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Research Journal of
Chemistry and Environment
Vol. 22 (Special Issue II), August 2018
E-ISSN: 2278 - 4527 ! PRINT-ISSN No. 0972 - 0626
Journal is indexed in SCOPUS andChemical Abstracts
ii
GOD IS.
E-ISSN: 2278 – 4527 PRINT-ISSN No. 0972-0626
RESEARCH JOURNAL OF CHEMISTRY AND ENVIRONMENT
An International Research Journal of Chemical Sciences and Environmental Sciences
Res. J. Chem. Environ., Volume 22(Special Issue II), Pages 1-350, August (2018)
Editor- in- Chief (Hon.)
Dr. D.K. Vardhan Mumbai, INDIA
Correspondence Address:
Research Journal of Chemistry and Environment
Sector AG/80, Scheme No. 54, Indore 452 010 (M.P.) INDIA
Phone and Fax: +91-731-4004000
Website: https://worldresearchersassociations.com
E-mail: info@worldresearchersassociations.com
CONTENTS
Research Papers:
1. Biosynthesis of Silver Nanoparticles using Jicama Extract and Its Application for Colorimetric
Sensing of Mercury Ions
- Firdaus M. Lutfi, Megarani Juwita, Purawiardi R. Ibrahim, Eddy Diana Rakhmawaty and Rahayu Iman
1-3
2. Synthesis and Structural Analysis of Aurivillius Phase, Ca1-xBi3+xNdTi4-xMnxO15
- Zulhadjri, Bilian Sahiga J.I., Wendari Tio Putra and Emriadi
4-9
3. Digital-Based Image Detection System in Simple Silver Nanoparticles-based Cyanide Assays
- Tambaru David, Nomi Anastasia Grandivoriana and Nitti Fidelis
10-14
4. Preparation of Mixed Matrix Polymeric Membrane for removing of contaminants in Crude Biodiesel
- Saiful, Ramli Muliadi, Maulana Ilham, Fadli and Yusuf M.
15-21
5. A new zinc (II) responsive MRI contrast agent
- Adhitiyawarman and Lowe Mark P.
22-30
6. Synthesis of Benzyl-Protected Robtein (2,3,4,4,5-Pentahydroxychalcone) as an Intermediate to (–)-
Robidanol
- Suharti, Achmadi Suminar S., Irfana Luthfan, Sugita Purwantiningsih, Wukirsari Tuti, Ilmiawati Auliya
and Arifin Budi
31-38
7. Antibacterial and antioxidant activities of pyrogallol and synthetic pyrogallol dimer
- Cynthia, Ignatia Florence, Sutanto Hery and Darmawan Akhmad
39-47
8. Preparation and Characterization of Titania/Bentonite Composite Application on the Degradation of
Naphthol Blue Black Dye
- Lubis Surya, Sheilatina, Putra Vicky Praja and Nika Syahrinta Sepia
48-53
9. Optimization of Light Exposure on Superoxide Scavenger Test of Manganese (III)-Salen Acetate
Complex - Nurrahmajanti, Asiyah Onggo, Djulia Deawati and Yusi
54-57
10. Application of Molecularly Imprinted Polymer Technique on Chitosan Membranes for Increasing
Creatinine Transport Effectiveness
- Sulistyaningsih Eka
58-64
iii
11. Total Phenolic and Flavonoid Contents and Antimicrobial activity of Acorus calamus L. Rhizome
Ethanol Extract
- Rita Wiwik Susanah, Kawuri Retno and Swantara I. Made Dira
65-70
12. Optimization of process parameters for inulinase production from endophytic fungi Fusarium solani
LBKURCC67, Humicola fuscoatra LBKURCC68 and Fusarium oxysporum LBKURCC69
- Devi Silvera, Illiyin Luthfin, Ardhi Aulia, Pratiwi Nova Wahyu and Saryono
71-78
13. Morphological identification and hydrolytic enzyme-producing abilities of fungi associated with
wilting banana plants (Musa sp.)
- Saryono, Piska Finna, Sari Nurmala, Pratiwi Nova Wahyu and Ardhi Aulia
79-86
14. 20-Epibryonolic Acid from Tetrameles nudiflora Leaves
- Darmawan Akhmad, Fajriah Sofa, Megawati and Lotulung Puspa Dewi N.
