INTRODUCTION:

Medicinal plants are of great importance to the health of individuals and communities. The most important of these bioactive constituents of plants are alkaloids, glycosides, flavanoids, carbohydrates, tri-terpenoides, tannins, and phenols¹. Plants play a vital role in traditional medicine and pharmacopoeial drugs.

A large proportion of the world population depends on traditional medicine because of the scarcity and high costs of orthodox medicine. Many plants consist of different phytopharmaceuticals. These are having important applications in the fields of agriculture, human and veterinary medicine. The natural products play an important role in the novel development for the treatment and prevention of diseases².

A large number of medicinal plants and their purified constituents have shown better therapeutic effects. In order to promote the use of medicinal plants as potential sources of anticarcinogenic agents, it is important to exhaustively inspect their composition and activity and thus validate their use. Some phytochemicals produced by plants have anticancer activity and used for the development of new anticarcinogenic drugs³. It has been shown that in-vitro screening methods could provide imparts the required preliminary observations to determine crude plant extracts with potentially useful properties for further chemical and pharmacological investigations⁴.

The determination of phytoconstituents is largely performed by relatively expensive and often laborious techniques such as gas (GC) and liquid (LC) chromatography combined with specific detection schemes. Analysis of small amounts of chemicals has become easier and more cost-effective owing to the development of hyphenated chromatographic techniques such as GC or LC-MS^5-10. GC-MS analysis can identify pure compounds present at less than 1gm. The geographic characteristics of Chloris barbataare plant and narrow, linear and lanceolate leaf blades, 10-20 cm long, 2-3 mm wide and usually bluish-green with rough edges. They often have long scattered hairs on the upper surface, near the base. In the last few years spectroscopic methods have become explosively found as a key technological platform for secondary metabolite profiling in both plant and non plant species. To create optimum interest and awareness amongst the cultivation of medicinal plants. Criteria for selection of Chloris barbata are easily available, lack literature and more active principles presented. Chloris barbata isan important medicinal plant and widely used in around the world but especially in the tropical and sub tropical regions. It is belongs to the family of poaceae and external used for the skin disorders. Chloris barbata leaves used in fever, diarrhea and diabetics^11-12. From literature revelled that lack of sufficient data about phytochemical evaluation of Chloris barbata plant extract was not published. From Literature survey confirms that there is no method available for phytochemical evaluation for chlorisbarbata plant extract by using UV-VIS, FTIR and GC-MS. Therefore, the present study was conducted to investigate the phytochemical constituents of Chloris barbatausing UV-VIS, FTIR and GC-MS.

MATERIALS AND METHODS:

Collection and processing of plant material:

The leaves of the plant Chloris barbata were collected from the natural habitats of west godavari district, Andhra Pradesh, India. The samples were washed with sterile distilled water. The leaves were cut, shade dried, ground into fine powder and stored in air tight polythene bags until use.

Sample plant extraction:

5 g of air dried powder of leaf sample was extracted with 100 ml of solvent such as methanol with gentle stirring for 48 h. The sample was kept in dark for 48 h with intermittent shaking. After incubation the solution was filtered through Whatmann No. 1 filter paper and the filtrate was collected (crude extract). It was then transferred to glass vials and kept at 4°C before use.

UV-VIS and FTIR Spectroscopic analysis:

The extracts were examined under visible and UV light for approaching analysis. For the UV-VIS and FTIR spectrophotometer analysis, the extracts were centrifuged at 3000 rpm for 10 min and filtered through Whatmann No.1 filter paper by using high pressure vacuum pump. The sample is diluted to 1:10 with the same solvent. The extracts were scanned in the wavelength ranging from 200-800 nm using Perkin Elmer Spectrophotometer and the characteristic peaks were detected. FT-IR analysis was conducted by using Perkin Elmer Spectrophotometer, which was used to identify the characteristic peaks and their functional groups. The peak values of the UV-VIS and FTIR were recorded.

