9 22.3

H-3 × DS-0 + SA-0 30.3 31.3 30.8

H-3 × DS + SA-0 18.6 15.6 17.1

H-3 × DS + SA-0.375 22.2 18.0 20.1

H-3 × DS + SA-0.75 26.1 20.5 23.3

H-3 × DS + SA-1.50 21.8 18.9 20.4

Means (GS) 25.9 A 23.1 B



DS=Drought Stress


15.0

20.0

25.0

30.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


306


Figure 3.34: Achene yield of drought stressed sunflower hybrids under foliar


Increase in achene yield by the exogenous application of SA (Fig. 3.34 a, b)

was found to be the direct result of improved number of achenes per head and 1000-

achene weight. Earlier, Fariduddin et al. 2003 reported that as compared to control

mustard plant yielded more number of pods per plant and seed yield when sprayed

with salicylic acid.

4.3.35 Oil Yield

Difference between two growth stages of salicylic acid application was

significant for oil yield of drought-stressed sunflower plants (Table 3.35). Spray of

SA at vegetative stage rendered greater oil yield (8.30 g/plant) as compared to that

at flowering stage (7.17 g/plant). Drought stressed three sunflower hybrids also

differed significantly with each other for oil yield under foliar application of salicylic

acid in different concentrations at two growth stages. The H-2 (NX00989) gained

the highest oil yield (8.61 g/plant) while H-3 (FH-352) produced the lowest one (6.68

g/plant). Likewise, there was significant difference among various concentrations of

15.0

20.0

25.0

30.0

35.0




307

salicylic acid application to drought-stressed sunflower plants. The highest oil yield

(11.82g/plant) was obtained from control treatment (DS- 0+SA-0), while the lowest

(5.22 g/plant) was under DS+SA-0 (drought stress without salicylic acid spray).

There was significant decrease in oil yield with the employment of drought stress,

which was highly countered by salicylic acid. Application of salicylic acid at the

concentration of 0.75 mM resulted the highest increase of oil yield in drought

stressed plants with a significant difference from other concentrations. Interactions

among various treatment factors were statistically significant for: sunflower hybrids

(H) ×conc. of salicylic acid (SA), and SA × GS.

Interactive effect of three sunflower hybrids with two growth stages of sunflower at

which SA foliar application was made, has been drawn in Figure 3.35 a. Sunflower

hybrids did not differ regarding oil yield as obtained through SA application either

at vegetative growth stage or at flowering stage, even all showing similar response.

The H-3 (FH-352) showed statistically inferior results, while H-2 (NX-00989)

performed the best and the difference was significant. Similarly, comparative

response of drought stressed sunflower hybrids to SA application at two growth

stages was statistically significant (Figure 3.35 b). The 0.75 mM concentration of

SA sprayed at both stages produced significantly higher oil yield in droughtstressed

plants as compared to that with other SA concentrations. However, control treatment

rendered the highest value for oil yield in non stressed plants, which was

significantly superior to all the treatments in drought-stressed sunflower hybrids.

Among DS treatment combinations with various SA concentrations, better response

of sunflower hybrids was with SA foliar application at vegetative growth stage.

Ameliorative effect of salicylic acid to counter drought stress in sunflower plants at

the highest by raising SA concentration to 0.75 mM, above which the impact was

reduced significantly.

308

Lower oil yield found upon the imposition of water stress in sunflower

hybrids (Table 3.35) might be due to reduced achene yield and oil contents

percentage (Table 3.34, 3.29). These findings are in line as Bajehbaj (2011) showed

that in four sunflower cultivars oil percentage and oil yield decreased significantly

upon the application of water deficit stress. Akhtar et al. (1993) observed that

sunflower grown without water stress had maximum seed oil content (41.3%) than

plants subjected to water stress at seed-setting which gave the minimum seed oil

content. Exogenous foliar application of SA (100 ppm) to

Table 3.35: Oil yield (g/plant) of drought stressed sunflower hybrids under





H-1 (NX-19012) 8.44 NS 7.39 7.91 B

H-2 (NX-00989) 9.24 7.99 8.61 A

H-3 (FH-352) 7.23 6.13 6.68 C


Control (DS-0 + SA-0) 12.02 a 11.61 a 11.82 A

DS + SA-0 5.66 e 4.79 f 5.22 D

DS + SA-0.375 7.47 c 6.11 de 6.79 C

DS + SA-0.75 9.01 b 6.97 c 7.99 B

DS + SA-1.50 7.35 c 6.37 d 6.86 C


H-1 × DS-0 + SA-0 12.09 NS 11.70 11.89 B

H-1 × DS + SA-0 5.81 5.08 5.44 H

H-1 × DS + SA-0.375 7.35 6.20 6.78 G

H-1 × DS + SA-0.75 9.48 7.15 8.31 DE

H-1 × DS + SA-1.50 7.45 6.84 7.14 G

H-2 × DS-0 + SA-0 13.82 13.01 13.41 A

H-2 × DS + SA-0 6.31 5.40 5.85 H

H-2 × DS + SA-0.375 8.59 7.10 7.84 EF

H-2 × DS + SA-0.75 9.49 7.67 8.58 D

H-2 × DS + SA-1.50 7.99 6.76 7.38 FG

H-3 × DS-0 + SA-0 10.16 10.13 10.14 C

H-3 × DS + SA-0 4.86 3.90 4.38 I

309

H-3 × DS + SA-0.375 6.46 5.03 5.74 H

H-3 × DS + SA-0.75 8.07 6.10 7.09 G

H-3 × DS + SA-1.50 6.60 5.51 6.05 H

Means (GS) 8.30 A 7.17 B


letter(s) are statistically similar at P≤0.05. H=Hybrid, SA=Salicylic acid

DS=Drought Stress


Figure 3.35: Oil yield of drought stressed sunflower hybrids under foliar



5.00

6.00

7.00

8.00

9.00

10.00

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


4.00

6.00

8.00

10.00

12.00

DS-0 + SA-0 DS + SA-0 DS + SA-0.375 DS + SA-0.75 DS + SA-1.50 Salicylic acid treatments (mM ) to droughted plants


310

sunflower at vegetative stage produced maximum achene oil (40.7 %), but lower

values with SA treatment at flowering stage (Ahmad et al., 2009). The SA treatment

caused significant increase in oil content in sunflower cultivars (Dawood et al.,

2012). These results confirmed our findings that SA increased the oil yield.

4.3.36 Biological Yield

Comparison between two growth stages of foliar SA application to

droughtstressed sunflower plants showed that biological yield was significantly

higher with

SA applied at vegetative stage (49.8 g/plant) vs. flowering stage (Table 3.36).

Among three drought-stressed sunflower hybrids, H-2 (NX-00989) produced the

highest biological yield (49.8 g/plant), which differed significantly from others,

while H-3 (FH-352) had the lowest biological yield (45.2/plant). Various

concentrations of salicylic acid applications to drought-stressed sunflower plants

also differed significantly. The highest biological yield (60.5 g/plant) was recorded

in control treatment without receiving both drought stress and salicylic acid, while

the lowest (40.3 g/plant) was that of drought stressed plants without receiving SA.

Application of salicylic acid at the concentration of 0.75 mM rendered the highest

increase in biological yield of drought-stressed plants with significant difference

from other concentrations. There was significant decrease in biological yield due to

drought stress, and its effect was significantly altered by salicylic acid application.

The only significant interaction between treatments was that of sunflower hybrids

(H) × growth stages (GS). Interaction between three sunflower hybrids and two

growth stages for SA foliar application to sunflower has been shown in Figure 3.36

a. It reflects that sunflower hybrids differed more if SA was applied at vegetative

growth stage even all giving greater biological yield as compared to that at Table

3.36: Biological yield (g/plant) of drought stressed sunflower hybrids under

foliar applied various concentrations of salicylic acid at two growth stages

311




H-1 (NX-19012) 50.52 NS 46.84 48.68 B

H-2 (NX-00989) 51.96 47.75 49.85 A

H-3 (FH-352) 47.13 43.42 45.27 C


Control (DS-0 + SA-0) 60.45 a 59.66 a 60.06 A

DS + SA-0 42.38 d 38.29 e 40.33 D

DS + SA-0.375 46.97 c 42.50 d 44.73 C

DS + SA-0.75 52.64 b 46.42 c 49.53 B

DS + SA-1.50 46.92 c 43.14 d 45.03 C


H-1 × DS-0 + SA-0 60.18NS 59.86 60.02NS

H-1 × DS + SA-0 42.49 38.94 40.72

H-1 × DS + SA-0.375 47.62 43.29 45.45

H-1 × DS + SA-0.75 54.98 47.29 51.13

H-1 × DS + SA-1.50 47.34 44.84 46.09

H-2 × DS-0 + SA-0 63.47 60.53 62.00

H-2 × DS + SA-0 45.44 40.58 43.01

H-2 × DS + SA-0.375 48.86 44.75 46.80

H-2 × DS + SA-0.75 53.76 48.65 51.20

H-2 × DS + SA-1.50 48.27 44.23 46.25

H-3 × DS-0 + SA-0 57.70 58.61 58.15

H-3 × DS + SA-0 39.20 35.34 37.27

H-3 × DS + SA-0.375 44.44 39.46 41.95

H-3 × DS + SA-0.75 49.17 43.32 46.24

H-3 × DS + SA-1.50 45.13 40.36 42.76

Means (GS) 49.87 A 46.00 B



DS=Drought Stress

312



Figure 3.36: Biological yield of drought stressed sunflower hybrids under foliar

applied various concentrations of salicylic acid at two growth stages. flowering

stage. H-2 (NX-00989) produced significantly higher yield, while H-3 (FH-352)

performed the least. Likewise, differential response of drought-stressed sunflower

hybrids to foliar application of SA at two growth stages was significant (Figure 3.36

26.0

30.0

34.0

38.0

42.0

46.0

50.0

54.0

H-1 (NX-19012) H-2 (NX-00989) H-3 (FH-352)

Sunflower hybrids


20

30

40

50

60

70


Salicylic acid treatments (mM) to droughted plantsAxis


313

b). SA application in 0.75 mM concentration at both stages produced significantly

higher biological yield of drought-stressed plants as compared to those receiving no

SA. However, control treatment rendered the highest value for biological yield,

which was significantly superior to all the treatments in droughtstressed sunflower

plants. Among all the treatment combinations, interaction of DS with various SA

concentrations showed better response of sunflower hybrids to SA foliar application

at vegetative growth stage. It was evident that the protective effect of salicylic acid

on drought-stressed sunflower plants enhanced with elevated SA concentration up

to 0.75 mM, beyond which it declined.

In this study biological yield of sunflower was greatly reduced by the water

deficit at both the stages (Table 3.36).The results are in complete agreement with the

findings of Nazariyan (2009) who showed that drought stress severely decreased

biological yield of sunflower. Water stress both at vegetative and flowering stages

badly affects the growth and yield components of crop but stress causes more

damage at flowering stage. Similar findings were recorded by Petcu et al., 2003 that

biological yield might be reduced by the decrease in leaf area which ultimately

resulted in reduction of photosynthetic activity in sunflower. Tahir and Mehdi 2001

observed reduction of biomass of all sunflower genotypes under water

stress.

Exogenous application of SA could significantly ameliorate the negative

effect of moisture stress at both stages (3.36 a, b). These results are in agreement

with the findings of De-Guang et al., (2001) who reported that the foliar application

of SA on maize had clear drought resistance and yield increasing effects. Among

different treatments of exogenous application of SA, maximum biological yield was

recorded, when foliar application of SA (100 ppm) was done at vegetative stage,

(Ahmad et al., 2009).

315

SUMMARY

The present study to investigate the possible role of exogenous application

of SA to mitigate the effects of moisture stress in autumn planted hybrid sunflower

was carried out at the National Agriculture Research Center, Islamabad, Pakistan

during autumn 2009 and 2010 and the entire analytical work was conducted at the

Stress Physiology Laboratory, NARC and Central lab of PMAS UAAR, Pakistan.

