Naga Vamsi Krishna A et al. IRJP 2012, 3 (3) · ANTIOXIDANT ACTIVITY AND GC-MS ANALYSIS OF...
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Naga Vamsi Krishna A et al. IRJP 2012, 3 (3) Page 252 INTERNATIONAL RESEARCH JOURNAL OF PHARMACY www.irjponline.com ISSN 2230 – 8407 Research Article ANTIOXIDANT ACTIVITY AND GC-MS ANALYSIS OF PHRAGMYTES VALLATORIA LEAF ETHANOLIC EXTRACT Naga Vamsi Krishna A* 1 , Venkata Raman B 2 , Kasetti Ramesh Babu 3 , Chippada Apparao 3 1 Department of Biochemistry, Achraya Nagarjuna University, Guntur, AP, India 2 Department of Biotechnology, K.L. University, Guntur, AP, India 3 Department of Biochemistry, Sri Venkateswara University, Tirupati, AP, India Article Received on: 05/01/12 Revised on: 14/02/12 Approved for publication: 16/03/12 *Email: [email protected] ABSTRACT Phragmytes vallatoria belongs to the family of poaceae. It has so many medicinal properties used mainly for wound healing, arthritis, antimetics, fabrifuges, rheumatism and diabetes. The plant containing polyphenols, flavonoids, alkaloids, proteins (6.6 %) and fibre (40.7 %). The present investigation deals with the GC-MS analysis and antioxidant activity of ethanolic extract of the leaves of the Phragmytes vallatoria plant. The antioxidant activity of ethanolic extract was evaluated by DPPH method. The results indicate that ethanolic extract of Phragmytes vallatoria possess considerable antioxidant activity. The highest radical scavenging activity was detected (IC50 = 735 µg/ML). Seven compounds were identified during GC-MS analysis. GC-MS analysis reveals that hexadeconic acid ethyl ester (30.88 %) was major component fallowed by 9, 12, 15- Octadecatrienoic acid, methyl ester (29.41) and 9, 12-Octadecadienoic acid (19.44%). This study reveals that Phragmytes vallatoria plant having attraction sources of fatty acids as well as of effective natural antioxidants. KEY WORDS: Phragmytes vallatoria, Antioxidant activity, GC-MS. INTRODUCTION Phragmytes vallatoria is belongs to the family of poaceae and it is spreaded throughout India. According to literature it has different types of application in medicine and agriculture products. For medicinally it had the properties of as diuretic, animistic, diaphoretic, wound healing, diabetes, arthritis, rheumatism, antemetics and febrifuges 1 . For agriculture using building materials, fibre, pulp and paper sisling apparatus. It is also having considerable antioxidant activity, through DPPH activity. Mainly these antioxidants are powerful allies in combating inflammation and lowering heart disease and cancer risk. Flavonoids and other plant phenolics, phenolic acids, stlibenes, tannins, lignans, and lignin, are especially common in leaves, flowering tissues, and woody parts such as stems and barks 2 . The antioxidant activity of plants might be due to their phenolics compounds 3 . The antioxidant activity of phenolics is mainly due to their redox properties, which allow them to act as reducing agents, hydrogen donators, and singlet oxygen quenchers. In addition, they have a metal chelation potential 4 . The present study aims to investigate the presence of phytochemicals in ethanolic extract of Phragmytes vallatoria and study the antioxidant activity. MATERIALS AND METHODS Collection of plant material Phragmytes vallatoria is obtained from chirala (Prakasam district, Andhra Pradesh, India). Leaves were collected and shade dried and powdered. Then powdered leaves were extracted with ethanol using soxhlet apparatus. Extract were concentrated by rotary evaporator under vacuum. Determination of antioxidant activity The antioxidant activity of ethanolic extract of Phragmytes vallatoria was assayed by the methanol radical scavenging activity by DPPH assay method 5 . GC-MS analysis GC-MS performed with GC Clarus 500 Perkin Elmer equipment. Compounds were separated in Elite-1 capillary column (100% Di methyl poly siloxane), 30 ×0.25×1µmdf.Samples were injected at a temperature of about 250 °C with a split ratio of 10: 1 with a flow rate of helium 1 ml/min.Mass detector turbo mass gold-perkin Elmer was used as detector. MS-programme The constituents were identified after comparison with those available in the computer library (NIST ver. year 2005) attached to the instrument and reported. RESULTS Antoxidant activity of ethanolic extract DPPH stable free radical method is rapid and sensitive method for assaying the antioxidant activity of a specific compound or plant extracts. The DPPH radical is considered to be a model for a lipophilic radical.A chain in lipophilic radicals was initiated by the lipid autoxidation.DPPH is a stable free radical at room temperature and accepts an electron or hydrogen radical to become a stable diamagnetic molecule 6 . The antoxidant activity of ethanolic extract was studied. The DPPH scavenging activity expressed as IC 50 value were presented in Table-1.The ethanolic extract of Phragmites vallatoria having lowest IC 50 value with efficient free radical scavenging activity. The reference antioxidant vitamin-C having higher activity than that of Phragmites vallatoria. The higher DPPH radical-scavenging activity is associated with lower IC 50 value. The results may arise from the other chemical constituents rather than fatty acids. GC-MS analysis Tables-2 represents the GC-MS analysis of the ethanolic extract of Phragmites vallatoria. Mainly seven prominent compounds with their corresponding peaks were identified. Two unsaturated methyl esters (Peak No.4, 5), two minor fatty acids (Peak No.2, 6), diisooctyl ester (Peak No. 6), 3, 7, 11, 15-Tetramethyl-2-hexadecen-1-ol (peak No.1) and phytol (Peak No.3) were identified. Major comprised compounds are Hexadecanoic acid ethyl ester, 9,12,15-Octadecatrienoic acid,
Transcript of Naga Vamsi Krishna A et al. IRJP 2012, 3 (3) · ANTIOXIDANT ACTIVITY AND GC-MS ANALYSIS OF...