GC-MS analysis:

10 g of powdered leaf sample is soaked with 30 ml methanol overnight and filtered through ash less filter paper with sodium sulphate (2g). The extract is concentrated to 1 ml by bubbling nitrogen into the solution. The extract contained both polar and non-polar phyto components. 2 μl of the methanolic extract of Chloris barbatawas utilized for GC-MS analysis. Agilent 5973N GC/MS system was used in the analysis utilized a HP 5 column [30 nm × 0.25 nm ID × 1μm df] and the components were separated using high pure helium as carrier gas at a constant flow of 1 ml/min. The 2 μl sample extract insert into the instrument was detected by the Photon multiplier tube with the aid of the Turbo mass 5.1 software. During the 12^th minute GC extraction process, the oven was maintained at a temperature of 250°C with 2 minutes holding. The injector temperature was set at 220°C (Quadrupole mass analyser). The different parameters involved in the operation of the Clarus 500 MS, were also standardized (Inlet line temperature: 200°C; Source temperature: 200°C). Mass spectra were taken at 70 eV; a scan interval of 0.5 s and fragments from 45 to 450 Da. The MS detection was completed in 18 minutes.

Components identification:

The relative percentage amount of each component was calculated by comparing its average peak area to the total areas. The detection employed the NIST (National Institute of Standards and Technology) Ver.2.0-Year 2005 library. The compound prediction is based on Dr. Duke’s Phytochemical and Ethno botanical Databases by Dr. Jim Duke of the Agricultural Research Service/USDA. Interpretation of GC-MS was conducted using the database of NIST having more than 62,000 patterns. The spectrum of the unknown component was compared with the spectrum of the known components stored in the NIST library. The name, molecular weight and structure of the components of the test materials were ascertained.

RESULTS AND DISCUSSION:

The qualitative UV-Vis spectrum profile of Chloris barbata, methanol extract was selected from 200 to 800 nm dueto sharpness of peaks and proper baseline. The profile showedthe peaks at 206 to 800 nm and the profile showed the peaks206.5, 296.5, 400 and 663 nm with absorption 1.378, 0.273,0.482, 0.116 respectively. The UV-Vis spectrum of Chloris barbata, the methanol extract was taken at the 1.378 and 0.273respectively and the results of UV were shown in Figure.1 and Table.1

Figure-1: UV-Visible spectrum of Methanol extract of Chloris barbata

Table-1: UV-VIS peak values of methanolic extracts of Chloris barbata

S. No	Wavelength (nm)	Absorption value
1	206.5	1.378
2	296.5	0.273
3	400	0.482
4	663	0.116

The FTIR spectrum was used to recognize the functional group of the active components based on the peak value in the region of infrared radiation. The results of FT-IR peak values and functional groups were represented in Figure.2 and Table 2. Conduct the next advanced phytochemical analysis technique of FT-IR the presence of various functional groups of different compounds was found. The solvent had its respective functional group like presence of hydrogen bonded acids, phenols, alcohols, amines, alkanes, alkenes, alkynes, aromatics, aldehydes and nitriles etc. Hence, the crude extracts treat with UV-VIS and FT-IR analysis is used for the identification of chemical constituents present in Chloris barbata. The results concern to GC-MS analysis leads to the identification of number of compounds from the GC fractions of the methanolic extract of Chloris barbata. GC-MS analysis is also issue the spectrum for the methanolic extract. The spectrum profile of GC-MS established the presence of 6 major components with the retention time 12.17, 13.26, 14.25, 15.33, 16.19, 17.9 respectively. This gas chromatogram manifest the relative concentrations of various compounds obtain to eluted as a function of retention time. The heights of the peak indicate the relative concentrations of the components present in Chloris barbata. These mass spectra are fingerprint of the compound which can be recognizing from the NIST data library. These compounds were identified through mass spectrometry attached with GC. These observations may be due to the nature of biological active components and the stronger extraction capacity of methanol could have been produced number of active constituents responsible for anticancer activity. The biological activities based on Dr. Duke’s Phytochemical and Ethnobotanical Databases were enclosed in table 3. The results disclose the presence of 6 different compounds namely were 2-carboxymethyl-3-methyl-cyclopentane carboxylic acid (11.57%), acetic acid, 3, 3, 6-trimethyl-4-oxo-3, 4-dihydro-2H-pyran-2-yl-ester (3.04%), ursodeoxycholic acid (33.58%), 1-octene, 2-methoxy (2.8), 2-propenimine, 3-[1-cyclohexenyl]-N-cyclohexyl, N-oxide (40.4%) and 1, 3, 12-Nonadecetriene (7.8%). The GC-MS results were shown in Figure.3 to 10 and Table.3.

