Phytochemical Composition of Methanolic Extract of ...

Research J. Pharm. and Tech. 9(5): May 2016

ISSN

0974-3618 (Print) 0974-360X (Online)

www.rjptonline.org

RESEARCH ARTICLE

Phytochemical Composition of Methanolic Extract of Phyllanthus acidus L (Skeels) Fresh Leaves by GC/MS Analysis 1

Rohan Sharadanand Phatak1*, Anup Subhash Hendre2, Pushpa Prakash Durgawale2 Directorate of Research, Krishna Institute of Medical Sciences Deemed University, Malkapur, Karad-415539, India, 2 Department of Biochemistry Krishna Institute of Medical Sciences, Malkapur, Karad-415539, India *Corresponding Author E-mail: [email protected]

ABSTRACT: Background: Secondary metabolites and different phytochemicals are present in the medicinal plant Phyllanthus acidus belonging to family Phyllanthaceae. Scientific evidence data related GC-MS analysis of methanolic extract of Phyllanthus acidus fresh leaves has not been reported till date during literature review. Objective: The study was aimed to investigate phytochemicals present in the methanolic extract of Phyllanthus acidus fresh leaves. Material and Methods: Methanolic extraction of fresh leaves Phyllanthus acidus was prepared by simple maceration and GC-MS analysis data was provided by IICPT, Thanjavur. Phytochemical investigation was carried out to identify the possible components from Phyllanthus acidus fresh leaves by evaluating retention time in the GC-MS chromatogram. Results: Eleven peaks have revealed in the GC-MS spectrum of the methanolic extract of Phyllanthus acidus. From the results, squalene is present as major bioactive compounds in methanolic extract of Phyllanthus acidus fresh leaves. Conclusion: The present data provides to find the molecular formula and weight of 11 biomolecules. Further investigation should be done to isolate bioactive compounds and their structural elucidation and screening of pharmacological activity in the drug development.

KEYWORDS: GC/MS analysis, Phytochemicals, Phyllanthus acidus INTRODUCTION: Phyllanthus acidus L. Skeels is a small tropical tree that grows up to 12 m tall, which bears pale-yellow waxy fruits in abundance on the branches, which are edible and tastes sour belonging to Phyllanthaceae family.[1] Phyllanthus acidus plant is rich source of phytochemicals with interesting biological activities such as antioxidant[2], hepatoprotective [3], antiinflammatory and antinociceptive[4] activities. Volatile constituents from the ripe fruit were analyzed by gas chromatography-mass spectrometry. Seventy-seven volatiles were identified: 45 terpenes, 18 esters, seven acids, four aldehydes, two phenols, and one alcohol, of which 66 were found for the first time in grosella.[5] Received on 15.03.2016 Accepted on 28.04.2016

Modified on 04.04.2016 © RJPT All right reserved

Research J. Pharm. and Tech. 2016; 9(5): 559-561. DOI: 10.5958/0974-360X.2016.00106.2

The scientific data regarding GC-MS analysis of methanolic extract of Phyllanthus acidus fresh leaves has few reported till date during literature review. Therefore, in our present study, it was undertaken into consideration to characterize GC-MS chromatogram for evaluation of bioactive phytochemicals from methanolic extract of the plant.

MATERIAL AND METHODS: Collection of plant material: Plant material was collected from the local areas of Karad, Satara District, Maharashtra, India. Preparation of extract: Fresh leaves of Phyllanthus acidus was collected, washed under running tap water and dried under shade and 2 gm of dried powder of leaves of Phyllanthus acidus was placed into a conical flask. 50ml of methanol

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was added to the conical flask and was macerated for 3 days in order to maximize the extraction. After 3 days it was filtered through Whatmann filter paper, prepared as methanolic stock solution and stored in a suitable container. 1.5ml of methanolic extract was pipetted into Eppendorf tube and sent to IICPT, Thanjavur for GCMS analysis.

2 min, and the total GC/MS running time was 36 min. The relative percentage amount of each component was calculated by comparing its average peak area to the total areas. The mass-detector used in this analysis was Turbo-Mass Gold-Perkin-Elmer, and the software adopted to handle mass spectra and chromatograms was a Turbo-Mass ver-5.2.