87-90
15. Synthesis of fluorescent compound, 7-Hydroxy-4-methyl-2H-chroman-2-one via Pechmann
condensation of citric acid and resorcinol
- Al Anshori Jamaludin, Rahayu Diah Siti, Hidayat Ace Tatang, Hidayat Ika Wiani and Zainuddin Achmad
91-96
16. Distinguishing of Oligosaccharide Linkage Isomers by Nanoparticle-assisted Fragmentation in
MALDI-TOF MS
- Primadona Indah, Dedi, Wang Yi-Sheng and Chen Yu-Ju
97-101
17. Recombinant Anti BNP-SCFV Production in Escherichia coli and its Application for the Detection of
Heart Failure by Electrochemical Immunosensor using Screen-Printed Carbon Electrode-Gold
Nanoparticles (SPCE-GNP)
- Gaffar Shabarni, Hartati Yeni Wahyuni, Susanti Rini and Subroto Toto
102-107
18. Chemical Compositions of Banana Peels (Musa sapientum) Fruits cultivated in Malaysia using
proximate analysis
- Hassan Pyar and Peh K.K.
108-113
19. Isolation and Identification of Quercetin Derivatives and their -Glucosidase Inhibitory Acitivities
from Bryophyllum pinnatum
- Fajriah Sofa, Darmawan Akhmad, Megawati Megawati and Hanafi Muhammad
114-119
20. Influence of synthesis time on Lanthanum Silicate Apatite (La9.33Si6O26) properties
- Noviyanti Atiek Rostika, Juliandri, Agustina Shofia Utari and Malik Yoga Trianzar
120-123
21. A comparison of the fermentation performance and stress tolerance of baker’s yeast cells grown in
media with or without magnesium addition
- Rachman Saadah D., Hidayat Risma W., Safari Agus and Ishmayana Safri
124-128
22. Investigating Aluminasilicate Source in Geopolymer Compressive Strength: Surface Morphology and
Crystal Phase Study
- Anggarini Ufafa and Sukmana Ndaru Candra
129-133
23.
Optimization Density of Geopolymer Concrete based on Taguchi Method
- Sukmana Ndaru Candra, Pambudi Andry Agung and Anggarini Ufafa
134-138
24. Synthesis of 5-(4’-Chlorobenzylidene) hydantoin and N-3 Substituted 5-(4’-Chlorobenzylidene)-3-
Methylhydantoin
- Naufal Muhammad, Hamdani Ulfi Dadan, Maharani Rani, Al Anshori Jamaludin and Hidayat Ika Wiani
139-145
25. In vitro antioxidant activity of Sonneratia ovata Backer extract
- Khumaidah Laili, Nurmalasari Fatati, Putri Fatmawati Inneke, Purnomo Adi Setyo, Santoso Mardi,
Ersam Taslim and Fatmawati Sri
146-150
iv
26. Effects of bacterium Ralstonia pickettii addition on DDT biodegradation by Daedalea dickinsii
- Purnomo Adi Setyo, Rizqi Hamdan Dwi, Fatmawati Sri, Putro Herdayanto Sulistryo and Kamei Ichiro
151-156
27. The effect of Calcination Temperature of Sulfated Zirconia Catalyst for Simultaneous Reactions in
Biodiesel Production
- Andrijanto Eko and Brown Rob
157-162
28. Synthesis of Zirconia 1-D Nanomaterials from Local Zircon-Based Zr(OH)4 mediated by PEG-6000
- Septawendar Rifki, Nuruddin Ahmad, Maryani Eneng, Sutardi Suhanda and Purwasasmita Bambang
Sunendar
163-171
29. Effects of the Precipitation pH of Sodium Silicate on the Amorphous Silica Characteristics and its
capability in the Pb and Cd Adsorption
- Maryani Eneng, Septawendar Rifki and Suhanda
172-178
30. Critical Parameters of Silver Nanoparticles (AgNPs) synthesized by Sodium Borohydride Reduction
- Sholikhah Umi Nur, Pujiyanto Anung, Lestari Enny, Sarmini Endang, Lubis Hotman, Widyaningrum
Triani and Febriana Siska
179-183
31. Antibacterial Activity of Prenylated Xanthones from Pericarp of Garcinia mangostana against
Persistent Dental Infection Microorganism Enterococcus faecalis
- Setiawan Arlette Suzy, Herawati Mieke, Dewi Warta and Supratman Unang
184-188
32. Identification of a DNA Polymerase I Gene Fragment from a Local Isolate (PLS 80) from an
Underwater Hot Spring
- Febriani, Helwati H., Velayati M.A. and Iqbalsyah T.M.
