Phytochemical screening and Gas chromatography - mass spectrometry analysis on Ischaemum pilosum (Kleinex Willd.)

Ischaemum pilosum (Kleinex Willd.) a weed among the grass is reported for ethno - medicinal practices for treatment of various treatments for human and domestic animals. The current work deals with phytochemical analysis in different parts of plants to find out bioactive compounds. The first - time reported results on I. pilosum reveal the significant phytochemicals by using preliminary phytochemical analysis, UV Visible spectral technique, FTIR analysis and GC - MS analysis. The preliminary phytochemical test confirms the presence of alkaloids, anthraquinone, cardiac glycosides, coumarins, flavonoids, glycosides, phenols, reducing sugars, saponins, steroids, tannin and triterpenes in Ischaemum pilosum. UV Visible spectra and FTIR gives the ranges of absorptions and functional group like Carboxylic acids (O - H) at 2956,92 cm - 1 , Alkanes (O - H) at 2849,89 cm - 1 , Aldehydes (C=O) at 1735,92 cm - 1 , Aromatic Rings (C=C) at 1462,95 cm - 1 , Alkanes (C - H) at 1377,97 cm - 1 , Esters (C - O) 1166,95 cm - 1 and Phenyl Ring (C - H) 758,97 cm - 1 . The GC - MS analysis related twenty - one compounds like Phenol, 4 - bis (1,1 - dimethylethyl), Pentanoic acid, 5 - hydroxy, 2,4 - di - t - butylphenyl esters, E - 15 - Heptadecenal, 1 - Hexadecanol, n - Hexadecanoic acid, l (+) - Ascorbic acid 2,6 - dihexadecanoate, Palmitic anhydride


Introduction
The medicinal value of plants lies in their specific chemical substances that produce a definite physiological action on the human and animal body (1)(2)(3).In many parts of the world plant-based medicine are used for human and animal illnesses from thousands of years due to its bioactive compounds (4)(5)(6).Plants used in ethno-medicinal practices are believed to have potential sources for effective and safe treatment due to their constituents (7).The most important bioactive constituents among these medicinal plants are alkaloids, flavonoids, tannins, and phenolic compounds.Due to great medicinal potential plants are used in various cultures and traditions and now-a-days in modern pharmacopoeial drugs (8)(9)(10).A large number of phytochemical investigations and therapeutics compound isolation are in practice (11)(12)(13).Plant-based different purified constituents and crude drugs have been reported for their valuable therapeutic potential (14).To develop and promote the use of medicinal plants as potential sources for different modes of disease it is important to investigate phytochemicals and validate their bioactivities (14)(15)(16).
Ischaemum pilosum belongs to the family Poaceae, locally named as Kunda, Konsand Khavon.It is perennial creeping grass with stolons that covers shoots at the base with dry striate leaf sheaths (Fig. 1).I. pilosum is found commonly as a weed in many states of India (17,18).Plant is a source of good lignocellulosic material for the preparation of ethanol (19,20).A decoction of root powder is given against urine stone for urine disorder.A teaspoon of root powder is used to increase the flow of milk in cattle (19).
The determination of phytochemical compounds is largely practiced by developing spectroscopic and chromatographic techniques in medicinal plants (8,21,22).GC-MS analysis can act as a valuable tool for identifying pure compounds present in plant extracts (23,(23)(24)(25)(26)(27).Similarly spectroscopic (UV-Vis, FTIR) methods together or separately can be used in this chemical analysis sense andalso act as supportive methods for phytoconstituents finding (28)(29)(30)(31).This study evaluates the phytochemical in Ischaemum pilosum and help to serve as a basis for the development of novel plant based drugs from weed plants.The work is alsoafirst-time report to validate the medicinal importance of plants by chemical extractions with phytochemical techniques.(17)(18)(19).

Plant collection and identification
Ischaemum pilosum (Klein.exWilld.)Wt. plants were collected from forests of Toranmal of Shahda Tahsil (21.840213°N, 74.456583° E) of Nandurbar district.The collected plant are taxonomically identified and the herbarium (Voucher number: ANP-2204) is stored in the Department of Botany, Jijamata College Nandurbar MS.

Preliminary phytochemical screening
Proximate analysis of leaves, stem and leaves powdered of Ischaemum pilosum are taken with the estimation of moisture content, dry matter content and ash content as determined using the AOAC method (35)(36)(37)(38).Leaves, stem and root parts'soxhlet extract in six different solvents like aqueous (Aq), methanol (M), ethanol (E), pet-ether (Et), chloroform (C) and acetone (A) were used to perform preliminary phytochemicals test to identify phytochemical constituents (Table 01.) (39,40).

Spectral Analysis
UV Visible spectral analyses were carried out for leaves, stem and root soxhlet extract in 60% concentration ethanol and chloroform solvents with 200 nm to 11000 nm range (28,(41)(42)(43).Fourier Transform Infrared Spectrophotometer (FTIR) is perhaps the most powerful tool for identifying the types of chemical bonds (functional groups) present in compounds.The wavelength of light absorbed is characteristic of the chemical bond as can be seen in the annotated spectrum.By interpreting the infrared absorption spectrum, acetone leaves samples of Ischaemum pilosum extracts were selected.10 mg of the dried extract powder was encapsulated in 100 mg of KBr pellet, in order to prepare translucent sample discs.The powdered sample of the plant was loaded in an FTIR spectroscope (Shimadzu, IR Affinity 1, Japan), with a scan range from 400 to 4000 cm 1 and a resolution of 4 cm 1 .The intense bands of FTIR results were compared with standard values to recognize the functional groups of chemical compounds present in the leaves sample.(40,(43)(44)(45)(46)(47).