Naga Vamsi Krishna A et al. IRJP 2012, 3 (3)
Page 252
INTERNATIONAL RESEARCH JOURNAL OF PHARMACY www.irjponline.com ISSN 2230 – 8407
Research Article
ANTIOXIDANT ACTIVITY AND GC-MS ANALYSIS OF PHRAGMYTES VALLATORIA LEAF ETHANOLIC EXTRACT
Naga Vamsi Krishna A*1, Venkata Raman B2, Kasetti Ramesh Babu3, Chippada Apparao3 1Department of Biochemistry, Achraya Nagarjuna University, Guntur, AP, India
2Department of Biotechnology, K.L. University, Guntur, AP, India 3Department of Biochemistry, Sri Venkateswara University, Tirupati, AP, India
Article Received on: 05/01/12 Revised on: 14/02/12 Approved for publication: 16/03/12
*Email: [email protected] ABSTRACT Phragmytes vallatoria belongs to the family of poaceae. It has so many medicinal properties used mainly for wound healing, arthritis, antimetics, fabrifuges, rheumatism and diabetes. The plant containing polyphenols, flavonoids, alkaloids, proteins (6.6 %) and fibre (40.7 %). The present investigation deals with the GC-MS analysis and antioxidant activity of ethanolic extract of the leaves of the Phragmytes vallatoria plant. The antioxidant activity of ethanolic extract was evaluated by DPPH method. The results indicate that ethanolic extract of Phragmytes vallatoria possess considerable antioxidant activity. The highest radical scavenging activity was detected (IC50 = 735 µg/ML). Seven compounds were identified during GC-MS analysis. GC-MS analysis reveals that hexadeconic acid ethyl ester (30.88 %) was major component fallowed by 9, 12, 15- Octadecatrienoic acid, methyl ester (29.41) and 9, 12-Octadecadienoic acid (19.44%). This study reveals that Phragmytes vallatoria plant having attraction sources of fatty acids as well as of effective natural antioxidants. KEY WORDS: Phragmytes vallatoria, Antioxidant activity, GC-MS. INTRODUCTION Phragmytes vallatoria is belongs to the family of poaceae and it is spreaded throughout India. According to literature it has different types of application in medicine and agriculture products. For medicinally it had the properties of as diuretic, animistic, diaphoretic, wound healing, diabetes, arthritis, rheumatism, antemetics and febrifuges1. For agriculture using building materials, fibre, pulp and paper sisling apparatus. It is also having considerable antioxidant activity, through DPPH activity. Mainly these antioxidants are powerful allies in combating inflammation and lowering heart disease and cancer risk. Flavonoids and other plant phenolics, phenolic acids, stlibenes, tannins, lignans, and lignin, are especially common in leaves, flowering tissues, and woody parts such as stems and barks 2. The antioxidant activity of plants might be due to their phenolics compounds3. The antioxidant activity of phenolics is mainly due to their redox properties, which allow them to act as reducing agents, hydrogen donators, and singlet oxygen quenchers. In addition, they have a metal chelation potential4. The present study aims to investigate the presence of phytochemicals in ethanolic extract of Phragmytes vallatoria and study the antioxidant activity. MATERIALS AND METHODS Collection of plant material Phragmytes vallatoria is obtained from chirala (Prakasam district, Andhra Pradesh, India). Leaves were collected and shade dried and powdered. Then powdered leaves were extracted with ethanol using soxhlet apparatus. Extract were concentrated by rotary evaporator under vacuum. Determination of antioxidant activity The antioxidant activity of ethanolic extract of Phragmytes vallatoria was assayed by the methanol radical scavenging activity by DPPH assay method5. GC-MS analysis GC-MS performed with GC Clarus 500 Perkin Elmer equipment. Compounds were separated in Elite-1 capillary
