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Dr. Sushrutha .C.K Dr. Ashok B.K.

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Mr. R. Giridharan

Honorary Members - Editorial Board

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INDEX – GJRMI, Vol.2, Iss. 7, July 2013

MEDICINAL PLANTS RESEARCH

Bio-Chemistry IN VITRO ANTIOXIDANT AND NITRIC OXIDE SCAVENGING ACTIVITIES OF PIPER

GUINEENSE SEEDS

Etim Okon E, Egbuna Chibuzor F, Odo Christian E, Udo Nsikan M, Awah Francis M 475–484

Bio- Technology PHARMACOGNOSTICAL INVESTIGATIONS ON DIFFERENT PARTS OF Clerodendrum inerme:

Chethana G S, Savitha H, Jyothi N, Hari Venkatesh K R, Gopinath S M 485–491

Bio-Chemistry

IDENTIFICATION OF HYDROCARBON DEGRADERS IN A CRUDE OIL POLLUTED VESSEL

AND POSSIBLE GROWTH INDUCING POTENTIAL OF AZADIRACHTA INDICA

Etim Okon E, Udosen Christiana I, Akara Priestine O N J

492–498

Review Article

ANALYSIS OF TRADITIONAL CHINESE MEDICINE INJECTIONS USED IN THE TREATMENT

OF RESPIRATORY SYSTEM-RELATED DISEASES BASED ON THE CHINESE MARKET

Zhi-Qiao Ma, Jin-Jian Lu, Wen-Shan Xu, Xiu-Ping Chen, Hao Hu, Yi-Tao Wang

499–508

Review Article

EXOTIC MEDICINAL PLANTS GROWING IN IRAN: A SYSTEMATIC AND

PHARMACOLOGICAL REVIEW

Mikaili Peyman, Aghajanshakeri Shahin, Moloudizargari Milad

509–524

Review Article

ETHNO MEDICINAL PRACTICES AMONG THE BINJHWAR TRIBE OF CHHATTISGARH,

INDIA

Shukla Rajesh, Chakravarty Moyna, Goutam M P

525–531

INDIGENOUS MEDICINE

Ayurveda – Dravya Guna EVALUATION OF ANTIARTHRITIC ACTIVITY OF LEPIDIUM SATIVUM LINN SEEDS

AGAINST FREUND’S ADJUVANT INDUCED ARTHRITIS IN RATS

Raval Nita D, Ashok B K, Ravishankar B

532–537

Ayurveda – Dravya Guna

DEVELOPMENT OF RANDOM AMPLIFIED POLYMORPHIC DNA MARKERS FOR

AUTHENTIFICATION OF OLAX SCANDENS ROXB.

Naik Raghavendra, Borkar Sneha D, Acharya R N, Harisha C R 538–545

Ayurveda – Review Article

ADVANCEMENTS IN INDIAN SYSTEM OF MEDICINE (ISM) INFORMATICS: AN OVERVIEW

Samal Janmejaya 546–553

COVER PAGE PHOTOGRAPHY: DR. HARI VENKATESH K R, PLANT ID – INFLORESCENCE OF

VIRATARU [DICHROSTACHYS CINEREA (L.) WIGHT & ARN], OF THE FAMILY MIMOSACEAE

PLACE – VRIDDHACHALAM, CUDDALLORE DISTRICT, TAMIL NADU, INDIA

Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 7 | July 2013 | 475–484

Global Journal of Research on Medicinal Plants & Indigenous Medicine || GJRMI ||

ISSN 2277-4289 | www.gjrmi.com | International, Peer reviewed, Open access, Monthly Online Journal

IN VITRO ANTIOXIDANT AND NITRIC OXIDE SCAVENGING

ACTIVITIES OF PIPER GUINEENSE SEEDS

Etim Okon E1*, Egbuna Chibuzor F2, Odo Christian E3, Udo Nsikan M4, Awah Francis M5

1, 2, 5Department of Biochemistry, Madonna University, Elele, Rivers state, Nigeria

3Department of Biochemistry, University of Nigeria, Nsukka, Nigeria

4Department of Pharmacology and Toxicology, University of Uyo, Uyo, Nigeria

*Corresponding author: E-mail: [email protected].

Received: 17/05/2013; Revised: 20/06/2013; Accepted: 28/06/2013

ABSTRACT

Piper guineense is a pepper widely consumed in some parts of West Africa especially Nigeria

and Ghana on account of its nutritional and medicinal properties. Indigenous people value the plant

for its ethno-medical uses, as well as its food spicing capabilities. In this study, the seeds of the fruits

were accessed for phyto-chemical constituents using spectro-photometric standard assays. The

antioxidant activity was accessed by determining its ability to scavenge the 2, 2-diphenyl-1-

picrylhydrazyl (DPPH) radical, superoxide ion radical and nitric oxide radical. The results showed

that the seeds were rich in flavonoids, flavonols and phenolic compounds. Total phenols were

estimated at 1.16 mg gallic acid equivalents while flavonoids were 1.28 mg rutin equivalents. The

seeds showed maximum inhibition of DPPH. radical of 66.4% at 500 µg/ml as compared to ascorbic

acid which inhibited 77.4% at the same concentration. Superoxide anion inhibition was maximally

70% at 1000 µg/ml as compared to rutin which inhibited 99.35% at the same concentration. The seed

extract was found to rapidly scavenge nitric oxide in vitro at different time intervals. The seeds could

therefore be employed as a natural antioxidant booster hence its relevance in food industry and

justification for its ethno-medicinal uses.

KEYWORDS: Piper guineense seeds, antioxidant activity, nitric oxide scavenging

Research article

Cite this article:

Etim Okon E, Egbuna Chibuzor F, Odo Christian E, Udo Nsikan M, Awah Francis M (2013), IN VITRO

ANTIOXIDANT AND NITRIC OXIDE SCAVENGING ACTIVITIES OF PIPER GUINEENSE SEEDS,

Global J Res. Med. Plants & Indigen. Med., Volume 2(7): 475–484

mailto:[email protected]



INTRODUCTION

Logically, human beings live in a highly

oxidative environment and many processes

involved in metabolism may result in the

production of more oxidants (Rui and Boyer,

2004). It has been estimated that there are more

than ten thousand oxidative hits to DNA per

cell per day in humans (Amnes et al., 1993).

For protection against free radicals, organisms

are endowed with endogenous (catalase, SOD,

gluthathione peroxidase / reductase) and

exogenous (vitamin C and E, ß-carotene, uric

acid) defense systems. However, in disease

conditions and in situations like contamination,

UV exposure etc. these systems are not

sufficient.

Most of the protective effects of plants on

living cells have been attributed to their non-

nutrient constituents e.g. carotenoid,

flavonoids, isoflavonoid and phenolic acids.

These and more different phyto-chemicals have

been shown to possess a range of activities

which may help in protecting against chronic

diseases like cancer, inflammatory diseases etc.

and also protects against lipid peroxidation

(Hollman and Katan, 1997; Liu, 2003). The use

of spice supplements in food has received

global attention due to their potential use as

anti-microbial, anti-helminthic, antioxidant,

anti-diabetic, neuro-protective, hypo-

cholesterolaemic, anti-hypertensive,anti-

inflammatory, cancer preventive and anti-

mutagenic agents (Kong et al., 2010; Mann,

2011).

Piper guineense Schumach. & Thonn,

popularly known as Ashanti pepper (Uziza), is

a climbing pepper belonging to the plant family

called Piperaceae (Rehm and Espig, 1991;

Amusan and Okorie, 2002; Asawalam, 2006).

This bush pepper is widely consumed in some

parts of West Africa especially in Nigeria and

Ghana on account of its nutritional and

medicinal properties (Negbenebor, 1999).

Studies have shown that apart from the use

of these plants as spices and condiments, they

have several other wide applications in the

local treatment and management of many

diseases. Indigenous people value the plants for

their ethno-medical uses as much as for spicing

foods (Stethberger, 1996). P. guineense is

used as an anti-convulsant (Pei, 1983; Abila et

al., 1993). Piper guineense provides oil used as

aromatic in the drink industry and also

medicinally (Rehm and Espig; Burkill, 1984).

The fruits contain the pungent Piperine, resin

and essential oils. The pungency of the pepper

is due to the presence in the fruits of various

resins particularly Chavicine and a yellow

alkaloid, piperine that contains 5–8% of the

weight of black pepper (Rehm and Espig; Lale,

1992). The fruit and leaves are used as spice for

preparing soup for post-natal women. Powder

from the dried fruits mixed with honey acts as a

carminative and reduces stomach aches. Extract

of black pepper has been reported to stimulate

digestion of foods by stimulating secretion of

digestive enzymes, pancreatic amylases, trypsin

and chymotrypsin (Platel and Srinivasan, 2000)

and is thus used for the treatment of digestive

disorders.

Despite these entire ethno-medicinal claims

about the potential uses of P. guineense, there

is little available scientific data to justify all the

claims. In this study we therefore investigated

the antioxidant potentials and the phyto-

chemical composition of the seeds.

MATERIAL AND METHODS

Chemicals

Folin-ciocalteu reagent (FCR), 2,2-

diphenyl-1-picrylhydrazyl (DPPH),

polyvinylpolyrrolidone (PVPP), riboflavin,

aluminum chloride, ferric chloride, sodium

nitrite, sulfanilamide, sodium nitroprusside

(SNP), methanol, L-ascorbic acid, phosphoric

acid, sodium bicarbonate, N-naphthyl-

ethylenediamine dihydrochloride, disodium

hydrogen phosphate, sodium chloride,

potassium chloride, nitroblue tetrazolium

(NBT), ethanol, sodium hydroxide, potassium

dihydrogen phosphate, sodium carbonate,

acetic acid, sodium acetate, methionine and

ethylene diamine tetraacetate (EDTA) were all

purchased from Sigma Aldrich, USA.



Preparation of Crude Extract

Seeds of Piper guineense were purchased

from a local market in Elele, Nigeria. The plant

was authenticated by Effiom O. Etim of Akwa

Ibom Agricultural Development Project

(AKADEP), Uyo, Nigeria. The seeds were air-

dried in ambient temperature. Dried seeds were

crushed to powder with mortar and pestle and

reduced to fine powder with a manual blender.

186.8 g of the resulting powder was subjected

to extraction with 80% ethanol, concentrated

using a rotator evaporator and stored at 4oC

until used.

Antioxidant Activity Assays

Quantitative DPPH Radical-scavenging

Assay

Scavenging activity on DPPH free radicals

by the extract were assessed for according to

the method reported by Awah et al., (Awah et

al., 2012). Briefly, a 2.0 ml solution of extract

at different concentration diluted in two fold in

methanol was mixed with 0.5 ml of 0.3 mM

DPPH in methanol. The mixtures were shaken

vigorously and allowed to stand at room

temperature in the dark for 25 min. Blank

solutions were prepared with each test sample

solutions (2.0 ml) and 0.5 ml of 0.3 mM DPPH

solutions plus 2.0 ml of methanol. L-ascorbic

acid was used as the positive control, thereafter,

the absorbance of the assay mixture were to be

measured at 517 nm against each blank with a

UV-visible spectrophotometer. DPPH radical

was calculated using the equation:

where (as in Awah et al., 2010) A0 is the

absorbance of the control, and As is the

absorbance of the tested sample. The IC50 value

represented the concentration of the extract that

caused 50% inhibition of DPPH radical and

was calculated by linear regression of plots,

where the abscissa represented the

concentration of tested sample and the ordinate

the average percent of inhibitory activity from

three replicates.

Superoxide Anion Radical (O2−)-Scavenging

Assay

This assay was based on the capacity of the

extract to inhibit the photochemical reduction

of nitro blue tetrazolium (NBT) as described by

Martinez et al., (Martinez et al., 2001) with

slight modifications (Awah et al., 2012).

Briefly, each 3.0 ml reaction mixture contained

0.1 M phosphate-buffered saline (PBS) (pH

7.8), 0.1M methionine, 1mM riboflavin, 1mM

EDTA, NBT (1mM) and 1.0 ml of test sample

solutions. The tubes were kept in front of a

fluorescent light and absorbance was read at

560 nm after 30 min. The entire reaction

assembly was enclosed in a box lined with

aluminum foil. Identical tubes containing

reaction mixture were kept in the dark and

served as blanks. The percentage inhibition of

superoxide generation was estimated by

comparing the absorbance of the control (rutin)

and those of the reaction mixture containing

test sample as per the equation:

Where A0 is the absorbance of the control, and

As is the absorbance of the tested sample (Awah

et al., 2010).

In vitro Nitric Oxide Radical (NO)

Scavenging Assay

NO generated from sodium nitroprusside

(SNP) was measured according to the method

modified by Awah (Awah et al., 2012). Briefly,

the reaction mixture (5.0 ml) containing SNP

(5 mM) in phosphate buffered saline. (pH 7.3),

with or without the plant extract at different

concentrations, was incubated at 25ºC for 180

min in front of a visible polychromatic light

source (25W tungsten lamp). The NO. radical

thus generated interacted with oxygen to

produce the nitrite ion (NO.) which was

assayed at 30 min intervals by mixing 1.0 ml of

incubation mixture with an equal amount of

Griess reagent (1% sulfanilamide in 5%



phosphoric acid and 0.1% xiv. N-naphtyl

ethylenediamine dihydrochloride). The

absorbance of the chromophore (purple azo

dye) formed during the diazotization of nitrite

ions with sulphanilamide and subsequent

coupling with N-napthyl ethylenediamine

dihydrochloride was measured at 546 nm. The

nitrite generated in the presence or absence of

the plant was estimated using a standard curve

based on sodium nitrite solutions of known

concentrations. Each experiment was carried

out in triplicates and the data presented as an

average of three independent determinations.

Phytochemical Analysis

Determination of total phenolic contents

Total phenolics were determined using

Folin-Coicalteu reagent (FCR) as described by

Velioglu et al., (Velioglu, 1998), with slight

modifications (Awah et al., 2012). FCR

consists of a yellow acidic solution containing

complex polymeric ions formed from

phosphomolybdic and phosphor tungistic

heteropoly acids. Dissociation of a phenolic

proton in a basic medium leads to a phenolate

anion which reduces FCR forming a blue

colored molybdenum oxide whose colour

intensity is directly proportional to the phenolic

contents. Briefly, 100 µl of the extract

dissolved in methanol (1 mg/ml) was mixed

with 750 µl of FCR (diluted 10-fold in dH2O)

and allowed to stand at 22ºC for 5 min; 750 µl

of Na2CO3 (60 g/l) solution was then added to

the mixture. Shaken to mix. After 90 minutes,

the absorbance was measured at 725 nm.

Determination of Tannins

Tannins content in each sample was

determined using insoluble polyvinyl-

polypirrolidone (PVPP), which binds tannins

(Makkar et al., 1993). Briefly, 1 ml of extract

dissolved in methanol (1 mg/ml), in which the

total phenolics were determined, was mixed

with 100 mg PVPP, vortexed, left for 15 minat

4ºC and then centrifuged for 10 min at 3000

rpm. In the clear supernatant, the non-tannin

phenolics were determined the same way as the

total phenolics (Velioglu et al., 1998). Tannin

content was calculated as difference between

total and non-tannin phenolic content.

Determination of Flavonoids and Flavonols

The flavonoids content was determined

according to the method described by Kumaran

and Karunakaran (Kumaran and Karunakaran,

2006) with slight modifications (Awah, 2012).

This method is based on the formation of a

flavonoid-aluminium complex which absorbs

maximally at 415 nm. Briefly, 100 µl of plant

extracts in methanol (10 mg/ml) was mixed

100 µl of 20% aluminium trichloride in

methanol and a drop of acetic acid, and then

diluted with methanol to 5 ml. The absorbance

at 415 nm was read after 40 min. Blank

samples were prepared from 100 µl of plant

extract and a drop of acetic acid, and then

diluted to 5 ml with methanol. The absorption

of standard rutin solution (0.5 mg/ml) in

methanol was measured under the same

conditions. The amount of flavonoids in the

plant extract in rutin equivalents (RE) was

calculated by the following formula:

where A is the absorption of plant extract

solution, Ao is the absorption of standard rutin

solution, m is the weight of plant extract, mg

and mo is the weight of rutin in the solution,

mg. The flavonoid content is expressed in mg

rutin equivalents/mg plant extract.

The content of flavonols was also

determined as described by Kumaran and

Karunakaran (Kumaran

and Karunakaran,

2006) with slight modifications (Awah, 2012).

Briefly, 1 ml of methanolic extract (10 mg/ml)

was mixed with 1 ml aluminium trichloride

(20 mg/ml) and 3 ml sodium acetate

(50 mg/ml). The absorbance at 440 nm was

read after 2.5 h. The absorbance of standard

rutin solution (0.5 mg/ml) in methanol was also

measured under the same conditions. The

amount of flavanols in the extract was

calculated by the same formula for flavonoids.



Statistical analysis

Linear regression plots were done using

Microsoft Excel for Windows Vista. All the

results are to be expressed as mean ± standard

error of the mean (SEM) (n = 3).

RESULTS

In vitro Free Radical Scavenging Activity

Effect of extract on DPPH. Radicals

The Piper guineense seed extracts showed

significant dose-dependent DPPH radical

scavenging capacity (Figure 1). The extract was

most efficient at 500 μg/ml, inhibiting 66.4 %

of DPPH radical compared to ascorbic acid

which inhibited 77.4% at the same

concentration.

Effect of extract on superoxide (O2-) anion

radical

The Piper guineense seed extracts inhibited

the formation of reduced NBT in a dose-related

manner. As shown in Figure 2, the extract

showed the maximal O2.- anion inhibitory

activity of 70.0 % at the concentration of 1000

μg/ml compared to the standard antioxidant

rutin 99.35% at 1000 μg/ml. The O2.-

scavenging effect of the extracts could

culminate in the prevention of .OH radical

formation since O2.- and H2O2 are required for

.OH radical generation.

IC50 for free radical inhibition

The concentration of the Piper guineense

seed extracts that inhibited 50% of the free

radicals and lipid peroxidation (IC50) was used

to determine the potency of the extracts. The

lower the IC50 value the better the extract

potency. As shown in Table 1 below, the Piper

guineense seed extracts were efficient

inhibitors of different free radicals compared to

standard anti-oxidants. The IC50 values for

DPPH radical was 227.66 μg/ml and that for

O2- anion 83.04 μg/ml.

Figure 1: DPPH radical scavenging activity of Piper guineense seed extract

Data represented as mean ± SEM (n = 3)



Figure 2: Superoxide anion radical (O2-) inhibition by Piper guineense seed extract.

Data represented as mean ± SEM (n = 3)

Table 1: Free radical and lipid peroxidation inhibitory potency (IC50)

IC50 value for inhibitory potential (μg/ml)

Extract DPPH. radical Superoxide anion (O2

.-)

Piper guineense seed 227.66 83.04

Standard anti-oxidant 18.638 * 22.2535

β

Data represented as mean ± SEM (n = 3) * compared to ascorbic acid

β compared to rutin.

Effect of extracts on nitric oxide (NO)

radical production

Nitric oxide (NO.) released from sodium

nitroprusside (SNP) has a strong NO character

which can alter the structure and function of

many cellular components. This study showed

that the Piper guineense extract in SNP

solution decreased levels of nitrite, a stable

oxidation product of NO. liberated from SNP

(Figure 3). The Piper guineense extract

exhibited strong NO. radical scavenging

activity leading to the reduction of the nitrite

concentration in the assay medium, a possible

protective effect against oxidative damage. The

NO. Scavenging capacity was concentration

dependent with 1000 µg/ml of the extracts

scavenging most efficiently.

Quantitative Analysis on Phyto-chemical

Constituents of Piper guineense seed extract.

Phenolic compounds were a major class of

bioactive components in the extracts. The

amount of total phenolics was 1.16 ± 0.17 mg

gallic acid equivalent (GAE)/mg of dry plant

extract and flavonoid contents was 1.28 ±

0.47 mg rutin equivalents / g dry weight plant

extract (Table 2).



Figure 3: Effect of Piper guineense seed extract on nitric oxide (NO.) radical production.

Table 2: Quantitative phytochemical constituents

Extract Phenolic contents * Total

flavonols ‡

Total

flavonoids ‡ Total Phenols Non-

tannins

Tannins

Piper

guineense seed

1.16 ± 0.17 0.95 ± 0.14 0.21 ± 0.04 0.53 ± 0.05 1.28 ±

0.47

Data represented as Mean ± SD (n = 3)

* Expressed as mg gallic acid equivalents (GAE) / mg dry weight Piper guineense seed extract ‡ Expressed as mg rutin equivalents (RE) / g dry weight Piper guineense seed extract

DISCUSSION

Free radicals are organic molecules

responsible for aging, tissue damage, and

implicated in a wide variety of diseases. These

molecules are very unstable, therefore they

look to bond with other molecules due to the

presence of unpaired electrons in their outer

shell,thereby causing mitochondria

malfunction,cell membrane damage and

eventually apoptosis. Antioxidants, present in

many foods, are molecules that prevent free

radicals from harming healthy tissue. The free

radicals are also involved in the normal

physiology of living organisms. In this study

the antioxidant activity of Piper guineense

extract was investigated using its DPPH radical

and nitric oxide scavenging potentials. DPPH

radicals are stable free radicals. This method

depends on the reduction of the purple DPPH

by accepting electrons from antioxidants to

stable coloured (yellow-coloured) DPPH. The

degree of colour change from purple to yellow

at different concentration was measured at

517 nm.