189-192
33. Biohydrogen Production through Separate Hydrolysis and Fermentation and Simultaneous
Saccharification and Fermentation of Empty Fruit Bunch of Palm Oil
- Kusmardini Diah, Prasetyo Joni, Saepudin Endang and Hudiyono Sumi
193-197
34. Study Cytotoxicity of Haarlem Oil by Crystal Violet Staining Assay
- Khairan Khairan, Jamier Vincent and Jacob Claus
198-202
35. Determination of Optimum Pressure for Gd3+ Separation from Gd-DTPA with Nanofiltration
Membrane
- Rahayu Iman, Winanti Anti, Juliandri, Noviyanti Atiek Rostika, Eddy Diana Rakhmawaty, Anggraeni Anni
and Bahti Husein H.
203-205
36. Cytotoxic Effects of Cassava (Manihot esculenta Crantz), Adira-2, Karikil and Sao Pedro Petro
Varieties against P-388 Murine Leukemia Cells
- Widiastuti Diana, Mulyati Ade Heri, Eka Herlina, Siti Warnasih, Suchyadi Yudhie, Triastinurmiatiningsih
and Supratman Unang
206-208
37. Coating of Nanotube Ti-6Al-4V Alloy with Hydroxyapatite-Chitosan Composite by Electrophoretic
Deposition Method
- Charlena, Suparto Irma Herawati and Puspitawati Tuti
209-213
38. pH Effect of Gold Nanoparticles Synthesis Encapsulation in Polyamide Amine Generation 3.0
(AuNP-PAMAM G3.0)
- Pujiyanto Anung, Lestari Enny, Kurniasih Dede, Mujinah, Sholikhah Umi Nur, Gunawan Adang Hardi,
Amalia Sarah S. and Mutalib Abdul
214-219
39. Mode of action of an insecticidal oleanane glycoside on digestive enzymes of Crocidolomia pavonana
- Maharani Rani, Rizqiani Anggun Fuji, Ishmayana Safri, Hidayat Yusup and Dono Danar
220-225
v
40. Comparative assessment of yeast fermentation performance, ethanol tolerance and membrane fluidity
- Ishmayana Safri, Kennedy Ursula J. and Learmonth Robert P.
226-235
41. Preparation of ZnO/SiO2 Composite using Silica isolated from Rice Husk
- Eddy Diana Rakhmawaty, Ishmayana Safri, Noviyanti Atiek Rostika and Wahyuni Astri
236-239
42. Separation of Radioisotope Iodine-131 from Radiotellurium-131 using Dowex-1x8 resin column
chromatography
- Setiawan Duyeh, Nugraha Nabil Aulia and Hastiawan Iwan
240-244
43. Prelimimary Study of using Palm Oil Mill Effluent to produce Bio Hydrogen as Biofuel
- Prasetyo Joni, Febijanto Irhan, Murti S.D. Sumbogo and Dewi Eniya L.
245-248
44. Analysis and Characterization of Complex Compound Gadolinium-(1,4,7,10-Tetraazacyclododecane,
1-4-7-10-Tetraacetic Acid)n-Poliamidoamine Generation 3-Trastuzumab as a Novel Contrast Agent
for Magnetic Resonance Imaging
- Kusuma Yenni Nugraha Wira, Mutalib Abdul, Anggraeni Anni, Ramli Martalena, Fauzia Retna Putri and
Bahti Husein H.