The study consisted of two laboratory experiments and the adult stage in Green

house. In the first lab experiment Screening was done with six sunflower hybrids

viz., Hyoleic-41, FH-352, NX-00989, Hysun-33, NX-19012 and Parsun-2 were

subjected to drought stress imposed at germination and seedling growth stages was

investigated in a laboratory experiment (25±30C). Four water stress levels of zero

(control), -0.6, -1.33, and -1.62 MPa were developed using polyethyleneglycol 8000

(PEG-8000). Complete randomized design with three replications was used for this

experiment. Germination stress tolerance index (GSI), plant height stress index

(PHSI), root length stress index (RLSI) and dry matter stress index (DMSI) were

used to evaluate the genotypic response to PEG-induced water stress. The second

lab experiment was conducted to evaluate the mode of application. It was divided in

two sub experiments: Two tolerant and one sensitive hybrid viz: NX19012 (H-1),

NX-00989 (H-2), and FH-352 (H-3) respectively, were

selected.Three levels of drought (PEG-8000) (0, 10%, 20%), four levels of salicylic

acid (0, 0.375, 0.75 and 1.50 mM) under laboratory conditions (25±30C) was

conducted. The experiment was laid out in a completely randomized design with

three replications. For foliar SA application: Sunflower‘s surface sterilized seeds

were grown in Hoagland‘s solution. Drought stress was applied to the 15 days old

316

312

plant + SA was applied as foliar spray to the plants. For SA seed treatment: Seeds

were soaked in SA solution for 10hrs. The soaked seeds were grown in Hoagland‘s

solution for 15 days. Drought stress was applied to the plants. Fresh and dry biomass,

plant water relations, physiological and biochemical attributes were assessedin both

experiments. Adult stage experiment was conducted in green house conditions, to

study the induced response of foliar application ofsalicylic acid on Helianthus

annuus hybrids NX-19012 (H-1), NX-00989 (H-2), and FH-352 (H-3) at drought

stress. A factorial experiment with a completely randomized design in three

replications, four concentrations of salicylic acid (0, 0.375, 0.75 and 1.5mM) as

foliar spray at vegetative and flowering stages under water stress was performed. To

one set of pots water stress was given at vegetative stage by withholding water till

wilting and sprayed with varying concentrations of SA (0, 0.375, 0.75 and 1.5 mM)

applied as a foliar spray to plants and rewatered. To second set of pots water was

with held at flowering stage till wilting and foliar sprayed with varying

concentrations of SA (0, 0.375, 0.75 and 1.5 mM then irrigated. The control plants

were irrigated normally till harvesting. Germination, plant height and dry matter

stress tolerance indices for all sunflower hybrids decreased with increasing water

stress. In contrast, an increase in RLSI was observed in all sunflower hybrids except

Hyoleic-41 and FH352. Sunflower hybrids NX-00989 and NX-19012 performed

better and were classified as drought tolerant and Hyoleic-41 and FH-352 were

categorized as sensitive. The variation among hybrids for DMSI was found to be a

reliable indicator of drought tolerance in sunflower. From the results of the seedling

experiment, it is clear that water stress caused a marked inhibitory effect on seedling

growth of sunflower hybrids. This adverse effect of drought stress on seedling

growth was more on H-3 as compared to H-1and H-2. Moisture stressinduced

reduction in seedling growth of hybrids was counteracted by 0.75mM concentration

317

of SA applied as foliar spray significantly as compared to other treatments and seed

soaking mode of application. Growth and biomass was greatly reduced by the

drought stress in both stressed and non stressed plants in both modes was addressed

by the application of SA better results by foliar spray in tolerant hybrids. Compatible

solutes played very significant role in osmotic adjustment. Water stress increased

the level of proline, sugar and amino acids but reduced proteins SA application

further enhanced the level and helped in maintaining leaf protein amount. Foliar

application showed higher values over pre treatment of seeds with SA

concentrations.

Water stress at both the stages significantly decreased the achene yield, oil

yield and yield contributing factors (head diameter, number of achenes per head and

1000-achene weight); water stress at flowering was the most critical. Exogenous

applications of SA at vegetative and flowering stages ameliorated the negative

effects of water stress on yield and all yield contributing factors, particularly at

vegetative stage. Biological yield was significantly decreased by imposing water

stress at vegetative (budding) and flowering stage. Water stress at vegetative

(budding) stage was the most critical and it was significantly improved by foliar

application of 0.75mM SA. Achene yield, oil yield and biological yield was higher

in drought tolerant hybrids than the sensitive one.

Shoot and root fresh and dry biomass, and leaf area, were decreased by water

stress at both stages but it was improved by the exogenous application of SA at

vegetative and flowering stages, leaf number had shown no alteration because of

stress and SA application.

Leaf relative water contents (%), leaf water potential, leaf osmotic potential

and leaf turgor potential were significantly decreased by imposing water stress both

at vegetative and flowering stages but reduction was more obvious when water stress

was applied at flowering. However all these water relation parameters were

318

improved by exogenous application of SA at vegetative (budding) and flowering

stages but the results were more pronounced by 0.75mM application at vegetative

stage.

Regarding biochemical attributes, leaf free proline, soluble sugars and free

amino acids and chlorophyll, contents accumulation was increased whereas leaf

proteins were decreased by imposing water stress but all parameters were increased

by application of SA. SA 0.75 mM had better affect when applied at vegetative stage

The activities of all antioxidant enzymes (SOD, POD and CAT) were

increased in the leaves of water stressed plants of all the sunflower hybrids and

enhanced more due to exogenous SA foliar application. Maximum increase was in

H-2 while minimum in H-3. Enzymes showed their higher values at flowering

stage.

It was observed that pronounced changes occur in fatty acid composition due

to imposition of drought stress in sunflower hybrids. Sunflower oil contains large

amounts of unsaturated fatty acids mainly linoleic acid and oleic acid. There was a

significant negative effect of moisture stress on the oleic acid and stearic acid in all

hybrids while linoleic acid and palmitic acid increased in sunflower seeds grown in

water stress.SA had shown no counteracting role in ameliorating the negative effects

of drought.

Achene protein contents (%) were significantly increased by water stress; maximum

achene protein contents were resulted when crop faced water stress at flowering.

Exogenous applications of SA at vegetative and flowering stages decreased the

achene protein contents significantly. Tolerant hybrids had higher values while

lower in sensitive at both stages of growth. Achene oil contents (%) were

significantly decreased by water stress; minimum achene oil contents were resulted

when crop faced water stress at flowering and maximum in control. Exogenous

applications of SA at vegetative and flowering stage slightly improve the oil

319

contents. H-2 showed increased oil contents in contrast H-1 and H-3 had lowest

values.

320

CONCLUSION

Physiological status of plants can directly affected by the water stress or

extremes of water accessibility which disturbed their metabolism, growth,

development and productivity. Moreover, physiological and biochemical changes

induced by water stress at the cellular level include turgor loss, changes in membrane

fluidity and composition, changes in solute concentration, and protein– protein and

protein–lipid interactions. In response, plants have developed different

physiological, biochemical and molecular mechanisms to meet water stress, such as

osmotic adjustment, enhancement in antioxidant activity. So, series of laboratory and

greenhouse pot experiments were conducted to assess the role of SA exogenous

application in inducing drought tolerance in three sunflower hybrids (two tolerant

and one sensitive), following conclusions can be drawn:

1. Sunflower germination and growth was severely hampered by PEG

induceddrought stress. All the studied parameters viz: GSI, PHSI and DMSI were

reduced under stress except RLSI.

2. Comparing different modes, foliar application was found better in mitigating

the harmful effects of PEG induced drought stress than seed soaking at seedling stage

of growth. Of various levels of salicylic acid, 0.75 mM concentration applied as

foliar or as a pre-soaking seed treatment was observed to be most effective in

enhancing growth under drought stress and non-stress conditions.

3. In pot experiment, reduction in growth of sunflower hybrids under drought

stress was alleviated by exogenous application of salicylic acid. Foliar application

of SA was showed to be most useful at vegetative stage in promoting growth and

yield under water stress.

321

317

4. Growth of drought stressed plants of sunflower was directly related with

increased photosynthetic rate. A positively correlation was observed between

photosynthesic activity with stomatal conductance and leaf area.

5. Improvement in photosynthetic efficiency was partially associated with

amount of photosynthetic pigments, may be due to osmo-protective role of SA on

chloroplast (thylakoid membranes).

6. Exogenous application of different concentrations of salicylic acid, particularly

0.75mM at vegetative stage, significantly mitigated theadverse effects of water stress

on leaf relative water contents, water potential, osmotic potential and turgor potential

by increasing the biosynthesis of osmoprotectants (proline and sugars etc).This could

be one of the reasons to reduce oxidative stress.

7. To counteract the adverse effects of stress due to over-production ROS, plants

have evolved some defense mechanisms in the form of production of some key

antioxidant enzymes peroxidase (POD), superoxide dismutase (SOD), catalase

(CAT). Reduction in the adverse effects of photosynthetic capacity of water stressed

plants due to exogenous application of SA could also be due to increased antioxidant

capacity by increasing activities of antioxidant enzymes (SOD, POD and CAT).

8. Exogenous use of Salicylic acid enhanced the endogenous levels of SA that

might have direct and indirect osmoprotective effect thereby mitigating oxidative

stress on photosynthetic machinery, which finally resulted in enhanced growth, yield

and yield quality.

9. Drought stress reduced seed quality of sunflower hybrids in terms of seed

protein, oil and fatty acid, which was also improved due to exogenous application of

SA. Exogenous application of SA also enhanced the seed oil quality by increasing

322

the oil un-saturation (in terms of oleic acid contents) that ultimately enhanced the oil

quality.

10. Water stress-induced reduction in number of grains, 1000 grain weight and

seed yield per plant was alleviated by exogenous application of SA as foliar spray

treatment by increasing source activity or preferential partitioning of assimilates.

11. Of different SA concentrations, 0.75 was found to be the most effective in

promoting growth, different physiological and biochemical attributes, grain yield,

seed and seed oil quality of sunflower hybrids.

Drought stress-induced reduction in growth of sunflower can be alleviated

by exogenous use of salicylic acid. However, efficacy of the SA was higher when

applied at vegetative growth stage, suggesting that vegetative phase before the onset

of reproductive stage is critical that overall determines plant growth and yield, and

0.75mM concentration best ameliorated the adverse effects of drought

stress.

LITERATURE CITED

A. O. A. C. 1990. Official Methods of Analysis. 15 th Ed. Association of Official

Analytical Chemists, Inc., Virginia, USA, pp: 770-771.

Abbate, P. E., J. L. Dardanellib, M. G. Cantareroc, M. Maturanoc, R. J. M.

Melchiorid and E. E. Sueroa. 2004. Climatic and water availability effects on

water-use efficiency in wheat. Crop Sci., 44: 474-483.

Abdel Rahim, E. A., M. A. Shallan, S. H. Ahmed and M. M. Farag. 2000. Effects of

some plant growth regulators on datura seeds oil. J. Agric. Sci. Mansoura Uni.,

25(12): 8249-8259.

323

Aberg, B. 1981. ―Plant growth regulators XLI. Monosubstituted benzoic acid‖.

Sweden J.Agric. Res., 11: 93–105.

Agarwal, S., R. K. Sairam, G. C. Srivastava and R. C. Meena. 2005. Changes in

antioxidant enzymes activity and oxidative stress by abscisic acid and salicylic

acid in wheat genotypes. Biol. Plantarum. 49: 541–550.

Ahmad, M. A., P. V. Murali and G. Marimuthu. 2014. Impact of salicylic acid on

growth, photosynthesis and compatible solute accumulation in Allium cepa L.

subjected to drought stress. Int. J. of Agri. and Food Sci., 4(1): 22-30.

Ahmad, S., R. Ahmad, M. Y. Ashraf, M. Ashraf and E. A. Waraich. 2009. Sunflower

(Helianthus annuus L.) response to drought stress at germination and seedling

growth stages. Pak. J. Bot., 41(2): 647-654.

320

Akhtar, M., M. Zubair, M. Saeed and R. Ahmad. 1993. Effect of planting geometery

and water stress on seed yield and quality of spring planted sunflower

(Helianthus annuus L.). Pak. J. Agric. Sci., 30: 73-76.

Albuquerque, F. M. C. de and N. M. de Carvalho. 2003. Effect of type of

environmental stress on the emergence of sunflower (Helianthus annuus L.),

soyabean (Glycine max L. Merril) and maize (Zea mays L.) seeds with different

levels of vigor. Seed Sci. Technol., 31: 465-467.

Al-Hakimi, A. M. A., 2001. Alleviation of the adverse effects of NaCl on gas

exchange and growth of wheat plants by ascorbic acid, thiamin and sodium

salicylate. Pak. J. Biol. Sci., 4(7): 765-770.

324

Ali, A. A., T. B. Ali and K. A. M. Nour. 2009. Antioxidants and some natural

compounds applications in relation to tomato growth, yield and chemical

constituents. Annu. Agric. Sci., Moshtohor., 47(4): 469-477.

Allakhverdiev, S. I., A. Sakamoto, Y. Nishiyama and N. Murata. 2000. Inactivation

of photosystems I and II in response to osmotic stress in Synechococcus.

Contribution of water channels. Plant Physiol., 122: 1201–1208.

Alscher, R. G. and J. L. Hess. 1993. Antioxidants in higher plants. CRC, Press,

Boca Raton, FL.

