Gas Chromatography-Mass Spectrometry Analysis of Chloroform Extract of Coccinia grandis Voigt

 

Y. H. Momin¹*, V. C. Yeligar²

¹Department of Pharmaceutical Chemistry, Annasaheb Dange College of B. Pharmacy, Ashta 416301, India.

²Department of Pharmaceutical Chemistry, Oxford College of Pharmacy, Bangalore, 560078, India.

 

ABSTRACT:

Background: Gas chromatography-Mass spectroscopy is comprehensive techniques for identification of bioactive components present in medicinal plants. Objective: Objective of present study was analysis of chloroform extract of stem part of Coccinia grandis Voigt plant belonging to Cucurbitaceae by Gas chromatography- Mass spectroscopy (GC-MS). Material and Methods: After Successive extraction with petroleum ether, chloroform and ethyl acetate for selected plant, further Chloroform stem extract was analyzed for different bioactive compounds by GC-MS method. Result and Discussion: The GC –MS data of chloroform extract of stem part of Coccinia grandis Voigt plant showed 24 bioactive phytoconstituents such as Spiro [4.5] dec-6-en-8-one, Hexadecanoic acid methyl ester, Cyclononasiloxane octadecamethyl, 9-Heptadecanone, 2, 4-Di-tertbutylphenol, 1, 7-dimethyl-4-(1-methylethyl), 7, 9-Di-tert-butyl-1-oxaspiro (4, 5) deca-6, 9-diene-2, 8 dione. Hexadecanoic acid and 2, 4-Di-tert-butylphenol are an antioxidant agents and Spiro [4.5] dec-6-en-8-one 1, 7- dimethyl-4-(1-methylethyl), 9 heptadecanone, Hexadecanoic acid methyl ester shown antimicrobial property.

Conclusion: Present study concluded that development of potential bioactive compounds identified from GC MS analysis could serve better approach for treatment of various diseases such as microbial infection, fungal infection, inflammation, diabetes mellitus.

 

 

 

INTRODUCTION:

Medicinal plants have become beneficial source for the treatment of different ailments. It can be observed from ancient literature about the utilization of natural medicinal plants or herbal products. The plants are rich wellspring of saponins, flavonoids, alkaloids, glycosides, sterol, and tannins. These phytoconstituents contribute different therapeutic properties. The phenolic compounds and flavonoids possesses antioxidant activity, anticancer activity etc.¹ Frequent utilization of synthetic drugs are not moderate and delivers adverse reactions, subsequently herbal remedies may supportive as economical and therapeutic.

 

Coccinia grandis Voigt is a perennial creeper or climber plant from Cucurbitaceae family. It is usually known as Ivy gourd or scarlet gourd or kowai fruit or kundru. This plant grows up to 13cm covering small trees, shafts, and buildings. It comprises of 5 lobed glabrous, broad leaves which is 5 to 10cm in length. The upper surfaces of leaves are hairless while lower surface is hairy with heart or pentagon shape. It consists of single tendrils. Juvenile fruit of Coccinia grandis Voigt contains white streaks and complete ripe fruit is of bright scarlet. Seeds are rounded at apex, yellowish grey in color, ovoid and compressed. Flowers are 2-3cm long, female and male flowers rise at axils on petiole and contain 3 stamens. Coccinia grandis Voigt plant has long, thick, tuberous root with fibrous fracture². The stem of these plants are smooth, green shading and slender when immature however as it develops it gets broad, swollen and glabrous in nature. Coccinia grandis Voigt has been used to treat diabetes mellitus³, bronchitis, urinary tract infection, ulcer, microbial infection, oxidative stress, jaundice.

Variety of techniques can be used for identification of phytoconstituents from medicinal plants. GC MS is one of the powerful techniques for identification of secondary metabolites present in medicinal plants which may supportive for isolation of phytoconstituents of interest.^4,5,6 This technique utilizes specific test which confer positivity in identification of compounds from plant extracts^7. Gas chromatography confers the separation of the components from mixture and mass spectroscopy analyzes single components individually⁸.