249-254
45. Comparison of Different Methods for Purification of Gold Nanoparticles– Polyamidoamine
(PAMAM) Generation 4 Dendrimer
- Narsa Angga Cipta, Pujiyanto Anung, Gunawan Adang H., Yusup Iyus Maolana, Listyowati Indrarini,
Fauzia Retna Putri, Bratadiredja Marline Abdassah, Mutalib Abdul, Soetikno Ristaniah D. and
Soedjanaatmadja R. Ukun M.S.
255-259
46. Utilization of Sericin on Modification of Cotton Dyeing using Acid Dyes
- Ramadhani Puri Awaliyah and Umam Khairul
260-266
47. Toxicity of Seed oil of Azadirachta indica, Calophyllum inophyllum and their Mixture against
Crocidolomia pavonana Larvae
- Kadarusman Ratna Puji Astuti, Widayani Neneng S., Dono Danar, Hidayat Yusup, Sunarto Toto,
Hartati Sri and Maharani Rani
267-273
48. Addition of mango essence to methyl eugenol more attracted to Bactrocera dorsalis Complex
(Diptera: Tephritidae) on mango plantation in Majalengka, West Java
- Susanto Agus, Djaya Luciana and Sudrajat Rian Aprianti
274-279
49. Solubility Improvement of Antimalaria Drug through Co-Crystalization with Malic Acid
- Gozali Dolih, Alatas Fikri and Sugandi Rani
280-284
50.
Simple method and environmentally friendly ash content removal process from rice husk char using
potassium carbonate solution
- Solihudin, Rustaman and Haryono
285-288
51. Polymer Electrolyte Membrane Fuel Cell based on Sulfonated Polystyrene and Phosphoric Acid with
Biocellulose as a Matrix
- Naumi Fadlinatin, Natanael Christi L., Rahayu Iman, Indrarti Lucia and Hendrana Sunit
289-293
52. A Voltammetric Immunosensor for Detection of HER2 using Gold Modified-Screen Printed Carbon
Electrode
- Hartati Yeni Wahyuni, Beladona Siti U. Misonia, Wyantuti Santhy and Gaffar Shabarni
294-301
53. Fast and Simultaneous Detection of Sm, Eu, Gd, Tb and Dy using combination of Voltammetry
Method and Multivariate Analysis
- Wyantuti Santhy, Pratomo Uji, Hartati Yeni Wahyuni, Anggraeni Ani and Bahti Husein H.
302-306
vi
54. Vannamei Shrimp (Litopenaeus vannamei, Boone, 1931) Performance with Bacillus and Lactobacillus
Probiotic Formulation Consortium in Dry Preparation
- Andriani Yuli, Safitri Ratu and Fauziah Shaiyanne
307-312
55. The Implementation of Natural Pigments of Pumpkin Meal to enhance the Color Quality of Koi Fish
(Cyprinus carpio)
- Yustiati Ayi, Arisya Yuni, Herawati Titin, Rachman Boedi and Andriani Yuli
313-318
56. Novel Mechanical Filter for reducing Ammonia Concentration of Silver Barb culture in a
Recirculating Aquaculture System (RAS)
- Hasan Zahidah, Rajibbusalam, Maulina Ine and Andriani Yuli
319-324
57. The Reduction of Lead in Fish Meat from Floating Net Cages in Cirata Reservoir by Citric Acid
- Junianto, Hasan Zahidah and Afrianto Eddy
325-329
58. Cytotoxic Sesquiterpenoid from the Stembark of Aglaia argentea (Meliaceae)
- Harneti Desi, Farabi Kindi, Nurlelasari, Maharani Rani, Supratman Unang and Shiono Yoshihito
330-333
59. Cellulose isolation from Gracilaria Genus and its Potential as Bioethanol Raw Material
- Satiyarti Rina Budi, Febrishaummy Indriani, Yun Yenny Febriyani and Mulyani Rahmaniar
334-336
60. Sn (IV) doped lanthanum silicate apatite structure (La9.33Si6-xSnxO26; x: 0.1; 0.3; 0.5) as an electrolyte
- Noviyanti Atiek Rostika, Eddy Diana Rakhmawaty, Hastiawan Iwan, Dzulfikar Muhammad
and Syarif Dani Gustaman
337-341
61. Determination of Individual Spectra of Sm, Eu, Gd, Tb and Dy from the UV-Vis Spectrum of
Mixture Solution
- Anggraeni Anni, Avian Mohamad Sepril, Purnama Andrew and Mutalib Abdul
342-346
62. Secondary Metabolites from Steambarks of Dysoxylum alliaceum
- Mayanti Tri, Nurcahyanti Ois, Darwati, Julaeha Euis, Farabi Kindi, Sumiarsa Dadan
and Dinata Deden Indra
347-350
Research Journal of Chemistry and Environment_______________________________Vol. 22(Special Issue II) August (2018) Res. J. Chem. Environ.