Ananieva, E. A., K. N. Christov and L. P. Popova. 2004. Exogenous treatment with

salicylic acid leads to increased antioxidant capacity in leaves of barley plants

exposed to paraquat. J. Plant Physiol., 161: 319-328.

Andrade, A., A. Vigliocco, S. Alemano, A. Llanes and G. Abdala. 2013.

Comparative morpho-biochemical responses of sunflower lines sensitive and

tolerant to water stress. Ame. J. Pl. Sci., 4: 156-167.

Anjum F., M. Yaseen, E. Rasul, A. Wahid and S. Anjum. 2003. Water stress in barley

(Hordeum vulgare L.). Effect on chemical composition and

chlorophyll contents, Pakistan J. Agr. Sci., 40: 45–49.

Anjum, S. A., L. C. Wang, M. Farooq, I. Khan, L. L. Xue and C. M. Zou. 2011.

Brassinolide application improves the drought tolerance in maize through

modulation of enzymatic antioxidants and leaf gas exchange. J. Agron. Crop

Sci., doi:10.1111/j.1439-037X.2010.00459.x.

Arfan, M., H. R. Athar and M. Ashraf. 2007. Does exogenous application of salicylic

acid through the rooting medium modulate growth and photosynthetic capacity

in two differently adapted spring wheat cultivars under salt stress? J. Plant.

Physiol., 164: 685-694.

Arnon, I. 1972. Crop Production in Dry Regions, Background and Principles. (Ed.):

N. Polunin. Leonard Hill Book, London, 1: 203-211.

http://www.scirp.org/journal/Home.aspx?JournalID=207

http://www.scirp.org/journal/Home.aspx?JournalID=207

325

Arora A., R. K. Sairam and G. C. Sriuastava. 2002. Oxidative stress and anti

oxidative system in plants. Current Science, 82: 1227–1238.

Arteca, R. N. 1996. Plant growth substances: Principles and applications. CBS

Publishers, New Delhi.

Ashraf, M. and M. R. Foolad. 2007. Roles of glycinebetaine and proline in improving

plant abiotic stress tolerance. Environ. Exp. Bot., 59: 206-216.

Ashraf, M. Y., K. Akhtar, F. Hussain and J. Iqbal. 2006. Screening of different

accession of three potential grass species from Cholistan desert for salt

tolerance. Pak. J. Bot., 38: 1589-1597.

Ashraf, M. and J. W. O, Leary. 1996. Effect of drought stress on growth, water

relations and gas exchange of two lines of sunflower differing in degree of salt

tolerance. Int. J. Plant Sci., 157: 729-732.

Ashraf, M. and S. Mehmood. 1990. Response of four Brassica species to drought

stress. Environ. Expt. Bot., 30: 93-100.

Asif, M., N. Hussain and M. B. Khan. 2001. Effect of K on growth and seed yield of

spring Sunflower (Helianthus annuusL.). Pak. J. Biol. Sci., 5(Suppl. Issue No.

1): 100-101.

Aspinall, D. and L. G. Paleg. 1981. Proline accumulation: Physiological aspects: In:

Pleg L.G. and Aspinall D. (Eds), Physiology and biochemistry of drought

resistance in plants. Academic Press, Sydney, Australia, 215-228.

Atteya A. M. 2003. Alteration of water relations and yield of corn genotypes in

response to drought stress, Bulg. J. Plant Physiol., 29: 63–76.

Awad, El- M. M. and S. A. A. Mansour. 2007. Growth, yield and quality of potato

as affected by some antioxidants. J. Agric. Sci., Mansoura, 32(8): 6661-6669.

Aydýn, B., B. Nalbantoglu. 2011. Effects of cold and SA treatments on nitrate

reductase activity in spinach leaves. Turk. J. Biol., 35: 443-448.

326

Azachi, M., A. Sadka, M. Fisher, P. Goldshlag, I. Gokhman and A. Zamir. 2002. Salt

induction of fatty acid elongase and membrane lipid modifications in the

extreme halotolerant Alga Dunaliella salina plant physiol., 129: 1320-1329.

Azimi, M. S., J. Daneshian, S. Sayfzadeh and S. Zare. 2013. Evaluation of Amino

Acid and Salicylic Acid application on yield and growth of wheat under water

deficit. Int. J. Agric. Crop. Sci., 5(8): 816-819.

Azooz, M. M. and M. M. Youssef. 2010. Evaluation of heat shock and salicylic acid

treatments as inducers of drought stress tolerance in Hassawi wheat. Am. J.

Plant Physiol., 5: 56-70.

Bacelar, E. A., J. M. Moutinho-Pereira, B. C. Goncalves, J. I. Lopes and C. M.

Correia. 2007. Physiological responses of different olive genotypes to drought

conditions. Acta. Physiol. Plant., 31(3): 611-621.

Bajehbaj, A. A. 2011. Effects of drought stress and different densities on oil yield

and biomass yield of sunflower varieties African J. of Biotech. 10(29):

56085613.

Bakhsh, I., I. U. Awan and M. S. Baloch. 1999. Effect of various irrigation

frequencies on the yield and yield components of sunflower. Pak. J. Bio. Sci.,

2: 194-195.

Baldini, M., R. Givanardi and G. P. Vanozzi. 2000. Effect of different water

availability on fatty acid composition of the oil in standard and high oleic

sunflower hyrids. Ital. J. Agron. 6: 119-126.

Bandurska, H. and A. Stroinski. 2003. ABA and proline accumulation in leaves and

roots of wild (Hordeum spontaneum) and cultivated (Hordeum vulgare

‗Maresi‘) barley genotypes under water deficit conditions. Acta Physiol.

Plant., 25: 55-61.

Barkosky, R. R. and F. A. Einhelling. 1993. Effect of salicylic acid on plant water

relationship. J. Chem. Ecol., 19: 237-247.

327

Barnett, N. M. and A. W. Naylor. 1966. Amino acid protein metabolism in

Bermuda grass during water stress. Plant Physiol., 41: 1222-1230.

Bates, L. S., R. P. Waldren and I. D. Teare. 1973. Rapid determination of free proline

for water stress studies. Plant Soil, 39: 205-207.

Ben Ahmed, H., A. Farouk, M. Arafet, M. Zid and H. Ezzeddine. 2009. Salicylic

acid induced changes on some physiological parameters in tomato grown under

salinity. The Proceedings of the International Plant Nutrition Colloquium.

Benavides-Mendoza, A., H. Ramirez-Rodriguez, V. Robledo-Torres, J.

HernandezDavila, J. G. Ramirez-Mezquitic, E. Bacopulos-Tellez, A.

Sandoval-Rangel and M. A. Bustamante-Garcia. 2002. Seed treatment with

salicylates modifies stomatal distribution, stomatal density and the tolerance to

cold stress in pepper seedlings. Proceedings of the 16th International Pepper

Conference (Tampico, Tamaulipas, Mexico, November 10th -12th).

Bezrukova, M. V., A. R. Kildibekova and A. M. Aval‘baev. 2004. Participation of

wheat germ agglutinin in regulation of cell division in apical root meristem of

wheat seedlings. J. Testol., 46: 8-35.

Bezrukova, M. V., R. Sakhabutdinova, R. A. Fatkhutdinova, I. Kyldiarova, and F.

Shakirova. 2001. The role of hormonal changes in protective action of salicylic

acid on growth of wheat seedlings under water deficit. Agrochemiya, 2: 51-54.

Borrell, A. K. and G. L. Hammer. 2000. Nitrogen dynamics and the physiological

basis of stay-green in sorghum. Crop Sci., 40: 1295–1307.

Boyer, J. S. 1982. Plant productivity and environment. Sci., 218: 443-448.

Bradford, M. 1976. A rapid and sensitive method for the quantitation of microgram

quantities of proteins utilizing the principle of protein dye-binding. Anal.

Bioche., 72: 248-254.

Bremner, J. M. 1964. Organic forms of nitrogen. In: C. A. Black et al. (eds)

Methods of soil analysis, Part-2. Am. Soc. of Agron., Inc., Medison,

328

Wisconson. 9: 1235-1255.

Bukhsh, M. A. H. A. A. U., M. Ishaque and S. H. Sadiq. 2009. Performance of

sunflower in response to exogenously applied salicylic acid under varying

irrigation regimes. J. Ani. Pl. Sci., 19(3): 130-134.

Cag, S., G. Cevahiroz, M. Sarsag and N. G. Saglam. 2009. Effect of salicylic acid

on pigment, Protein content and peroxidase activity in excised sunflower

cotyledons. Pak. J. Bot., 41(5): 2297-2303.

Canakci, S. 2011. Effects of salicylic acid on growth, biochemical constituents in

pepper (Capsicum annuum L.) seedlings. Pakistan. J. Bio. Sci.,14: 300- 304.

Chadha, K. C. and S. A. Brown. 1974. Biosynthesis of phenolic acids in tomato

plants infected with grobacterium tumefaciens.Can. J. Bot. 52: 2041-2047.

Chaitanya, K. V., D. Sundar, P. P. Jutur and A. R. Reddy. 2003. Water stress effects

on photosynthesis in different mulberry cultivars. Plant Growth Regul., 40: 75-

80.

Chance. B. and A. C. Maehly.1955. Assay of catalase and peroxidases. Methods

Enzymol. 2: 764-775.

Chaves, M. M. and M. M. Oliveira. 2004. Mechanisms underlying plant resilience

to water deficits: prospects for water-saving agriculture. J Exp. Bot., 55: 2365–

2384.

Chaves, M. M., J. P. Maroco and J. S. Pereira. 2003. Understanding Plant response

to drought: from genes to the whole plant. Funct. Plant. Biol., 30: 239-264.

Chen, H., C. Hou, J. Kuc and Y. Lin. 2001. Ca2+ -dependent and Ca2+ -independent

excretion modes of salycilic acid in tobacco cell suspension culture. J. of Exp.

Bot., 52: 1219-1226

Cho, U. H. and N. H. Seo. 2005. Oxidative stress in Arabidopsis thalianaexposed to

cadmium is due to hydrogen peroxide accumulation, Plant Sci., 168: 113– 120.

329

Chou, I. T., C. T. Chen, C. H. Kao. 1991. Characteristics of the induction of the

accumulation of proline by abscisic acid and isobutyric acid in detached rice

leaves. Plant Cell Physiol., 32: 269-272.

ChuanJie, C., C. ShanNa, Y. Mei and Z. Qiao. 2003. Effect of salicylic acid on some

enzyme activities related tostress resistance and content of MDA in Vanilla

planifolia . Acta Bot. Yunnanica, 25: 700-704.

Clarke, S. F., P. L. Guy, D. J. Buritt and P. E. Jameson. 2002. Changes in the

activities of antioxidant enzymes in response to virus infection and hormone

treatment. Physiol. Pl., 114: 157–64.

Dat, J. F., D. H. Lopez, C. H. Foyer and I. M. Scott. 2000. Effects of salicylic acid

on oxidative stress and thermo tolerance in tobacco. J. Plant Physiol., 156: 659–

665.

Dawood, M. G., M. S. Sadak and M. Hozayen. 2012. Physiological role of salicylic

acid in improving performance, yield and some biochemical aspects of

sunflower plant grown under newly reclaimed sandy soil. Aust., J. Basic Appl.,

Sci., 6 (4): 82-89.

Debaeke, P., M. Cabelguenne, A. Hilaire and D. Raffaillac. 1998. Crop management

systems for rainfed and irrigated sunflower (Helianthus annuus) in South

Western France. J. Agric. Sci. Cambridge, 131: 171-185.

De-Guang, Y., S. Xiu Ying, Z. Tian Hong and Y. WenChu. 2001. Drought- resistant

effect of exogenous oxygen remover on maize. Beijing Agri. Sci.,

19 (5): 25-27.

Dehkhoda, A., M. Naderidarbaghshahi, A. Rezaei and B. Majdnasiri. 2013. Effect

of water deficiency stress on yield and yield components of sunflower

cultivars in Isfahan. Int. J. Far. All. Sci., (2): 1319-1324.

Del Blanco, I. A., S. Rajaram, W. E. Kronstad and M. P. Reynolds. 2000.

Physiological performance of synthetic hexaploid wheat–derived populations.

Crop Sci., 40: 1257–1263.

330

Demir, A. O., A. T. Goksoy, H. buyukcangaz, Z. M. Turan and E. S. Koksal. 2006.

Deficit irrigation of sunflower in a humid climate. Irrig. Sci., 24: 279-289.

Demirevska, K., D. Zasheva, R. Dimitrov, L. Simova-Stoilov, M. Stamenova and

U. Feller. 2009. Drought stress effects on Rubisco in wheat: changes in the

Rubisco large subunit. Acta Physiol. Plant, 31: 1129-1138.