 

Present study has been made to investigate phytoconstituents from stem chloroform extract of Coccinia grandis Voigt plant by Gas chromatography- Mass spectroscopy method. This study has not been carried earlier as per literature survey.

 

MATERIALS AND METHODS:

Collection and authentication of plant

The stem part of Coccinia grandis Voigt was gathered from Sangli and nearby towns of Sangli in the month of October. The authentication of plant was carried out from Botanical Survey of India Pune by Dr. Priyanka Ingale (Scientist C). The voucher specimen number is BSI/WRC/IDEN.CER./2019/H3/ 138. The stem was cut into small pieces, washed thoroughly with water to remove dust, and dried under the shade at room temperature for 10 to 15 days.

 

Preparation of extract:

The dried pieces of stem of plant were crushed in an electrical processor and made coarse powder for further extraction. Two Kilogram coarse powder was exposed to continuous hot soxhlet extraction at 60⁰C for 20 hours for petroleum ether; chloroform and ethyl acetate solvents respectively. The filtrates obtained from continuous hot extraction for each solvent were evaporated to dryness. The yield of petroleum ether, chloroform, ethyl acetate extracts was 16 g, 15 g, and 9 g respectively. These extracts were stored at 0-4⁰C in the refrigerator. Further chloroform stem extract was used for GC MS analysis. Because further study aims for isolation of non polar potent phytoconstituents and chloroform is the one of non polar solvent which can dissolve non polar phytochemicals abundantly.

 

Phytochemical screening:

The phytochemical screening was carried out for chloroform stem extract of Coccinia grandis Voigt plant to check presence of various classes of bioactive compounds such as sterol, alkaloids, glycosides, tannins, flavonoids, phenols, saponins, carbohydrates.^{9, 10, 11}

Identification of bioactive compounds by GC-MS:

GC-MS analysis for chloroform stem extract of Coccinia grandis Voigt was determined by GC-MS (Schimadzu) at Shivaji University Kolhapur, Maharashtra, India. The column temperature was programmed from 30⁰C initially and then heated up to 300⁰C at 10⁰C/5 min. highly pure helium was used as carrier gas at constant flow rate of 1.0 ml/ min^12,13,14. One µl sample was injected. Sample was run for 40 minutes. The compound identification was based on retention time and integral peak area.

 

RESULT AND DISCUSSION:

Phytochemical screening:

Phytochemical screening of Coccinia grandis Voigt stem chloroform extract revealed the presence of alkaloids, glycosides, carbohydrates, phenols, sterols, saponins, fatty acids esters which have potential to treat the various ailments. Table 1. exhibited presence of phytoconstituents.

 

Table 1: phytochemical screening of stem chloroform extract of Coccinia grandis Voigt plant.

Sr. No.	Phytochemical Tests	Chloroform stem Extract of Coccinia grandis Voigt
1.	Test of Sterol	+
2.	Test of Flavonoids	-
3.	Test of Tannins	-
4.	Test of Glycosides	+
5.	Test of Carbohydrates	+
6.	Test of Alkaloids	+
7.	Test of Saponin	+
8.	Test of Phenol	+

+ Sign indicates presence of test, - sign indicates absence of test.

 

Identification of bioactive compounds by GC MS:

GC MS is one of the powerful techniques for determination of bioactive compounds from medicinal pants. Abundantly compounds present in chloroform stem extract of Coccinia grandis Voigt plant were Spiro [4.5] dec-6-en-8-one, Hexadecanoic acid methyl ester, Methyl stearate, Cyclononasiloxane octadecamethyl9- Heptadecanone, 2,4-Di-tert-butylphenol, 1,7dimethyl-4-(1-methylethyl),7,9-Di-tert-butyl-1-oxaspiro (4,5) deca-6,9-diene-2,8 dione. From GC MS data, this stem chloroform extract revealed the presence of phenolic compounds, terpenoids, hydrocarbons, fatty acids which possesses antioxidant activity, antidiabetic activity and antimicrobial activity. GC-MS data confirmed the various phytoconstituents present with different retention times as described in table 2. This data may further help to isolate bioactive constituents of interest. The figure no.1 exhibited bioactive constituents present in Coccinia grandis Voigt stem chloroform extract.