column (100% Di methyl poly siloxane), 30 ×0.25×1µmdf.Samples were injected at a temperature of about 250 °C with a split ratio of 10: 1 with a flow rate of helium 1 ml/min.Mass detector turbo mass gold-perkin Elmer was used as detector. MS-programme The constituents were identified after comparison with those available in the computer library (NIST ver. year 2005) attached to the instrument and reported. RESULTS Antoxidant activity of ethanolic extract DPPH stable free radical method is rapid and sensitive method for assaying the antioxidant activity of a specific compound or plant extracts. The DPPH radical is considered to be a model for a lipophilic radical.A chain in lipophilic radicals was initiated by the lipid autoxidation.DPPH is a stable free radical at room temperature and accepts an electron or hydrogen radical to become a stable diamagnetic molecule6. The antoxidant activity of ethanolic extract was studied. The DPPH scavenging activity expressed as IC50 value were presented in Table-1.The ethanolic extract of Phragmites vallatoria having lowest IC50 value with efficient free radical scavenging activity. The reference antioxidant vitamin-C having higher activity than that of Phragmites vallatoria. The higher DPPH radical-scavenging activity is associated with lower IC50 value. The results may arise from the other chemical constituents rather than fatty acids. GC-MS analysis Tables-2 represents the GC-MS analysis of the ethanolic extract of Phragmites vallatoria. Mainly seven prominent compounds with their corresponding peaks were identified. Two unsaturated methyl esters (Peak No.4, 5), two minor fatty acids (Peak No.2, 6), diisooctyl ester (Peak No. 6), 3, 7, 11, 15-Tetramethyl-2-hexadecen-1-ol (peak No.1) and phytol (Peak No.3) were identified. Major comprised compounds are Hexadecanoic acid ethyl ester, 9,12,15-Octadecatrienoic acid,
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methyl ester (z,z,z)-, 9,12-Octadecadienoic acid (z,z)-, and minor compounds such as Phytol, Octadecanoic acid ethyl ester, 1,2-Benzenedicarboxylic acid diisooctyl ester and 3,7,11,15-Tetramethyl-2-hexadecen-1-ol were also identified. Phytol is one part of the chlorophyll and important in plant biosynthesis. Hexadecanoic acid, ethyl ester was predominant in relation to other fatty acids. In the investigated extract α-linolenic and linoleic acid, which belong to the group of so called essential fatty acids (EFAs). These two EFAs are precursors in the biosynthesis of biologically important lipids and have also been reported in the reduction of coronary heart disease7. Terpinen-4-ol, neophytadiene, hexahydrofarnesyl acetone, vitamin E, phytol and squalene were identified for the first time in the species from the genus Veronica. Several investigations revealed in vitro antioxidant activity of monoterpenes (γ-terpinene) and diterpenes (vitamin E, phytol) or essential oils8. Meanwhile, the polyunsaturated fatty acid (Z, Z)-9, 12-octadecadienoic acid (LA), a conjugated linoleic acid known as an antioxidant that can protect membranes from harm9. Among the identified phytochemicals, hexadecanoic acid have the property of antioxidant and antimicrobial activities10. 9,12,octadecanoic acid (z,z)-has the property of anti-inflammatory and anti arthritic as reported by earlier worker11. When humans as well as rodents are fed free phytol, a high proportion is absorbed and converted in vivo to phytonic acid12-15. The phytol conversion of phytonic acid is a natural rexinoid. It shows antidiabetic activity in type-II diabetic patients16 linoleic acid, and their conjugates also can normalize impaired glucose tolerance in zucker diabetic fatty acids17. All chlorophyll derivatives are having antioxidant properties18. DISCUSSION In this study, we demonstrate that the antioxidant activity of Phragmytes vallatoria having the most efficient free radical scavenger by the lowest Ic 50 value. The activity of the reference antioxidant (vitamin-c) was much higher than compare with the ethanolic extract. The GC-MS analysis of ethanolic extract of Phragmytes vallatoria reveals the presence of fatty acids and plastisizer compounds. By this study it can be conclude that ethanolic extract of Phragmytes vallatoria was good source for natural antioxidants. Medicinally it has the properties of diuretic, animistic, diaphoretic; wound healing, diabetes, arthritis, rheumatism, antiemetic and febrifuges 19, 1.