Desnigh, R. and G. Kanagaraj. 2007. Influence of salinity stress on photosynthesis

and ant oxidative systems in two cotton varieties. Gen. Appl. Plant Physiol.,

33(3-4): 221-234.

Dhindsa, R. S. and R. E. Cleland. 1975. Water stress and protein synthesis. II.

Interaction between water stress, hydrostatic pressure, and abscisic acid on the

pattern of protein synthesis in Avena coleoptiles. Plant Physiol., 55: 782785.

Din, J., S. U. Khan, I. Ali and A. R. Gurmani. 2011. Physiological and agronomic

response of canola varieties to drought stress. J. Ani. P. Sci., 21 (1): 78-82.

Djibril, S., O. K. Mohamed, D. Diaga, D. Diégane, B.F. Abaye, S. Maurice and B.

Alain. 2005. Growth and development of date palm (Phoenix dactylifera L.)

seedlings under drought and salinity stresses. African J. Biotechnol., 4: 968–

972.

Dubios, M. K., J. K. Gilles, P. A. Robers and F. Smith. 1951. Calorimetric

determination of sugar and related substance. Analyt. Chem., 26: 351-356.

Ehdaie, B., G. A. Alloush and J. G. Waines. 2008. Genotypic variation in linear rate of

grain growth and contribution of stem reserves to grain yield in wheat. Field

crops Res., 106, 34–43.

El-Midaoui, M., A. Talouizte, M. Benbella, H. Serieys, Y. Griveau and A. Berville.

2001. Effect of osmotic pressure on germination of sunflower seeds

(Helianthus annuus L.). Helia, 24: 129-134.

El-Shraiy, A. M. and A. M. Hegazi. 2009. Effect of acetylsalicylic acid, Indole-3-

bytric acid and gibberellic acid on plant growth and yield of pea (Pisum

sativum L.) Australian J. Basic and Appli. Sci., 3(4): 3514- 3523.

331

El-Tayeb, M. A. 2005. Response of barley Gains to the interactive effect of salinity

and salicylic acid. Plant Growth Regul., 45:215-225.

El-Tayeb, M. A. and L. A. Naglaa. 2010. Response of Wheat cultivars to drought

and Salicylic acid. American-Eurasian J. of Agro., 3(1): 01-07.

Elwana, M. W. M. and M. A. M. El-Hamahmyb. 2009. Improved productivity and

quality associated with salicylic acid application in greenhouse pepper.

Scientia Horticulturae, 122(4): 521-526.

Emam, Y., A. Shekoofa, F. Salehi and A. H. Jalali. 2010. Water stress effects on two

common bean cultivars with contrasting growth habits. AmericanEurasian J.

Agric. Environ. Sci. 9(5): 495-499.

Enyedi, A. J., N. Yalpani, P. Silverman and I. Raskin.1992. Localization,

conjugation, and function of salicylic acid in tobacco during the hypersensitive

reaction to tobacco mosaic virus. — Proc. Nat. Acad. Sci.

USA. 89: 2480–2484.

Eraslan, F., A. Inal, A. Gunes and M. Alpaslan. 2007. Impact of exogenous salicylic

acid on growth, antioxidant activity and physiology of carrot plants subjected

to combined salinity and boron toxicity. Sci. Hortic., 113:120-128.

Erickson, I. J., D. L. Ketring and J. F. Stone. 1991. Response of internal tissue water

balance of peanut to soil water. Agron. J., 72: 73-80.

Ezzat-Ollah, E., M. R. Shakiba, S. A. Mahboob, A. Hoshang and T. Mahmood. 2007.

Water stress, antioxidant enzyme activity and lipid peroxidation in wheat

seedling. Inter. J. Food Agri. Environ., 5(2007): 149-153.

Fariduddin, Q., S. Hayat, and A. Ahmad. 2003. Salicylic acid influence net

photosynthesis rate, carboxylation efficiency, nitrate reductase activity and

seed yield in Brassica juncea. Photosynthetica 41: 281- 284.

Farooq, M., A. Wahid, D. J. Lee, S. A. Cheema and T. Aziz. 2010. Comparative time

course action of the foliar applied glycinebetaine, salicylic acid, nitrous oxide,

332

brassinosteroids and spermine in improving drought resistance of rice. J.

Agron. Crop Sci. 196: 336-345.

Farooq, M., A. Wahid, N. Kobayashi, D. Fujita and S. M. A. Basra. 2009. Plant

drought stress: effects, mechanisms and management. Agron. Sustain. Dev.,

29: 185–212.

Farooq, M., A. Wahid, S. M. A. Basra and I. U. Din, 2009a. Improving water

relations and gas exchange with brassinosteroids in rice under drought stress.J.

Agron. Crop Sci.,195: 262–269.

Farooq, M., S. M. A. Basra, A. Wahid, N. Ahmad and B. A. Saleem, 2009b.

Improving the drought tolerance in rice (Oryza sativaL.) by exogenous

application of salicylic acid. J. Agron. Crop Sci.,195: 237–246.

Farzane, M. H., R. Monem, S. M. Mirtaheri and S. F. Kashani. 2014. Effect of

salicylic acid on germination and growth seedling of 10 variety barley

(Hordeum Vulgare L.) under drought Stress Int. J. Biosci., 5(1): 445-448.

Fischer, C. and W. Höll. 1991. Food reserves in Scots pine (Pinus sylvestris L.).

Seasonal changes in the carbohydrate and fat reserves of pine needles. Trees,

5: 187-195.

Flexas, J. and H. Medrano. 2002. Drought-inhibition of photosynthesis in C-3 plants:

Stomatal and nonstomatal limitation revisited. Ann. Bot., 89: 1831890.

Foolad, M. R. and G. Y. Lin. 2001. Genetic analysis of cold tolerance during

vegetative growth in tomato, Lycopersicon esculentum Mill. Euphytica. 122:

105–111.

Foyer, C. H. and J. M. Fletcher. 2001. Plant antioxidants: colour me healthy.

Biologist 48, 115–120.

Fu, J. and B. Huang. 2001. Involvement of antioxidants and lipid peroxidation in the

adaptation of two cool-season grasses to localized drought stress. Environ.

Exp. Bot., 45: 105-114.

333

Galle, A., I. Florez-Sarasa, T. Afwa, R. Paepe, V. Flexas and M. Ribas-Carbo. 2010.

Effects of drought stress and subsequent rewatering on photosynthetic and

respiratory in Nicotiana sylvestris wild type and the mitochondrial complex I-

deficient CMSII mutant. J. of Exp. Bot., 61(3): 765-775.

Galle, A., J. Csiszar, T. Irma and L. Erdei, 2002. Changes in water and chlorophyll

fluorescence parameters under osmotic stress in wheat cultivars. Proc. of the

7th Hungarian Congress on Plant Physiology, S2–PO5, 85–6.

Gao, X. P., X. F. Wang, Y. F. Lu, L. Y. Zhang, Y. Y. Shen, Z. Liang and D. P.

Zhang. 2004. Jasmonic acid is involved in the water-stress-induced betaine

accumulation in pearleaves plant.Plant Cell Environ., 27: 497-507.

Gharib, F. A. L. 2006. Effect of salicylic acid on the growth, metabolic activities and

oil content of basil and marjoram. Int. J. Agri. Biol., 4: 485-492.

Giannopolitis, C. N. and S. K. Ries. 1977. Superoxide dismutase I. Occurrence in

higher plants. Plant Physiol., 59: 309-314.

Glass, A. D. M. 1974. Influence of phenolic acids on ion uptake. II. Inhibition of

potassium absorption. J. Exp. Bot., 25: 1104-1113.

Golbashy, M., M. Ebrahimi, S. K. Khorasani and R. Choukan. 2010. Evaluation of

drought tolerance of some corn (Zea mays L.) hybrids in Iran. African J.

Agricultural Res., 5 (19): 2714-2719.

Govt. of Pakistan. 2013. Economic Survey of Pakistan. Pakistan Oilseed

Development Board, Islamabad, Pakistan.

Govt. of Pakistan. 2008. Pakistan Economic Survey 2007-08. Econ. Advisor‘s

Wing, Finance Div., Islamabad, Pakistan.

Gratao, P. L., A. Polle, P. J. Lea and R. A. Azevedo. 2005. Making the life of heavy

metal-stressed plants a little easier. Functional Plant Biol, 32: 481–494.

Gunes, A., A. Inal, M. Alpaslan, F. Eraslan, E. G. Bagci and N. Cicek. 2007.

Salicylic acid induced changes on some physiological parameters

334

symptomatic for oxidative stress and mineral nutrition in maize (Zea mays L.)

grown under salinity. J. Plant Physiol., 164: 728-736.

Gussakovsky, E. E., B. A. Salakhutdinov and Y. Shaha K. 2002. Chiral

macroaggregates of LHCII detected by circularly polarized luminescence in

intact pea leaves are sensitive to drought stress. Funct. Plant Biol., 29: 955-

963.

Haider, S. K. and Saifullah. 2001. Effect of foliar and drench application of acetyl

acetic acid on control of Rhizoctoin asolani and on dry matter production and

portioning of potato. J. Biolo. Scis.11: 1074-1077.

Hamada, A. M. and Al-Hakimi, A. M. A., 2001. Salicylic acid versus salinitydrought

induced stress on wheat seedlings. Rostlinna Vyroba, 47: 444-450.

Hamayun, M., S. A. Khan, Z. K. Shinwari, A. L. Khan, N. Ahmad and I. J. Lee.

2010. Effect of polyethylene glycol induced drought stress on physiohormonal

attributes of soybean. Pakistan J. Bot., 42: 977–986.

Harris, D., R. S. Tripathi and A. Joshi. 2002. On-farm seed priming to improve crop

establishment and yield in dry direct-seeded rice, in: Pandey S., Mortimer M.,

Wade L., Tuong T.P., Lopes K., Hardy B. (Eds.), Direct seeding: Research

Strategies and Opportunities, International Research Institute, Manila,

Philippines, 231-240.

Hasegawa, P. M., R. A Bressan., J. K. Zhu and H. J Bohnert. 2000. Plant cellular

and molecular responses to high salinity. Annu. Rev. Plant Physiol. Plant

Mol. Biol., 51: 463-499.

Hatam, M. and G. Q. Abbasi. 1994. Oilseed Crops. In: Crop production. (Eds): M.

S. Nazir and E. Bashir. 343 p. National Book Foundation, Islamabad, Pakistan.

Hayat, Q., S. Hayat, M. Irfan and A. Ahmad. 2010. Effect of exogenous salicylic

acid under changing environment: a review. Environ. Exp. Bot. 68: 14-25.

Hayat, S., A. Ahmad and M. N. Alyemeni. 2013. SALICYLIC ACID: Plant

335

Growth and Development Springer Science+Business Media Dordrecht. p.

66-77.

Hayat, S., Q. Fariduddin, B. Ali and A. Ahmad. 2005. Effect of salicylic acid on

growth and enzyme activities of wheat seedlings. Acta. Agron. Hung. 53: 433-

437.

Hayat, S., S. A. Hasan, Q. Farriduddun and A. Ahmad., 2008. Growth of tomato

(Lycopersicon esculentum) in response to salicylic acid under water stress. J.

Plant Int., 3 (4): 297-304.

He, Y., Y. Liu, W. Cao, M. Huai, B. Xu, and B. Huang. 2005. Effects of salicylic

acid on heat tolerance associated with antioxidant metabolism in Kentucky

bluegrass. Crop Sci. 45: 988–995.

Heshmat, S., Aldesuquy1, A. Samy, Abo- Hamed, A. Mohme, Abbas, Abeer and H.

Elhakem. 2012. Role of glycine betaine and salicylic acid in improving growth

vigour and physiological aspects of droughted wheat cultivars. J. of Stress

Physiol. and Biochem., 8: 149-171.

Hiscox, J. D. and G. F. Israelstam. 1979. A method for the extraction of chlorophyll

from leaf tissue without maceration. Canadian Journal of Botany.

57: 1332 – 1334.

Hoekstra, F. A and J. Buitink, 2000. Mechanisms of plant dessiccation tolerance.

Trends in Plant Sci. 8: 431-438.

Horvath, E., G. Szalai and T. Janda. 2007. Induction of abiotic stress tolerance by

salicylic acid signalling. J Plant Growth Regul., 26:290-300.

Hsiao and Xu. 2000. Sensitivity of growth of roots versus leaves to water

stress:biophysical analysis and relation to water transport. J. Expt. Bot., 51: 1595-

1616.

Hsieh, T. H., J. T. Lee, Y. Y. Charng and M. T. Chan. 2002. Tomato Plants

ectopically expressing Arabidopsis CBF 1 show enhanced resistance to water

deficit stress. Plant Physiol., 130: 618-626.