Figure-2: IR spectrum of Methanol extract of Chloris barbata

Table-2: FTIR peak values and functional groups of methanolic extracts of Chloris barbata

S. No	Peak value	Stretching’s	Functional Group
1	3463.92	N-H	Amines
2	3383.96	N-H	Amines
3	3270.87	O-H	Hydrogen bonded Acids, Phenols, Alcohols
4	2926.37	C-H	Alkanes
5	2786.68	CHO	Aldehydes
6	2686.85	CHO	Aldehydes
7	2571.93	O-H	Hydrogen bonded Acids, Phenols, Alcohols
8	2448.18	C=C	Alkynes
9	2210.56	C=N	Nitriles
10	2057.36	C=C	Aromatics
11	1844.50	=C-H	Aromatics
12	1627.73	C=C	Alkenes
13	1491.00	C=C	Aromatics
14	1401.47	C-O	Phenols
15	1338.02	C-O	Phenols
16	1266.64	S=O	Thiol

Figure-3: Gas chromatography and mass spectroscopy (GC-MS) analysis of methanol sample of Chloris barbata

Table-3: Phytocomponents identified in methanolic plant extract of Chloris barbata

S. No	Retention time (min)	Name of the compound	Molecular formula	Molecular weight (gm/ml)	Peak area (%)	Nature of the compound	Biological activity
1	12.17	-2-Carboxymethyl-3-methyl-cyclopentan carboxylic acid	C₈H₁₁O₄	171.172	11.57	Carboxylic acids	Glutamine synthesis Asparagines synthesis Glycinamide Ribonucleotide synthesise
2	13.26	Acetic acid 3,3,6-trimethyl-4-oxo-3,4-dihydro-2H-pyran-2-yl-ester	C₁₀H₁₄O₆	230.216	3.04	Esters	Antimicrobial Activity
3	14.25	Ursodeoxycholic acid	C₂₄H₄O₄	392.58	33.58	Bile acids	Gene expression and Cellular function
4	15.33	1-octene,2-methoxy	C₉H₁₈O	142.242	2.8	Alkene	Antioxidative, Antidiarrhoeal,Antimicrobial
5	16.19	2-propenimine,3-(1-cyclohexenyl)-N-cyclohexyl, N-oxide	C₁₅H₂₄NO	234.363	40.4
6	17.9	1,3,12-Nonadecatriene	C₁₉H₃₄O₂	294.479	7.8	Alkene	Antioxidative, Antidiarrhoeal,Antimicrobial

Figure-4: Mass spectrum of 2-carboxymethyl-3-methyl-cyclopentane carboxylic acid

Figure-5: Mass spectrum acetic acid, 3,3,6-trimethyl-4-oxo-3,4-dihydro-2H-pyran-2-yl-ester

Figure-6: Mass spectrum ursodeoxycholic acid

Figure-7: Mass spectrum of 1-octene, 2-methoxy

Figure-8: Mass spectrum of 2-propenimine, 3-[1-cyclohexenyl]-N-cyclohexyl,N-oxide

Figure-9: Mass spectrum of 1, 3, 12-Nonadecetriene

SUMMARY AND CONCLUSION:

The phytochemical screening of the Chloris barbatawas studied and showed that the leaves were rich in alkaloids, flavonoids, carbohydrates, glycosides, tannins, phenols and triterpeniods. GC-MS analysis was done using the organic solvent methanol and it shows the presence of six different chemical compounds present in the Chloris barbata leave sample. Hence, the identified phytocomponents in Chloris barbataby using GC-MS can be used as a pharmacognostical tool for the identification of adulterants. It covers the way for the development of several treatment regime based on this extract. In addition, further research is necessary to identify and purify the active compounds responsible for therapeutic activity for Chloris barbataplant.

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Received on 14.08.2019 Modified on 30.08.2019

Asian J. Research Chem. 2019; 12(5):268-273.

DOI: 10.5958/0974-4150.2019.00050.6