The extract showed a potent DPPH radical

scavenging potential inhibiting 66.4% of DPPH



at 500 μg/ml compared to the standard ascorbic

acid 77.4%. The antioxidant in the extract

neutralized the free radical character of DPPH

by transferring either electrons or hydrogen

atoms to DPPH (Naik et al., 2003), thereby

giving rise to the colour change. This

interaction depends on the structural

conformation of the bioactive compounds

present in the plant extract of which the

hydroxyl groups of flavonoids are highly

favorable (El-sayed, 2009). The degree of

discoloration indicated the scavenging potential

of the extract in terms of hydrogen donating

ability (Mosquera et al., 2007).

Though having its beneficial effect, nitric

oxide (NO) contribution to oxidative damage is

increasingly becoming evident. Excess

production of NO has been associated with

several ailments such as carcinomas, juvenile

diabetes, multiple sclerosis, arthritis and

ulcerative colitis (Hazra., et al., 2008). The

Piper guineense extract scavenged nitric oxide

in a time dependent manner figure 3, the nitrite

oxide levels were higher in the SNP only tubes

compared to SNP and extract mediums at the

different time interval. This suggests that SNP

generated nitric oxide, but the extract with its

potent nitric oxide scavenging activity, was

able to mop up the radicals.

Phenolic compounds, flavonoids and

flavonols which are known to possess good

medicinal values (Desta, 1993), were assayed

for in this extract. These phyto-chemicals have

a lot of pharmacological properties as proved

by earlier studies (Ajali, 2004). The observed

presence of tannins could be of great medicinal

importance since tannins serve as a good

antioxidant (Gulein, 2005). Therefore Piper

guineense extracts are good source of

antioxidants, which are widely believed to be

an important line defense against oxidative

stress leading to a lot of diseases like insomnia,

diabetes etc.

Flavonoids were also observed to be

present in the plant which could at the same

time contribute extensively to some biological

properties that promote human health and

reduce the risk of various diseases. Flavonoids

extend the activity of vitamin C, acts as

antioxidant, protects LDL cholesterol from

oxidation to unsafe cholesterol, reduce the risk

of cancer and also have anti-inflammatory

action (Ajali, 2004).

Plant phenolics present have received

considerable attention because of the potential

antioxidant activity. Phenolic compounds are

effective hydrogen donors which make them

good antioxidants. As shown in Table 1 the

total phenolic content of the extract was

moderately high.

This study therefore portrays that ethanol

extract of piper guineense seeds exhibited high

antioxidant and free radical scavenging

activities. Some reactive oxygen species (ROS)

are associated with the pathogenesis of

inflammatory diseases as such the free radical

inhibitory effect of the extract justifies its

ethno-medical use in the treatment of different

disease conditions.

CONCLUSION

Piper guineense being a plant used as

spices and condiments, have several other wide

applications in the local treatment and

management of many diseases. It has

significant antioxidant activity owing to its free

radical scavenging potential as shown in the

results. Therefore, this plant could be relevant

in the prevention and treatment of diseases

whose pathogenesis could implicate the

oxidative stress as well as in the food industry

as a good preservative owing to its anti-

oxidative potential.

ACKNOWLEDGEMENTS

The authors acknowledge the chancellor;

Madonna University, Elele, Very Rev. Prof. E.

M. P. Edeh for his support and Prof. A. A.

Uwakwe of Department of Biochemistry,

University of Port Harcourt, Nigeria, for his

useful suggestions.



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Source of Support: Nil Conflict of Interest: None Declared




PHARMACOGNOSTICAL INVESTIGATIONS ON DIFFERENT PARTS OF

CLERODENDRUM INERME

Chethana G S1, Savitha H

2, Jyothi N

3, Hari Venkatesh K R

4, Gopinath S M

5*

1Research Associate, Sri Sri Ayurveda Trust, Bangalore

2PG Scholar, ALN Rao Memorial Ayurvedic Medical College, Koppa-577126, Chikkamagalur District,

Karnataka, India 3Research Assistant, ALN Rao Memorial Ayurvedic Medical College, Koppa-577126, Chikkamagalur District,

Karnataka, India 4Head, R & D, Sri Sri Ayurveda Trust, Udayapura, Bangalore District, Karnataka, India

5HOD, Department of Biotechnology, Acharya Institute of Technology, Bangalore District, Karnataka, India

*Corresponding Author; Email Id:[email protected]


ABSTRACT

Clerodendrum inerme is a hedge plant belongs to the Verbenaceae family, traditionally used for

ornamental purpose in home gardens. Clerodendrum inerme is used in many places in landscaping,

as a ground cover or a hedge plant, especially near the sea, as it tolerates the salt spray. The different

parts of the plant are used as folk medicines against many diseases. The plant has many active

components like alkaloids, flavonoids, terpenes etc. having a wide range of application in the field of

medicine. Being a weed, with innumerable therapeutically useful components is a boon to the plant

to be explored & established for its medicinal potential. The present study was undertaken with an

aim to explore the pharmacognostic aspect of the plant which forms the very basic part of a drug

evaluation. The microscopic and macroscopic observations made have been documented which

might be an eye opener to future researches on this plant.

KEYWORDS: Clerodendrum inerme, leaf, stem, root, pharmacognosy

Research article

Cite this article:

Chethana G S, Savitha H, Jyothi N, Hari Venkatesh K R, Gopinath S M (2013),

PHARMACOGNOSTICAL INVESTIGATIONS ON DIFFERENT PARTS OF CLERODENDRUM

INERME, Global J Res. Med. Plants & Indigen. Med., Volume 2(7): 485–491



INTRODUCTION

Clerodendrum inerme is valued in

landscaping as a groundcover or hedge plant. It

has attractive evergreen foliage and fragrant

white flowers that form in clusters and are

accented by delicate red protruding stamens.

Seaside Clerodendrum, as its name suggests,

grows well along the beach tolerating the salt

spray of the ocean and the harsh rays of the

sun. It is a versatile plant and can be grown as

topiary or as a bonsai (S.R. Harish and K.

Murugan, 2011), (Forest Starr et al., 2003).

Clerodendrum inerme belongs to the family

Verbenaceae. The genus Clerodendrum

includes over 450 species of tropical regions.

The name Clerodendrum is derived from the

Greek kleros, meaning chance or fate, and

dendron, meaning tree, in reference to the

uncertain medicinal qualities of some of the

plants (Forest Starr et al., 2003).

Evergreen sprawling shrub, 1–1.8 meter

tall. Stems (Fig 1 & 2) woody, smooth. Leaves

(Fig 1 & 2) ovate to elliptical (5–10 cm) long,

acute to acuminate tip, green, smooth, slightly

shiny upper surface, pinnate venation, margins

entire, leaves opposite, simple. Cyme or umbel

usually comprised of 3 flowers joined at a

common base point; corolla white, fused, with

5 lobes; stamens 4, reddish to purple and

upwardly curved. Fruit green turning black, 1–

1.5 cm long, obovoid (S.R. Harish and K.

Murugan, 2011), (Forest Starr et al., 2003).

The different parts of the plant have shown

therapeutically significant activities like

antinemtidalcidal activity, antimicrobial

activity, and anti-hepatotoxic activity. It has

also been used for curing skin diseases,

rheumatism etc. The methanolic extract of root

contains verbanoside, which possess analgesic

and anti microbial activities which is prescribed

conforming the traditional use of this plant as

medicine. The crude extract of the leaf also

have shown the anti microbial activity.

(Chethana G.S et al., 2013). This plant

Clerodendrum inerme is known to have many

active principles like alkaloids, flavanoids,

terpenes etc (Chethana G.S et al., 2013)

It is an important medicinal plant reported

to be used in the treatment of skin diseases,

venereal infections, elephantiasis, asthma,

topical burns and for rheumatism. It is also

used as a substitute of quinine. In Siddha

medicine, it is used under the names of

‘Chankan kuppi’ and ‘Pechagnan’. A glycoside

ester namely Verbascoside has been isolated

from the root of this plant, which has analgesic

and antimicrobial properties. The antioxidant

activity of the plant extract may be due to the

presence of polyphenols which are reported as

strong antioxidants (Forest Starr et al., 2003).

Fig. 1 Clerodendrum inerme in its habitat Fig. 2 Clerodendrum inerme in its habitat



The leaves and stems contain a number of

triterpenes, diterpenoids, sterols and flavones.

The leaves yielded the flavanolid, friedelin,

salvigenin (5-hydroxy-6, 7, 4’- methoxy

flavones), acacetin, cirisimaritin,

pectolinarigenin, apigenin (5, 7-dihydroxy-4’

mathoxy flavaone) and amethyl flavones,

cleroflavone (7-hydroxy 5, 4’ dimethoxy-6-

methyl flavanone). The leaves also yielded

diterpenes clerodendrin B. the leaves exhibited

growth inhibition and anti-feedant activities in

house flies and mosquitoes. (KS Krishnan

Marg, 2010). Clerodendron inerme as a

febrifugal and uterine stimulant, a pest control

agent and antiseptic, to arrest bleeding,

treatment of asthma, hepatitis, ringworm,

stomach pains (Chellaiah Muthu et al., 2006).

The aim of the present study was to explore

the plant pharmacognostically by the study of

the macroscopic and microscopic observation

of internal and external parts of the plant.

MATERIALS AND METHODS

The whole plant, Clerodendrum inerme was

collected from Harihar TQ, Davangere dist,

Karnataka, India during the month of April

2013. The botanical identity was confirmed by

the Taxonomical experts in the R&D wing, Sri

Sri Ayurveda Trust, Udayapura, Bangalore-82,

Karnataka, India. The voucher no. of the

specimen is 129 maintained in Acharya College

of Technology, Bangalore, Karnataka, India.

Macroscopical evaluation

The sample was cleaned and macroscopic

evaluation of root, leaf, and stem was carried

out. The leaf, stem and root were then

separated and individual macroscopic

characters like size, shape, texture were noted

in detail.

Microscopical evaluation

Free hand sections of leaf, stem and root

were taken and washed with chloral hydrate

solution. Sections were first observed in

distilled water then stained with Saffranin red.

Photomicrographs were taken by Carl zeiss

trinocular microscope.

RESULTS AND DISCUSSION

Macroscopic characters of root:

Roots are cylindrical in shape, woody; cut

pieces 10 cm in length, 2–3 cm breadth.

Externally dark brown and internally brownish

cream in colour. Surface is rough and at places

longitudinal striations and wrinkles seen, at

some place it is exfoliated. Fracture short and

splintery. Odour is slight aromatic, bland in

taste. Fibrous in Texture.

Microscopic characters of root:

The transverse section of root shows, 10–12

rows of tangentially elongated and radially

arranged cork cells [Fig 4.1] followed by cortex

formed with 8-10 layers of oval to round

shaped parenchyma cells which are compactly

arranged with prisms of calcium oxalate [Fig

4.2]. The xylem vessels are of varying size,

lignified, found isolated or in the group of 2–3

[Fig 4.3]. Medullary rays are 2–3 seriate and

the cells are pitted and lignified [Fig 4.4].

Starch is found in wood only.

Macroscopic characters of stem:

Stem is cylindrical in shape, woody; cut

pieces 15 cm in length, 0.2–0.3 mm in breadth.

Externally light brown in colour & internally

whitish to light green in colour. Surface is

nearly smooth, pubescent with white oval

shaped lenticels present on it. Fracture short

and splintery. Odour is disagreeable with bland

taste. Fibrous in Texture.

Microscopic characters of stem:

The transverse section of stem shows,

single layer of epidermis with 2–3 celled

covering trichomes followed by 8–10 layers of

cortex consisting of oval shaped

parenchymatous cells which are compactly

arranged with prisms of calcium oxalate.

Patches of lignified fibers are scattered at

places in the outer few layers of cortex. Within

the cortex, there is an interrupted layer of

lignified fibers followed by phloem and xylem

vessels. Pith is very large consisting of oval

shaped parenchymatous cells.



Fig 3: T.S. of Root of Clerodendrum inerme

Fig 4: Microscopy of different root parts of Clerodendrum inerme

4.1 Cork, 4.2 Cortex, 4.3 Vascular bundle, 4.4 Medullary rays, 4.5 Absence of Pith



Fig 5: T.S of Stem of of Clerodendrum inerme

Macroscopic characters of leaf:

Leaves are simple, opposite or ternate,

elliptic or obovate in shape, size is 5–6 cm long

and 3–3.8 cm broad, apex is obtuse or

mucronate, slightly attenuate at base, margin

entire, reticulate venation, glabrous. Odour

disagreeable, coriaceous in texture.

Microscopic characters of leaf:

Cross section of midrib shows thick walled

circular cells of upper and lower epidermis with

cuticle [Fig 7.1 & Fig 7.6]. Just below the

upper epidermis we can see two types of

parenchyma cells i.e., 2–3 layer oval shape

parenchyma cells followed by elongated cells.

Above the lower epidermis 4–6 layer of oval

shaped parenchyma cells can be seen. The

vascular bundle present in middle of midrib is

semicircular [Fig 7.4]. Just below the vascular

bundle, lignified fibers can be seen [Fig 7.5].

Cross section showed both the epidermis

(Upper and Lower) in the lamina consisting of

fairly thick walled circular cells with thick

cuticle layer. In both epidermises we can

observe glandular trichomes which are sub

sessile and situated in a shallow cavity. The

mesophyll consists of two to three layered

palisade cells [Fig 7.2] just below the upper

epidermis and four or five layers of spongy

parenchyma cells with arenchymatous cells

[Fig 7.3] at places seen below the palisade cells

which extend up to lower epidermis. The lateral

veins occur in the median position and has

small collateral vascular bundle.



Fig 6: T.S of Leaf of Clerodendrum inerme

Fig 7: Microscopy of different leaf parts of Clerodendrum inerme

7.1 Upper epidermis with cuticle, 7.2 Palisade parenchyma, 7.3 Spongy parenchyma with aerenchyma, 7.4 Vascular

bundle, 7.5 Lignified fibres, 7.6 Lower epidermis with cuticle



CONCLUSION

From this work on Clerodendrum inerme ,it

is helpful in the study of physiology of each

cell structure of different parts of the plant. It is

very important to understand the physiology of

a plant cell like the presence of lignified cell

which supports the plant and many more. The

reason for undertaking this study is, it reported

that this plant has a significance usage in

treatment of fever, wounds, skin disease etc. It

is experimentally proven to show the activity of

some of the properties of the Clerodendrum

inerme like antimicrobial activity which

correlates the usage as folk medicines like

applying for wounds etc. Much more work has

to be carried out regarding the medicinal value

of this plant available in the treatment of the

diseases.

REFERENCES

S.R. Harish and K. Murugan., (2011).

Biochemical and Genetical Variation in

the Mangrove Associate (L) Gaertn.

Under Different Habitats of Kerala.

Asian J. Exp. Biol. Sci. 2(4): 553–561.

Forest Starr, Kim Starr, and Lloyd Loope.,

(2003). Clerodendrum inerme Seaside

Clerodendrum Verbenaceae. United

States Geological Survey--Biological

Resources Division Haleakala Field

Station, Maui, Hawai'i January.1–3.

Chellaiah Muthu, Muniappan Ayyanar,

Nagappan Raja and Savarimuthu

Ignacimuthu., (2006). Medicinal plants

used by traditional healers in

Kancheepuram District of Tamil Nadu.

India Journal of Ethnobiology and

Ethnomedicine.2:43.

Chethana G.S., Hari Venktatesh K.R., S.M

Gopinath., (2013). Review on

Clerodendrum inerme. JPSI. 2(2):38–40

Chethana G.S., Hari Venktatesh K.R., S.M

Gopinath (2013). Phyto Chemical

Analysis of Clerodendrum inerme.

IRJP. 4(5):208–209

KS Krishnan Marg (2001). The Wealth of

India. National Institute of Science

Communication, CSIR, New Delhi.

2:67–68.





IDENTIFICATION OF HYDROCARBON DEGRADERS IN A CRUDE OIL

POLLUTED VESSEL AND POSSIBLE GROWTH INDUCING POTENTIAL

OF AZADIRACHTA INDICA

Etim Okon E1*, Udosen Christiana I

2, Akara Priestine O N J

3

1, 3Department of Biochemistry, Madonna University, Elele, Nigeria

2Department of Microbiology, University of Uyo, Uyo, Nigeria.

*Corresponding author: E-mail: [email protected]


ABSTRACT

Polycyclic aromatic hydrocarbons (PAHs) are a class of organic compounds that have

accumulated in the natural environment mainly as a result of anthropogenic activities such as the

combustion of fossil fuels. Some microorganisms, mainly from the genera Pseudomonas and

Mycobacteria were found to be capable of transforming and degrading PAHs. Bioremediation is one

approach that has been used to remediate contaminated soil and waters, and at the same time

promotes the natural attenuation of the contaminants using microbial community of the site.

However, the aim of this study was to investigate the possible effect of Azadirachta indica leaves on

hydrocarbon degraders in crude oil polluted cotton wool vessel. The study was carried out using

pumpkin seeds, Azadirachta indica, and crude oil polluted cotton wool vessel. The result from this

study showed that the vessel with the highest quantity of Azadirachta indica leaves had the highest

growth of hydrocarbon degrading microorganism.

KEYWORDS: Azadirachta indica, Hydrocarbon degraders, polycyclic aromatic hydrocarbons.

Research article

Cite this article:

Etim Okon E, Udosen Christiana I, Akara Priestine O N J (2013), IDENTIFICATION OF

HYDROCARBON DEGRADERS IN A CRUDE OIL POLLUTED VESSEL AND POSSIBLE GROWTH

INDUCING POTENTIAL OF AZADIRACHTA INDICA, Global J Res. Med. Plants & Indigen. Med.,

Volume 2(7): 492–498




INTRODUCTION

Petrochemical industries and petroleum

refineries generate large amounts of priority

pollutants. The major pollutants found in these

industries are petroleum hydrocarbons (Chavan

and Mukherji, 2008).

Petroleum is a complex mixture of

hydrocarbons derived from the geologic

transformation and decomposition of plants and

animals that lived hundreds of millions of years

ago. Petroleum consists mostly of hydrocarbon

molecules. Crude oil and natural gas are the

most important primary fossil fuels. Polycyclic

aromatic hydrocarbons (PAHs), also known as

poly-aromatic hydrocarbons or polynuclear

aromatic hydrocarbons, are potent atmospheric

pollutants that consist of fused aromatic rings

and do not contain hetero-atoms or carry

substituent (Fetzer, 2000).

Bioremediation is the process whereby

organic wastes are biologically degraded under

controlled conditions to an innocuous state. The

main principle of this technique is to remove

pollutants from the natural environment or

convert the pollutants to a less harmful product

using the indigenous microbiological

community of the contaminated environment

(Mueller et al., 1997). Interest in the microbial

biodegradation of pollutants has intensified in

recent years as mankind strives to find

sustainable ways to clean up contaminated

environments. These bioremediation and

biotransformation methods use the naturally

occurring, microbial catabolic diversity to

degrade, transform or accumulate a huge range

of compounds including hydrocarbons,

polychlorinated biphenyls (PCBs),

polyaromatic hydrocarbons (PAHs),

pharmaceutical substances, radionuclide’s and

metals. The concentration of PAHs in the

environment varies widely depending on the

level of industrial development, proximity of

the contaminated sites to the production source

and the mode of PAHs transport. Reported soil

and sediment PAHs contaminations range from

1 µg/kg to over 300 g/ kg (Kanaly, 2000).

Anaerobic metabolism of PAHs is thought

to occur via the hydrogenation of the aromatic

ring. The basis of this mechanism is the

oxidation of the aromatic ring, followed by the

systematic breakdown of the compound to

PAH metabolites or carbon dioxide.

PAH-degrading microorganisms are

ubiquitously distributed in the natural

environment, such as in soils (bacteria and non-

ligninolytic fungi) and woody materials

(ligninolytic fungi). Many PAH contaminated

soils and sediments host active populations of

PAH-degrading bacteria (Tam et al., 2002).

Medicinal plants are part and parcel of

human society to combat diseases, from the

dawn of civilization. Azadirachta indica A.