336

Huang, R. H., J. H. Liu, Y. M. Lu and R. X. Xia. 2008. Effect of salicylic acid on

the antioxidant system in the pulp of ‗Cara cara‘ navel orange (Citrus sinensis

(L.) Osbeck) at different storage temperatures. Postharvest Biology and

Technology, 47: 168-175.

Hussain, M. 2008. Mitigating the effects of moisture stress in sunflower (Helianthus

annuus L.) by exogenous applications of glycinebetaine and salicylic acid.

(unpublished) Ph.D thesis. Agri. Univ. Fbd.

Hussain, M., M. A. Malik, M. Farooq, M. B. Khan, M. Akram and M. F. Saleem,

2009. Exogenous glycinebetaine and salicylic acid application improves water

relations, allometry and quality of hybrid sunflower under water deficit

conditions. J. Agron. Crop Sci., 195: 98–109.

Hussain, M., M. A. Malik, M. Farooq, M. Y. Ashraf and M. A. Cheema. 2008.

Improving drought tolerance by exogenous application of glycinebetaine and

salicylic acid in sunflower. J. Agron. Crop Sci. 194: 193-199..

Hussein, M. M., L. K. Balbaa and M. S. Gaballah. 2007. Salicylic Acid and Salinity

Effects on Growth of Maize Plants. Res. J. Agric. Biol. Sci., 3 (4): 321-328.

Inal, A. 2002. Growth praline accumulation and ionic relations of tomato

(licopersicum esculentum L.) as influence by NaCl and Na2SO4 salinity.

Turk. J. Bot. 26: 285-290.

Iqbal, M. and M. Ashraf. 2006. Wheat seed priming in relation to salt tolerance:

growth, yield and levels of free salicylic acid and polyamines. Ann. Bot.

Fennici 43: 250-259.

Iqbal, N. 2004. Influence of exogenous glycinebetaine on drought tolerance of

sunflower (Helianthus annuus L.). Ph. D. Thesis, Dept. Bot., Univ. Agric. Fsd.

Pakistan.

337

Iqbal, N., M. Y. Ashraf and M. Ashraf. 2005. Influence of water stress and

exogenous glycinebetaine on sunflower achene weight and oil percentage. Int.

J. Environ. Sci. Tech., 2: 155-160.

Izquierado, N., L. Aguirrezabal, F. Andrade and V. Pereyra. 2002. Night temperature

affects fatty acid composition in sunflower oil depending on the hybrid and the

phenological stage. Field Crop Research, 77: 115-126.

Jaleel, C. A., B. Sankar, P. V. Murali, M. Gomathinayagam, G. M. A. Lakshmanan

and R. Panneerselvam, 2008e. Water deficit stress effects on reactive oxygen

metabolism in Catharanthus roseus;impacts on ajmalicine accumulation.

Colloids Surf. B: Biointerfaces 62: 105–111.


and R. Panneerselvam, 2008c. Alterations in morphological parameters and

photosynthetic pigment responses of Catharanthus roseus under soil water

deficits. Colloids Surf. B: Biointerfaces, 61: 298–303.


and R. Panneerselvam, 2008d. Antioxidant potential and indole alkaloid profile

variations with water deficits along different parts of two varieties of

Catharanthus roseus Colloids Surf. B: Biointerfaces, 62: 312–318.

Jaleel, C. A., P. Manivannan, A.Wahid, M. Farooq, H. J. Al-Juburi, R.

Somasundaram and R. Panneerselvam. 2009. Drought stress in plants: A

review on morphological characteristics and pigments composition. Int. J.

Agric. Biol., 11(1): 100-105.

Jaleel, C. A., R. Gopi, B. Sankar, M. Gomathinayagam and R Panneerselvam,

2008a. Differential responses in water use efficiency in two varieties of

Catharanthus roseus under drought stress. Comp. Rend. Biol., 331: 42–47.

Janda, T., G. Szalai, I. Tari and E. Paldi. 1999. Hydroponic treatment with salicylic

acid decreases the effects of chilling injury in maize (Zea mays L.) plants,

Planta, 208: 175-180.

338

Javed, S., M. Y. Ashraf, S. Mahmood, S. A. Bukhari, M. Meraj and A. Perveen.

2013. Comparative Evaluation of Biochemical Changes in Different

Safflower Varieties (Carthamus tinctorius L.) under Water Deficit. J. Food

Process Technol., 4: 270.

Jensen, H. E. and V. P. Mogensen. 1984. Yield and nutrient contents of spring wheat

subjected to water stress at various growth stages. Acta. Agric. Seandinar. 34:

527–533.

Jiang, R. and N. Huang. 2001. Drought and heat stress injure to two cool season turf

grass in relation to antioxidant metabolism and lipid peroxidation. Crop Sci.,

41: 436-442.

Johansen, C., B. Baldev, J. B. Brouwer, W. Erskine, W. A. Jermyn, L. Li- Juan,

B. A. Malik, A. Ahad Miah and S. N. Silim. 1992. Biotic and abiotic stresses

constraining productivity of cool season food legumesin Asia, Africa and

Oceania. In: Muehlbauer, F.J., W.J. Kaiser (eds.), Expanding the production

and use of cool season food legumes, pp: 75–194. Kluwer academic publishers,

Dordrecht, The Netherlands.

Jonson, B. L. 2003. Dwarf sunflower response to row spacing, stand reduction and

defolation at growth stages. Plant Sci., 83:319-329.

Kabiri, R., F. Nasibi, H. Farahbakhsh. 2014. Effect of exogenous salicylic acid on

some physiological parameters and alleviation of drought stress in Nigella

sativa plant under hydroponic culture. Plant Protect. Sci., 50: 43–51.

Kadioglu, A., N. Saruhan, A. Sağlam, R. Terzi and T. Acet. 2011. Exogenous

salicylic acid alleviates effects of long term drought stress and delays leaf

rolling by inducing antioxidant system. Plant Growth Regul., 64: 27-37.

Kage, H., M. Kochler and H. Stützel. 2004. Root growth and dry matter partitioning

of cauliflower under drought stress condi-tions: measurement and simulation.

European J. Agron., 20: 379–394.

339

Kang, G., C. Wang, G. Sun, Z. Wang. 2003. Salicylic acid changes activities of

H2O2-metabolizing enzymes and increases the chilling tolerance of banana

seedlings. Environ. Exp. Bot., 50: 9-15.

Kang, H. M., M. E. Saltveit. 2002. Chilling tolerance of maize, cucumber and rice

seedling leaves and roots are differentially affected by salicylic acid. Physiol.

Plant, 11(5): 571–576.

Karlidag, H., E. Yildirim and M. Turan. 2009. Salicylic acid ameliorates the adverse

effect of salt stress on strawberry. J. Agric. Sci., 66: 271-278.

Karuppaiah, P., S. Rameshkumar, K. Shah and R. Mari-musthu. 2003. Effect of

antitranspirants on growth, photosyhthetic rate and yield characters brinjal.

Indian J. of Plant Physiol., 8(2): 189-192.

Kauser, R., H. R. Athar and M. Ashraf. 2006. Chlorophyll Fluorescence: A potential

indicator for rapid assessment of water stress tolerance in canola (Brassica

Napus L.). Pak. J. Bot., 38: 1501-1509.

Kawakami, J., K. Iwama and Y. Jitsuyama. 2006. Soil water stress and the growth

and yield of potato plants grown from micro tubers and conventional seed

tubers, Field Crops Res., 95: 89-96.

Kaya, M. D., G. Okçub, M. Ataka, Y. Çıkılıc and O. Kolsarıcıa. 2006. Seed

treatments to overcome salt and drought stress during germination in sunflower

(Helianthus annuus L.). Eur. J. Agron., 24: 291-295.

Keller, F., M. M., Ludlow. 1993. Carbohydrate metabolism in drought-stressed

leaves of pigeonpea (Cajanus cajan). J. Exp. Bot. 44, 1351–1359.

Keyvan, S. 2010. The effect of drought stress on yield, relative water content,

proline, soluble carbohydrates and chlorophyll of bread wheat cultivars. J.

Animal Plant Sci., 8(3): 1051-1060

Khalilvand, B. and M.Yarnia. 2007. The effects of water deficit stress on some of

the physiological characters of sunflower in the different density of planting.

J. of Agric. Sci., 1(2): 37-52.

340

Khan, A. L., M. Hamayun, S. A. Khan, Z. K. Shinwari, M. Kamaran, Sang-Mo

Kang, Jong-Guk Kim and In-Jung Lee. 2011. Pure culture of Metarhizium

anisopliae LHL07 reporgrams soybean to higher growth and mitigates salt

stress. World J. Microb Biotech., 28(4): 1483-94.

Khan, A. L., Z. K. Shinwari, Yoon-Ha Kim, M. Waqas, M. Hamayun, M. Kamran

and In-Jung Lee. 2012. Isolation and detection of Gibberellins and indole acetic

acid from Endophyte Chaetomium globosum LK4 growing drought

stressed plant. Pak. J. Bot., 44(5): 1601-1607.

Khan, S. and J. Khan. 2010. Drought tolerant wheat cultivar (Raj) for rainfed areas

of KPK, Pakistan. Pak. J. Agri. Sci., 47: 355-359.

Khan, S. U., A. Bano, J. U. Din and A. R. Gurmani. 2012. Abscisic acid and

Salicylic acid seed treatment as potent inducer of drought tolerance in wheat

(Triticum aestivum L). Pak. J. Bot., 44: 43-49.

Khan, W., B. Prithiviraj and D. L. Smith. 2003. Photosynthetic responses of corn

and soybean to foliar application of salicylates. J. Plant Physiol., 160: 485492.

Khodary, S. E. A. 2004. Effect of salicylic acid on the growth, photosynthesis and

carbohydrate metabolism in salt stressed maize plants. Int. J. Agri. Biol., 01:

5-8.

Kiani, S. P., P. P. Talia, Maury, P. Grieu, R. Heinz, A. Perrault, V. Nishinakamasu,

Hopp, E. Gentzbittel, I. Paniego and A. Sarrafi. 2007. Genetic analysis of plant

water status and osmotic adjustment in recombinant inbred lines of sunflower

under water treatments. Plant Sci., 172(94): 773-787.

Kmal, A. K., E. A. Amen, and A. M. Al-Said. 2006. Response of snap bean

(Phaseolus vulgaris L.) to some salicylic acid derivatives and selenium under

high temperature stress. J. Agric. Sci., Mansoura Univ., 31(11): 7321-7328.

Korkmaz, A., M. Uzunlu and A. R. Demirkiran. 2007. Treatment with acetyl

salicylic acid protects muskmelon seedlings against drought stress, Acta

Physiol. Plant. 29: 503–508.

341

Krantev, A., R. Yordanova, T. Janda, G. Szalai, L. Popova. 2007. Treatment with

salicylic acid decreases the effect of cadmium on photosynthesis in maize

plants, J. Plant Physiol., (in press).

Kumar, B., D. M. Pandey, C. L. Goswami and S. Jain. 2001. Effect of growth

regulators on photosynthesis, transpiration and related parameters in water

stressed cotton. Biol. Plant., 44: 475-478.

Kumar, S. G., A. M. Reddy and C. Sudhakar. 2003. NaCl effects on proline

metabolism in two high yielding genotypes of mulberry (Morus alba L.) with

contrasting salt tolerance, Plant Sci., 165: 1245–1251.

Kumar, S. V., G. Tan, S. C. Quah and K. Yusoff. 2002. Isolation of microsatellite

markers in mung bean, Vigna radiate. Molecular Ecology Notes: 2: 96-98.

Kumar, B., D. M. Pandey, C. L. Goswami and S. Jain. 2001. Effect of growth

regulators on photosynthesis, transpiration and related parameters in water

stressed cotton. Biol. Plant., 44: 475-478.

Kusaka, M., M. Ohta and T. Fujimura. 2005. Contribution of inorganic components

to osmotic adjustment and leaf folding for drought tolerance in pearl millet.

Physiol. Plant. 125: 474–489.

Lafitte, H. R., G. Yongsheng, S. Yan and Z. K. Li. 2007. Whole plant responses, key

processes and adaptation to drought stress: the case of rice. J. Exp. Bot., 58:

169–175.

Laghrab, M. H., F. Nouri and H. Z. Abianeh. 2003. Effect of the reduction of drought

stress using supplementary irrigation for sunflower in dry farming conditions.

In Agronomy and Horticulture. 59: 81-86.

Lazaridou, M., A. Kirilov, B. Noitsakis, N. Todorov and I. Katerov. 2003. The effect

of water deficit on yield and water use efficiency of lucerne. Optimal forage

systems for animal production and the environment. Proceedings of

342

the 12th Symposium of the Europea Grassland Federation, Pleven, Bulgaria,

26–28 May 2003.

