Int J Pharm Pharm Sci, Vol 13, Issue 2, 7-13Original Article

ANALYSIS OF BIO-ACTIVE COMPOUNDS PRESENT IN THE LEAVES AND STEM OF TRICHOSANTHES ROXB. USING GC-MS TECHNIQUE WITH RESPECT TO ITS ANTI-INFLAMMATORY ACTION

AGHIL SOORYA ARAVINDAKSHAN*, SEKAR THANGAVEL

Department of Botany, Bharathiar University, Coimbatore-46, Tamil Nadu, India
*Email: aghilsoorya@gmail.com

Received: 12 Nov 2020, Revised and Accepted: 28 Dec 2020

ABSTRACT

Objective: Structural elucidation studies on Trichosanthes lobata ethyl acetate and methanol extracts of leaf and stem parts through Gas Chromatography-Mass Spectrometry (GC-MS) technique with respect to anti-inflammatory potential.

Methods: Extracts obtained with shade dried and powdered samples in successive solvent extraction using ethyl acetate and methanol by Soxhlet apparatus and subjected to GC-MS analysis and interpreted for its anti-inflammatory compounds.

Results: The study revealed that the extraction solvent used was able to recover compound of classes such as organic acid esters and conjugated alkaloids in larger quantities than other classes of compounds and they varied with leaf and stem and also with the polarity of solvents used. In total compounds identified, GC-MS profile of the Ethyl Acetate leaf extract of T. lobata contained 41 compounds, stem extract contained 45 compounds which have reported bioassays in PubChem. Whereas GC-MS profile of methanol leaf extract of T. lobata contained 66 compounds and stem extract contained 46 compounds having bioassay reports in PubChem. A large number of phytochemical peaks with good area percentage were found in methanolic extract. We were also able to find out potent anti-inflammatory compounds including Octanoic acid, Dodecanoic acid, Octadecane, Enoic acid, Hexanoic acid, Quinazolin-8-one, Ilicic acid, Pentadecanoic acid, Oxaspiro, Benzeneacetic acid, etc. from the extracts.

Conclusion: T. lobata contains phytocompounds against inflammation which may serve as a new drug lead of natural products origin in future and make it employable in modern pharmacological practices.

Keywords: Ethyl Acetate, Methanol, Trichosanthes lobata, GC-MS, Phytochemicals, Leaf, Stem

© 2021 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (https://creativecommons.org/licenses/by/4.0/)
DOI: https://dx.doi.org/10.22159/ijpps.2021v13i2.40236. Journal homepage: https://innovareacademics.in/journals/index.php/ijpps.

INTRODUCTION

Basically, nature is the first source of medicinal compounds, from which bigger and accurate molecules are being formed. Since ancient times the natural substrates have been used as a source of various products applied in food, drug, cosmetic, textile and energy [1]. These naturally available products of higher plants serve as traditional medicine, possibly due to scientifically not proven mechanisms of action. The natural compounds exhibit various beneficial biological activities such as antimicrobial, antioxidant, anticancer, anti-inflammatory, anti-obesity, anti-angiogenic, neuroprotective activities, etc. Therefore, various phytochemicals isolated from plant sources have attracted much attention in the field of pharmacology [2]. Trichosanthes species belongs to Cucurbitaceae family. Generally, fruits of this species are proved to have good sources of phenolics and antioxidants, also possesses anticancer, antiproliferative, cardioprotective, antioxidant properties, etc. Many research studies have reported that it contains important protein called Ribosome Inactivating Protein which helps in the formulation of anticancer agents. In general, biological components present in plants are considered as a stimulant for its medical properties like anti-inflammatory, antidiabetic, antiulcer and cardioprotective activities. Similarly, there are various herbal preparations from Trichosanthes species being prescribed widely for the treatment of inflammatory conditions. One such plant is Trichosanthes lobata Roxb. Commonly called as patolam, kaypanpatolam, kattupatolam and peppatolam. It is a monoecious climber, an indigenous species therapeutically used in the tribes of Idukki hills, Kerala, India. These species have shown the presence of active constituents like flavonoids, phenolics, carotenoids, saponins, triterpenoids, etc. Although the extracts of this plant have been traditionally used in the treatment of inflammations, there is no scientific evidence which supports this therapeutic use. Our earlier research has documented its acute and chronic in vivo anti-inflammatory activities. The overall results obtained confirmed that the extracts of Trichosanthes lobata contain anti-inflammatory substances.

Inflammation refers to the reaction of living tissues to overcome injury, infection or irritation. Lysosomal enzymes are released during inflammation producing a variety of disorders leading to tissue injury by damaging the macromolecules and lipid peroxidation of membranes. These activities are presumed to be responsible for certain pathological conditions such as heart attacks, septic shocks, rheumatoid arthritis, etc. The extracellular activity of these enzymes is said to be related to acute or chronic inflammation [3]. Thus, there is a need for natural anti-inflammatory agents to achieve increased pharmacological response and less side effects. In this regard, complementary, alternative and traditional medicine play a pivotal role in medication. While proving them through scientific methods gives more authentication to the practices followed. Fulzule et al. [4] studied anti-inflammatory activity of Trichosanthes cucumerina and it was evaluated by use of the carrageenan-induced paw edema model in Wistar rats. Also, the underlying mechanism by which T. cucumerina mediates the anti-inflammatory activity was assessed by determining its effects on membrane stabilizing activity and nitric oxide inhibitory activity. Inhibitions of nitric oxide (NO) production and membrane stabilization activities are probably the mechanisms by which ligand molecules mediate its anti-inflammatory actions. Those findings rationalized the traditional practice of this plant as an anti-inflammatory agent. They suspect that membrane stabilizing abilities and the absence of inhibitory action are possible mechanisms through which T. cucumerina arbitrates its anti-inflammatory action.