GC-MS analysis: GC-MS analysis of methanolic extract of Phyllanthus acidus was performed using a Perkin–Elmer GC Clarus 500 system comprising an AOC-20i auto-sampler and a Gas Chromatograph interfaced to a Mass Spectrometer (GC-MS) equipped with a Elite-5MS (5% diphenyl/95% dimethyl poly siloxane) fused a capillary column (30 × 0.25 μm ID × 0.25 μm df). For GC -MS detection, an electron ionization system was operated in electron impact mode with ionization energy of 70 eV. Helium gas (99.999%) was used as a carrier gas at a constant flow rate of 1 ml/min, and an injection volume of 2 μl was employed (a split ratio of 10:1). The injector temperature was maintained at 250 °C, the ion-source temperature was 200 °C, the oven temperature was programmed from 110 °C (isothermal for 2 min), with an increase of 10 °C/min to 200°C, then 5 °C/min to 280°C, ending with a 9 min isothermal at 280 °C. Mass spectra were taken at 70 eV; a scan interval of 0.5 s and fragments from 45 to 450 Da. The solvent delay was 0 to

Identification of phytochemical constituents: GC-MS mass-spectrum was interpreted by using the database of National Institute Standard and Technology (NIST). Mass spectrum of the unknown components was compared with the spectrum of known components stored in the NIST library. Three major peaks of GC-MS spectrum were ascertained while two phytochemical components were remained unidentified.

RESULTS AND DISCUSSION: GC-MS chromatogram analysis of the methanolic extract of Phyllanthus acidus [Fig. 1] showed eleven peaks. In accordance to reference mass spectra of the constituents available in the NIST library, eleven peaks of GC-MS spectrum revealed that squalene and others were identified as sugar, and fatty acids [Table 1].

Figure 1: GC-MS chromatogram analysis of the methanolic extract of Phyllanthus acidus Table 1: Phytoconstituents of methanolic extract of Phyllanthus acidus Sr. No RT Name of Compound 1 3.87 4H-Pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl 2 6.17 -D-Glucopyranose, 4-o--D-galactopyranosyl 3 8.07 2-Decanynoic acid 4 11.25 -D-Glucopyranoside, O--D-galactopyranosyl-(1.fwdarw.3)--Dfructofuranosyl 5 12.13 n-Hexadecanoic acid 6 13.24 9,12-Octadecadienoic acid, methyl ester, (E,E)7 14.30 9,12,15-Octadecatrienoic acid, methyl ester, (Z,Z,Z)8 16.19 Lactose 9 18.82 1,2-Benzenedicarboxylic acid, mono(2-ethylhexyl)ester 10 22.16 Squalene 11 27.92 Z,Z,Z-4,6,9 Nonadecatriene

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MF C6H8O4 C12H22O11 C10H16O2 C18H32O16

MW 144 342 168 504

Peak Area % 23.65 31.91 20.58 2.23

C16H32O2 C19H34O2 C19H32O2 C12H22O11 C16H22O4 C30H50 C19H34

256 294 292 342 278 410 262

10.09 0.37 7.91 2.37 0.21 0.47 0.22


Eleven peaks have revealed in the GC-MS spectrum of the methanolic extract of Phyllanthus acidus. From the results, squalene is present as major bioactive compounds in methanolic extract of Phyllanthus acidus fresh leaves. Other phytocomponents are found to be fatty acids, lipid, sugar derivatives. Although it is need to fractionate extract by using different solvents which may helpful to explicate isolated phyto-compound and its pharmacological assays. In our previous studies, we have observed that squalene, 9, 12, 15-Octadecatrienoic acid and Hexadecanoic acid are commonly found in the most plants. [6], [7]

CONCLUSION: The present data provides to find the molecular formula and weight of 11 biomolecules. Further investigation should be done to isolate bioactive compounds and their structural elucidation and screening of pharmacological activity in the drug development.

ACKNOWLEDGEMENT: Authors are gratefully acknowledged to Dr. S. Kumaravel, Quality Manager, Food Testing Laboratory and the Director, Indian Institute of Crop Processing Technology (IICPT), Thanjavur for providing GC-MS facility to carry out the work.

CONFLICT OF INTEREST: None declared.

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