Lazcano-Ferrat, I. and C. J. Lovatt. 1999. Relationship between relative water

content, nitrogen pools, and growth of Phaseolus vulgaris L. and P. acutifolius

Gray during water deficit. Crop Sci., 39: 467-475.

Lenzi, A., M. Fambrini, S. Barotti, C. Pugliesi and P. Vernieri. 1995. Seed

germination and seedling growth in a wilty mutant of sunflower (Helianthus

annuus L.): effect of absisic acid and osmotic potential. Environ. Exp. Bot.,

35:427-434.

Leport, L., N. C. Turner, R. J. French, M. D. Barr, R. Duda and S. L. Davies. 2006.

Physiological responses of chickpea genotypes to terminal drought in a

mediterranean-type environment. Eur. J. Agron., 11: 279-291.

Lian, B., X. Zhou, M. Miransari and D. L. Smith. 2000. Effects of salicylic acid on

the development and root nodulation of soybean seedlings. J. Agron. Crop Sci.,

185 (3): 187-192.

Lindberg, S. and M. Greger. 2002. Plant genotypic differences under metal deficient

and enriched conditions. In: Prasad, M.N.V., Kazimierz, S. (Eds.),

Physiology and Biochemistry of Metal Toxicity and Tolerance in Plants.

Kluwer Academic Publishers, Dordrecht, The Netherlands, pp. 357-393.

Lio, H. S., M. L. Feng and X. Hao. 2004. Deficiency of water can enhance root

respiration rate of drought-sensitive but not drought tolerant spring wheat.

Agric. Water Manage. 64: 41-48.

Liu Xini, Li Yun and Z. Shuqiu. 2000. Effect of salicylic acid on growth and content

of IAA and IPA of broad bean seedlings. Plant communications. 36

(6): 512-514.

Lopez, C. M. L., H. Takahashi and S. Yamazaki. 2002. Plantwater relations of

kidney bean plants treated with NaCl and foliarly applied glycinebetaine. J.

Agron. Crop Sci., 188: 73–80.

343

Ludlow, M. M. and R. C. Muchow. 1990. A critical evaluation of traits for improving

crop yields in water-limited environments, Adv. Agron. 43, 107– 153.

Ludlow, M. M. 1985. Photosynthesis and dry matter production in C3 and C4 pasture

plants, with special emphasis on tropical C3 legumes and C4 grasses. Aust. J.

Agr. Res. 12: 557-572.

Mafakheri, A., B. Siosemardeh, P. C. Bahramnejad and Y. S. Struik. 2010. Effect of

drought stress on yield, proline and chlorophyll contents in three chickpea

cultivars. AJCS. 4 (8): 580-585.

Maibangsa, S., M. Thangaraj and R. Stephen. 2001. Effect of brassinosteroid and

salicylic acid on rice (Oryza sativa L.). Ind. J. Agric. Res., 34: 258-260.

Maibangsa, S., M. Thangaraj and R. Stephen. 2000. Effect of brassinosteriod and

salicylic acid on rice grown under low irradiance condition. Ind. J. Agric. Res.,

34: 258-260.

Makoto, T. B., B. F. Carver, R. C. Johnson and E. L. Smith. 1990. Relationship

between water content and grain yield during reproductive period in winter

wheat. Euphytica, 49: 255-262.

Mamenko, T. P. and O. A. Iaroshenko. 2009. Effect of salicylic acid on water

potential, ethylene secretion and activity of antioxidative processes in the

winter wheat leaves under drought conditions. Ukr. Biokhim. Zh., 81(2): 117-

124.

Manikavelu, A., N. Nadarajan, S. K. Ganesh, R. P. Gnanamalar and R. C. Babu.

2006. Drought tolerance in rice: morphological and molecular genetic

consideration. Plant Growth Regul., 50: 121-138.

Manivannan, P., A. Jaleel, B. Sankar, A. Kishorekumar, R. Somasundaram, G. M.

A. Lakshmanan and R. Panneerselvam. 2007. Growth biochemical

modifications and proline metabolism in Helianthus annuus L. as induced by

drought stress, Colloids Surf. B: Biointerfaces, 59: 141–149.

344

Manivannan, P., C. A. Jaleel, A. Kishorekumar, B. Sankar, R. Somasundaram, R.

Sridharan and R. Panneerselvam. 2007a. Propiconazole induced changes in

antioxidant metabolism and drought stress amelioration in Vigna unguiculata

(L.) Walp, Colloids Surf. B: Biointerfaces, 57: 69–74.

Manivannan, P., C. A. Jaleel, B. Sankar, A. Kishorekumar, R. Somasundaram, G.

M. A. Lakshmanan and R. Panneerselvam. 2007b. Growth biochemical

modifications and proline metabolism in Helianthus annuus L. as induced by

drought stress, Colloids Surf. B: Biointerfaces, 59: 141–149.

Marcińska, I., I. Czyczyło-Mysza, E. Skrzypek, M. Grzesiak, F. Janowiak F. and M.

Filek. 2013. Alleviation of osmotic stress effects by exogenous application of

salicylic or abscisic acid on wheat seedlings. Int. J. Mol. Sci. 14: 13171-13193.

Martinez, J. P., H. Silva, J. F. Ledent and M. Pinto. 2007. Effect of drought stress on

the osmotic adjustment, cell wall elasticity and cell volume of six cultivars of

common beans (Phaseolus vulgaris L.), Euro. J. Agron., 26: 30-38.

Martin-Mex, R., A. Nexticapan-Garces and A. Larque-Saavedra. 2005b. Effect of

salicylic acid in sex expression in Carica papaya L. In: 10th International

Symposium on Plant Bioregulators in Fruit Production, June 26–30, Saltillo,

Coahuila, Mexico, p. 113.

Martin-Mex, R., E. Villanueva-Couoh, T. Herrera-Campos and A. LarqueSaavedra.

2005a. Positive effect of salicylates on the flowering of African violet. Sci.

Hortic., 103: 499-502.

Mathur, N. and A. Vyas. 2007. Physiological effect of some bioregulators on

vegetative growth, yield and chemical constituents of pearl millet Pennisetum

typhoides (Burm, Stapf. and Hubb). Int. J. Agric. Res. 2(3): 238-245.

Mazahery-Laghab, H., F. Nouri and H. Z. Abianeh. 2003. Effects of the reduction of

drought stress using supplementary irrigation for sunflower (Helianthus

345

annuus L.) in dry farming conditions. Pajouhesh va Sazandegi Agron. Hort.,

59: 81-86.

Mckeon, T. A. and P. K. Stumpf. 1982. Purification and characterization of the

stearoil- acyl carrier protein desaturase and the acyl-acyl protein thioesterase

from mature seeds of sunflower. The J. of Biochem., 257(20): 12141–12147.

Meek, C., D. M. Oosterhuis and J. Gorham. 2003. Effects of foliar-applied and

endogenous levels of glycine betaine on cotton growth and yield. J. Crop

Management. p.9.

Meo, A. A. 1999. Influence of fertilizer and water stress on leaf area of sunflower

(Helianthus Annuus L.). Pak. J. Agri. Sci., 36: 1-2.

Metwally, A., I. Finkermeier, M. Georgi and K. J. Dietz. 2003. Salicylic acid

alleviates the cadmium toxicity in barley seedlings, Plant Physiol., 132:

272281.

Mittler, R. 2002. Oxidative stress, antioxidants and stress tolerance. Trends in Plant

Science, 7: 405–410.

Moayedi, A. A., A. N. Boyce and S. S. Barakbah. 2009. Study on osmotic stress

tolerance in promising durum wheat genotypes using drought stress indices.

Res. J. Agri. Bio. Sci., 5 (5): 603-607.

Mohammad, M. El., M. Benbella and A. Talouizete. 2002. Effect of sodium chloride

on sunflower (Helianthus annuus L.) seed germination. Helia, 37:

51-58.

Mohammadian, R., M. Moghaddam, H. Rahimian and S.Y. Sadeghian. 2005. Effect

of early season drought stress on growth characteristics of sugar beet

genotypes. Turkisk J. Bot., 29: 357–368.

Mohammadkhani, N. and R. Heidari. 2008. Drought-induced accumulation of

soluble sugars and proline in two maize varieties. World Appl. Sci. J., 33: 448-

453.

346

Moller, I. M., P. E. Jensen and A. Hansson. 2007. Oxidative modifications to cellular

components in plants. Annual Review of Plant Biology, 58: 459–481.

Monakhova, O. F. and I. I. Chernyadev. 2002. Protective role of kartolin-4 in wheat

plants exposed to soil drought. Appl. Biochem. Micro., 38: 373-380.

Morgan, J. M. 1984. Osmoregulation and water stress in higher plants. Annu. Rev.

Plant Physiol., 35: 299–319.

Moussa, H. R. and S. M. El-Gamel. 2010. Effect of salicylic acid pre-treatment on

cadmium toxicity in wheat. Biol Plant., 54: 315–320.

Mujtaba, S. M. and S. M. Alam. 2002. Drought phenomenon and crop

growth.http://www.Pakistan economist .com/issue/ issue13/I and e4.htm.

Munné-Bosch, S. and J. Penuelas. 2003. Photo and antioxidative protection, and a

role for salicylic acid during drought and recovery in field grown Phillyrea

angustifolia plants, Planta. 217: 758–766.

Natali, L., T. Giordani and A. Cavallini. 2003. Sequence variability of a dehydrin

gene of a Helianthus annuus. Theor. Appl. Genet., 106(5): 811-818.

Nayyar, H. and D. Gupta. 2006. Differential sensitivity of C3 and C4 plants to water

deficit stress: association with oxidative stress and antioxidants. Environ. Exp.

Bot. 58: 106-113.

Nayyar, H. and D. P. Walia. 2003. Water stress induced proline accumulation in

contrasting wheat genotypes as affected by calcium and abscisic acid. Biol.

Plant., 46: 275-279.

Nazariyan, G., M. Mehrpooyanb and M. Khiyavic. 2009. Study of effects of drought

stress on yield and yield components of four sunflower cultivars in Zanjan, Iran

Plant Ecophysiol. 3: 135-139.

Nazarli, H., M. R. Zardashti, R. Darvishzadeh and M. Mohammad. 2011. Change in

activity of antioxidative enzymes in young leaves of sunflower (Helianthus

347

annuus L.) by application of super absorbent synthetic polymers under drought

stress condition. AJCS., 5 (11): 1334-1338.

Németh, M., E. Horváth T. Janda, E. Páldi and G. Szalai. 2002. Exogenous salicylic

acid increases polyamine content but may decrease drought tolerance in maize.

Plant Sci., 162: 569–574.

Niakan, M., A. Jahanbani and M. Ghorbanli. 2010. Spraying effect of salicylate

different concentrations on growth parameters, amount of photosynthetic

pigments anthocyanin, flavonoids and solution sugars of Coriandrum sativum

L. J. Plant Sci. Researches, 18(2): 10-18.

Nielsen, D. C. and N. O. Nelson. 1998. Black bean sensitivity to water stress at

various growth stages. Crop Sci. 38: 422-427.

Nizami, H. R., Z. Khazaei, B. Rezazadeh and A. Hosseini. 2008. Effects of drought

stress and defoliation on sunflower (Helianthus annuus) in controlled

conditions Desert, 12: 99-104.

Nobel, P. S. 1991. Physicochem. and Environ. Plant Physio., Academic Press, San

Diego.

Noctor, G., C. H. Foyer. 1998. Ascorbate and glutathione keeping active oxygen

under control. Annu. Rev. Plant Physiol. Plant Mol. Biol., 49: 249-279.

Nonami, H. 1998. Plant water relations and control of cell elongation at low water

potentials. J. Plant Res., 111: 373–382.

Noreen, S. and M. Ashraf, 2010. Modulation of salt (NaCl)-induced effects on oil

composition and fatty acid profile of sunflower (Helianthus annuus L.) by

exogenous application of salicylic acid. J. Sci. of Food and Agric., 90(15):

2608-2616.

Okcu, G. M., D. Kaya and M. Atak. 2005. Effects of salt and drought stresses on

germination and seedling growth of pea (Pisum sativum L.) Turk. J. Agron.,

29: 237–242.

348

Okoko, N. E. K., M. J. Mahasi, N. Kidula, M. Ojowi and F. Makini. 2008.

Participatory sunflower production, technology dissemination and value

addition in South West Kenya. Afric. J. Agric. Res. 3(6): 396-399.

Pal, M., E. Horvath, T. Janda, E. Paldi, and G. Szalai. 2005. Cadmium stimulates the

accumulation of salicylic acid and its putative precursors in maize (Zea mays)

plants. Physiol. Plantarum, 125: 356-364

Parida, A. K., S. Vipin , Dagaonkar, S. Manoj, Phalak, G. V. Umalkar, P. Laxman

and P. Aurangabadkar. 2007. Alterations in photosynthetic pigments, protein

and osmotic components in cotton genotypes subjected to short-term drought

stress followed by recovery Plant Biotechnol Rep. 1: 37–48.