65
Total Phenolic and Flavonoid Contents and Antimicrobial activity of Acorus calamus L. Rhizome
Ethanol Extract Rita Wiwik Susanah1*, Kawuri Retno2 and Swantara I. Made Dira1
1. Chemistry Department, Faculty of Mathematics and Natural Sciences, Udayana University, INDONESIA
2. Biology Department, Faculty of Mathematics and Natural Sciences, Udayana University, INDONESIA
*susanah.rita@unud.ac.id
Abstract The rhizomes of Acorus calamus L. are widely used as
medicinal plant for diseases caused by microbes. The
antimicrobial activity of rhizomes is suspected to have
a positive correlation to the content of phenolic and
flavonoid compounds. This study aims to determine the
total phenolic and flavonoid contents and to evaluate
the antimicrobial activity of ethanol extract of Acorus
calamus L. rhizome. An extraction process was carried
out at room temperature by maceration methods.
The determination of total phenolic and flavonoid
contents was conducted by a UV-Vis Spectrofotometer
using standard gallic acid and quercetin respectively.
Additionally, the antimicrobial activity was evaluated
by the agar disc diffusion method. The total phenolic
and flavonoid contents were successively 2398.40 mg
GAE/100g and 190.46 mg QE/100g dry rhizome
extract. The extracts demonstrated antibacterial
activity against Escherichia coli and Staphylococcus
aureus and antifungal activity against Candida
albicans. Minimum inhibitory concentration of the
extracts against Escherichia coli, Staphylococcus
aureus and Candida albicans was 2, 3 and 3%
respectively with the inhibition zone of 9.80, 9.50 and
8.67 mm.
Keywords: Acorus calamus L., Antimicobial Activity,
Flavonoid contents, Phenolic contents.
Introduction The environment plays a major role in improving public
health. However, poor public health conditions such as the
incidence of various diseases are also influenced by the
surrounding environment. An inadequate environment can
potentially have a direct impact on human health, it can
spread dangerous agents, or indirectly by disrupting the
ecosystems that sustain life1. A dirty environment is good for
breeding various disease vectors such as microbes, including
both bacteria and fungi. Microbial pathogens can cause
various diseases in living organisms by infection. The
microbes that often cause the disease are Escherichia coli, Staphylococcus aureus and Candida albicans.
Medicines such as tetracycline, amoxicillin, miconazole are
often applied to treat infectious diseases. However, these
medicines can lose effectiveness due to microbial
resistance2. Hence, new medicines from natural sources such
as antibiotics from plants are required. A variety of plant
species are traditionally used in some countries for treatment
of infectious diseases such as Allium sativum, Bunium persicum, Oryza sativa3, Vitex negundo, Piper nigrum,
Duranta repens and Acorus calamus4.
Among the plant widely used as traditional medicine to treat
diseases caused by microbials is A. calamus L. This plant has
many uses. Rahamoz-Haghighi et al5 reported that A.
calamus rizhome ethanol and methanol extracts could inhibit
the growth of S. aureus, Staphylococcus epidermidis, E. coli with similar effects. Rawal et al6 revealed that acetone,
aqueus, ethanol and petroleum ether extracts of A. calamus
rizhome could potentially inhibit the growth of Fusarium oxysporum f.sp. lycopersici with Minimum Inhibitory
Concentration (MIC) of 500, 750, 250, 250 mg/mL
respectively. Funde7 investigated anticancer, antioxidants
and antimicrobial activity of A. calamus and stated that A.
calamus was useful for multi-diseases therapeutic research.
Anisah et al8 reported that ethanol extract of A. calamus
rhizome contains alkaloids, flavonoids and polyphenols.