Gas chromatography-mass spectrometry (GC-MS) analysis
GC-MS is one of the hyphenated analytical techniques used for Ischaemum pilosum acetone leaves extract by thesoxhlet extraction method.GC-MS study was conducted as per the Crop Processing Research Institute, Tanjore.The Clarus 500 GC used in the analysis used a fused silica column loaded with Elite-1 (100% dimethyl siloxane poly, 30 nm x 0.25 nm DI x 1 μmdf) and the components were separated using helium as a carrier gas at a constant rate of 1 mL/min.The Turbo Gold mass detectors (Perkin Elmer) were used to detect the 2 μl leaves acetone extract injected into the instrument using the Turbo mass 5.1 software.The injector temperature was set at 250 °C (mass analyzer).The parameters involved in the operation of the Clarus 500 MS have been standardized (input line temperature: 200 °C, source temperature: 200°C ). Mass spectra were taken at 70 eV; a scan interval of 0.5 s and fragments of 45 to 450 Da and MS detection was completed in 36 min (23,48).

Identification of compounds
GC-MS interpretation was performed using the NIST database (Multi format version of NIST (NIST20MF)) with more than 62,000 models.The spectrum of the unknown component was compared with the spectrum of known components stored in the NIST library.Prediction of the compound from the GCMS library was done with the help of Dr. Duke's phytochemical and ethnobotanical database by Dr. Jim Duke, Agricultural Research Service / USDA.The name, molecular weight and structure of the compounds were determined for acetone leaves extract (40,(49)(50)(51)(52).

Result Preliminary phytochemicals
Proximate Analysis for moistures content, dry matter content and ash content is performed for Ischaemum pilosum leaves, stem and root parts as shows overall 52.16 % of moisture content, 47.33 % dry matter content and 30.33 % ash content in the plant (Table 02).Preliminary phytochemical test for the leaf, stem and root parts extract of I. pilosum for alkaloids, anthraquinone, cardiac glycosides, coumarins, flavonoids, glycosides, phenols, reducing sugars, saponins, steroids, tannin and triterpenes were found in different solvents (Table 02).

GC-MS profiling of Ischaemum pilosum leaves acetone extract
GC-MS test for leaves acetone extract were carried out for chemical compounds present in samples.Twenty-one compounds were listed from sample of Ischaemum pilosum leaves and identified by GC-MS analysis (Fig. 5).The listed, identified compounds are like Phenol, 4-bis (

Discussion
Preliminary phytochemical screening of Ischaemum pilosum extracts reveals the presence of flavonoids that have been reported for antioxidant activities (10).The presence of flavonoids is a highly effective scavenger in various free radicals implicated in many diseases (53,54).The presence of alkaloid and terpenoids are reported for gastric ulcers (55).
Terpenoidsare also important in cardiovascular muscle relaxation (56).Steroid's presence in leaves sample helps to relieve swelling and anti-inflammation properties (57,58).The presence of tannins have been reported as healing agents for inflammation and burn (7).Other phytochemicals present like anthraquinone, cardiac glycosides, coumarins, glycosides, phenols, reducing sugars and saponins are also reported in preliminary phytochemical reports are having various biological activities (7,59).Moreover, UV spectral and FTIR studies give absorbance peaks that reveals the functional group alkanes, aldehydes, aromatic rings and phenyl ring helps to confirm the structure (43,60).
The identification and comparison of the GC-MS analyzed compounds were performed using established databases such as NIST and Dr. Duke's Phytochemical and Ethno-botanical databases (68).This allowed for accurate matching of the mass spectra and retention times of the compounds, aiding in their identification.The results of

Conclusion
This study first time reported Phytoconstituents from Ischaemum pilosum with GC-MS reports.The conformation tests of phytochemicals for alkaloids, phenolic etc helps to support the ethno-medical importance of the plant reported in the literature.Further detailed subsequent phytochemical analysis up to GC-MS analysis found 21 different phytoconstituents.The compounds like Phenol, 4 -bis (1, 1-dimethylethyl), Eicosane, n-Hexadecanoic acid, 2, 6-bis (1,1-dimethylethyl) phenol and heptadecane are having bioactive and therapeutic abilities as reported by various authors.In addition, many other compounds found in leaves' acetone extracts could have biological activities.We suggest the utilization of this weed for further pharmaceutical processing to make plant-based medicine from weed.This research contributes to the growing body of knowledge on plant-derived compounds and their potential applications in various fields, including medicine and drug discovery.Further investigations and clinical studies are needed to harness the full therapeutic potential of Ischaemum pilosum.

Table 02 .
Proximate analysis test

Table 04 .
FTIR spectral peak values and functional groups

Table 05 .
Phytochemical Components found in acetone leaves extract of Ischaemum pilosum by GC-MS results

(g/mol) Structure of Compound
Ischaemum pilosum.The antimicrobial, antifungal, antitumor, antibacterial, antiinflammatory, antimalarial, antioxidant, and cytotoxic activities associated with these compounds indicate their potential as valuable therapeutic agents of plant.Further studies are warranted to explore their mechanisms of action and potential applications in the field of medicine.