Patakas, A. and B. Noitsakis. 2001. Leaf age effects on solute accumulation in water-

stressed grapevines. Plant Physiol. 158: 63-69.

Petcu, E., A. Arsintescu and D. Stanciu. 2003. Studies regarding the hydric

stresseffect on sunflower plants. Analele Institutuli de Cereale si Plante

Technice, Fundulea., 70: 347-356.

Petropoulos, S. A., D. Daferera, M. G. Polissiou and H. C. Passam. 2008. The effect

of water deficit stress on the growth, yield and composition of essential oils of

parsley. Sci. Hort., 115: 393–397.

Philipson, J. J. 2003. Optimal conditions for inducing coning of container-grown

(Picea sitchensis) grafts. The effects of applying different quantities of GA

4/7, timing and duration of heat and drought treatment, and girdling. Forest.

Ecol. Manag., 53: 39–52.

Pinhero, R. G., M. V. Rao, G. Palyath, D. P. Murr and R. A. Fletcher. 2001. Changes

in the activities of antioxidant enzymes and their relationship to genetic and

paclobutrazol-induced chilling tolerance of maize seedlings. Plant Physiol.,

114: 695-704.

349

Pirjol-Sovulescu, L., C. I. Milica and V. Vranceanu. 1974. A study of drought

resistance during different growth phase in sunflower. Olegineux Abst., 29:

869 (Pl. Br. Abst., 45: 2222 1975).

Plaut, Z. 2003. Plant exposure to water stress during specific growth stages.

Encycl. of Water Sci., p. 673– 675.

Poór, P., K. Gomes, ç. Szepesi, F. Horvath, L. M. Simon and I. Tari. 2011.

Salicylic acid treatment via the rooting medium interferes with the stomatal

response, CO2 fixation rate and carbohydrate metabolism in tomato and decreases

the harmful effects of subsequent salt stress. Plant Biology, 13:105114.

Popova, L., E. Ananieva, V. Haristova, K. Christov, K. Georgiera, E. Alexieva and

Z. H. Stoinov. 2003. Salicylic acid and methyl jasmonate-induced protection

on photosynthesis to paraquat oxidative stress. Bulg. J. Plant Physiol. 133152.

Poroyko, V., L. G. Hejlek, W. G. Spollen, G. K. Springer, H. T. Nguyen, R. E. Sharp

and H. J. Bohnert. 2005. The maize root transcriptome by serial analysis of

gene expression. Plant Physiol., 138: 1700–1710.

Rajaram, S., H. J. Braun and M. Van. Ginkel. 1996. CIMMYT's approach to breed

for drought tolerance. Euphytica., 92 (1/2): 147-153.

Ramakrishna, Y. S., G. G. S. N. Rao, R. Kesava and P. K. Vijaya. 2000. Weather

resources management. In: Natural resource management for agricultural

production India. (Eds. JSP Yadav and GB. Singh): 247-370.

Ramanjulu, S. and D. Bartels. 2002. Drought and dessication-induced modulation of

gene expression in plants. Plant Cell Environ., 25: 141– 151.

Rao, S. R., A. Qayyum, A. Razzaq, M. Ahmad, I. Mahmood and A. Sher. 2012.

Application of salicylic acid and l-tryptophan in drought tolerance of maize. J.

Anim. Plant Sci., 22 (3): 768-772.

Raskin I., 1992. Role of salicylic acid in plants. Ann. Rev. Plant Physiol. Plant

Mol. Biol. 43: 439-463.

350

Raskin, I., H. Skubatz, W. Tang and B. Meeuse. 1990. Salicylic acid levels in

thermogenic and non-thermogenic plants. Ann Bot. 66: 369–373.

Rathore, P. S. 2001. Techniques and managements of field crop

production.Publisher. Jodhpur: Agrobios India.

Reddy, A. R., K. V. Chaitanya and M. Vivekanandan. 2004. Drought-induced

responses of photosynthesis and antioxidant metabolism in higher plants. J.

Plant Physiol., 161: 1189-1202.

Reddy, G. K. M., K. S. Dangi, S. S. Kumar and A. Reddy. 2003. Effect of moisture

stress on seed yield and quality in sunflower. Ind. J. oilseeds Research, 20(2):

282-283.

Reynolds, M. P., A. M. Kazi, and M. Sawkins. 2005. Prospects for utilizing plant

adaptive mechanisms to improve wheat and other crops in drought and salinity

prone environments. Annals. Appl. Biol., 146: 239-259.

Sadeghipour, O. and P. Aghaei. 2012. Impact of exogenous salicylic acid application

on some traits of common bean (Phaseolus vulgaris l.) under water stress

conditions. Int. J. Agric. Crop. Sci., 11: 685-690.

Sadras, V. O., F. J. Villalobes and E. Fereres. 1993. Leaf expansion in field-grown

sunflower in response to soil and leaf water status. Agron. J., 85: 564-570.

Saei, A., Z. Zamani, A. Talaie and R. Fatahi. 2006. Influence of drought stress

periods on olive (Olea europaea L. cv. Zard) leaves stomata. Int J Agric Boil

4: 430-433.

Saensee, K., T. Machikowa and N. Muangsan. 2012. Comparative Performance of

Sunflower Synthetic Varieties under Drought Stress. Int. J. Agric. Biol., 14(6):

929-934.

Sajjan, A. S., V. P. Badanur and G. M. Sajjanar. 1999. Effect of external water

potential on seed germination, seedling growth and vigor index in some

genotypes of sunflower. In: Proc. Symp. Recent Advances in Management of

351

Arid Ecosystem, (Eds.): S.A. Faroda, N.L. Joshi, S. Kathju and A. Kar. pp.

215-218.

Sakamoto, A. and N. Murata. 2002. The role of glycine betaine in the protection of

plants from stress: clues from transgenic plants. Plant, Cell Environ., 25:

163171.

Sakhabutdinova, A. R., D. R. Fatkhutdinova, M. V. Bezrukova and F. M. Shakirova.

2003. Salicylic acid prevents the damaging action of stress factors on wheat

plants. Bulg. J. Plant Physiol., Special Issue. pp: 314-319.

Samarah, N. H., A. M. Alqudah, J. A. Amayreh and G. M. Mcandrews. 2009. The

effect of late-terminal drought stress on yield components of four barley

cultivars. J. Agron. Crop. Sci., 195: 427-441.

Sanaa, A. M. Z., S. I. Ibrahim and H. A. M. S. Eldeen. 2001. The effect of

naphthalene acetic acid (NAA), salicylic acid on growth, fruit setting, yield and

some correlated components in dry bean. Ann. Agric. Sci., Cairo. 46(2): 451-

463.

Sankar, B., C. A. Jaleel, P. Manivannan, A. Kishorekumar, R. Somasundram and R.

Panneerselvam. 2007. Drought-induced biochemical modifications and proline

metabolism in Abelmoschus esculentus (L.) Moench. Acta Bot. Croat. 66:43-

56.

Sankar, B., C. A. Jaleel, P. Manivannan, A. Kishorekumar, R. Somasundaram and

R. Panneerselvam. 2007. Effect of paclobutrazol on water stress amelioration

through antioxidants and free radical scavenging enzymes in Arachis hypogaea

L. Colloids Surf. B: Biointerfaces, 60: 229–235.

Sankar, B., C. A. Jaleel, P. Manivannan, A. Kishorekumar, R. Somasundaram and

R. Panneerselvam, 2008. Relative efficacy of water use in five varieties of

Abelmoschus esculentus(L.) Moench under water-limited conditions. Colloids Surf.

B: Biointerfaces, 62: 125–129.

352

Sarangthem, K. and T. H. N. Singh. 2003. Efficacy of salicylic acid on growth,

nitrogen metabolism and flowering of Phaseolus vulgaris. Crop Res., 26: 355-

360.

Satyabrata, M., M. R. Hedge and S. B. Chattopodhay. 1988. Hand Book of annual

oilseed crops. Oxford IBH Pub. Co. (Pvt.) Ltd. New Delhi p. 176.

Sayyari, M., M. Ghavami, F. Ghanbari and S. Kordi. 2013. Assessment of salicylic

acid impacts on growth rate and some physiological parameters of lettuce

plants under drought stress conditions, Int. J. of Agri. and Crop Sci., 5(17):

1951-1957.

Scholander, P., H. Hammel, E. Bradstreet and E. Hemmingsen. 1965. Sap pressure

in vascular plants. Science, 148: 339-345.

Schonfeld, M. A., R. C. Johnson, B. F. Carver and D. W. Mornhinweg. 1988.

Water relations in winter wheat as drought resistance indicator. Crop Sci., 28:

526-531.

Schuppler, U., P. H. He, P.C.L. John and R. Munns. 1998. Effects of water stress on

the cell division and cell-division-cycle-2-like cell-cyclekinase activity in

wheat leaves. Plant Physiol., 117: 667–678.

Senaratna, T., D. Touchell, E. Bunn and K. Dixon. 2000. Acetyl salicylic acid

(aspirin) and salicylic acid induce multiple stress tolerance in bean and tomato

plants, Plant Growth Regul., 30: 157-161.

Sgherri, C. L. M. and F. Navari-Izzo. 1995. Sunflower seedlings subjected to

increasing water deficit stress: oxidative stress and defence mechanisms.

Physiologia Plantarum, 93: 25-30.

Sgherri, C. L. M., Pinzino, C. and Navari-Izzo, F. 1996. Sunflower seedlings

subjected to increasing water stress by water deficit: Changes in O2- production

related to the composition of thylakoid membranes. Physiologia Plantarum, 96:

446-452.

353

Shah, A. N., H. Shah and N. Akmal. 2005. Sunflower area and production variability

in Pakistan: Opportunities and Constraints. HELIA. 28 (43): 165178..

Shakeel, S. and S. Mansoor. 2012. Salicylic acid prevents damaging action of salt in

mung bean (Vigna radiate L.) seedlings. Pak. J. Bot., 44 (2): 559-562.

Shakirova, F. M., A. R. Sakhabutdinova, M. V. Bezrukova, R. A. Fatkhutdinova and

D. R. Fatkhutdinova. 2003. Changes in the hormonal status of wheat seedlings

induced by salicylic acid and salinity, Plant Sci. 164: 317–322.

Shao, H. B., L. Y. Chu, C. A. Jaleel, P. Manivannan, R. Panneerselvam, M. A. Shao.

2009. Understanding water deficit stress-induced changes in the basic

metabolism of higher plants-biotechnologically and sustainably improving

agriculture and the ecoenvironment in arid regions of the globe. Crit. Rev.

Biotechnol., 29: 131-151.

Shao, H. B., L. Y. Chu, M. A. Shao, C. Abdul Jaleel and M. Hong-Mei. 2008. Higher

plant antioxidants and redox signaling under environmental stresses.

Comp.Rend. Biol. 331: 433–441.

Shao, H. B., L. Y. Chuc, G. Wu, J. H. Zhang, Z. H. Lua and Y.C. Hug. 2007. Changes

of some anti-oxidative physiological indices under soil water deficits among

10 wheat (TriticumaestivumL.) genotypes at tillering stage.

Colloids Surf. B: Biointerfaces, 54: 143–149.

Sharma, K. and S. Kaur. 2003. Effect of salicylic acid, caffeic acid and light intensity

on yield and yield contributing parameters in soybean. Environ. Ecol., 21 (2):

332-335.

Sharp, R. E., W. K. Silk and T. C. Hsiao. 1988. Growth of maize primary root at low

water potential. I. Spatial distribution of expansive growth. Plant Physiol., 87:

50–57.

Siddiqi, E. H. and M. Ashraf. 2008. Can leaf water relation parameters be used as

selection criteria for salt in Safflower (Carthamus Tinctorius L.) Pak. J. Bot.,

40(1): 221-228.

354

Siddique, M. R. B., A. Hamid and M. S. Islam. 2000. Drought stress effects on water

relations of wheat. Bot. Bull. Acad. Sin. 41: 35-39.

Singh, B. and K. Usha. 2003. Salicylic acid induced physiological and biochemical

changes in wheat seedlings under water stress. Plant Growth Reg., 39: 137141.

Sivakumar, R., M. K. Kalarani, V. Mallika and K. B. Sujattha. 2001. Effect of growth

regulators on biochemical attributes, grain yield and quality in pearl millet

(Pennisetum glaucum L. R. Br.). Madras Agric. J., 88: 256-259.

Sivakumar. D., N. Hewarathnagamagae, K. Wilson, R. S. Wijetatnam and R. L. C.