Juss (syn. Melia azadirachta) is well known in

India and its neighbouring countries for more

than 2000 years as one of the most versatile

medicinal plant showing a wide spectrum of

biological activity. A. indica A. Juss and M.

azedarach are two closely related species of the

family Meliaceae. The former is popularly

known as Indian neem (margosa tree) or Indian

lilac, and the latter is known as Persian lilac.

Neem is an evergreen tree, cultivated in various

parts of the Indian subcontinent. Every part of

the tree has been used as traditional medicine

for household remedy against various human

ailments, from antiquity (Chatterjee and

Pakrashi, 1994).

This study is therefore aimed at using

grounded neem tree leaves to remediate crude

oil polluted vessel. The result will ascertain the

effectiveness of these leaves in bioremediation

process.


Chemicals

Crystal violet, Lugol iodine, Ethanol,

Hydrogen peroxide, Urea solution, K2HPO4,

Methyl red, α –naphtol, KOH, Methyl-P-

phenylene diamine hydrochloride, Mineral salt

medium, Sabouraud dextrose agar, Nutrient

agar, Urea agar, Citrate agar, Nutrient broth,

and Peptone were all purchased from Sigma

Aldrich, USA.

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

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

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

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



Experimental design:

Vessel 1 ------- Pumpkin seeds + water only

Vessel 2 -------- Pumpkin seeds + 200 ml of

crude oil + water

Vessel 3 ------- Pumpkin seeds + 200 ml of

crude oil + 150 g of grounded Azadirachta

indica Leaves + water

Vessel 4 ------- Pumpkin seeds + 200 ml of

crude oil + 300 g of grounded Azadirachta

indica Leaves + water

The pumpkin seeds were planted and

monitored for eight weeks using sterilized

cotton wool as soil. After which the cotton

wool polluted with crude oil was collected and

used for the microbiological assay.

Isolation of hydrocarbon utilizing organisms

from cotton wool polluted with crude oil.

30 g of each polluted cotton wool was

soaked in saline water (100 ml) for 1 hour, after

which 1 ml from each of the soaked container

was pipette and mixed with 9 ml of sterile

water in a test tube and the dilution were

prepared up to 10-5

dilution.

Enumeration of crude oil utilizing fungi

This was done using the surface spreading

technique (Mbagwu, 1992)

1 ml from 10-5

of each sample was used to

seed each sterile plate in triplicates. 20 ml of

the mineral salt agar medium at 45oC

supplemented with 1% Lactic acid was poured

into the seeded plates and swirled.

Enumeration of crude oil utilizing bacteria

20 ml of mineral salt medium was

supplemented with nystatin was poured in the

seeded plates and swirled. These plates were

left on the bench to set. Filter sterile paper were

soaked in sterile crude oil and placed on the

lids of the cultured plates, after which the plates

were incubate for 14 days at room temperature.

Colony that was developed on the plates was

counted with colony counter and was recorded

as colony forming unit per gram (Okpowasili et

al., 1988).

Identification of bacteria

The procedure of Hamamura et al., (2006)

was used.

Pure culture that was obtained was sub-

cultured from the primary culture on Nutrient

agar plates and was incubated at room

temperature for 24 hours. Discrete colonies

were stocked in nutrient agar slants and labeled

accordingly. Further characterization and

identification were carried out base on

microscopic examination, gram staining and

some biochemical assays.

A smear of the culture was placed on a

clean grease-free slide, which was air-dried and

fixed. The slide was then stained with crystal

violet solution (primary stain) for 1 minute,

after which it was washed with tap water and

well drained. Iodine solution (mordant) was

used to flood the slide for 30seconds, after

which the slide was washed with tap water.

Decolourization was done using acetone and

washed with water. The slide was flooded with

safranin (counter staining) for 60 secs, after

which the slide was washed and allowed to dry.

It was examined under oil immersion objective.

Gram positive organisms retained the colour of

the primary stain (purple) while the gram

negative retained the secondary stain (pink)

(Fowole and Oso, 1988).

Biochemical test

The biochemical test for identification of

bacteria isolates were carried out as described

by Prescott et al., (2005)

Catalase test

A drop of 30% H2O2 was placed on a glass

slide using a wire loop, a little inoculums was

removed and mixed with the H2O2 on the slide.

A positive test was indicated by bubbling; the

enzyme present on catalase positive organisms

degrades hydrogen peroxide and releases O2

which is detected as effervescence.



Motility test

This test was used to determine the presence or absence of Flagella. The motility medium in tubes were inoculated by making a fine stab of the isolates with an inoculating needle to the depth of about 1–2 cm short of the tubes bottom and were incubated for 24 hours at 37

oC.

Motile organisms grow outside the line of stabbing, while the non-motile organisms grow on the line of stabbing.

Urease test

This test is used to study the ability of an organism to utilize Citrate present in Simmon’s media as a sole source of carbon for growth. In order to identify an organism that is able to use citrate as carbon source, the test organisms were inoculated into Simmon citrate agar slant and incubated for 24–72 hours. The development of a blue colour indicates a positive test.

Triple sugar iron test (TSI)

This test is based on the fermentation of the sugars, lactose, dextrose and the production of gas and hydrogen sulphide. A sterilized wire loop was used to pick the inoculums and stab the butt of the triple sugar iron agar medium with the test organism; these tubes were inoculated for 24 hour.

Oxidase test

The test indicates the presence of cytochrome oxidase which catalyses the oxidation of reduced cytochrome by oxygen. It indicates the ability of microbes to oxidize amine.

A freshly prepared 1% solution of oxidase reagent was soaked on a piece of filter paper and with a sterile loop the test organisms were smeared on the area impregnated with the oxidase reagent. Deep purple coloration after a few seconds shows a positive test.

Screen test of isolates for ability to utilize

hydrocarbons

Bacterial and fungal isolates were tested for

their ability to utilize hydrocarbon using

turbidity method. Bacterial isolates were

cultured on Nutrient broth and incubated for 24

hours at 28oC. Fungal isolates were inoculated

into malt extract broth and incubated at room

temperature (Nweke and Okpowasili, 2011).

1 ml and 1 g each from bacterial and fungal

isolates were inoculated into mineral salt broth

and 1ml of sterile crude oil was added into the

inoculated tubes .The control tubes were

incubated at room temperature under stationary

condition for 7 days. The growths of the

inoculates were determined by visual

observation of the mineral salt broth medium

turbidity as compared with the un-inoculated

control tubes.

RESULTS

Effect of Azadirachta indica Leaves on the

Levels of Bacterial Hydrocarbon Degraders

in Crude Oil Polluted Cotton Wool Vessel.

As shown in Figure 1 below, the bacterial

counts of the crude oil polluted cotton wool

medium was significantly affected by

bioremediation with the A. indica leaves in a

concentration dependent manner. At all

concentrations of A. indica the bacterial total

hydrocarbon degraders levels were

significantly higher (p<0.05) compared to the

levels in the control. However, at 150 g of

neem leaves, there was no significant

difference (p > 0.05) in the bacterial counts

(2.0 × 105cfu/g) compared to the vessel with

plant and crude oil only (2.1 × 105cfu/g).

Effect of Azadirachta indica Leaves on the

Levels of Fungal Hydrocarbon Degraders in

Crude Oil Polluted Cotton Wool Vessel.

Figure 2 shows that the fungal counts of the

crude oil polluted cotton wool was significantly

improved by bioremediation with the leaves of

A. indica in a concentration dependent manner.

At all concentrations the fungal total

hydrocarbon degraders levels were

significantly higher (p < 0.05) in the A. indica

treated medium compared to the levels in the

control (1.0 × 104cfu/g).



Figure 1: Effect of Azadirachta indica leaves on the Levels of Bacteria Hydrocarbon Degraders

in Crude oil Polluted Cotton Wool Vessel.

Figure 2: Effect of Azadirachta indica Leaves on the Levels of Fungal Hydrocarbon Degraders

in Crude oil Polluted Cotton Wool Vessel.

0

0.5

1

1.5

2

2.5

3

3.5

Control Plant + Crude only Plant + Crude + Neem (150g) Plant + Crude + Neem (300g)

Bac

teri

al C

ou

nts

(x

105

cfu

/g)

0

0.5

1

1.5

2

2.5

Control Plant + Crude only

Plant + Crude + Neem (150g)

Plant + Crude + Neem (300g)

Fun

gal C

ou

nts

(x

10

5cf

u/g

)



DISCUSSION

The results of the present study confirmed

the matter that many of bacterial strains,

especially gram-negative bacteria were found

to degrade poly aromatic hydrocarbons (PAHs)

compounds at various extents (Frick et al.,

1999; Cerniglia 1992 ‘Sutherland et al.,

1995).this explains the significant growth of

these degraders as seen in vessels rich in crude

oil compared to control that received normal

water. All of PAHs degrading bacterial strains

which have, been identified in the present study

were gram negative, which agreed with the

results indicated that most efficient of the

PAHs degrading bacteria were belong to the

genus Pseudomonas (Frick et al., 1999).

Microorganisms and plants have

complementary roles in phyto-remediation of

the polluted soil.

Phyto-remediation refers to the use of

plants to clean contaminated soil. Increase in

biodegradation of organic contaminants in the

rhizosphere soil, the zone of soil directly

adjacent to and under the influence of plant

roots has been reported (Frick et al., 1999).

For successful phyto-remediation, both

plants and microorganisms must survive and

grow in crude oil contaminated soil. Phyto-

remediation can be applied at moderate

contamination levels or after the application of

other remediation measures as a polishing step

to further degrade residual hydrocarbons and

improve soil quality (Frick et al., 1999).

From the result in figure 1 and 2, leaves of

Azadirachta indica has been able to enhance

the growth of hydrocarbon degraders according

to the work of Frick et al., (1999) showing their

complementary role in bioremediation. There

was high microbial growth in vessel 4 with

300 g of Azadirachta leaves compared to other

vessels. The biostimulation potential of

Azadirachta Indica leaves in increasing the

microbial population in crude oil contaminated

vessel maybe due to the nitrogen and

phosphorus content of the leaves. Okpkwasilli

(1994), observed that the use of NPK fertilizer,

urea fertilizer and poultry droppings effectively

stimulated bacterial growth into utilization of

crude oil. The high total hydrocarbon degraders

(THD) bacteria in vessel 4 with 300 g of

grounded leaves (3-2 × 105cfu/g) and that of

total fungal count(2.0 × 105cfu/g) compared to

control (1.0 × 104cfu/g) respectively suggests

its effectiveness in bioremediation process.

CONCLUSION

The biodegradation of polycyclic aromatic

hydrocarbons in the environment is a complex

process, microorganisms such as bacteria and

fungi are the key agents of bioremediation,

with bacteria assuming the dominant role in

marine ecosystems and fungi becoming more

important in freshwater and terrestrial

environments (Leahy and Colwell, 1990). The

result from this study shows that the vessel

with the highest quantity of Azadirachta indica

leaves in it had the highest growth of

hydrocarbon degraders in it, which means that

the presence of A. Indica leaves enhanced the

growth of the hydrocarbon degraders. It

therefore suggests that A. indica leaves could

act as growth inducers to enhance the growth of

hydrocarbon degraders (bacteria and fungi) for

effective bioremediation process in a

hydrocarbon polluted environment.

REFERENCES

Cerniglia, C.E.,(1992). Biodegradation of

polycyclic aromatic hydrocarbons.

Biodegradation, 3: 351–368.

Chatterjee, A. and Pakrashi, S. (1994). The

Treatise on Indian Medicinal Plants,

vol. 3,p. 76 (eds)

Chavan,A.and Mukherji.(2008).Treatment of

hydrocarbon rich wastewater using oil

degrading bacteria and phototropic

microorganisms in rotating biological

contactor; Effect of N:P

ratio.J.Hazardous Materials.154(1–3)

63–72.



Fetzer, J.C. (2000) The Chemistry and analysis

of the large Polycyclic hydrocarbons.

Polycyclic Aromatic Compounds (New

York: Wiley) 27: 2–143.

Fowole M.O, and Oso, B.A (1988) Laboratory

Manual of Microbiology.

Frick, C.M., R.E. Farrell and J.J. Germida

(1999). Assessment of

phytoremediation as in-situ technique

for cleaning oil-contaminated sites, Pet.

Tech. All. Can. Calgary, AB.

Hamamum.N., S.H.Olson., D.M.Ward and

W.P.Inskeep. (2006). Appl. Environ.

Microbiol. 72:6316–6324.

Kanaly, R.A. and S. Harayama, 2000.

Biodegradation of high-molecular-

weight polycyclic aromatic

hydrocarbons by bacteria. J. Bacteriol.,

182:2059–2067.

Kausik Biswas, Ishita Chattopadhyay, Ranajit

K. Banerjee and Uday Bandyopadhyay

(2002). Biological activities and

medicinal properties of neem

(Azadirachta indica) Current Science,

82 (11).

Leahy, J.G and Colwell, R.R (1990) Microbial

degradation of hydrocarbons in

environment Microbiol.Rev.3:305.

Mbagwu J.S.C. (1992). Bioresource

technology.42:167–175.

Mueller JG, Devereux R, Santavy DL, Lantz

SE,Willis SG and Pritchard PH, (1997)

Phylogenetic and physiological

comparisons of PAH-degrading bacteria

from physiologically diverse

soils.Antonie Leeuwenhoek 71:329–343.

Nweke C. O. and Okpowasili G.C, (2011)

Inhibition of β-galactosidase and α-

glucosidase bact. Of zinc and cadmium

Journal of Env.chem & ecotoxicology.

Okpokwassilli,G.C, (1994). Pollution control:

The increasing role of bioremediation:

In:R.A. Borofice (ed.) Biotechnology in

National Development : National

Agency for science and Engineering

infrastructure (NASENI)

Prescott,L.M.,J.P.Harley and D.A.Klein.(2005).

Microbiology;Mcgraw-Hill Higher

Education.

Sutherland, J.B., F. Rafii, A.A. Khan and

C.E.Cerniglia, (1995). Mechanisms of

polycyclic aromatic hydrocarbon

degradation. Microbial transformation

and degradation of toxic organic

chemicals. L.Y. Young and C.E.

Cerniglia (Eds.), Wiley-Liss,New York,

N.Y., pp: 169–306.

Tam NFY, Guo CL, Yau WY and Wong YS,

(2002) Preliminary study on

biodegradation of phenanthrene by

bacteria isolated from mangrove

sediments in Hong Kong. Mar Pollut

Bull 45:316–324.





ANALYSIS OF TRADITIONAL CHINESE MEDICINE INJECTIONS USED

IN THE TREATMENT OF RESPIRATORY SYSTEM-RELATED DISEASES

BASED ON THE CHINESE MARKET

Zhi-Qiao Ma1, Jin-Jian Lu

2, Wen-Shan Xu

3, Xiu-Ping Chen

4, Hao Hu

5, Yi-Tao Wang

6*

1, 2, 3, 4, 5, 6State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical

Sciences, University of Macau, Macao, China

*Corresponding Author: Email: [email protected]


ABSTRACT

Traditional Chinese medicine (TCM) injections (TCMIs) comprise TCM formulations with vital

functions in the treatment of cancer, cardiovascular diseases, respiratory system-related diseases

(RSRDs) and so on. This article analyzes TCMIs used in the treatment of RSRDs in the Chinese

market with special emphasis on the Xiyanping, Tanreqing, Xuebijing, Yanhuning, Reduning,

Chuankezhi, Chuanhuning, and Shuanghuanglian (Chinese names of these TCMIs) injections, which

present excellent market performances. Analysis of the clinical applications, herbal and chemical

compositions, and pharmacological activities of these TCMIs were also conducted. TCMIs have vital

functions in the treatment of RSRDs. This article aims to explore the market prospects and

development of TCMIs used to treat RSRDs.

KEYWORDS: Traditional Chinese medicine injections (TCMIs), Respiratory system

related-diseases (RSRDs), Qingre, adverse reactions, Chinese market

Review article

Cite this article:

Zhi-Qiao Ma, Jin-Jian Lu, Wen-Shan Xu, Xiu-Ping Chen, Hao Hu, Yi-Tao Wang (2013), ANALYSIS

OF TRADITIONAL CHINESE MEDICINE INJECTIONS USED IN THE TREATMENT OF

RESPIRATORY SYSTEM-RELATED DISEASES BASED ON THE CHINESE MARKET, Global

J Res. Med. Plants & Indigen. Med., Volume 2(7): 499–508




INTRODUCTION

Many respiratory system-related diseases

(RSRDs), such as asthma, chronic obstructive

pulmonary disease and pneumonia are common

diseases. According to a data from the World

Health Organization (WHO), at least 3000

million people have died because of chronic

respiratory diseases and more than 90% of the

patients with such diseases come from low- or

middle-income countries (WHO, 2012a).

Lower respiratory tract infections are ranked

third in the top ten global death reasons and the

first in low-income countries. Almost 14

million children below five years old are killed

every year by pneumonia, a classic lower

respiratory tract infection (WHO, 2012b).

According to data from the 2010 Chinese

health statistical yearbook, the morbidity of

RSRDs in China is about 6.94%, totaling over

80 million people, and RSRDs rank as the

fourth leading cause of death in urban and rural

areas (Ministry of Health of the People’s

Republic of China, 2010).

Besides chemical drugs used for standard

treatment of RSRDs, traditional Chinese

medicine (TCM) use is increasing in China

because of its excellent clinical effects (Wang

et al., 2006). TCMs have important functions in

preventing serious infections, such as SARS

and H1N1, as they have multiple functions,

which include relieving cough, diminishing

inflammation, eliminating phlegm, and

relieving asthma (Chang et al., 2011a).

TCM injections (TCMIs) are sterile

formulations (emulsion, powder, or thick liquid)

prepared for injecting into the body and are

made after extraction and purification (National

Pharmacopoeia Committee, 2005). TCMIs are

becoming increasingly valued in China due to

their positive clinical effects (Yao, 2007). This

article aims to explore the market prospects and

development of TCMIs used to treat RSRDs.

MATERIAL AND METHODS

Data sources

Data on the sales of TCMIs in the whole

Chinese market were obtained from the

Chinese Clinical Medicine Terminal

Competition Pattern Database of the State Food

and Drug Administration (SFDA), Southern

Medicine Economic Research Institute

(SMERI), (China Medicine Economic

Information Network, 2012), which is

calculated by the TCM purchasing practices of

150 sample hospitals from nine cities (i.e.,

Beijing, Guangzhou, Nanjing, Chongqing,

Chengdu, Xian, Haerbin, Shenyang, and

Zhengzhou) in China. Thirteen large categories

and 75 small categories of drugs are listed

based on the WHO therapeutic classification,

Anatomical Therapeutic Chemical Code, and

national essential drugs list classification for

TCM. Data on clinical applications, herbal and

chemical compositions and pharmacological

activities were searched from China National

Knowledge Infrastructure (CNKI) database

and/or Pubmed database.

Data analysis

The annual sales and sales growth rate of

TCMIs were analyzed: For market share

calculations, the formula SN/Stotal × 100% was

used; for sales growth rate, the formula

(SN-SN-1)/SN × 100% was used, where SN is the

sales of one injection at the year N and Stotal are

the total sales of the drugs, including the

injections.

The top eight selling TCMIs used in the

treatment of RSRDs in the Chinese market

were chosen for analysis. The names of these

injections were used as key words to search for



their clinical applications, herbal and chemical

compositions, and pharmacological activities in

the CNKI database. The chemical names of the

injections were further used as key words to

perform searches in Pubmed database. All of

the studies included in this work were

published from 1994 to 2012.

RESULTS

TCM for RSRDs treatment in the Chinese

market

The sales of chemical drugs and TCMs

accounted for 82.6% and 17.4% of all drugs

sales in 2010, respectively, based on the data

from SFDA SMERI (China Medicine

Economic Information Network, 2012). The

sales of chemical drugs and TCMs for the

treatment of RSRDs accounted for 54.5% and

45.5% of all sales, respectively, in 2010

(excluding systemic anti-infective drugs, which

are mostly treated by antibiotics in China),

indicating the importance of TCMs in treating

RSRDs in China. The sales of TCMs for

RSRDs increased from 2006 to 2011, with an

average growth rate of 34% (Figure 1), the

annual growth rates from 2007 to 2011 are 24%,

17%, 29%, 16% and 25% respectively.

Figure 1. The sales of TCM for the treatment of RSRDs from 2006 to 2011



TCMIs for RSRDs treatment in the Chinese

market

At present, 1,269 TCMIs formulations are

registered in SFDA, including 524 species

(41.3%) for RSRDs treatment (China Medicine

Economic Information Network, 2012). Among

these injections, 365 species were listed in the

National Health Insurance Directory and 135

species were listed in the national essential

drugs list. These injections account for 70%

and 26% of the total number of TCMIs used in

the treatment of RSRDs, respectively.