 

Table 2: GC MS analysis of chloroform stem extracts of Coccinia grandis Voigt plant.

PEAK	RETENTION TIME	MOL. FORMULA	MOL. WT	AREA %	NAME OF COMPOUNDS
1	14.054	C12H24	168	0.40	Cyclohexane, 1,5-diethyl-2,3-dimethyl-
2	14.131	C14H28	196	1.30	3-Tetradecene, (E)-
3	14.515	C16H30	222	1.06	Cyclohexane,1,1'-(1,2-dimethyl-1,2-ethanediyl) bis
4	15.616	C12H24	168	1.20	Cyclohexane, hexyl-
5	18.272	C14H28	196	0.93	Cyclohexane, 1,2,4,5-tetraethyl-
6	18.787	C16H30	222	1.03	2,2-Dicyclohexylbutane
7	19.967	C15H30	210	3.90	1-Pentadecene
8	20.409	C15H30	210	1.06	1,7-Dimethyl-4-(1-methylethyl) cyclodecane
9	21.528	C14H28	196	1.49	Cyclohexane, octyl-
10	23.269	C14H22O	206	4.05	2,4-Di-tert-butylphenol
11	23.828	C14H28	196	0.65	Cyclohexane, 1,2,4,5 tetraethyl
12	25.173	C15H30	210	6.37	1-Pentadecene
13	26.780	C17H34	238	1.92	Cyclohexane, undecyl-
14	27.740	C15H24O	220	12.06	Spiro [4.5] dec-6-en-8-one, 1,7-dimethyl-4-(1-methylethyl)-
15	29.848	C19H38	266	7.09	1-Nonadecene
16	31.606	C17H34O	254	6.82	9-Heptadecanone
17	32.416	C17H24O3	276	3.24	7,9-Di-tert-butyl-1-oxaspiro (4,5) deca-6,9-diene-2,8- dione
18	32.685	C17H34O2	270	9.48	Hexadecanoic acid, methyl ester
19	34.075	C19H38	266	6.39	1-Nonadecene
20	36.673	C19H38O2	298	8.91	Methyl stearate
21	37.938	C19H38	266	8.87	1-Nonadecene
22	40.365	C18H54O9Si9	666	7.94	Cyclononasiloxane, octadecamethyl-
23	41.506	C27H56O	396	2.73	1-Heptacosanol
24	41.596	C21H44	296	1.12	Heneicosane
				100.00

Mol Formula: molecular Formula and Mol. Wt: Molecular Weight.

 

 

Fig. 1 Gas Chromatography- Mass Spectroscopy spectrum of chloroform extract of Stem part of Coccinia grandis Voigt.

 

CONCLUSION:

The GC MS analysis of stem part of Coccinia grandis Voigt chloroform extract revealed presence of 24 constituents and further investigation for isolation study could be helpful for treatment of various ailments because isolated and optimized phytoconstituents from the plants could serve as a better candidate in drug discovery. Hexadecanoic acid¹⁵ serves as antioxidant agent, 2, 4-Di-tert-butylphenol is an antioxidant and antifungal agents, ¹⁶ Spiro [4.5] dec-6-en-8-one 1, 7-dimethyl-4-(1-methylethyl), 9 heptadecanone, Hexadecanoic acid methyl ester shown antimicrobial, anti-inflammatory property¹⁷ from reported literatures. Thus further isolation and findings for mechanism is necessary which could open the new avenues in the drug development process.

 

ACKNOWLEDGEMENTS:

The author thankful to Annasahebji Dange, Founder President, Prof. R.A. Kanai Sir, Executive director of Annasaheb Dange College of B. Pharmacy, Ashta and Principal Dr. R.B. Jadhav Sir for providing facilities and giving support to carry out this research work. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

 

CONFLICT OF INTEREST STATEMENT:

There is no conflict of interest.

 

FUNDING:

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sector.

 

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Received on 30.12.2019           Modified on 20.03.2020

Accepted on 09.05.2020         © RJPT All right reserved