Wijesundara. 2002. Effect of ammonium carbonate and sodium bicarbonate on

anthracnose o f papaya. Phytoparasitica 30(5): 486-492.

Smirnoff, N. 1993. The role of active oxygen in the response of plants to water deficit

and desiccation. New Phytol., 125:27-58.

Souza, R. P., E. C. Machado, J. A. B. Silva, A. M. M. A. Lagoa and J. A. G. Silveira.

2004. Photosynthetic gas exchanges in cowpea (Vigna unguiculata) during

water stress and recovery. Environ. Exp. Bot., 51: 45-56.

Specht, J. E., K. Chase, M. Macrander, G. L. Graef, J. Chung, J. P. Markwell, M.

Germann, J. H. Orf and K. G. Lark. 2001. Soybean response to water. A QTL

analysis of drought tolerance. Crop Sci., 41: 493–509.

Steel, R. G. D., J. H. Torrie and D. A. Deekey. 1997. Principles and procedures of

statistics A Biometrical Approach. 3rd ed. McGraw Hill Book Co. Inc. New

York. pp. 400-428.

Stefanoudaki, M., M. Williams, C. Chartzoulakis and J. Harwood. 2009. Orchard

irrigation with saline water. J. Agric. Food Chem., 57: 1421-1425

Stymme, S. and L. A. Appelqvist. 1980. The biosynthesis of linoleate and linolenate

in homogenate from developing soya been cotyledons. Plant Sci. Lett., 17: 287

– 297.

355

Szalai, G., S. Horgosi, V. Soós, I. Majláth, E. Balázs and T. Janda. 2010. Salicylic

acid treatment of pea seeds induces its de novo synthesis. J. Plant Physiol., 168:

213-219.

Szalai, G., I. Tari, T. Janda, A. Pestenacz and E. Paldi. 2000. Effects of cold

acclimation and salicylic acid on changes in ACC and MACC contents in

maize during chilling.Biologia Plantarium, 43: 637-640.

Szepesi, A. 2006. Salicylic acid improves the acclimation of Lycopersicon

esculentum Mill. L. to high salinity by approximating itssalt stress response to

that of the wild species L. Pennellii. Acta. Biologica. Szegediensis. 50(3-4):

177.

Tahir, M. H. N. and S. S. Mehdi. 2001. Evaluation of open pollinated sunflower

(Helianthus annuus L.) populations under water stress and normal conditions.

Int. J. Agric. Biol., 3: 236-238.

Tahir, M. H. N., M. Imran and M. K. Hussain. 2002. Evaluation of sunflower

(Helianthus annuus L.) inbred lines for drought tolerance. Int. J. Agric. Biol.,

3: 398–400.

Taiz, L. and E. Zeiger. 2006. Plant Physiology, 4th Ed., Sinauer Associates Inc.

Publishers, Massachusetts.

Talieva, M. N. and V. V. Kondrat Eva. 2002. Influence of exogenous salicylic acid

on the level of phytohormones in tissues of Phlox paniculata and Phlox setacea

leaves with special reference to resistance against the powdery mildew

causative agent Erysiphe cichoracearum DC. f. phlogis Jacz. Biol Bull 29: 551-

554.

Tambussi, E. A., C. G. Bartoli, J. Bettran, J. J. Guiamet and J. C. Araus. 2000.

Oxidative damage to thylakoids proteins in water stressed leaves of wheat

(Triticum aestivum L.) Physiol. Plant., 108: 398-404.

356

Tamura, T., K. Hara, Y. Yamaguchi, N. Koizumi and H. Sano. 2003.Osmotic stress

tolerance of transgenic tobacco expressing a gene encoding a membranelocated

receptor- like protein from tobacco plants. Plant Physiol., 131:454–

62.

Tas, S. and B. Tas. 2007. Some physiological responses of drought stress in wheat

genotypes with different ploidity in Turkiye. World J. Agricultural Sci., 3: 178-

183.

Tasgin, E., O. Atici, B. Nulbantoglu and L. P. Popova. 2006. Effects of salicylic acid

and cold treatment on protein levels and on the activities of antioxidant

enzymes in the apoplast of winter wheat leaves. Phytochemistry, 67: 710-

715.

Tayebe, A. and P. Hassan. 2010. Antioxidant enzyme changes in response to drought

stress in ten cultivars of oilseed rape (Brassica napus L). J. Genet.

Plant Breed., 46 (1): 27–34.

Thomas, H. and C. J. Howarth. 2000. Five ways to stay green. J. Exp. Bot., 51: 329–

337.

Treshow, M. 1970. Disorders associated with adverse water relations. Environment

and plant response. McGraw-Hill book company, U.S.A., 153-174.

Tripathy, J. N., J. Zhang, S. Robin, T. T. Nguyen and H. T. Nguyen. 2000. QTLs

for cell-membrane stability mapped in rice (Oryza sativa L.) under drought

stress, Theor. Appl. Genet. 100: 1197–1202.

Ullah, F., A. Bano and A. Nosheen. 2012. Effects of plant growth regulators on

growth and oil quality of canola (Brassica napus L.) under drought stress. Pak.

J. Bot., 44 (6): 1873-1880.

Uzunova, A. N. and L. P. Popova. 2000. Effect of salicylic acid on leaf anatomy and

chloroplast ultrastructure of barley plants. Photosynthetica, 38: 243–250.

Uzunova, K. and Z. Zlatev. 2013. Drought-induced changes in photosynthesis of

young cowpea plants. Agricultural science and technology. 5(1): 32-34.

357

Van Ginkel, M., D. S. Calhoun, G. Gebeyehu, A. Miranda, C. Tianyou, R. Pargas

Lara, R. M. Trethowan, K. Sayre, L. Crossa and S. Rajaram. 1998. Plant traits

related to yield of wheat in early, late, or continuous drought conditions.

Euphytica, 100: 109–121.

Vanacker, H., H. Lu, D. N. Rate and J. T. Greenberg. 2001. A role for salicylic acid

and NPR1 in regulating cell growth in Arabidopsis. Plant J., 28:209-216.

Vanaja, M., S. K. Yadav, G. Archana, N. Jyothi Lakshmi, P. R. Ram Reddy, P.

Vagheera, S. K. A. Razak, M. Maheswari and B. Venkateswarlu. 2011.

Response of C4 (maize) and C3 (sunflower) crop plants to drought stress and

enhanced carbon dioxide concentration. Plant Soil Environ., 57: 207 215.

Verberne, M. C., N. Brouwer, F. Delbianco, H. J. Linthorst, J. F. Bol and R.

Verpoorte. 2002. Method for the extraction of the volatile compound salicylic

acid from tobacco leaf material. Phytochem. Anal., 13: 45-50.

Vrânceanu, A. V. 2000. Floarea-soarelui hibrida Editura Ceres. Bucharest,

Romania.

Vrânceanu, A. V., D. Stanciu, M. Stanciu, M. Pacureanu-Joita, I. Sorega, and I.

Mantu. 2005. Jupiter - a new Orobanche resistant sunflower hybrid.

Romanian Agricultural Research, 22: 19-22.

Wahid, A and E. Rasul. 2005. Photosynthesis in leaf, stem, flower and fruit, in:

Pessarakli M. (Ed.), Handbook of Photosynthesis, 2nd ed., CRC Press, Florida,

pp. 479–497.

Wang, L. J., L. Fan, W. Loescher, W. Duan, G. J. Liu and J. S. Cheng. 2010. Salicylic

acid alleviates decreases in photosynthesis under heat stress and accelerates

recovery in grapevine leaves. BMC Plant Biol., 10: 34.

Wang, Z. Y., F. M. Li, Y. C. Xiong and B. C. Xu. 2008. Soil-water threshold range

of chemical signals and drought tolerance was mediated by ROS homeostasis

358

in winter wheat during progressive soil drying, J. Plant Growth Regul., 27:

309-319.

Waseem, M., H. R. Athar and M. Ashraf. 2006. Effect of salicylic acid applied

through rooting medium on drought tolerance of wheat. Pak J. Bot. 38 (4):

1127-1136.

Webber, M., J. Barnett., B. Finlayson and M. Wang. 2006. Pricing China‘s

Irrigation Water. Working Paper, School of Anthropology, Geography and

Environmental Studies, The University of Melbourne, Victoria, Australia.

Wu, Q. S., R. X. Xia and Y. N. Zou. 2008. Improved soil structure and citrus growth

after inoculation with three arbuscular mycorrhizal fungi under drought stress.

European J. Soil Biol., 44: 122–128.

Wullschleger, S. D., T. M. Yin, S. P. DiFazio, T. J. Tschaplinski, L. E. Gunter, M.

F. Davis and G. A.Tuskan. 2005. Phenotypic variation in growth and biomass

distribution for two advanced-generation pedigrees of hybrid poplar.

Canadian J. for. Res., 35: 1779–1789.

Xu, P. L., Y. K. Guo, J. G. Bai, L. Shang and X. J. Wang. 2008. Effects of longterm

chilling on ultrastructure and antioxidant activity in leaves of two cucumber

cultivars under low light. Physiologia Plantarum, 132: 467–478.

Yadav, S. K., N. J. Lakshmi, M. Maheswari, M. Vanaja and B. Venkateswarlu. 2005.

Influence of water deficit at vegetative, anthesis and grain filling stages on

water relation and grain yield in sorghum. Indian J. Plant Physiol., 10: 20-

24.

Yalpani, N., A. J. Enyedi, J. Leon and I. Raskin. 1994. Ultraviolet light and ozone

stimulate accumulation of salicylic acid and pathogenesis related proteins and

virus resistance in tobacco. Planta, 193: 373-376.

Yang, J., J. Zhang, Z. Wang, G. Xu and Q. Zhu. 2004. Activities of key enzymes in

sucrose-to-starch conversion in wheat grains subjected to water deficit during

grain filling. Plant Physiol., 135: 1621-1629.

359

Yazdanpanah, S., A. Baghizadeh and F. Abbassi. 2011. The interaction between

drought stress and salicylic and ascorbic acids on some biochemical

characteristics of Saturejahortensis. Afr. J. Agric. Res., 6: 798-807.

Yeo, A. 1998. Molecular biology of salt tolerance in the context of whole-plant

physiology J. Expt. Bot., 49: 913–929.

Yildirim, E., M. Turan and I. Guvenc. 2008. Effect of foliar salicylic acid

applications on growth, chlorophyll and mineral content of cucumber

(Cucumis sativus L.) grown under salt stress. J. Plant Nutr., 31: 593-612.

Yordanov, I., V. Velikova and T. Tsonev. 2000. Plant responses to drought,

acclimation, and stress tolerance. Photosynthetica, 38:171-186.

Zaghlool, S. A. M. 2002. The effect of gibberellic acid (GA3), salicylic acid (SA),

sperimidine (Spd) and methods of application on growth, yield, some chemical

constituents and some phytohormones in mungbean (Vigna radiate L.). Arab

Univ. J. Agric. Sci. Ain Shams Univ. Cairo, 10 (2): 493-504.

Zang, X. and S. Komatsu. 2007. A proteomic approach for identifying osmoticstress-

related proteins in rice. Phytochem., 68: 426-437.

Zeid, I. M. and Z. A. Shedeed. 2006. Response of alfalfa to putrescine treatment

under drought stress, Biol. Plant. 50: 635–640.

Zhang, M., L. Duan, Z. Zhai, J. Li, X. Tian, B. Wang, Z. He and Z. Li. 2004. Effects

of plant growth regulators on water deficit-induced yield loss in soybean.

Proceedings of the 4th Inter. Crop Sci. Congress, Brisbane, Australia.

Zhao, H. J., X. W. Lin, H. Z. Shi and S. M. Chang. 1995. The regulating effect of

phenolic compounds on the physiological characteristics and yield of

soybeans. Acta Agron Sin., 21: 351-355.

Zhao, Y., S. K. Christensen, C. Fankhauser, J. R. Cashman, J. D. Cohen, D. Weigel

and J. Chory. 2001. A role for flavin monooxygenase-like enzymes in auxin

biosynthesis. Science 291: 306–309.

360

Zhu, J. K. 2003. Regulation of ion homeostasis under salt stress. Curr. Opin. Plant

Biol., 6: 441–445.

Zhu, J. K. 2002. Salt and drought stress signal transduction in plants, Annu. Rev.

Plant Biol. 53: 247–273.

Zhu, J. K. 2001. Plant salt tolerance. Trends in Plant Sci. 6: 66–71.

Zhu, J. K., P. M. Hasegawa and R. A. Bressan. 1997. Molecular aspects of osmotic

stress in plants. Critical Rev. of Plant Sci. 16: 253–277.

salicylic acid induced adaptive response of …prr.hec.gov.pk/jspui/bitstream/123456789/7681/1/husn...

Documents