The brand concentration of TCMIs for

RSRDs treatment is high. In 2011, five species

of injections were observed in the top ten sales

of TCMs for RSRDs treatment and their sales

accounted for 38% of the total sales of TCMs

for RSRDs. Among these injections, the sales

of three species added up to one billion (China

Medicine Economic Information Network,

2012). The top five selling TCMIs for RSRDs

treatment are Xiyanping, Tanreqing, Xuebijing,

Yanhuning, and Reduning injections (Chinese

names of some TCMIs) (see in Table 1). All of

the injections maintained a positive growth

trend, except for the Yanhuning injection, the

growth rate of which declined by 1% (Figure 2).

Xiyanping, Tanreqing, and Reduning injections

are all listed in the national health insurance

directory. The sales of Xiyanping injection

increased to 0.32 billion yuan (RMB) in 2011,

almost twice that of Bailing capsules, the best

seller of other TCM formulations for RSRDs

treatment (China Medicine Economic

Information Network, 2012).

Figure 2. The change of TCMIs’ sales for the treatment of RSRDs from 2006 to 2011



Table 1. The rank of TCMIs’ sales for the treatment of RSRDs in 2011

Rank Injections Chinese herbal

medicine

compositions

Main chemical

compositions

Market

share of

RSRDs

medicine

Sales

(million,

RMB)

Growth

rate

(%)

1 Xiyanping

injection

Herba

Andrographisis

Andrographolide, etc. 14.7% 323 73

2 Tanreqing

injection

Radix Scutellariae,

Bear bile powder,

Goat horn, Flos

Lonicerae

Japonicae, Fructus

Forsythiae

Baicalin, Chlorogenic

acid,

Ursodeoxycholic acid

etc.

9.3% 204 22

3 Xuebijing

injection

Flos Carthami,

Radix Paeoniae

Rubra, Rhizoma

Chuanxiong, Radix

Angelica Sinensis,

Radix Salviae

Miltiorrhizae

Danshensu, Safflower

yellow pigment A,

Tetramethylpyrazine,

Ferulic acid,

Paeoniflorin,

Protocatechuic

aldehyde etc.

6.5% 142 23

4 Yanhuning

injection

Herba

Andrographisis

Andrographolide, etc. 4.5% 98.2 -1

5 Reduning

injection

Herba Artemisiae

Annuae, Flos

Lonicerae

Japonicae,

Fructus

Gardeniae

Chlorogenicacid,

Geniposide,

Artemisinin etc.

2.2% 47.5 11

6 Chuankezhi

injection

Herba Epimedii,

Radix Morinda

Officinalis

Epimedium

polysaccharide,

Icariin,Epimedium

flavonoids etc.

0.4% 9.6 150

7 Chuanhuning

injection

Herba

Andrographisis

Andrographolide ,

etc.

0.4% 8.1 -32

8 Shuanghuanglian

injection

Flos Lonicerae

Japonicae, Radix

Scutellariae,

Fructus Forsythiae

Caffeic acid,

Chlorogenic acid,

Baicalin, Forsythin

etc.

0.1% 2.6 -17



Clinical applications of TCMIs for RSRDs

Both Xiyanping and Reduning injections are

used to treat infectious diseases and mainly

focus on children (Liu et al., 2007; Zhang &

Yang, 2012). These injections are also used to

treat hand, foot, and mouth disease in babies,

infantile diarrhoea, acute icteric model hepatitis,

chronic prostatitis, and old herpes zoster,

among others, in addition to the treatment of

RSRDs (Liu et al., 2007; Zhang & Yang, 2012).

Tanreqing, Yanhuning, and Chuankezhi

injections are mostly used to treat bronchitis

and various types of pneumonia (Feng, 2007;

He, 2008; Liang, 2006). Besides treating

bronchitis and the upper respiratory tract

infections, both Chuanhuning and

Shuanghuanglian injections are used to treat

gastrointestinal tract inflammation, urinary tract

infection, and myocarditis (Li, 2005; Liu & Xu,

2005). Xuebijing injection is significantly

different from the aforementioned injections, as

it is generally used for emergency and critical

care, such as systemic inflammatory response

syndrome caused by acute respiratory distress

syndrome and sepsis (Zhang, 2008). Therefore,

TCMIs have wide clinical applications and

excellent performance in the market for RSRDs

treatment.

Herbal compositions of TCMIs used for

RSRDs treatment

Among the TCMIs mentioned in Table 1,

most belong to the Qingre area (clearing heat)

in TCM. For example, the major compositions

of Xiyanping, Yanhuning, and Chuanhuning

injections are herbs of Andrographis (Ma &

Kuang, 2010; Zhong, Zeng, & Guo, 2010).

Both Tanreqing and Shuanghuanglian

injections contain Flos Lonicerae Japonicae,

Radix Scutellariae and Fructus Forsythiae (Li

& Li, 2011; Tu & Huang, 2008). Reduning

injection contains Herba Artemisiae Annuae,

Flos Lonicerae Japonicae and Fructus

Gardeniae (Jiang, et al.,2008; Liang, Huang,

& Cai, 2008). The main herbs that compose

Xuebijing injection are Flos Carthami, Radix

Paeoniae Rubra, Rhizoma Chuanxiong, Radix

Angelica Sinensis and Radix Salviae

Miltiorrhizae, all of which have the effect of

Huoxue (promoting blood circulation) (Chang

et al., 2011b; Yu, 2011). The main herbs in

Chuankezhi injection are Herba Epimedii and

Radix Morinda Officinalis, which have the

effect of Buyi (tonic) (Li et al., 2009). No herb

belongs to the Huatan Zhike Pingchuan class

(preventing phlegm from forming, stopping

coughing and relieving asthma) of TCM, which

is the most-represented TCM for RSRDs

treatment among the top eight ranking

injections. The absence of an herbal component

is due to the dosage form, as syrup is the main

form for the Huatan Zhike Pingchuan class of

TCM.

Chemical compositions and pharmacological

activities of TCMIs used for RSRDs

treatment

We further analyzed the main chemical

compositions and pharmacological activities of

the top eight injections listed in Table 1. Three

injections contained andrographolide or its

derivatives. Andrographolide presents excellent

antibacterial, antiviral, anti-inflammatory, and

anticancer effects (Chen et al., 2009; Ji, 2011;

Lim et al., 2012). Chlorogenic acid and

baicalin have also emerged in numerous

injections. Chlorogenic acid has

anti-inflammatory, antioxidative, antibacterial,

cardiovascular protective effects, and so on (Ji,

2011; Wang et al., 2011). Baicalin has

antibacterial, antiviral, antioxidative,

anti-inflammatory, sedation, and immune

system regulation effects (Ji, 2011; Srinivas,

2010; Zhu et al., 2012). Several injections

contain geniposide, danshensu, safflower



yellow A, phillyrin, and icariin, among others,

all of which have numerous pharmacological

activities (Ji, 2011).

DISCUSSION

Chemical drugs continue to enjoy several

advantages in the Chinese market. However,

the markets for TCM and chemical drugs are

similar in RSRDs treatment if the influence of

antibiotics is excluded. The TCM market in

RSRDs treatment has continuously grown in

recent years. If the Chinese government

continues to enforce numerous policies to

reduce antibiotic abuse, the advantages of TCM

will significantly increase.

TCMIs can enter human tissues, blood, or

organs directly, and can be absorbed faster than

other forms of TCM. Thus, TCMIs overcome

several disadvantages of TCM dosing, such as

dose inaccuracy, slow effect and low

bioavailability. Compared with anti-neoplastic

TCMIs, which are mostly used for adjuvant

therapy (Lai et al., 2012), TCMIs have more

important functions in RSRDs treatment. From

the herbal and chemical composition analysis,

most TCMIs for RSRDs treatment have Qingre

effects (clearing heat).

Classifying TCM against the standards of

Western medicine is difficult due to the

different theories behind Chinese and Western

medicine. For example, Xuebijing injection is

also used to treat cardiovascular diseases.

Clinical applications of cross treatments could

result in greater potential for TCMIs. With

more and more TCMIs used in the treatment of

RSRDs are into the national health insurance

directory, along with the new medical reform

policy is carried out constantly, the TCMIs

market will continue to expand in China.

Although the TCMIs market for RSRDs

treatment is increasing significantly, the entire

market scale remains small. The market

concentration is extremely high such that few

products occupy most of the market. The

increased market is mainly attributed to these

products, which are almost exclusively

manufactured or monopolized by a few

companies. Thus, the medicine market and

patients are cautious about the TCMIs used in

RSRDs treatment. Besides, the credibility of

TCMIs has decreased because numerous

patients have reported adverse reactions (Liang

& Shi, 2012; Ma & Kuang, 2010).

Chemical compositions, clinical

applications, and individual differences are the

three main reasons for the adverse reactions of

TCMIs. TCMIs have several features, such as

complex compositions, herb quality

unsteadiness, and residual impurity, and these

factors are the main sources of adverse events

(Ma & Kuang, 2010; Wu et al., 2012).

Irrational clinical applications also result in

adverse reactions (Ma & Kuang, 2010; Wu et

al., 2012). For such adverse events, legal and

administrative regulations should be enhanced.

The SFDA carried out a series of specialized

policies for the quality of TCMIs and safety

evaluation from 2007 to 2009. These policies

have increased the threshold of TCMIs research

and development and advanced efforts for the

safety evaluation of TCMIs. Identifying and

controlling the effective and toxic compositions

of the injections are necessary. Technological

studies and clinical compatibility studies for

TCMIs should be given more attention.

CONCLUSION

TCMIs have highly important functions in

RSRDs treatment in China. From the market’s

perspective, TCMIs for RSRDs treatment show

a positive trend and great potential, with rapid



increasing sales and expanding market demand.

In clinical view, TCMIs show the predominant

and effective feature, by combining some

advantages between TCM and Western

medicine. Development of TCMIs is a

complicated issue with traditional theory and

advanced technology, so research and

development institutes, manufacturers,

hospitals, and regulation departments must

cooperate with one another to establish a

positive outlook for the development of

TCMIs.

ACKNOWLEDGMENTS:

This study is supported by the research

funding of University of Macau

(UL016/09Y4/CMS/WYT01/ICMS,

MYRG208(Y2-L4)-ICMS11-WYT, and

MYRG160(Y2-L2)-ICMS11-HH).

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Source of Support: University of Macau, China Conflict of Interest: None Declared




EXOTIC MEDICINAL PLANTS GROWING IN IRAN: A SYSTEMATIC AND

PHARMACOLOGICAL REVIEW

Mikaili Peyman1, Aghajanshakeri Shahin

2*, Moloudizargari Milad

3,

Javaherypour Soheil4

1Department of Pharmacology, Faculty of Pharmacy, Urmia University of Medical Sciences, Urmia, Iran

2, 3, 4Student of Veterinary Medicine, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran

*Corresponding Author: E-mail: [email protected]


ABSTRACT

Medicinal plants are being used as desirable natural sources of various preparations made in

traditional herbal therapy worldwide. Since ancient times, traditional remedies have been trusted by

Iranian people. Plants of various families can be found in the prescription of rural healers in all

regions of Iran. Some of these are native to Iran but many others are known as exotic plants which

only grow in especial regions worldwide. This study for the first time illustrates the presence and the

growth of several families of these plants in the climate of Iran along with some of their most

important pharmacological effects and their active constituents.

KEYWORDS: Exotic medicinal plants; Iran; systematic study; pharmacological study

Review article

Cite this article:

Mikaili. P., Aghajanshakeri. S., Moloudizargari. M., Javaherypour. S (2013), EXOTIC

MEDICINAL PLANTS GROWING IN IRAN: A SYSTEMATIC AND PHARMACOLOGICAL

REVIEW, Global J Res. Med. Plants & Indigen. Med., Volume 2(7): 509–524




INTRODUCTION

The use of phytomedicines in the Iranian

traditional medicine has a long history. Since

ancient times, rural native-healers and drug

sellers in different parts of Iran have used

different herbs in their prescriptions that grow

in Iran. Present article consists of 15 exotic

medicinal plants that grow in Iran along with

some of their important pharmacological

properties. These species include:

Buxushyrcana, Cercissiliquastrum, Cestrum

nocturnum, Chamaerops humilis, Cotinus

coggygria, Cupressus arizonica, Cupressus

sempervirens, Diospyros khaki, Eriobotrya

japonica, Citrus aurantium, Citrus limon,

Citrus paradise, Citrus reticulata, Citrus

sinensis and Citrus unshiu. The use of these

plants in traditional medicine has been proved

and their pharmacological properties have been

already confirmed in different in vitro studies

by many researchers. This study illustrates the

potential of these plants to be employed as

novel medicinal sources for the development of

new drugs in Iran and other countries. Present

work is the pioneer study that shows the growth

of these exotic species in Iran.


In order to gather the required information,

the authors of the study traveled to Mazandaran

province located in North of Iran and

documented images of the exotic species which

grow in that region. Systematic review of

literature was performed and a summary of the

important botanical and pharmacological

properties of each plant was presented.

RESULTS

[1] Buxus hyrcana

Description

B. hyrcana (common box) exists as tree and

is distributed throughout the world (M. Mesaik

et al., 2010). B. hyrcana (Fig. 1) is locally

known as "šemšād-jangalī" in Iran and the

medicinal parts of the plant including its dried

leaves and the woody aerial parts have been

traditionally used among Iranians (Mikaili et

al., 2012; Esmaeili et al., 2009). The main

compounds of B. hyrcana responsible for a

variety of pharmacological effects of this plant

are steroidal alkaloids such as Buxidin and E-

Buxenone (Mesaik et al., 2010) and

triterpenoidal alkaloids such as 17-oxo-3-

benzoylbuxadine and buxhyrcamine (Ata et al.,

2010).

Pharmacological effects

B. hyrcana possesses hypotensive effect

(Mikaili et al., 2012), antiplasmodial activity

(Esmaeili et al., 2009), acetylcholinesterase,

butyrylcholinesterase and glutathione S-

transferase inhibitory activity (Ata et al., 2010;

Bahar, Ata, & Meshkatalsadat, 2006; Chouhary

et al., 2003). Inhibitory effect on the growth of

Mycobacterium tuberculosis attributable to the

cycloprotobuxine present in the plant (Mikaili

et al., 2012). Immunosuppressive activity due

to the inhibition of IL-2 by steroidal alkaloids

(M. A. Mesaik et al., 2010). Some

triterpenoidal alkaloids from this plant are

shown to have weak anti-leishmanial and

modest anti-fungal effects (Ata et al., 2010).

Figure 1. Buxus hyrcana Figure 2. Cercis siliquastrum



Figure No. 3. Chamaerops humilis Figure 4. Cotinus coggygria

Figure 5. Cupressus arizonica Figure 6. Diospyros khaki

Figure 7. Eriobotrya japonica Figure 8. Citrus aurantium



[2] Cercis siliquastrum

Description

C. siliquastrum (Fabaceae) commonly

known as Judas tree is a small deciduous tree

from Southern Europe and Western Asia which

is noted for its prolific display of deep-pink

flowers in spring. It is one of the main

components of a Unani herbal tea called

"Zahraa" (Carmona, Llorach, Obon, & Rivera,

2005). According to a study three flavonoids

including 3-O-monoglycosides of kaempferol,

quercetin and myricetin were obtained from the

genus Cercis (Salatino et al., 2000).


It shows anti-malarial activity (Kaiser et al.,

2007). Leaf extracts of C. siliquastrum (Fig. 2)

strongly inhibit 2C-methyl-D-erythritol 4-

phosphate synthase (IspC), an enzyme

catalyzing the first step of the non-mevalonate

pathway of isoprenoid biosynthesis. Since this

pathway serves as the unique source of

terpenoids in numerous pathogenic eubacteria

and in apicoplast-type protozoa which is absent

in mammalian cells, therefore it can be an

attractive target for anti-infective chemotherapy

(Kaiser et al., 2007).

[3] Cestrum nocturnum

Description

C. nocturnum is a garden shrub from the

family Solanaceae, commonly known as "lady

of the night" which is used as a remedy for

different health disorders. (Perez-Saad &

Buznego, 2008). The crude extract of C.

nocturtum has bioactive phytochemicals with

predominance of saponins(Patil, Patil, Salunke,

& Salunkhe, 2011). Spirostanolsaponin,

furostanolsaponin, pseudo-furostanolsaponin,

pregnane glycosides, cholestane glycosides,

pregnane-carboxylic acid gamma-lactone

glycoside, and spirostanol glycosides have been

isolated from the leaves of C. nocturnum

(Mimaki, Watanabe, Sakagami, & Sashida,

2002).


Its active substances possess analgesic activity provided through a peripheral action mechanism (Perez-Saad & Buznego, 2008). Its Mosquito larvicidal activity has been established (Patil et al., 2011). The n-butanol and polysaccharides extracts of C. nocturnum inhibited tumor growth in tumor-bearing mice in a dose dependent manner (Zhong et al., 2008). Steroidal saponins from this plant have been shown to be cytotoxic against squamous cell carcinoma-(HSC-2) cells and normal human gingival fibroblasts (Mimaki et al., 2001).

[4] Chamaerops humilis

Description

C. humilis (Fig. 3) family Arecaceae (Palm family) is known as the Mediterranean Fan Palm native to the Mediterranean coast (Mayoral, Torres, Munoz, Bartolome, & Blanca, 2006). Fruits are eaten in Morocco; heart "palmito" is consumed in Spain and its young suckers are cooked and eaten in Italy (Haynes & McLaughlin, 2000). It has been used for ornamental purposes in Japan and as a medicinal plant to treat diabetes in Morocco (Gaamoussi et al., 2010). It is also used to make brooms in Italy (vernacular name: Palma nana) (Nedelcheva et al., 2007). Some of its main constituents include: procyanidins (fruits), leucoanthocyanidin, diosgenin, flavone C-glycoside and tricin (leaves and root), and methyl proto-dioscin (stem) (Hirai et al., 1985).


This plant is probably an uncommon cause of allergy because it is only consumed in its natural form in a few rural areas of the Mediterranean region (Mayoral et al., 2006). Its extract inhibited the growth of calcium oxalate monohydrate crystals in vitro (Beghalia et al., 2007). The aqueous leaf extract decreases total cholesterol and triglycerides (Gaamoussi et al., 2010). It was suggested in one study that the plant may become a good source of antidiabetic medication and may also be useful in the management of secondary complications of diabetes (dyslipidemia) (Gaamoussi et al., 2010).



[5] Cotinus coggygria

Description

C. coggygria (syn. Rhus cotinus, family

Anacardiaceae) is a shrub that extends from

southern Europe, the Mediterranean, Moldova

and the Caucasus, to central China and the

Himalayas. It is also known as the "smoke

tree". Plants of the family Anacardiaceae have

a long history of use by various peoples for

medicinal and other purposes such as its usage

as a dyestuff. In folk medicine, C. coggygria

(Fig. 4) is routinely used as an antiseptic, anti-

inflammatory, antimicrobial and anti-

haemorrhagic agent in wound-healing, as well

as for countering diarrhea, paradontosis and

gastric and duodenal ulcers (Matic et al., 2011;

Valianou et al., 2009).


Alcoholic extract of the plant has high

antioxidant (313 and 231 mg AA/g dry extract),

as well as DPPH free-radical scavenging

property (IC (50) =9 and 99 μg/ml), inhibitory

activity toward lipid peroxidation (IC (50) =3

and 17 μg/ml) and reducing power (Niciforovic

et al., 2010; Savikin et al., 2009). Isolated

compounds mainly responsible for its

antioxidant activity include: disulfuretin,

sulfuretin, sulfurein, gallic acid, methyl gallate,

and pentagalloyl glucose (Westenburg et al.,

2000). It has in vitro cytotoxic activity towards

HeLa and LS174 human cancer cell lines

(Savikin et al., 2009). In one study, the

methanol extract of C. coggygria in a

concentration of 5% induced recessive lethal

mutations on X-chromosome on Drosophila

melanogaster in all broods indicating its

mutagenesity (Stanic et al., 2011).

[6] Cupressus arizonica

Description

C. arizonica (Fig. 5) (family Cupressaceae) is

an aromatic evergreen coniferous plant with

great importance in urban horticulture and in

the pharmaceutical and fragrance industries

(Hassanpouraghdam, 2011; Rehfeldt, 1997). It

is native to North and Central America and is

widespread in Mediterranean countries because

of their optimal conditions for growth. It was

introduced to Iran in 1954 and has been

commonly cultivated in many parts of the

country. This plant is becoming an increasingly

frequent cause of allergic diseases (Sanchez-

Lopez, Asturias, Enrique, Suarez-Cervera, &

Bartra, 2011; Sedaghat et al., 2011). α-pinene,

limonene and umbellulone has been determined

as major components of the oil of C. arizonica

(Sedaghat et al., 2011).