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969-978. 3. Cook NC, Samman S. Flavanoids- chemistry, metabolism, cardio
protective effects, and dietary sources. J Nutr Biochem 1996; 7: 66-76. 4. Rice-Evans CA, Miller NJ, Bolwell PG, Bramley PM, Pridam JB.The
relative antioxidant activities of plant derived polyphenolic flavonoids. Free Radic Res 1995; 22: 375-383.
5. Cheung LM, Cheung PCK, Ooi VEC. Antioxidant activity and total phenolics of edible mushroom extracts. Food. Chem 2003; 81: 249–255.
6. Soares JR, Dinis TCP, Cunha AP, Almeida LM. Antioxidant activities of some extracts of Thymus Zygi. Free Radic Res 1997; 26: 469-478.
7. Oliver MF. It is more important to increase the intake of unsaturated fats than to decrease the intake of saturated fats: evidence from clinical trials relating to ischemic heart disease. Am J Clin Nutr 1997; 66: 980S-986S.
8. Grassman J. Terpenoids as plant antioxidants.Vitam Horm 2005; 72: 505-535.
9. Ha YL, Storkson J, Pariza MW. Inhibition of benzo (a) pyrene-induced mouse forestomach neoplasia by conjugated dienoic derivatives of linoleic acid. Cancer Res 1990; 50: 1097– 1101.
10. Bodoprost J, Rosemeyer H. Analysis of Phenacylester derivatives fatty acids from human skin surface sebum by reversed-phase HPTLC: chromatographic mobility as a function of Physicochemical properties. Int J Mol Sci 2007; 8: 1111-1124.
11. Jones PJ. Clinical nutrition: 7. Functional foods-more than just nutrition. CMAJ 2002; 166: 1555-1563.
12. Steinberg D, Avigan J, Mize C, Eldjarn L, Try K, Refsum S. Conversion of U-C14- phytol to phytanic acid and its oxidation in heredopathia atactica polyneuritiformis. Biochem Biophys Res Commun 1965; 19: 783–789.
13. Steinberg D, Avigan J, Mize CE, Baxter JH, Cammermeyer J, Fales HM. Effects of dietary phytol and phytanic acid in animals. J Lipid Res 1966; 7: 684–691.
14. Mize CE, Avigan J, Baxter JH, Fales HM, Steinberg D. Metabolism of phytol U-14C and phytanic acid-U-14C in the rat. J Lipid Res 1966; 7: 692–697.
15. Sezille G, Jaillard J, Scherpereel P, Biserte G. Biosynthesis of glyceryl triphytanate in rabbit receiving a diet enriche with phytol. Clin Chim Acta 1970; 29: 335–337.
16. McCarty MF. The chlorophyll metabolite phytanic acid is a natural rexinoid-potential for treatment and prevention of diabetes. Medical Hypothesis 2001; 56: 217-219.
17. Houseknecht KL, Vanden Heuvel JP, Moya-Camarena SY, Portocarrero CP, Peck LW, Nickel KP et al. Dietary conjugated linoleic acid normalizes impaired glucose tolerance in the Zucker diabetic fatty fa/fa rat Biochem Biophys Res Commun 1998; 244: 678–682.
18. Jyotsna M, Srivastava RK, Shukla SV, Raghav CS. Antioxidants in Aromatic and Medicinal plants. Science Tech Entrepreneur 2007; Fragrance and Flavour Development Centre, Kannauj.
19. Nagavamsikrishna A, Ramgopal M, Venkataraman B, Balaji M. Anti-diabetic efficacy of ethanolic extract of Phragmites vallatoria on STZ induced diabetic rats. Int J Pharm Pharm Sci 2012; 4: 118-12.
Table 1: DPPH free radical scavenging activity of Phragmites vallatoria leaf ethanolic extract Sample IC50 value (µg/ml)
Phragmites vallatoria Vitamin-C
735 23
Table 2: Phyto-components identified in the Phragmites vallatoria leaf ethanolic extract
No. RT Name of the compound Molecular formula
Molecular weight
Peak area %
1. 14.36 3,7,11,15-Tetramethyl-2-hexadecen-1-ol C20H40O 296 4.08 2. 16.43 Hexadecanoic acid, ethyl ester C18H36O2 284 30.88 3. 18.39 Phytol C20H40O 296 5.72 4. 18.91 9,12-Octadecadienoic acid (z,z)- C18H32O2 280 19.44 5. 19.06 9,12,15-Octadecatrienoic acid, methyl ester, (z,z,z)- C19H32O2 292 29.41 6. 19.39 Octadecanoic acid, ethyl ester C20H40O2 312 5.23 7. 23.92 1,2-Benzenedicarboxylic acid, diisooctyl ester C24H38O4 390 5.23
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Figure 1: GC-MS chromatogram of Phragmites vallatoria ethanolic leaf extract
Source of support: Nil, Conflict of interest: None Declared
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