Rahamoz-Haghighi et al9 reported that A. calamus ethanol
extract contains phenyl propanoids, monoterpenes,
sesquiterpenes and -asarone. The compounds
demonstrated antibacterial activity. Based on preliminary
tests, the methanol extract of rhizome collected in Denpasar
contains triterpenoids, steroids, flavonoids, polyphenols and
alkaloids. The antimicrobial activity of A. calamus rhizomes
is probably caused by the content of the compounds in the
rhizomes such as essential oils, flavonoids and polyphenols.
Rita et al10 reported that the essential oils of A. calamus
could potentially inhibit the growth of C. albicans with the
minimum inhibitory concentration (MIC) of 1%. The
essential oils also inhibit the growth of Fusarium solani, a
pathogenic fungus causing stem rot diseases on dragon fruit
stems11,12. The results demonstrate that the oils strongly
inhibited F. solani at a concentration of 10% with an
inhibition zone of 10 mm, MIC of 2 %. The results also
indicate that the growth of colony, spores and fungal
biomass increased with the increase of essential oil
concentration.
Besides essential oils, the antimicrobial activity of A.
calamus rizhome was associated with flavonoid and
phenolic content. Mahboubi et al13 revealed that the
Research Journal of Chemistry and Environment_______________________________Vol. 22(Special Issue II) August (2018) Res. J. Chem. Environ.
66
antimicrobial efficacy of the plant extracts is correlated with
their phenolic and flavonoid contents. Hence, it is necessary
to investigate antimicrobial activity to ethanol extract of the
rhizomes collected in Bali and to determine the total content
of phenolic and flavonoid compounds.
Material and Methods Plant Material: A. calamus L. was identified at LIPI-UPT
Center for Plant Conservation Botanical Garden "Eka
Karya" Bali. Rhizomes of A. calamus were collected around
Denpasar Bali. The rhizomes were respectively cleaned, cut
and dried at room temperature for 15 days. Next, they were
powdered and stored for later analysis.
Microbial Agents: Two strains bacteria E. coli (Gram-
negative) and S. aureus (Gram-positive) and a fungal
pathogen, C. albicans, were applied to the antimicrobial
assay. These microorganisms were obtained from culture
collection of Laboratory of Microbiology, Department of
Biology, Faculty of Mathematics and Natural Sciences,
Udayana University. The isolates were purified and
maintained at 4°C until later use.
Extraction: Around 750 g of A. calamus rizhome powder
was extracted with 10 L of ethanol 96% for 24 h at room
temperature (25°C). The extract was filtered through a
Whatmann filter paper and then evaporated under vacuum
and stored at 4°C until further analysis.
Determination of Total Phenolic and Flavonoid Contents
Total Phenolic contents: Folin-Ciocalteu reagent was
applied for the determination of total phenolic contents14. A
total of 0.1158 gram extract was dissolved in 96% ethanol to
obtain a volume of 5 mL. Dilution was performed 100 times;
1 μL of filtrate was dissolved in ethanol to obtain 100 μL of
solution. 100 μL of Folin-Ciocalteu reagent and 800 μL of
5% sodium carbonate were added so the total solution
volume became 1000 μL. The mixture was then allowed to
stand for 90 minutes, before the absorbance was measured at
a wavelength of 760 nm. A series of gallic acid solutions
with various concentrations were also prepared.
The absorbance of each concentration was measured at a
wavelength of 760 nm. From the standard gallic acids
standards, a calibration curve was made to get the equation
line of y = ax + b. The total phenolic contents were expressed
as mg gallic acid equivalents /100 g of extract. The total
phenols can be determined by using the following formula:
where F1 = total phenol, C = equality of gallic acid (g/mL),
V = total volume of extract (mL), F = the dilution factor and m = weight of sample (g).
Total Flavonoid contents: Total flavonoids were
determined by the aluminum chloride method14. A total of
0.1035 grams of samples were dissolved in 96% ethanol to
obtain a volume of 5 mL. Dilution was performed 5 times.
100 μL of filtrate was dissolved in ethanol to obtain 500 μL
of solution. 500 μL of 2% aluminum chloride was added, so
the total volume of the solution became 1000 μL. The
mixture was then allowed to stand for 90 minutes before the
absorbance was measured at a wavelength of 415 nm.