The plant has shown larvicidal activity

against malaria vector Anopheles stephensi

(with the highest dose of 160 ppm essential oil)

(Sedaghat et al., 2011), in vivo and in vitro

allergenic activity (Ariano et al., 2006),

sensitization (symptoms include: allergic

rhinitis and asthma) (Sanchez-Morillas et al.,

2005).

[7] Cupressus semperviren

Description

C. sempervirens (family Cupressaceae) is a

tree that grows up to 30 meter height. The plant

is indigenous to Turkey and is cultivated

throughout the Mediterranean region. Chief

compounds include: alpha-pinene, D-

camphene, D-silvestrene, p-cymene, L-

cadinene, cedrol, terpinenol-4, terpineol,

acetyl- and isovalerianyl esters of monoterpene

alcohols. The drug is used externally for head

cold, cough and bronchitis (PDR for Herbal

Medicines, 2000; Rehfeldt, 1997).


Cytotoxic activity against amelanotic

melanoma C32 in vitro (IC (50) value of

104.90 microg/mL) (Loizzo et al., 2008),

inhibition of glucose-6-phosphatase glycogen

phosphorylase enzymes (Rawat et al., 2010),

and sensitisation (symptoms: rhinitis and

asthma) (Sposato & Scalese, 2011). Cypress

also acts as an expectorant (PDR for Herbal

Medicines, 2000).



[8] Diospyros khaki

Description

D. kaki (Fig. 6) (family Ebenaceae) known

as young persimmon is widely distributed in

East Asia and its leaves are traditionally used

for the treatment of hypertension, angina and

internal haemorrhage. Flavonoids are the main

therapeutic components of D. khaki (Bei et al.,

2009). Triterpeneaglycone is also an important

constituent of this plant that has drawn

attentions of many researchers (Chen et al.,

2009; Matsumoto et al., 2010). It has good

antioxidant activity due to its phenolic content

(Celep et al., 2012). Main compounds include:

triterpenoids (Chen et al., 2009).


D. kaki possesses neuroprotective effects

attributable to its flavonoids (Bei et al., 2009)

and hypolipidemic activity through an acid-

binding (bile acid) mechanism (Matsumoto et

al., 2010). In one study, the acetone extract of

the peel of persimmon inhibited melanin

biosynthesis in mouse B16 melanoma cells.

The inhibitory effects were found to be

mediated by suppression of tyrosinase

expression. (Fukai et al., 2009; Ohguchi et al.,

2010). The presence of 2-methoxy-4-

vinylphenol has led to high antioxidant and free

radical scavenging activity of the plant (Fukai

et al., 2009; Sun et al., 2011). It was shown in

an in vitro study that the oral administration of

starch with polyphenol concentrate of

persimmon leaf tea can dose-dependently

decrease the blood glucose level in Wistar rats

due to inhibition of pancreas alpha-amylase

(Kawakami et al., 2010).

[9] Eriobotrya japonica

Description

Loquat (family Rosaceae) is a perennial

subtropical fruit tree. The fruits can be

consumed fresh or processed into jam, juice,

wine, syrup, or candied fruits. The flowers

(inflorescences) and leaves have been widely

used as Traditional Chinese Medicine for

treatment of cold, cough, gastro-enteric

disorders, diabetes mellitus, chronic bronchitis

and asthma. Many studies demonstrated that

large amounts of flavonoids and phenolics were

found in the fruit and leaf of loquat (Alshaker

et al., 2011; Zhou et al., 2011).


In one study, E. japonica (Fig. 7) showed

potent inhibitory effect on the inflammatory

mediators including nitric oxide, iNOS, COX-

2, TNF-α and IL-6 via the attenuation of NF-

κB translocation to the nucleus (Cha et al.,

2011). The plant has antinociceptive activity

via both central and peripheral mechanisms as a

weak opioid agonist (Cha et al., 2011). It also

possesses antioxidant activity due to its

flavonoids and phenolics (Xu and Chen, 2011;

Zhou et al., 2011). Moreover, the plant is a

potent anti-metastatic agent (Zhou et al., 2011).

[10] Citrus aurantium

Description

Bitter orange (family Rutaceae) is

indigenous to tropical Asia but is widely

cultivated in other regions nowadays, such as

the Mediterranean. Preparations of Bitter

Orange flower and flower oil are used as a

sedative and a preventive measure for gastric

and nervous complaints, gout, sore throat,

anxiety and sleeplessness. Fruit peel is used for

the treatment of pain in the epigastrum,

vomiting and anorexia as folk medicine in

China. Limonoids and Flavonoids are the chief

constituents of the plant (PDR for Herbal

Medicines, 2000).


C. aurantium (Fig. 8) is used as an

alternative medicine in some countries to treat

anxiety, and recently the anxiolytic role of this

medicinal plant has been already established in

an animal model study. Blossoms of the plant

may also be effective in terms of reduction in

preoperative anxiety before minor operation

(Akhlaghi et al., 2011). Since ephedra-

containing dietary supplements (herbal weight-

loss products) were banned from the US

market, manufacturers changed their



formulations by eliminating ephedra and

replacing with other botanicals, including bitter

orange. It contains, among other compounds,

synephrine (Oxedrine), a chemical that is

chemically similar to ephedrine. It was shown

in a study that this compound has noticeably

less effect than ephedrine on the blood pressure

and heart rate (Han et al., 2012). It was shown

in an study that crude methanol extracts of the

peels of C. aurantium L. may induce caspase-

dependent apoptosis at least in part through Akt

inhibition, providing evidence that CMEs have

anticancer activity on human leukemia cells

(Fugh-Berman & Myers, 2004). Although

seville orange (C. aurantium) extracts are being

marketed as a safe alternative to ephedra in

herbal weight-loss products, it may also have

the potential to cause adverse health effects. C.

aurantium contains 69, 79-

dihydroxybergamottin and bergapten, both of

which inhibit cytochrome P450-3A, and would

be expected to increase serum levels of many

drugs. Synephrine has lipolytic effects in

human fat cells only at high doses (Hansen et

al., 2012). Flavonoids isolated from C.

aurantium L. induced G2/M phase arrest

through the modulation of cell cycle related

proteins and apoptosis through activation

caspase. These finding suggest flavonoids

isolated from C. aurantium L. were useful

agent for the chemoprevention of gastric cancer

(Hansen, Juliar, White, & Pellicore, 2011).

Doses of up to 100 mg synephrine/kg body

weight did not produce developmental toxicity

in Sprague-Dawley rats (Lee et al., 2012).

[11] Citrus limon

Description

C. limon (family Rutaceae) is a tree

indigenous to northern India, cultivated in

Mediterranean regions and in subtropical

regions of the world. In folk medicine, lemon

juice is recommended as a drink in fever, as a

remedy for acute rheumatism and as an antidote

to intoxicants, particularly opium. Flavonoids

such as hesperidin, rutin and ericitrim are the

chief constituents of the plant (PDR for Herbal

Medicines, 2000).


C. limon (Fig. 9) essential oil (EO)

possesses a strong antioxidant potential.

Moreover, it presented scavenger activity

against all in vitro tests. Oral EO (50, 100, and

150 mg/kg) significantly reduced the number of

writhes, and at highest doses, it reduced the

number of paw licks whereas naloxone

antagonized the antinociceptive action of EO

(highest doses). This suggested the

participation of the opioid system (Campelo et

al., 2011). Essential oil of C. limon may

suppress the growth of Acinetobacter (a

multidrug-resistant) species and could be a

source of metabolites with antibacterial

modifying activity (Guerra et al., 2011). The

essential oil is also a safe and effective

penetration enhancer for topical administration

of lipid- and water-soluble vitamins (especially

vitamin E). Since topical bioavailability of

lipid- and water-soluble vitamins is a critical

issue for protection or anti-ageing formulations.

It might be useful to enhance the permeability

of the skin to different vitamins (Valgimigli et

al., 2012). There is an evidence of sedative and

anxiolytic effects of the essential oil of the

plant that might involve in action on

benzodiazepine-type receptors, and also an

antidepressant effect where noradrenergic and

serotoninergic mechanisms will probably play a

role (C et al., 2011).

[12]Citrus paradise

Description

Grapefruit (C. paradise Macf., family

Rutacaeae) (Fig. 10) is a popular plant

worldwide, not only because of its taste and

nutritive value, but it is also considered to be a

functional food that promotes good health

(Owira & Ojewole, 2010). Grapefruit's peel

flour contains high levels of ascorbic acid and

carotenoid. It is also a good source of dietary

fiber and phenolic compounds. Possessing all

these properties it could be useful in the

formulations of functional food, taking

advantage of the presence of dietary fiber and

antioxidant compounds in one ingredient

(Rincon et al., 2005).




It was found in different studies that this

plant has got significant cardiovascular effects.

In an in vitro study C. paradise peel extract

decreased coronary vascular resistance and

mean arterial pressure in dog’s isolated heart.

This effect of the plant was blocked when the

isolated hearts were pre-treated with L-NAME.

Cardiovascular effects of this plant have also

been shown in human cases. C. paradise juice

decreases diastolic and systolic arterial pressure

both in normotensive and hypertensive

participants (Diaz-Juarez et al., 2009). Alcohol

decoction of C. paradise seed is reputed for the

local management of array of human diseases

including, anemia diabetes mellitus and

obesity. The results of various in vitro studies

also support these effects of C. paradise. For

example, in a study on alloxan-induced diabetic

rats, oral treatment with 100 - 600 mg/kg/day

of methanolic extract of the plant, for 30 days,

resulted in significant (p < 0.05, p < 0.01, p <

0.001) reductions in FPG, TG, TC, LDL-c

VLDL-c in the diabetic rats, effects which were

comparable to that of metformin. The extract

also caused significant (p < 0.05, p < 0.01) rise

in HDL-c values in the alloxan diabetic rats

(Adeneye, 2008). Oral treatment with the seeds

of C. paradise for two weeks (5 to 6 seeds

every 8 hours) significantly reduced the profuse

growth of the bacteria in patients with urinary

infections. The isolated bacterial species from

the blood samples of the patients include

Pseudomonas aeruginosa, Klebsiella species,

Staphylococcus aureus, and Escherichia.

Moreover, ingestion of the seeds of the plant

altered the antibiotic resistance of P.

aeruginosa in one of the patients (Oyelami et

al., 2005). Glyceric extract of the seeds of C.

paradise is also shown to have good

antioxidant activity specially when utilized as

aqueous solutions (Giamperi, Fraternale,

Bucchini, & Ricci, 2004).

[13] Citrus reticulata

Description

C. reticulata (Tanegrine) (Fig. 11) family Rutaceae is well known for its various pharmacological effects (Zhou et al., 2012).


Since Tangerine's peel is rich in

magnesium, carotenoid and extractable

polyphenols, it may be suitable, to reduce risk

of some diseases such as cardiovascular and

some associated to lipid oxidation (Rincon,

Vasquez, & Padilla, 2005). The hexane and

chloroform extracts of C. reticulate (specially

the alcohol-soluble fraction) are shown to have

significant antibacterial activity against both

gram positive and gram negative bacteria. The

antibacterial effect of the plant is said to be due

to the presence of three polymethoxylated

flavones, namely desmethylnobiletin, nobiletin

and tangeretin (Jayaprakasha et al., 2000). It

was shown in a study that the essential oil of C.

reticulate has inhibitory activity on

proliferation of human embryonic lung

fibroblasts. It also showed preventive effects on

bleomycininduced pulmonary fibrosis in rats.

The mechanism may be via adjusting the

unbalance of oxidation and antioxidation,

down-regulating CTGF protein and mRNA

expressions and reducing collagen deposition

and fibrosis (Zhou et al., 2012). This plant is

also reported to have anaphylactic effects. The

lipid transfer protein allergen from mandarin

fruit was isolated and recognized as the cause

to an IgE-mediated allergy in a patient with

anaphylaxis from mandarin (Ebo et al., 2007).

[14] Citrus sinensis

Description

C. sinensis (family Rutaceae) (Fig. 12) is

indigenous to Asia and is cultivated in the

Mediterranean and other subtropical regions in

many parts of the world. The fragrant flowers

are arranged single or in short, limp racemes.

The medicinal parts of the plant are the fresh

and dried peel as well as the oil extracted from

the peel. Flavonoids are the main chemical

compounds of this plant (PDR for Herbal

Medicines, 2000).


Proximate analysis of sweet orange (C.

sinensis) seed flour showed a composition of

54.2% fat, 28.5% carbohydrate, 5.5% crude



fiber, 3.1% crude protein and 2.5% ash for the

dehulled orange seed flour (dry weight)

(Akpata & Akubor, 1999). The peel extract of

C. sinensis dose-dependently inhibited

H(2)O(2)-induced lipid peroxidation in red

blood cells of rats, in an in vitro study. In the in

vivo investigation 25 mg/kg C of the extract

maximally inhibited hepatic LPO. Besides this

inhibitory effect, the extract exhibited

antithyroidal, hypoglycemic, and insulin

stimulatory properties, which suggest its

potential to ameliorate both hyperthyroidism

and diabetes mellitus (Parmar & Kar, 2008).

Sweet orange is also said to have insecticidal

activity. One study showed significant activity

of C. sinensis essential oil against larvae and

pupae of housefly (Muscadomestica) (Kumar,

Mishra, Malik, & Satya, 2012). It also

promotes gastric juice secretion (PDR for

Herbal Medicines, 2000).

[15] Citrus unshiu

Description

C. unshiu (Fig. 13) known as Satsuma

Mandarin belongs to the Rutaceae family and is

well known all over the world especially in

Japan. It has been long used as a traditional

medicine in eastern Asian countries (Ozaki et

al., 2000).


This plant is a good source of antioxidants

(Ma et al., 2008). In an study on antitumor

activity of the extract of C. unshiu in tumor-

bearing murine models. It was shown that this

plant possesses significant antitumor activity

probably via a mechanism of boosting

cytokines such as IFN-γ and TNF-α and

enhancing immune-mediated anti-tumor

properties (Lee et al., 2011). C. unshiu has anti-

allergic activity. It was shown in a study that

fifty percent methanol extract of C. unshiu

powder showed potent inhibitory activity

against histamine release from basophils of

patients suffering from seasonal allergic rhinitis

to ceder pollen. The most potent flavonoid

responsible for this effect of the plant was

hesperetin. Suppression of IgE-mediated

stimulation of basophils through PI3-K

pathway was assumed as the possible

mechanism by which flavonoids inhibited the

degranulation process (Kobayashi and Tanabe,

2006). It has been also shown that water and

ethyl acetate extracts of C. unshiu peel

decreases hepatitis C virus absorption in

MOLT-4 cells. The active ingredient that

markedly inhibited HCV infection was

nobiletin (Suzuki et al., 2005).

Figure 9. Citrus limon Figure 10. Citrus paradise



Figure 11. Citrus reticulata Figure 12. Citrus sinensis

Figure 13. Citrus unshiu

CONCLUSION

Present review study reveals that the

mentioned species possess important

pharmacological effects of the current interest

in pharmaceutical sciences. Since they can

grow in the climate of Iran, they have the

potential to be further studied and be employed

for their various medicinal and

pharmacological properties in Iran and

elsewhere.

ACKNOWLEDGMENT

Hereby we express our gratitude to Mr. Ali

Aghajanshakeri whose help and guidance made

the preparation of this manuscript possible.



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ETHNO MEDICINAL PRACTICES AMONG THE BINJHWAR TRIBE OF

CHHATTISGARH, INDIA

Shukla Rajesh1*, Chakravarty Moyna

2, Goutam M P

3

1Senior Executive Technical - Head, Chhattisgarh State Minor Forest Produce (T&D) Cooperative

Federation Limited, A 25 VIP Estate Khamardeeh, Shankar nagar, Raipur - 429007 C.G., India 2Reader, School of Studies in Anthropology, Pt. Ravishankar Shukla University, Raipur (C.G.)., India

3Retd. Director, State Forensic Science Laboratory, Penshion bada Raipur (C.G.), India

*Corresponding Author: Email- [email protected], [email protected]


ABSTRACT

The present paper deals with the Ethno medicinal practices among the Binjhwar tribe of Raipur

division of Chhattisgarh State, India. Objective of the present paper was to document the oral

tradition of medicine, mode of treatment, awareness towards modern medicinal system and toxic

effect of herbs used by the Binjhwar tribe. The information were collected by using various

anthropological techniques of data collection. The most frequently occurring diseases among the

tribe were skin infections, fever, eye infection etc. which is being treated by using medicinal plants

as well as magico-religious performances. Traditional healers use herbs, shrubs, climbers, trees,

animal remains and minerals for the preparations of medicines for common ailments.

KEYWORDS: Traditional medicine, Magico-religious practices, Binjhwar, Chhattisgarh

Review article

Cite this article:

Shukla Rajesh, Chakravarty. M., Goutam.M. P., (2013), ETHNO MEDICINAL PRACTICES

AMONG THE BINJHWAR TRIBE OF CHHATTISGARH, INDIA,






INTRODUCTION

The tribal of India have preserved a huge

knowledge of traditional medicinal uses of

plants growing around them. Since the vedic

times, importance of medicinal plants always

has the same graph. Medicinal plants play a

vital role in traditional as well as modern health

care system. Ethno medicine (traditional) is the

mother of all other system of medicines. Ethno

medicinal study consists of evaluation of health

outcomes and results from the exercise of

traditional beliefs and behaviors. Traditional

herbal medicine is practiced in several parts of

the world especially in Australia, Africa,

Bangladesh, Brazil, China, Caribbean states,

Europe, Spain; North and South America,

Russia, Pacific islands where large ethnic

communities still live. The World Health

Organization (WHO), 1978 has estimated that

80% of the populations of developing countries

rely on traditional medicines, mostly plant

drugs, for their primary health care needs. In

India 65 % of the population relies on ethno

medicine which is the only source of their

primary health care needs (Rajasekharan et al.,

1996).. The Indian traditional medicine can be

categorized into two streams; I. The Classical

Health Traditions like Ayurveda and Siddha

which are highly organized, classified and

codified and has a sophisticated conceptual and

theoretical foundations and philosophical

explanations; II. The oral health tradition which

is very rich and diverse, but not codified or

organized (Rajasekharan et al.,1996). The

present study on Ethno medicinal practices

among the Binjhwar tribe, conceptually comes

under the second stream of Indian Traditional

Medicine. Some sporadic studies are available

on the Indian tribes such as; the Abors (Dunbar,

1915), Folk medicine of Bastar has been

carried out by (Hemadri et al., 1975), The

Vaidus of Maharasthra (Kurian et al., 1980),

Oraon and Korwa tribes of Sarguja and Raigarh

district, Madhya Pradesh in central India was

carried out by (Maheshwari et al., 1990),

Khairwars of Siddhi district Madhya Pradesh

(Pandey et al., 1999), The Birhors of Madhya

Pradesh (Pandey et al., 2000), The Munda of

Bangladesh (Sharmeen, 2005), Traditional

Health Practices of Raj-Gond (Shukla et al.,

2006), Health seeking behaviour among

Santhal of Orissa (Sonowal et al., 2007),

Indigenous medicine for Gynecological

Disorders by the tribal of Chhattisgarh (Shukla

et al., 2008).

Binjhwar is a civilized Dravidian tribe

found in Raipur and Mahasamund district of

Chhattisgarh and adjoining Orissa state. They

are landholding sections of Baigas, like Raj-

Gonds among the Gonds. Binjhwar is derived

from the Vindhya hills; the tribes still worship

the goddess Vindhyabasini. They have four

sub-divisions; Sonjharas, Birjhias, Binjhias and

Binjhwar proper. Binjhwar have separate Jyaati

Panchayat headed by Jyaati Panchayat

President at Rajadeori, governed by number of

rules to solve disputes and problems within the

community. The total population of Binjhwar

in the state ranges from 1,00,692 to 1,04,718

(Naik, 1972).

The present study aims to document the

oral traditions of tribal health, attitude towards

modern or allopathic medical system, identify

and document plants, animals and minerals

used for medicinal purposes and identify the

plants having toxic and harmful effects used as

folk medicine.


For the present study survey was conducted

in 13 tribal villages during September 2003 to

January 2005. During the course of the study

regular field visits were carried out in the study

area. Various methods of sampling were used

for area selection and primary data collection.

Purposive sampling method was used for

village selection from Kasdol, Sankra and

Pithora blocks of Raipur and Mahasamund

District of Chhattisgarh, India. Binjhwar

dominating villages of three blocks were

selected for the present study and 275

households were surveyed randomly. Interview

schedule was used for household survey to

collect information related with education, food

habit, health, occupation and social structure of

Binjhwars. Information about the use of



medicinal plants, mode of administration,

dosage and technique of diagnosing the

diseases were collected through interview from

the traditional healers (Baiga, Vaidhraj and

priests). Secondary data were collected from

journals, books, reports and government offices

to verify the health infrastructure facilities

provided by the government.