A series of quercetin solutions with various concentrations
were also prepared. The absorbance of each concentration
was measured at a wavelength of 415 nm. From the
quercetin standards, a calibration curve was made to obtain
equation line of Y = ax + b. The total flavonoid contents
were expressed as mg quercetin equivalents/100 mg extract.
The total flavonoids can be determined by using the
following formula:
where F1 = total flavonoids, C = equality of quercetin
(g/mL), V = total volume of extract (mL), F = the dilution
factor and m = weight of sample (g).
Antimicrobial Activity Assay: Antimicrobial activity assay
of A. calamus rhizome ethanol extract was conducted by the
well diffusion method at various concentrations of 0
(negative control), namely, 0.5, 1, 2, 3, 4, 6, 8 and 10% with
three repetitions10. In order to determine the optimum
concentration, the assay was performed on the extracts with
various concentrations greater than 10. The concentrations
applied were 12, 14 and 16%.
Petri dish containing 10 mL of PDA (Potato Dextro Agar)
media and 200 μL of suspension of the microbes (E. coli, S.
aureus and C. albicans) were allowed to solidify. After the
suspension was solid, the diffusion wells were made using a
cork borer and each well was filled with 20 μL of the extract
and incubated at a temperature of 37°C for 48 hours. The
antimicrobial activity was determined by the diameter of the
inhibition zone.
Results and Discussion Determination of Total Phenolic and Flavonoid Contents
The curves of the calibration to determine total phenolic and
flavonoid contents are presented in figures 1 and 2. Based on
the calibration curves, the calibration equation of gallic acid
(figure 1) obtained was y=0.047x + 0.0635 (R2=0.9899),
while that of quercetin (figure 2) was y=0.0313x+0.0272
(R2= 0.9961). The calculation to determine the total phenolic
and flavonoid contents is summarized at table 1. According
to table 2, the total phenolic and flavonoid contents were 2398.40 mg GAE/100g and 190.46 mg QE/100g dry
rhizome extract successively.
F1C.V.F. 10-6
m100%= ..................................... (1)
F2C.V.F. 10-6
m100%= ..................................... (2)
(2)
Research Journal of Chemistry and Environment_______________________________Vol. 22(Special Issue II) August (2018) Res. J. Chem. Environ.
67
The data showed that the total phenol was higher than total
flavonoids, likely due to only a few phenol compounds being
flavonoids especially quercetin. The most likely compound
is tannin, especially hydrolysable tannis. According to
Khanbabaee and Ree15, tannins consist of hydrolysable and
condensed tannins. The hydrolysable tannins include both
the gallotannins and the ellagitannins. The condensed
tannins consist of cathecin (flavan-3-ol) units.
Antimicrobial Activity Assay: Antimicrobial activity assay
against E. coli, S. aureus and C. albicans of A. calamus
rhizome ethanol extract was performed at concentrations of
0 (negative control), namely, 0.5, 1, 2, 3, 4, 6, 8 10, 12, 14
and 16 %. The antimicrobial assay results are shown in
table 3.
The data showed that the inhibition towards the microbes
increased with the increase of the concentration applied.
Rhoades and Roller16 revealed that in general, the inhibition
tends to increase with the increase of the extracts
concentration. Minimum inhibitory concentration (MIC) of
the extract against E. coli, S. aureus and C. albicans was 2,
3 and 3% respectively with the inhibition zone of 9.80, 9.50
and 8.67 mm.
Overall, E.coli was more sensitive to the extract compared
to S. aureus and C. albicans, except at a concentration of 6%.
This indicates that gram-negatives bacteria are more
sensitive compared to gram positive. These findings are in
contrast to the findings of Okigbo and Mmeka17 who
reported that S. aureus is the most susceptible to the plant
extracts followed by E. coli and then C. albicans. The reason
why E.coli is more susceptible to the extracts than other
microbes is unclear.