RESULTS

Ethno-medicinal practice among the

Binjhwar is complex containing different

treatment patterns i.e. herbal medicine, rituals,

magico-religious treatment and allopathic

medicines. They have their own concepts for a

disease, cause of illness, diagnosis of disease

and treatment of ailments.

The most frequently occurring disease

among the Binjhwar is cough and cold i.e.

21.82%, skin disease 14.91 %, fever and eye

infection 10.55 % and tuberculosis and stone

(0.73 %) each and other diseases like jaundice,

dysentery etc were reported. shown in fig. 01.

Specialists like herbalists, bone setters, mid

wives and pujari / priest are ethno-medicinal

service providers among the Binjhwars.

Traditional healers are expert in diagnosing the

disease by calculating pulse from various body

parts, observation of eye colour, tongue and

neck. Local traditional healers may be

categorized into two types according to their

nature of work; first, the people performing

magico-religious acts for diagnosing the

disease who are called pujari and second, use

organoleptic observation and also they may or

may not use some magical formulae for

diagnosing the disease.

Treatment among Binjhwar consists of

herbal remedies, magico-religious acts and

modern medical facilities available nearby. A

variety of medical specialists co-exists whose

services may be availed by the Binjhwar. These

healers are expert in treating various ailments

like fever, tuberculosis, asthma, jaundice,

gynaecological disorder and so on by using

different plant species given in Table 01. The

healers collect various parts like root, tuber,

rhizome, stem, bark, leaf, flower, seed and

whole plant for preparation of medicine.

Healers use 34 % roots, 13 % seeds and 12 %

barks and some animal remains or body parts

and minerals in trace quantity i.e. 05 % in

addition to herbs. For preparing of the

medicines, 52 Plant species belonging to 35

families are used by Binjhwars. Some species

are used in multiple medicinal preparations,

Keo-kand (Costus speciosus) in 7 ailments,

Arjun (Terminalia arjuna) in 5 ailments,

Mahua (Madhuca latifolia) in 6 ailments,

Dhaora (Anogeissus latifolia) in 3 ailments and

tendu (Diospyros melanoxylon) in 2 ailments.

Besides these wild plants several spices are also

used by them such as; Elaichi, Bade-elaichi,

Fenu greek leaves (Methi), Black pepper,

Garlic etc. are also mixed in trace quantities.

Fig 1. Person Suffering from Disease in One Calendar Year

21.82

10.55

3.27

10.91

2.181.09

9.45

6.91

0.73

10.55

6.91

14.91

0.730

5

10

15

20

25

Cough and cold

Fever

Indigesti

on / Acid

ity

Join

t Pain

Diarrh

oea

Dysente

ry

Gyneco

logic

al diso

rder

Jaundice

Tuberculosis

Eye Infe

ction

Stom

ach ach

e

Skin D

isease

Stone

Perc

en

tag

e



Table 1. Botanical Name, Family and Uses of Plant Species used by Binjhwar Tribes

Sl. Plant Name Botanical Name Family Disease

1 Adusa Adhatoda vasica

Nees.

Acanthaceae Itching, Fever

2 Agusti Sesbania grandiflora

Pers.

Fabaceae Loose Motion

3 Akarkara Spilanthes acmella Asteraceae Paralysis, Tooth ache, Epilepsy

4 Amaltas Cassia Fistula Linn. Caesalpiniaceae Tuberculosis

5 Amarbel Cuscuta reflexa Roxb. Convolvulaceae Jaundice

6 Ami haldi Curcuma amada

Roxb.

Zingiberaceae Painful menses / Blood discharge

7 Arjun, Kahua Terminalia arjuna Combertaceae Acidity, Weakness, Chest pain,

Arthritis, Diabetes

8 Bach,

Devnasan

Acorus calamus Linn. Araceae Epilepsy, Joint Pain

9 Ban tulsi Ocimum gratissimum

Linn.

Lamiaceae Joint Pain

10 Banrakash

(Doodh

Kochai)

Alocasia indica

(Roxb.) Schott.

Araceae Arthritis (Baat)

11 Behra Terminalia bellirica

Roxb.

Combertaceae Acidity, Loose Motion, Piles

12 Bhelwa Semicarpus

anacardium Linn.

Anacardiaceae Tuberculosis

13 Bhui aonla Phyllanthus niruri Euphorbiaceae Diarrhoea

14 Bhui neem Andrographis

paniculata Nees.

Acanthaceae Fever and Malaria

15 Chitawar Plumbago zeylanica

Linn.

Plumbaginaceae Moti jhira, Itching

16 Dahiman Cordia macloidii Boraginaceae Snake Bite

17 Dhaora Anogissus latifolia

Wall.

Combertaceae Diabetes, Dysentery, Loose

Motion

18 Doodhi Euphorbia hirta Euphorbiaceae Adequate milk Secretion

19 Girola Indigofera cassioides Fabaceae Vitality after Delivery

20 Gular Ficus racemosa Moraceae Diabetes

21 Gurmar Gymnema sylvestre R.

Br.

Asclepiadaceae Diabetes

22 Gursukhri Grewia hirsute Vahli. Asclepiadaceae Fracture, Wound

23 Gurud Stereospermum

suaveolens DC.

Bignoniaceae Fever and Malaria

24 Harra Terminalia chebula

Retz.

Combertaceae Acidity, Piles

25 Harsinghar Nyctanthes arbor-

tristis Linn.

Oleaceae Fracture

26 Hinglaj Balanites aegyptiaca Balanitaceae Typhoid



27 Kalimusli Curculigo orchrides

Gaertn.

Hypoxidaceae Promote sexual desire and

strengthening

28 Karanj Pongamia pinnata

Pierre.

Fabaceae Itching, Dog bite

29 Kaya Strychnos potatorum

Linn.

Loganiaceae Eye Infection

30 Keo-kand Costus speciosus

(Koeing.) Sm.

Zingiberaceae Indigestion, Arthritis, Joint Pain,

Head-ache, Strengthening,

Leucorrhoea, Piles

31 Kulthi Dolichos biflorus Fabaceae Body ache, Stone

32 Kurma Leea aspera Leeaceae Chest pain / Swelling

33 Kusum Schleichera oleosa

(Lour) Oken.

Sapinadaceae Wound

34 Lajwanti Mimosa pudica Linn. Mimosaceae Eye Infection and redness, Fever

35 Maha neem Melia azedarach Linn. Meliaceae Fever, Jaundice

36 Mahua Madhuca latifolia Sapotaceae Arthritis, Dysentery, Jaundice,

Piles, Snake Bite

37 Maida Litsea glutinosa

(Lour.) C. B. Robins

Lauraceae Fracture

38 Panarijadi Aristolochia indica

Linn.

Aristolochiaceae Piles

39 Pat koria

(Patha)

Cissampelos pareira

Linn.

Menispermaceae Fever and Malaria

40 Patal Kumrah,

Bhui tumba

Pueraria tuberosa

DC.

Fabaceae Bemchi, Weakness

41 Pipli Piper longum Linn. Piperaceae Tuberculosis, Asthma

42 Ramdatoon Smilex zeylanica Liliaceae Leucorrhoea,White discharge

43 Rasna Blepharispermum

subsessile

Asteraceae Arthritis (Baat)

44 Saan Crotalaria orixensis Fabaceae Jaundice

45 Sarphoonka Tephrosia purpurea

Pers.

Fabaceae Stone, Jaundice

46 Semhar Samalia malabarica

Schott & Endl.

Sterculiaceae Irregular Menses

47 Tejraj Peucedanum

nagpurense

Apiaceae Promote sexual desire, Male

Impotency

48 Tendu Diospyros

melanoxylon

Ebenaceae Painful menses / Blood discharge

49 Thelkajari Alangium salviifolium

(Linn.f.) Wang.

Alangiaceae Male Impotency

51 Tilai Wendlandia heynei Asteraceae Piles, Body ache

52 Tinsa Ougeinia oojeinensis

(Roxb.) Hochr.

Fabaceae Dysentery, Loose Motion



Fig 2. First Preference for the Health Care needs

Binjhwar first approach the traditional medicine men for primary health care i.e. 47.64 % followed by 25.82 % people approach either traditional or allopathic medicine and only 10.18 % people access allopathic treatment for primary health care. The reasons for non utilization of modern health care system are low literacy, poor economic status, and lack of awareness, cultural factors and distance of health centers, doctor - patient relationship and health facilities available from the administrative side.

CONCLUSION

It can be concluded that the Ethno-medicinal practices of the Binjhwars are more affordable, acceptable, culturally appropriate and available as compared to modern medicines. Even today Binjhwars accept traditional medicines in remote or inaccessible villages due to their dependence on and confidence in traditional medicine men. Therefore, proper education and mass communication might be helpful in creating an awareness of the Binjhwars towards modern health care system. The Binjhwars have a pluralistic medical situation in which allopathic, ayurvedic, homoeopathic and

indigenous system of medicine exist side by side.

In the present study some plants have been selected which have ethno botanical significance. During the course of field work the therapeutic and toxic effects on the users were recorded. It could be concluded that users of mentioned plants for treatment of various kinds are not aware of their toxicity and toxic chemical constituents. Terminalia bellarica Lam., Phyllanthus niruri Lam., Melia azedarach L., Madhuca latifolia Koenig., Plumbago zeylanica L., Semecarpus anacardium L. were considered to know their toxic effects.

Community based awareness programme should be organized to protect this community with the over dosage, accidental poisoning and chance contamination of these drugs. Local government officers should also establish a team of subject experts including local vaidhya, medical practitioners, botanist and anthropologist so that they can prepare a list of such plants giving details regarding their vernacular names, botanical names, toxicity of the particular plant part, method of reducing toxic effect (Sodhan) and dosages. Authors would recommend that a bridge should be

47.64

25.82

9.453.27 3.64

10.18

0

10

20

30

40

50

60

Traditi

onal Tre

atm

ent

Traditi

onal / A

llopat

hic

Jhad-p

hook/Allo

pathic

Jhad-p

hook

Allopath

ic/Tra

ditional

Allopath

ic

perc

en

tag

e



developed between Binjhwars traditional medicine and Modern medical system, which will help us to protect and conserve the traditional medical heritage as well as improve the utilization of modern medical facilities. Phyto-chemical or pharmacological investigation, nutritional analysis and clinical trials should be carried out to validate the claims. These informations may help the policy

makers for adopting the proper healthcare measures and may provide a lead in the development of new drugs.

Note: This paper is presented in the International Conferences on “Conservation, Marketing and Patenting of Medicinal Plants” organized by Chhattisgarh State Medicinal Plant Board Raipur, India. 14–15 March 2010.

REFERENCES

Dunbar D S G (1915). Abors and Garllongs. The Asiatic society, Culcutta, 5: 1–19.

Hemadri K, Rao S S (1975). Folk medicine of Bastar. Journal of Ethnobotany, 1: 61–66.

Kurian Joy C, B V Bhanu (1980). Ethnomedicine: A Study of the Nomadic Vaidus of Maharastra. The Eastern Anthropologist, 33(1): 71–78.

Maheshawari J K, Painuli R M, Dwivedi R P (1990). Notes on Ethnobotany of Oraon and Korwa Tribes of M.P. Contribution to Indian Ethnobotany: 75–90.

Naik, T. B. (1972): “Barah Bhai Binjhwar”: Madhya Pradesh Hindi Granth Academy, Bhopal (M.P.), India.

Pandey G D, Roy J (1999). A Study of Health Seeking Behavior among the Khairwars and Non-Khairwars of Sidhi District. Tribal Health Bulletin, 5(1): 11–16.

Pandey G D, Tirkey V R, Tiwary R S (2000). Some aspects of Health Seeking Behavior in Birhors - A Primitive Tribe of M.P. Man in India, 79(3&4): 291–299.

Rajasekharan S, Pushpangadan P,Biju S D (1996). Folk Medicines of Kerala – A Study on Native Traditional Folk Healing Art and its Practitioners. New Delhi: Deep Publications, New Delhi (ed. S. K. Jain).

Sharmeen S, Sharmeen T (2005). Traditional Health Practices of Munda Women: Continuity and Change. Man and life, 31 (1&2): 23–36.

Shukla R, Chakravarty M (2006). Anthropological Study on Traditional Health Practices of Raj-Gonds, Tribal Health Bulletin, Vol.12 (1&2), 2006: 61–66.

Shukla R, Chakravarty M, Gautam M P (2008). Indigenous medicine used for the treatment of Gynecological Disorders by the tribal of Chhattisgarh, India. Journal of Medicinal Plant Research, 2 (12): 356–360.

Sonowal C J, Praharaj P (2007). Tradition Vs Transition: Acceptance of Health Care Systems among the Santhals of Orissa. Studies on Ethno-medicine, 1(2):135–146.

World Health Organization (1978): “The Promotion and Development of Traditional Medicine”: Technical Report Series 622, Geneva.





EVALUATION OF ANTIARTHRITIC ACTIVITY OF LEPIDIUM SATIVUM LINN SEEDS

AGAINST FREUND’S ADJUVANT INDUCED ARTHRITIS IN RATS

Raval Nita D1*, Ashok B K

2, Ravishankar B

3

1Lecturer, Department of Dravyaguna, Government Ayurved College, Junagadh,

2Drug discovery lab, R & D, Himalaya drug company, Bangalore, Karnataka, India

3Director, SDM Research Centre for Ayurveda and Allied Sciences, Kuthpady, Udupi. Karnataka – 574118,

India

*Corresponding Author: Email: [email protected]; Mobile: +919898340450


ABSTRACT

Garden cress (Lepidium sativum Linn.) leaves and seeds are used in India as food supplement and

also as medicine. In traditional system of medicine, its seeds were used for various ailments like

inflammation, joint pain, backache etc. the present study was carried out to explore the folkloric use

of the seeds on Freund's adjuvant induced arthritic rats. The present study concludes the effect of the

seed powder on the Freund's complete adjuvant induced arthritic rat paw oedema in both developing

and developed phases of arthritis. Histopathology of proximal inter-phalangeal joints and radiology

of hind legs were studied. Significant changes were observed in drug treated group in radiological

study. In histo-pathological study bone and cartilage degeneration with bone erosion and

inflammatory changes were observed in the affected joint. These changes were not significantly

improved by the administration of test drugs. Result data suggested that at a lower dose, moderate

anti-arthritic effect was produced by the test drug while at a higher dose it had a tendency to produce

inconsistent effect.

KEY WORDS: anti arthritic, Fruend’s adjuvant, paw oedema, Lepidium sativum Linn.

Research article

Cite this article:

Raval Nita. D., Ashok. B.K,, Ravishankar. B., (2013) EVALUATION OF ANTIARTHRITIC

ACTIVITY OF LEPIDIUM SATIVUM LINN SEEDS AGAINST FREUND’S ADJUVANT

INDUCED ARTHRITIS IN RATS, Global J Res. Med. Plants & Indigen. Med.,

Volume 2(7): 532–537




INTRODUCTION:

Rheumatoid arthritis (RA) is a systemic

autoimmune disease of unknown aetiology.

The disease is characterized by articular

inflammation and by the formation of an

inflammatory and invasive tissue, rheumatoid

pannus that eventually leads to the destruction

of joints (Stephen J. 2008). Analgesics and

NSAIDs are used to suppress the symptoms

while disease-modifying anti-rheumatic drugs

(DMARDs) and newer therapies such as anti-

tumour necrosis factor (TNF)-α therapy

(etanercept, infliximab and adalimumab), anti-

CD20 therapy (rituximab) and abatacept are

often required to inhibit or halt the underlying

immune process (Anthony S. Fauci. et al.,

2008). NSAIDs are widely prescribed all over

the world to the patients with Rheumatoid

arthritis, Osteoarthritis etc. However the

prolonged use of these drugs has its own

drawbacks. Research works on NSAIDs

suggest that these drugs can increase the risk of

chronic renal disease (P Ejaz et al., 2004).

Garden cress is also known as Asalio or

Chandrashura in local languages and it is an

important medicinal crop in India (Tiwari et al,

2004). Garden cress (Lepidium sativum Linn.)

is an annual fast growing edible herb, belongs

to the family Brassicaceae. Seeds, leaves and

roots of it are of economic importance;

however, the crop is mainly cultivated for

seeds. It can be grown at all elevations,

throughout the year, but the best crop is

obtained in the winter season (Anonymous,

1962, CSIR, New Delhi.). Garden cress seeds

and leaves are used in food preparations (Datta

PK et al., 2011). Leaves are diuretic and gently

stimulant (Maghrani et al., 2005). To

investigate the folkloric use of the seeds the

present study was carried out on Freund’s

adjuvant induced arthritic rats.

MATERIALS AND METHODS:

Collection of sample (Lepidium sativum Linn

seeds)

The seeds for the proposed study were

collected from the campus of Gujarat Ayurved

University, Jamnagar in the month of April

2004 and it was authenticated by the expert of

botany from the Department of

Pharmacognosy, Gujarat Ayurved University,

Jamnagar, Gujarat, India

Experimental models:

Twenty four Charles Foster strain albino

rats weighing 180 ± 10g were obtained from

animal house attached to Pharmacology

laboratory, Gujarat Ayurved University,

Jamnagar Gujarat. Six rats were housed in each

cage made up of poly-propylene with stainless

steel top grill. The dry wheat (post hulled)

waste was used as bedding material and was

changed every morning. The animals were

acclimatized for seven days before

commencement of the experiment in standard

laboratory conditions 12 ± 01 hour day and

night rhythm, maintained at 25 ± 3oC and 50–

70% humidity as per CPCSEA guidelines.

Animals were provided with balanced food

(Amrut brand rat pellet feed supplied by Pranav

Agro Mills Pvt. Limited) and water ad libitum.

Protocol used in this study for the use of

animals was approved by the institutional

animal ethics committee (IAEC 04-

05/01/PhD.03).

Dose selection:

The dose fixation for the experimental

animals was done on the basis of body surface

area ratio by referring to the standard table of

(Paget and Barnes, 1964). The adult human

dose (6 g per day) (Chunekar K.C, 1999) was

converted to animal dose. On this basis the

calculated dose was fixed to be 550 mg/kg for

rats. The suspension of Lepidium seed powder

was made with sufficient quantity of distilled

water according to the required dose and

administered orally with the help of gastric

catheter sleeved to syringe.

Experimental study (Rosenthale, 1970):

The selected animals were grouped into

three groups of 6 rats each. To the first group

only water was administered and treated as

normal control. To second and third group, test

drug was administered in the dose of 550



mg/kg (TED) and 1100 mg/kg (TED × 02)

respectively. The drug was administered for 30

consecutive days. On day one, the complete

Fruend's adjuvant was made into fine emulsion

with the help of a syringe and 0.1 ml of it was

injected beneath the plantar aponeurosis in the

left hind paw and 0.05 ml subcutaneously into

the root of the tail. The volumes of both the

hind paws were measured with the help of

Plethysmometer. The paw volume of left hind

limb was measured at 2nd

, 3rd

, 5th

, 10th

and 15th

days, while of right hind limb on 15th

, 20th

, 25th

and 30th

days. Paw volume of the 0 (initial)

days were taken as the reference value for

determining the increase in paw volume on the

subsequent days. On 31st day animals were

weighed and sacrificed by over dose of ether

anesthesia and both right and left synovial

joints were dissected out and the histo-

pathological slides were prepared by referring

to standard procedure of (Raghuramulu et al.,

1983). The slides were viewed under trinocular

research Carl-Zeiss’s microscope at various

magnifications to note down the changes in the

microscopic features.

Statistical analysis:

The data were expressed as mean ±

standard error mean (SEM). The Significance

of differences among the test drugs and control

groups was assessed using one way analysis of

variance (ANOVA) and the test followed by

Dunnet’s test. The difference was considered

significant when p < 0.05.

RESULTS AND DISCUSSION:

The data pertaining to the effect of test drug

on Friend’s adjuvant induced arthritis in rats

have been tabulated in Table 1 & Table 2

A moderate decrease in both primary and

secondary edema was observed in therapeutic

dose, and double dose group in comparison to

control group. The suppression of primary

edema in therapeutic dose was as follows. After

48 hrs 14%, 5th day 34.32%, 10th day 49.01%,

15th day 45.40%, 20th day 33.21%, 25th day

82.32%, 30th day 33.84%. Whereas marginal

increase was observed after 24 hrs. In double

dose group the suppression of primary edema

was after 24 hrs 15.88%, 48hr 4.96%, 5th day

29.15%, 10th day 32.64%, 15th day 51.82%,

30th day 12.34%. Here the increase was

observed on 20th and 25th day.

In secondary edema the suppression

observed in therapeutic dose group was as

follows on 15th day 40.07%, 20th day 41.21%,

25th day 45.33%, 30th day 40.45%. In double

dose group 15th day 6.92%, 25th day 2.86%,

30th day 28.57% suppression of edema was

observed where as on 20th day marginal

increase was observed in double dose group.

Table – 1: Effect on primary paw oedema (Oedema of left hind paw)

Data: Mean SEM, **ANOVA test (F = 35.45, P < 0.01; df (2, 15) (F = 7.01; P < 0.01 df, (2, 15)

Table – 2: Effect on secondary paw oedema (Oedema of right hind paw)

Groups 15th

day 20th

day 25th

day 30th

day

Control 29.17 7.09 20.6 4.60 23.05 10.04 20.44 7.78

TED 17.48 4.51 12.11 5.72* 12.60 4.71 12.17 5.28

TED×02 31.19 7.43 31.82 7.38* 22.39 8.23 14.60 5.74 Data: Mean SEM, *ANOVA F=3.48; P < 0.10; df (12, 15)

Groups Percentage increase in paw oedema when compared to initial paw volume

2nd

day 48 hrs 5th

day 10th

day 15th

day

Control 68.49 9.30 45.33 7.58 39.86 6.73 20.77 6.40 11.21 2.36

TED 70.55 11.57 38.98 6.43 26.18 5.43 10.59 3.67** 06.12 2.12

TED×02 57.61 6.34 43.08 7.23 28.24 7.67 13.99 3.96** 05.4 2.44



Figure 1: Images depicting Radiology of hind legs in adjuvant induced arthritic rats

Figure 2: Images depicting histopathology of proximal interphalangeal joint in adjuvant

induced arthritic rats (1X 50 magnifications)

Radiology of hind legs in adjuvant induced

arthritic rats:

In arthritic disease radiographic changes are

useful diagnostic measures which indicate the

severity of the disease. In the early stage soft

tissue swelling can be seen, whereas severe

radiological changes like bony erosion and

joint space narrowing can be observed in

developed stage of arthritis. In adjuvant

induced arthritic rats soft tissue swelling and

cartilage erosions were observed. The degrees

of degenerative changes were less in test drug

treated group in comparison to control group.

The radiographic features of the rat joints in

adjuvant induced arthritic rats are shown in

Figure 1.

Effect on histopathology of joints:

Figure 2 shows representative sections of

inter-phalangeal joint from different groups.

Bone and cartilage degeneration with bone

erosion and inflammatory changes were

observed in the knee joint. These changes were

not significantly improved by the

administration of test drugs.



The main aim of the present study was to

find out the scientific basis for the reported

efficacy of test drug in arthritis. Hence it was

evaluated against Fruend's adjuvant induced

arthritis in rats. Injection of adjuvant elicits

immune response of cell-mediated type and

produces an arthritic syndrome. Arthritic rats

show swelling of the soft tissue around ankle

joint during initial phases of arthritis. This is

mainly due to edema of periarticular tissues

such as ligaments and joint capsules

(Sanmugapriya et al., 2010). Like human RA, it

is also characterized by synovitis, infiltration of

neutrophils and proliferative response leading

to synovial fibroplasias and periosteal bone

deposition (Hazeena et al., 1980).

In the present study a persistent moderate

suppression of primary oedema ranging

between 14 to 49% was observed at lower dose

up to 20th

day of observation. The secondary

oedema represents oedema of immunological

origin (Yoshikawa et al., 1985). In the

secondary oedema a consistent 40 to 45%

suppression was observed at lower dose

indicating that the test drug at this dose has

moderate suppression effect on oedema of

immunological origin. At higher dose

inconsistent effect was observed. The

suppression ranged between 5.74% to 54%.

Radiographic changes in RA conditions are

useful diagnostic measures which indicate the

severity of the disease. Soft tissue swelling is

the earlier radiographic sign, whereas

prominent radiographic changes like bony

erosions and narrowing of joint spaces can be

observed only in the developed stages (final

stages) of arthritis (Harris ED, 1990). The

radiographic features of the rat joints in

adjuvant induced arthritic model are shown in

plate 2. The degrees of degenerative changes

were less in test drug treated group in

comparison to control group.

CONCLUSION

At lower dose level moderate anti-arthritic

activity was observed in FA rats. Higher dose

level produced inconsistent effect. Hence it

would be necessary to identify suitable dose.

This indicates the complex nature of the seed

powder. The study confirms the anti-

inflammatory and anti-arthritic activity

potential of the plant material however; further

refinement in the form in which it is

administered along with optimum dose fixation

is required for optimum therapeutic application

of this plant. It would also be interesting to

study the other parts of the plant. It is possible

that they may also express significant anti-

inflammatory and anti-arthritic activity.

REFERENCES

Anonymous. (1962). The Wealth of India, Raw

Materials. VI. Publication and

information Directorate, CSIR, New

Delhi. India, p.71–73.

Anthony S. Fauci. and Dr. Longo(2008)

Harrison’s Principles of Internal

Medicine, Section 2, 17th

edition, part

14, section 2 chapters 314.

Chunekar K.C., (1999) G.S.Pandey (eds)

Bhavprakash nighantu, haritakyadi

Verga, Chaukhambha orientalia

publication.pp 39.

Datta PK, Diwakar BT, Viswanatha S, Murthy

KN, Naidu KA, (Mar-April 2011).

Safety evaluation studies on Garden

cress Lepidium sativum L. seeds in

Wistar rats. IJARNP-HS Publications

Vol. 4 (1), pp. 37–43.

Dr. Yograj Sharma, (2003–2004). Efficacy of

sowing methods, fertilizer application

and growth regulations in cultivation of

Lepidium sativum Linn. Published

thesis (MSc), Gujarat Ayurved

University, Jamnagar.



Harris ED (1990). Rheumatoid arthritis.

Pathophysiology and implications for

therapy. N Eng J Med, 322(18):1277–

1289.

Hazeena Begum, V., Sadique, J. (1980). Ind.

J.exp. Biol., 26,877.

Maghrani M, Zeggwagh NA, Michel JB.

Eddonks M. (2005). Antihypertensive

effect of Lepidium sativum L. In

spontaneously hypertensive rats. J

Ethnopharmacol 100: 193–197.

P Ejaz, K Bhojani, VR Joshi, (2004). NSAIDs

and kidney, JAPI, vol.52, pp-632.

Paget, G.E., Barnes, J.M. (1969). In Evaluation

of drug activity; Pharmacometrics

(Laurence, D.R., Bacharacha, A.L.)

Vol. 1, Academic Press, New York

Raghuramulu N, Nair KM and

Kalyanasundaram S (1983). A manual

of laboratory techniques, (National

Institute of Nutrition, Hyderabad).pp.

246–53.

Rosenthale, M.E. (1970). Arch. Int.

Pharmacodyn. 188. In: Dahanukar, S.,

Sharma, S., Karandikar, S.M.(1983).

Indian Drugs, 20 (10), 405.

Sanmugapriya Ekambram, Senthamil Selvan

Perumal, and Venkataraman

Subramanian, (2010). Evaluation of

antiarthritic activity of Strychnos

potatorum Linn seeds in Freund’s

adjuvant induced arthritic rat

model.BMC, Complementry and

Alternative medicine, vol.10.

Stephen J. Mc Phee, (2008). Current Medical

diagnosis & treatment, Maxine

papadakis publication. Mc Graw hill

Lange Chapter Musculo skeletal

disorder.

Tiwari PN, Kulmi GS. (2004). Performance of

Chandrasur (Lepidium sativum) under

different levels of nitrogen and

phosphorus. J Med Arom Pl Sci 26:

479–481.

Yoshikawa T, Tanaka H, Kondo M (1985).

The increase in lipid peroxidation in rat

adjuvant arthritis and its inhibition by

Superoxide dismutase. Biochem. Med,

33:320–326.


http://www.biomedcentral.com/sfx_links?ui=1472-6882-10-56&bibl=B20




DEVELOPMENT OF RANDOM AMPLIFIED POLYMORPHIC DNA

MARKERS FOR AUTHENTICATION OF OLAX SCANDENS ROXB.

Naik Raghavendra1*, Borkar Sneha D

2, Acharya R N

3, Harisha C R

4

1,2P G Scholar, Dravyaguna Department, IPGT&RA, Gujarat Ayurved University, Jamnagar, Gujarat, India

3Associate Professor, Dravyaguna Department, IPGT&RA, Gujarat Ayurved University, Jamnagar, Gujarat,

India 4Head, Pharmacognosy Laboratory, IPGT&RA, Gujarat Ayurved University, Jamnagar, Gujarat, India

*Corresponding Author: Email: [email protected]; Mobile: +918866679088


ABSTRACT

Olax scandens Roxb. (Olacaceae), commonly known as “Badru” is a shrub or small tree. Fruits

and leaves of this plant are used for medicinal and food purpose. The present study deals with the

pharmacognostical study of stem and molecular characterization of young leaves by random

amplified polymorphic DNA markers. Microscopic study of stem shows the presence of single layer

of epidermis with unicellular uniseriate, horn shaped trichomes, single layered, radially arranged

hypodermis, 6–7 layers of cortex, uniseriate to triseriate medullary rays, parenchyma cells with oil

globules, prismatic and rhomboidal crystals. All the primers gave good band patterns. Prominent

band of ~0.4 kb was obtained with OPA-02 and ~0.9 Kb prominent band was obtained with OPC-06

primer. These microscopic observations and the unique bright and light bands obtained in

polymerase chain reaction (PCR) amplification could serve as a measure for authentication and

standardization of the plant.

KEY WORDS: Olax scandens, Pharmacognosy, RAPD

Research article

Cite this article:

Naik Raghavendra, Borkar Sneha. D, Acharya. R.N., Harisha. C.R., (2013)

DEVELOPMENT OF RANDOM AMPLIFIED POLYMORPHIC

DNA MARKERS FOR AUTHENTICATION OF OLAX SCANDENS ROXB.,





INTRODUCTION

Historical background of traditional

medical systems explains how the herbal drugs

have made a great contribution in maintaining

human health. Even today, in most of the

developing countries a large proportion of the

population relies heavily on traditional

practitioners and medicinal plants to meet their

health care needs (Patel H et al., 2013). Olax

scandens Roxb. (Olacaceae), an ethno

medicinal plant, commonly known as “Badru”,

used for food and medicinal purposes (Rekha

Sinha et al., 2005), (Anonymous, 1996),

decoction of stem bark is used to cure fever and

cough (Veeramuthu et al., 2006), (Kirtikar &

Basu, 2003), leaves applied externally to cure

headache (Anonymous, 1990). Though the

plant has been reported for many biological

activities, till now no detailed study has been

reported regarding its pharmacognostical

characters except on leaves (Naik R. et al.,

2013). Documentation and standardization of

such biological resources is important for their

identification, authentication and utilization.

Identification of biological resources through

their molecular characterization is one of the

authentic methods of their standardization.

Recently certain plants have been reported for

their molecular characterization through

Random amplified polymorphic DNA markers

(Borkar S D et al., 2013). Hence the present

study was undertaken to study the microscopic

characters of stem and establish the molecular

characterization of Olax scandens Roxb. by

random amplified polymorphic DNA markers.


Collection and preservation of the sample

Olax scandens Roxb. was collected from its

natural habitat Balangir, Odisha, during

September 2012 identified and authenticated by

local taxonomist with the help of botanical

flora (Saxena H.O, 1995). A sample specimen

was preserved in Pharmacognosy lab of IPGT

& RA Jamnagar (SPECIMEN NO- PHM

6062/21/09/2012) and the stem was preserved

in a solution prepared from 70% ethyl alcohol:

glacial acetic acid: formalin (AAF) in the ratio

of 90:5:5 (Johnson Alexander Donald, 1940).

Pharmacognostic studies

Detailed microscopic characters were

studied by taking free hand thin transverse

section. Sections were stained with

Phloroglucinol and Hydrochloric acid to notice

the lignified elements like fibers, vessels etc

(Khandelwal K R, 2008). Photographs of the

section were taken with the help of Canon

digital camera attached to Zeiss microscope.

Powder characters were studied as per the

guidelines of Ayurvedic Pharmacopoeia of

India (Anonymous, 1999). The histo-chemical

tests were carried out according to the standard

guidelines of practical pharmacognosy

(Krishnamurty K V, 1988).

Molecular characterization (DNA

fingerprints)

Fresh leaves were used for molecular

characterization and DNA fingerprints, by

standard and most convenient RAPD method

(Baum BR, Mechanda S, 2001). The RAPD

reaction was performed following standard

procedures at Aristogene Biosciences Pvt. Ltd,

Bangalore.

DNA isolation:

Young leaves were selected, cut into small

pieces without cutting the veins. They were

washed with distilled water and ethanol. Frozen

with dry ice and crushed. To that, 2 ml of plant

DNA extraction buffer was added. The samples

were ground thoroughly, transferred into

centrifuge tube and added 10 ml plant DNA

extraction buffer. 50 μl of BME added to each

tube mixed well. Incubated at 65ºC for 1 hour

with intermittent mixing and centrifuged for 15

minutes at 10 K (10000 rpm). Supernatant was

transferred carefully into fresh tube and added

equal volume of chloroform and mixed well.

Centrifuged for 15 minutes at 10 K (10000

rpm). Aqueous layer was carefully pipetted into

fresh tube and precipitated with isopropanol.

DNA pellet suspended in 300 μl of TE and

subjected to column purification.



Column purification

Silica spin columns and buffers were from

Qiagen

The column was placed in collection tube,

400 μl of equilibration buffer was added to the

column and centrifuged at 10000 rpm for 1min.

Collected buffer was discarded. 400 μl of

equilibration buffer was added to the DNA

samples, mixed and loaded into the column

(This step was repeated till the DNA sample

was completed). Flow through was collected.

500 μl of wash buffer 1 was added, centrifuged

at 10000 rpm for 1minute and buffer was

collected. 500 μl of wash buffer 2 was added,

centrifuged at 10000 rpm for 1minute and

buffer was collected. The empty column was

centrifuged with collection tube to completely

remove the wash buffer for 2 minute. 50 μl of

elution buffer was added to the column placed

in new collection tube. Incubated at room

temperature for 2 minutes and centrifuge at

10000 rpm for 1minute and eluted sample was

saved (elution 1). Previous step was repeated

(elution 2). Quantization of eluted DNA

samples was done by loading into the agars gel

(Table 1–2).

PCR Conditions

38 µl of amplified DNA was aliquot into 2

different labeled vials and to this 2 µl of

respective template DNA was added and PCR

was set. One cycle given At 94˚C for 2

minutes, again 40 cycles given at 94˚C, 45˚C

and 72˚C for 30 sec, 60 sec. and 90 sec.

respectively and finally one cycle given at 72˚C

for 7 min (Table 3).

RAPD PCR

Table 1: Sequences of primers used

OPA-02 TGCCGAGCTG

OPB-10 CTGCTGGGAC

OPC-06 GAACGGACTC

Table 2: Reactions were set up with PCR master mix and respective Random primer

Table 3: PCR Conditions.

OPA-02 OPC-06 OPB-10 Notes

Double distilled water 18 µl 18 µl 18 µl

2X PCR master mix 20 µl 20 µl 20 µl 1X Contains 100µM each of dATP,

dGTP, dCTP and dTTP. Assay

buffer with 15mM MgCl2,

3U/reaction Taq Polymerase.

DNA sample 1 µl 1 µl 1 µl 10 pM used for each reaction

Random primer 1 µl 1 µl 1 µl

Total volume 40 µl 40 µl 40 µl

Temperature Time No. of cycles

94˚C 2 minutes 1

94˚C 30 seconds 40

45˚C 1 minute

72˚C 1min 30 sec

72˚C 7 minutes 1



RESULTS AND DISCUSSION

Microscopic examination of stem

T. S of the young stem was circular in

outline with uneven surface and showed the

following characters (Plate A1).

Epidermis

T.S showed outer most, single layered,

compactly arranged epidermis with somewhat

barrel shaped cells covered with cuticle without

any intercellular spaces (Plate A2). Epidermal

cells were interrupted by unicellular horn

shaped trichomes (Plate A3). Beneath the

epidermis, compactly arranged, single layered

angular cells, forming hypodermis (Plate A4).

Inner to the hypodermis 6–7 layers of

parenchyma cells forming cortex, loaded with

oil globules and prismatic crystals of calcium

oxalate (Plate A5, 6, 9). Pericyclic fibers are

lignified, arranged circularly forming an arch

shaped bundles (Plate A8, 12). Vascular

bundles open and collateral, arranged radially.

Metaxylem facing towards periphery and

protoxylem facing towards pith region (Plate

A7, 10). Xylem consists xylem parenchyma

and fibers. Phloem situated above the xylem

and consists phloem fibers and sieve elements.

Xylem vessels were separated by uniseriate to

triseriate medullary rays (Plate A11). Cells

were somewhat elongated barrel shaped with

starch grains, prismatic crystals and oil

globules. Pith occupies the central part of the

stem. Pith consist parenchyma cells. Cells were

thick, lignified and pitted near the tail of

vascular bundles. Most of the parenchyma cells

consists oil globules, prismatic crystals and

rhomboidal crystals.

PLATE A

1.Olax scandens Roxb. 2. T. S of stem in outline. 3. Trichomes.

4. Epidermis and hypodermis. 5. Rhomboidal crystal. 6. Prismatic crystals.



7. vascular bundles and pith. 8. Pericyclic fibers. 9. Crystals and oil globules.

10. T. S after staining 11.Medullary rays 12. Lignified pericyclic fibers.

Powder microscopy

Organoleptic characters showed that

powder was straw colored, with characteristic

odour and bitter taste.

Diagnostic character of stem powder

showed unicellular horn shaped trichomes of

epidermis, prismatic and rhomboidal crystals of

calcium oxalate, tannin content, cork, annular

and border pitted vessels of vascular bundles,

lignified pericyclic fibers and oil globules

(PLATE B1– B7).

Histochemical test

To confirm the presence and absence of the

chemical constituents the material was

subjected to various tests. Lignified cells,

starch, calcium oxalate crystals, tannin, oil

globules were present in the stem (Table 4).

PLATE B

1. Trichome 2. Annular vessels 3. Crystals and tannin content



4. Prismatic crystals 5. Lignified fiber. 6. Border pitted vessels.

7. Cork

1 2 3 R

8. DNA Fingerprints

Table 4: Histochemical tests for Olax scandens Roxb fruit.

Sr.

No.

Reagents Observation Characteristics Results

1. Phloroglucinol+

Conc. HCl

Red Lignified cells ++

2. Iodine Blue Starch ++

3. Phloroglucinol+

Conc. HCl

Dissolved Calcium oxalate

crystals

++

4. Fecl3 solution Dark blue to

black

Tannin cells ++

5. Sudan III Red Oil globules ++



DNA finger printing

Marker used was mid range ruler with 10

bands of 100, 200. 300. 600, 1000, 1500, 2000,

2500, 3000 and 3500 bp. In the gel photograph,

R-marker, lanes 1to 3- patterns obtained from

random primers.

All the primers gave good band pattern.

Prominent band of ~0.4 kb was obtained with

OPA-02 and ~0.9 Kb prominent band was

obtained with OPC-06 primer (PLATE B8).

CONCLUSION

Presence of microscopic characters like

unicellular, horn shaped trichomes, prismatic

and rhomboidal crystals of calcium oxalate, oil

globules are the key identification characters of

Olax scandens Roxb stem. The unique bands

obtained in Polymerase Chain Reaction (PCR)

amplification are clearly discriminated having,

many bright and light bands indicating the

genuinity of the plant. The observed

pharmacognostical characters and DNA finger

prints may be useful to establish the botanical

standards for identification and standardization

of Olax scandens Roxb. stem.

ACKNOWLEDGEMENT

The authors like to acknowledge the

administrative authorities of the institute IPGT

& RA, Jamnagar for providing facilities during

work. Authors also express their sincere thanks

to Mr. Pareshvar Sahoo, Taxonomist, SSN

Ayurvedic college Paikmal Odisha, Mr. B. N.

Hota, Rtd. DFO, Govt. of Odisha who helped

us during drug collection and Dr. Sudha,

Director, Aristogene Biosciences Pvt Ltd,

Bangalore for their co-operation for DNA

RAPD study of the plant.

REFERENCES

Anonymous, (1990), Glimpses of medico-

botany of Bastar district, Madhya

Pradesh, CCRAS publication, pp. 114.

Anonymous, (1996), Botanical exploration of

Phulbani and Korapat district of Orissa,

CCRAS publication, pp. 132.

Anonymous, (1999), The Ayurvedic

Pharmacopoeia of India, Govt. of India

publication New Delhi, 1st edition, Vol.

I, Appendix 2.

Baum BR Mechanda S, Livesey JF, Binns SE,

Arnason JT. (2001) Predicting

quantitative phytochemical markers in

single Echinacea plants or clones from

their DNA fingerprints. Phytochemistry.