The data in table 2 shows that the optimum concentration of
the extract to inhibit the growth of E. coli was 8.0 % because
the inhibition zone was not significantly different from the
concentration of 8.0 to 16%. Meanwhile, the optimum
concentration of that to inhibit S. aureus and C. albicans was
12.0 and 10.0% respectively. Figure 3 shows the inhibitory
zone of the extract with various concentrations towards E.
coli, S. aureus and C. albicans. The extract at a concentration
of 10% could strongly inhibit the growth of all microbes
used (figure 4). The inhibition zone less than 5 mm was
categorized as weak inhibition, between 5 and10 mm was
moderate, larger than 10 to 20 mm was strong and higher
than 20 mm was very strong inhibition14.
Camargo et al18 reported the antimicrobial effects of
phenolic acids and flavonoids of peanut by-products. Their
results showed that phenolic acid-rich extracts showed the
lowest minimum inhibitory capacity (MIC) which means
that the antibacterial effect is highest. Meanwhile, Xuan
et al19 studied the total phenolic and flavonoid contents of
commercial vegetable edible oils marketed in Japan. All the
oils studied possess antimicrobial activity on both S. aureus
and E. coli.
The total phenolic and flavonoid contents contribute to the
antimicrobial activity of this A. calamus rhizome. The
antimicrobial activities of the phenolic compounds are
associated to the ability to bind extracellular and soluble
proteins, thus enabling complexation with bacterial cell
walls20.
Figure 1: Calibration Curve of Standard Gallic Acid
Research Journal of Chemistry and Environment_______________________________Vol. 22(Special Issue II) August (2018) Res. J. Chem. Environ.
68
Figure 2: Calibration Curve of Standard Quercetin
Figure 3: Graph of Inhibitory Activity of A. calamus Rizhome Ethanol Extract against
E. coli, S. aureus and C. albicans
Figure 4: Inhibitory Activity of A. calamus Rizhome Ethanol Extract against
A) E. coli, B) S. aureus and C) C. albicans at concentration of 10% with three repetitions
(0: negative control; 1: the 1st repeat; 2: the 2nd repeat; 3: the 3rd repeat)
3 3 3
2
2
1 0 1 0 1 2
0
3
Research Journal of Chemistry and Environment_______________________________Vol. 22(Special Issue II) August (2018) Res. J. Chem. Environ.
69
Table 1
Total Flavonoid and Phenolic Contents of A. calamus Rhizome Ethanol Extract
Comp. Sample
Weight Abs
(Y)
Cons(x) Volume Dilution
Contents
(g) mg/L (mL) % mg/100g
Phenols 0.1158 0.325 5.5651 5.00 10 2.3984 2398.40
Flavonoids 0.1035 0.274 7.8850 5.00 5 0.1905 190.46
Table 2
Inhibition Zone of The Growth of E. coli, S. aureus and C. albicans of A. calamus Rhizome Ethanol Extract
at Various Concentrations
Treatment Avarage of Inhibition Zone (mm)
(%) E. coli S. aureus C. albicans
0 (negative control) 0a 0a 0a
0.5 0a 0a 0a
1.0 0a 0a 0a
2.0 9.80b 0a 0a
3.0 10.17b 9.50b 8.67b
4.0 11.17bc 11.00c 11.43c
6.0 12.17c 12.33d 13.60d
8.0 15.70d 13.27e 14.10d
10.0 16.60d 13.60e 15.27e
12.0 16.67d 14.53f 15.70e
14.0 16.70d 14.60f 15.73e
16.0 16.73d 14.63f 15.77e
* Values followed by the same letters in the same column are not significantly different according to the
Duncan’s Multiple Range Test at P<5%.
Conclusion The total phenolic and flavonoid contents of A. calamus
rizhome ethanol extract were successively 2398.40 mg
GAE/100g and 190.46 mg QE/100g dry rhizome extract.
The extract possessed the ability to inhibit the three
pathogens. Minimum inhibitory concentration of the extracts
against E. coli, S. aureus and C. albicans was 2, 3 and 3%
respectively with the inhibition zone of 9.80, 9.50 and 8.67
mm. The optimum concentration to inhibit E. coli, S. aureus
and C. albicans was 8, 12 and 10% respectively.
Acknowledgement We thank Udayana University for providing the financial
grant to conduct the research work. We express our gratitude
to the Head of Research and Community Institutions of
Udayana University, facilitating all the needs in the
disbursement of the research funds.
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