56 (6):543–9

Borkar S D, Naik R, Harisha C R, Acharya R N

(2013), Development of Random

Amplified Polymorphic DNA markers

for authentification of Rivea

hypocrateriformis (Desr.) Choisy,

Global J Res. Med. Plants & Indigen.

Med.,Volume 2(5): pp. 348–356

Johnson Alexander Donald, (1940), Plant

Micro techniques, Macgrow Hill Book

Company, New York, London. pp. 105.

Khandelwal K.R. (2008), Practical

Pharmacognosy Techniques and

Experiments, 19th Ed, Nirali Prakashan;

2008. pp . 15–18.

Krishnamurty K.V. (1988). Methods in the plant

histochemistry, Vishwanadhan Pvt

Limited, Madras, pp.1–77.



Naik Raghavendra, Borkar Sneha D, Harisha C

R, Acharya R N, (2013), A detailed

pharmacognostical evaluation on leaf of

Olax scandens Roxb, Global J Res.

Med. Plants & Indigen. Med, Volume 2,

(4): 189–203

Patel H, Ingalhalli R, (2013), A brief review on

Ginkgo biloba L. (maidenhair tree) a

rare multipurpose medicinal plant,

Global J Res. Med. Plants & Indigen.

Med., Volume 2(6): 418–427.

Rekha Sinha, Valeria Lakra, (2005), Wild tribal

food plants of Orissa, Indian journal of

traditional knowledge, Vol. 4(3), pp.

246–252.Saxena H.O, (1995), The Flora

of Orissa, Regional Research

Laboratory, Bhubaneshwar, 1st edition,

pp. 288.

Veeramuthu Duraipandiyan, Muniappan

Ayyanar, Savarimuthu Ignacimuthu,

(2006), Antimicrobial activity of some

ethnomedicinal plants used by Paliyar

tribe from Tamil Nadu, India, BMC

Complementary and Alternative

Medicine, 6:35 doi:10.1186/1472-6882-

6-35

Wallis TE, (1985), Textbook of

Pharmacognosy, London Churchill

Publication, pp. 572–82.





ADVANCEMENTS IN INDIAN SYSTEM OF MEDICINE (ISM)

INFORMATICS: AN OVERVIEW

Samal Janmejaya1*

1District Epidemiologist, Dist. Health Office, Gadchiroli, Maharashtra, India

*Corresponding Author: Email: [email protected]; [email protected];

Mob: +919438323843, +919901316384


ABSTRACT

Indian system of medicine has its root in India, evolved through a continuous process of

transformation from its original Vedic form to the modern day Indian System of Medicine. This

system of medicine has crossed the Indian Territory and made its presence in different parts of the

globe. More often the system remains unnoticed owing to its aboriginal facets. Digitalization is one

way to go about the problem which makes it accessible to those who need to know about it. This is

the need of the hour as computers can store and retrieve large pool of data which is not the case with

manual approach. This can be used for several purposes be it present or future. Digitalization or

computerization could be mere online web resources or software for information and decision

making. The entire gamut of these developments could be clubbed together in to a domain known as

ISM informatics. This can be considered as one of the tributaries of the broader umbrella of health

informatics and the same will help to elucidate the former in a better way. The term Indian System of

Medicine embraces many different systems of medicine practiced in India that includes Ayurveda,

Yoga & Naturopathy, Unani, Siddha and Homoeopathy. The present article tries to make a review of

the recent developments in the field of ISM informatics with a special focus on Ayurveda-

informatics.

KEYWORDS: Ayurveda, Indian Systems of Medicine, Informatics

Review article

Cite this article:

Samal Janmejaya (2013), ADVANCEMENTS IN INDIAN SYSTEM OF MEDICINE (ISM)

INFORMATICS: AN OVERVIEW, Global J Res. Med. Plants & Indigen. Med., Volume 2(7): 546–553





INTRODUCTION

Indian System of Medicine is the ethnic

bequest deeply buried in the traditional believes

of the people that are evolved through a

continuous process of transformation from its

original Vedic form to the modern day Indian

System of Medicine. It has witnessed a

paradigm shift from its oldest version of

“Guru-Sishya-Ashram” tradition (a way of

learning, by a disciple, from a Spiritual Mentor

in an Ashram) to the modern day medical

education system, formally taught in school of

Ayurveda with use of all modern amenities.

The present day Ayurveda has undergone many

changes as per the need of the day but

important principles have remained unchanged

(Janmejaya Samal, 2013). This system of

medicine has crossed the Indian Territory and

made its presence in different parts of the

globe. But the biggest problem in its

accessibility is its aboriginal facets. Owing to

its growing popularity it’s high time to

digitalize this system to place it before the

global audience. Several endeavors have been

made from different segments to develop some

of the user friendly digital versions/software of

this oldest and classical system of medicine but

still it has to traverse a long way to meet the

growing pace of time. This is highly essential

because the world is getting digital day by day

and the capacity of computers to store and

retrieve a large pool of data easily which is not

the case with manual approach. Computers

have changed human approach in every field

and health care sector is also a prey to it. ISM

informatics can be considered as one of the

tributaries of the broader umbrella of health

informatics and the same will help to elucidate

the former in a better way. Health Informatics

is as much a result of evolution as planned

philosophy, having its roots in the histories of

information technology and medicine (Cesnik

B, 2010).

ISM informatics is a judicious

integration of Information Technology and

Ayurveda, and other allied disciplines of Indian

medicine. Broadly the development of health

informatics in India can bring three different

benefits; firstly as an instrument in continuing

education, they enable health workers to be

informed and trained in advanced medical and

health sciences; secondly, they deliver health

services to the poor at rural and remote

locations; thirdly, they can increase the

transparency and efficiency of governance,

which should in turn improve the availability

and delivery of publicly provided health

services (C P Chandrasekhar, 2001). Again the

adoption of ICT (Information and

Communication Technology) in Ayurveda will

enhance the interaction between modern

medicine and Ayurveda (Sushant Sud, 2013).

In this context a diligent effort has been made

to review the recent developments in Indian

System of Medicine (ISM) informatics under

the broad umbrella of health informatics.

METHODOLOGY

Be it pure health informatics or ISM

informatics, the discipline becomes purely

technical in nature which is a judicious blend of

information and communication technology

and health or in the present context Indian

System of Medicine. Practically speaking this

goes beyond the scope of this article hence the

same has not been dealt with. This article

delineates a snapshot or an overview of the

developments in India System of Medicine

informatics based on systematic literature

review. Various literatures such as published

articles, books and monographs in the said

domain have been reviewed systematically and

the same has also been applied to web related

resources for the purpose of this study.

DISCUSSION

ISM informatics is somehow similar to

other advanced form of medical informatics but

the domain is very limited, as it cannot include

the advanced areas such as computer assisted

surgery or robotic surgery, neuro-computers

and Artificial neural networks etc. Different

organizations are working towards this

direction and more recently the Institute of

Ayurveda and Integrative Medicine, Bangalore,

India has opened up a center known as Center

for ISM informatics and Theoretical

Foundations which was started in 1995 to give

increased focus for the modernization of Indian



systems of medicine in order to improve access

for variety of research purposes (Institute of

Ayurveda and Integrative Medicine, 2013). The

major achievements of this center are as

follows:

CD on Medicinal plants on Siddha

System of medicine: This CD contains

33,350 Tamil names of plants recorded in

26 well known texts pertaining to the

Siddha system of medicine. These Tamil

names have been correlated to 1600

botanical entities. The nomenclature section

also incorporates correlation of these plant

species with around 66,000 vernacular

names in different Indian languages.

Detailed information has been compiled

and incorporated for 500 plants

recommended for BSMS curriculum of

Siddha system. It was released in 2008

(Institute of Ayurveda and Integrative

Medicine, 2013).

CD on Medicinal plants on Unani System

of medicine: This CD contains information

on 307 plants (including the 275 plants

recommended for BUMS curriculum) in

Unani system of medicine. Five texts of

Unani medicine have been used for this

literary research. A total of 380 plants have

been identified as sources of Unani

medicine from these texts. Around 500

plant entities have been estimated in Unani

system of medicine. Five Classical text of

Unani medicine have been used for this

literary research. This was released in 2008

(Institute of Ayurveda and Integrative

Medicine, 2013).

CD on Medicinal plants on

Homoeopathy: A thorough review and data

compilation from available literature (in

India) relating to plants used in

Homoeopathy has generated a master list of

550 botanical names. After establishing

proper synonym linkages, this number has

got reduced to 506 plant species. Each of

these 506 species has been analyzed for its

geographical distribution. This has

generated a list of 326 species which have

been identified as “recorded in India”. It

incorporates such species, which are wild in

India including naturalized ones and/ or

cultivated/ planted in India. The Materia

Medica section deals with the effect of 375

plant drugs (263 Indian & 112 exotic) on

various body organs including the mind.

Data has been derived from more than 23

published works dealing with Materia

Medica of Homoeopathy. It was released in

2008 (Institute of Ayurveda and Integrative

Medicine, 2013).

CD on Medicinal plants on Ayurveda:

“Dravya guna shastra” (Medicinal Plant

Science) or Ayurvedic Materia Medica on

plants is the complete science on

pharmacognosy of the plants used in Indian

medical heritage. This CD contains

comprehensive information about 370

plants recommended for BAMS syllabus. It

includes around 2300 Sanskrit slokas

(Hymns) with their translation in English. It

contains 800 plant images, botanical

correlation of Sanskrit names based on

accepted studies and current understanding.

Source of data embedded in this CD are

compiled from around 20 Ayurvedic

classical sources starting from 1500 BC to

1986 AD. It includes a glossary of around

3000 Sanskrit technical terms and popular

articles. It was released in 2005. Besides

this institute has also released some of other

CDs such as neighborhood plants of

Bangalore City in 2008, CDs on Indian

medicinal plants in Trade in 2005, CD on

plants of Charaka Samhita in 2003, CD on

clinically important plants of Ayurveda in

2002, CD on Prakriti (How to analyze body

constitution) in 2003, CD on medicinal

plants of Karnataka, Kerala and Tamil

Nadu in 2000. (Institute of Ayurveda and

Integrative Medicine, 2013).

Similarly another organization contributing

to this endeavor is the center for development

of advanced computing, Pune, India which is a

premier Research & Development (R&D)

organization of the Department of Electronics

and Information Technology, Ministry of

Communications & Information Technology

(MCIT) for carrying out R&D activities in IT,

Electronics and associated areas. This



organization has created innovative software on

various functionalities of Ayurveda known as

AyuSoft (Center for Development of Advanced

Computing, 2013).

AyuSoft:

This interactive software has been

developed in collaboration with C-DAC, Pune;

Interdisciplinary School of Health Sciences and

Department of Ayurveda, University of Pune;

Jnana Prabodhini, NGO, Pune, India. It is a

pioneering multidimensional effort for Indian

traditional medical system that provides end-to-

end medical solutions based on traditional

medicines and helps in making health decisions

that are expected to be more informed, more

accurate and quicker. The target end user for

this software may include hospitals,

practitioners and researchers. It has wide range

of applications including disease diagnostics

and treatment, diet and life style advice,

personal management information system,

multimedia based encyclopedia, and textual

and analytical report tool. It has the following

components: (Center for Development of

Advanced Computing, 2013)

Vaidya Sanmitra: - This application covers

most of the clinical requirements. It

includes Maanusha Vritta (Personal

management information system), case

analysis, Master data management, Vyadhi

Nidaana (Disease Diagnosis) (Center for

Development of Advanced Computing,

2013).

Prakriti Vichaya: - Prakriti (constitution) is

a unique concept of Ayurveda that seeks to

explain the element of individuality by

expressing the unique trait of an individual

that is defined by the specific and

permanent composition of their Dosha

(Humour) at conception. Prakriti plays an

important role in the prevention, diagnosis

and treatment of diseases. This module of

AyuSoft covers Dosha Prakriti, Maanas

Prakriti (Psychic Constitution), and

Dhaatusaarataa (Tissue System) (Center

for Development of Advanced Computing,

2013).

AayurVidnyaana: - A knowledge base

encompassing all the aspects of Ayurveda.

It includes articles, research papers of the

stalwarts from all over India; Collection of

information related to different aspects of

Ayurveda; Digitalized Brihattrayee (Three

major treatises of Ayurveda constituting

Charaka Samhita, Sushrut Samhita,

Astanga Hridaya) in Devanaagaree Script;

Video Clips of Therapeutic Procedures like

Medicated emesis, Medicated massage,

Blood Letting etc.; Photographs of different

herbs, Instruments and Diseases; Audio

Files of Dhanvantaree Stavana (Chants on

God Dhantwantari) and other Mantra

(Hymns)(Center for Development of

Advanced Computing, 2013).

Anveshaka: - Anveshaka is an interactive

and high-speed data-mining tool useful for

analyzing Ayurveda related data. It is useful

to researchers and students for carrying out

precise analytical search of classical data. It

comprises of the common data report,

Vyaadhi (Disease) diagnosis report, and

Vyaadhi (Disease) treatment report (Center

for Development of Advanced Computing,

2013).

Vyaadhinidaan: - Provides decision support

for probable diagnosis and treatment. It is

based on comprehensive diagnostic and

treatment data. In includes two important

features i.e. Diagnostic features and

Treatment features (Center for

Development of Advanced Computing,

2013).

EasyAYURVEDA:

This has been developed by VHCA herbals

which portray an excellent blend of Ayurveda

and Information Technology. There is a whole

unlimited bunch of literature in Ayurveda and

the demand for such a tool that can make

reading, searching easier is very high. Suppose

a researcher is interested in herbs with sheeta

virya (Potency) from the ancient texts of

Ayurveda, the process may be so tedious and

slow that output of the exercise shall fade away

with the passage of time. But software like

EasyAYURVEDA can make the process



relatively easier and faster. EasyAYURVEDA

is a Easy to use Ayurvedic software; contains

database of more than 500 Medicinal Plants

formulations; helps in advanced searching of

herbs & formulations and herb’s name in all

Indian languages; Multiple string searching &

single string searching, Indications on the basis

both Ayurveda & modern diagnostic principles

and terminologies. This software is the first of

its kind in the history of Ayurveda and is the

largest information portal of Ayurveda on

internet (VHCA Herbals, 2013).

PRAKES:

This is one of the innovative software

developed by Resource Center for Indian

Language Technology Solutions- Malayalam,

Center for Development of Advanced

Computing, Thiruvanantapuram, Kerala, India

and is available in both English and Malayalam

version. It is an interactive menu driven

interface. It helps in examining the Lakshanas

(Symptomatology) and assessing the

dominance of Tridoshas (Three Humors).

Advices on preventive promotive health care

services depending on humeral dominance. The

system records the interactions and results

along with the bio-data for future references. It

helps generate the hard copy of these

recordings (Malayalam Resource Centre,

2013).

BODY TUNE: (Computerized Ayurvedic

Medicare/CAM):

This software has been developed by

Dr.M.A Shajahan in 1983 in Govt. College of

Indian Medicine, Bangalore in collaboration

with Indian Institute of Science, Bangalore

which was proved clinically successful by

Gujarat Ayurveda University. A newer version

of the software was developed in collaboration

with Cyberveda Technologies in 1988. This

particular software neither replaces a doctor nor

avoids the importance of doctor-patient

relationship. It helps organize diagnostic

methods in a classical way envisaged by Indian

sages of Ayurveda. This user friendly

interactive software promotes accurate

diagnosis in a faster and organized way

(Shajahan, M.A, 1993; VHCA Herbals, 2013).

PRAKRTI:

This innovative and expert software has

been designed and developed by Chaitanya

Consultancy, Pune in 1989. It renders services

on different functionalities of Ayurveda such as

Prakrati (Constitution), dietary advices,

advices on daily regimens, likelihood of an

illness and its precautionary measures.

PILEX:

As the name indicates, this particular

software deals with entirety of piles. It is

intended for the diagnosis, prognosis,

complications and treatments of piles. It was

designed and developed by Gujarat Ayurveda

University, Jamnagar, Gujarat in 1990.

RASEX:

This innovative software was designed and

developed by Government Ayurveda College,

Trivandrum in collaboration with CIRA

(Center for Information Research and Action),

and C-DAC (Center for Development of

Advanced Computing), Thiruvanantapuram in

1992. This package attempts to correlate the

pharmacological properties with that of

therapeutic properties with the help of

computer. A database was created after

collecting, organizing and storing all the

pharmacological and therapeutic properties of

single rasa drug using DBase III plus. A list of

drugs, which conforms to the physician’s

specifications is collected and displayed in this

package (Sushant Sud, 2013).

Ayurinformatics:

Ayur-informatics is a science dealing with

the application of bioinformatics to the field of

Ayurveda. It is the application of modern drug

designing technology and bioinformatics to the

field of traditional Ayurvedic system of

medicine. This area is growing by leaps and

bounds with some impressible and noticeable



developments. Some of the recent works in this

domain are as follows:

An in-silico work based on Homology

Modeling for the proteins of RAS, MYC,

SRC, BRAC1, P53, and EGFR using

Modeller9v7 software for establishing an

Ayurvedic remedy for bronchial carcinoma

was done by Preenon Bagchi et al., It

reflects the use of classical Ayurveda drugs

having importance in the cure of lung

tumors (Preenon Bagchi et al.,2011).

Another in-silico work based on COMT

(Catechol-O-methyltransferase) gene was

done by Preenon Bagchi et al., The work

highlights the use of anti-psychotic herbal

drugs as agonist to COMT mutation gene

which inhibits the excessive dopamine

production and cures Schizophrenia. The

study reflects the use of herbs like

Chandana (Santalum album),

Shankhpushpi (Canscora decussata) and

Jatamansi (Nardostachys jatamansi) which

have mental regenerative & antipsychotic

property, their active compounds such as β-

santalol, xanthone and nardal respectively

are docked with COMT mutation protein

(Preenon Bagchi et al., 2012).

An in-silico work based on Homology

Modeling for the proteins of APP, APOE,

Presenilin 1 and Presenilin 2 using

Modeller9v7 and other software for

establishing an in-silico Ayurvedic

medication for Alzheimer’s disease was

carried out by Mohini Gore Active

compounds of medicinal herbs like

Sankhapuspi (Canscora decussata),

Jatamansi (Nardostachys jatamansi) and

Kapikachu (Mucuna pruriens) have been

used for the purpose of study (Mohini Gore

2010).

Ample of work is happening in the field of

Ayurvedic education, teaching and training.

This has not been reflected in the case with

practice. A list of 4 PubMed indexed journals,

38 non PubMed indexed journals, 4 Hindi

Ayurveda Journals, 26 Journals of

Complementary and Alternative Medicine and

11 magazines of Ayurveda have been

documented (Ayurbhishak 2013). Many of the

classical Ayurveda treatises such as Charaka

Samhita, Sushrut Samhita, Astanga Sangraha,

Astanga Hridaya, Sharangadhara Samhita,

Madhav Nidan, Harita Samhita, Bhela Samhita

and Kasyapa Samhita have been digitalized and

are available on line (Ayurbhishak 2013). This

academic and practice gap can be minimized by

working on these issues in collaboration with

renowned ISM practitioners with that of IT

professionals.

CONCLUSION:

This era is rightly called as the digital age.

Computer has immensely influenced human

life. Computer has to do with every walk of

human life so also the domain of medicine and

health care. The medical informatics has grown

by leaps and bounds but the same is not the

case with ISM informatics. Things are

relatively better and initiatives have already

been started elsewhere but more and more

endeavors are needed to place it on a noticeable

height. As a specialized field this particular

domain requires an integrative approach from

both the field of Ayurveda and Information

Technology. This judicious blend will

definitely be of great help in different facets of

Ayurveda be it clinical medicine, biomedical

research or information storage and retrieval.

Wider acceptance of these systems owing to

their safe and efficacious therapeutics on many

of human diseases has again created an urgent

need for the development of ISM informatics.

It has been widely observed that websites are

mushrooming imparting information, education

and communication in matters related to Indian

systems of medicine but the authenticity of the

same is skeptical which needs to be controlled

with governmental support. Again there are lots

of virgin areas which could be explored and

worked out for better access, operation and

above all better utility of Indian Systems of

medicine. This can lead to the development of

some work in the following domains of Indian

System of Medicine. These may include;

Computer based medical information

retrieval



Computers in Clinical Laboratory

Computer assisted medical decision making

Hospital information system

Clinical information system

Nursing information system

Dietetic information system

Computerized patient records

Computerized prescriptions for patients

Computer Assisted Patient Education and

Health care information

Telemedicine

Computer Assisted Drug Discovery and

Development

Computer Assisted Instruction in Medicine

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Call for Papers – Vol. 2, Issue 9, September 2013

Submit your manuscripts (Research articles, Review

articles, Short Communications, Letters to the Editor,

Book Reviews) to Global Journal of Research on

Medicinal plants & Indigenous medicine – GJRMI

Submit it online through www.gjrmi.com or mail it to

[email protected] on or before

August 10th 2013.

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