In this study, we have analysed the chemical profiling of ethyl acetate and methanol leaf and stem extracts of T. lobata, with special reference to their anti-inflammatory effects using by Gas Chromatography-Mass Spectrometry (GC-MS). The obtained compounds were checked for its anti-inflammatory properties in PubChem and tried to report the possible effect of plant extracts against inflammation.

MATERIALS AND METHODS

Plant collection and preparation of plant extracts

Fresh, healthy and matured leaves and stem of Trichosanthes lobata Roxb. were collected from Vattavada Panchayat (10 °10ʹ38.7ʺN 77 °15ʹ33.ʺE), Koviloor Post, Idukki District, Kerala state of India. The authenticity of the selected plant species was confirmed from the Botanical Survey of India, Southern Circle, Coimbatore (Vide No: BSI/SRC/5/23/2016/Tech./213). The leaves and stem were cleaned, dried in shade and ground well to a fine powder. About 50 g of dry powders were subjected to successive solvent extraction with ethyl acetate and methanol using Soxhlet apparatus. The filtered and dried extracts were subjected for GC-MS analysis.

Gas chromatography-mass spectrometry (GC-MS) analysis

The GC-MS analysis was carried out using the instrument Thermo GC-Trace Ultra Ver: 5.0, Thermo MS DSQ II with Column: DB 35-MS Capillary Standard Non-Polar Column possessing Dimension: 30 Mts, ID: 0.25 mm, Film: 0.25 μm. The initial temperature of the instrument was set to 110 °C and maintained for 2 min. At the end of this period, the oven temperature was raised up to 260 °C, at the rate of an increase of 6 °C/min, and maintained for 9 min. The temperature of injection port was ensured as 250 °C and the flow rate of Helium as 1 ml/min. An injection volume of 1 µl of sample is considered in the analysis. The ionization voltage was maintained to be 70 eV and samples were injected in split mode in the ratio 10:1. Mass Spectral scan range was set at 45-450 (MHz).

The chemical constituents were identified by GC-MS. The fragmentation patterns obtained from mass spectra were compared and analysed with spectrometer database using National Institute of Standards and Technology Mass Spectral database (NIST-MS). From the relative peak area of each component in the chromatogram the percentage of each component was calculated.

Table 1: Showing the list of compounds having pharmaceutical importance obtained from GC-MS profile of Ethyl acetate leaf extract of Trichosanthes lobata

S. No. RT (min) Compound name Molecular
formula		 Mol.
weight (g/Mol)		 Area % Biological activity*
1 14.43 Tetradecanoic acid C14H28O2 228 7.84 Antagonistic activity against Escherichia coli 
2 14.43 Octanoic acid C8H16O2 144 7.84 Prostaglandin E2 inhibitors, and Cox2
3 14.43 Dodecanoic acid C12H24O2 200 7.84 Antimicrobial and anticancer activity
4 14.43 D-Glucuronic acid C6H10O7 194 7.84 Apoptotic and cytotoxic activity
5 14.43 Dodecanoic acid C12H24O2 200 7.84 Blood pressure regulation and vascular inflammation
6 26.8 Trimethylsilyl C18H34O4Si3 398 4.21 Fungicidal activity
7 26.8 Phenazine, 5,10-dioxide C12H8N2O2 212 4.21 Anti-trypanosomal activity
8 28.8 Docosane C22H46 310 3.06 Antimicrobial activity
9 28.8 Octadecane C18H38 254 3.06 Immuno-stimulating properties, Anti-inflammatory activity
10 30.51 Benzoic acid C16H14O3 254 1.74 Drug metabolism in human liver
11 30.51 Benzopyran-4-One C16H14O3 254 1.74 Antiestrogenic activity
12 30.51 Fluoroquinazoline C15H11FN2O 254 1.74 Antifungal activity
13 33.50 Phthalic acid C23H30O4 370 3.19 Insecticidal, Antibacterial, Gastroprotective and cytotoxic activity
14 33.95 Diphenylpyridine C23H25N 315 3.98 Antiviral activity, Cytotoxicity
15 33.95 Quinone C24H14O6 398 3.98 Biomarker for cancer chemoprevention
16 34.28 Dimethylisoquinolin-8-ol C31H33NO4 483 4.94 Antibacterial activity and Cytotoxicity
17 34.28 Tetramethyl C26H28O8 468 4.94 Antimalarial activity, cox2 inhibition
18 34.28 Benzothiazin C11H13NOS 207 4.94 Antitubercular activity
19 34.28 Morphinane C22H17F6N3O2 469 4.94 Cell protective activity
20 35.09 Benzenepropanoic acid C11H14O3 194 2.73 Inhibitory activity against platelet aggregation
21 35.09 Tetrafluoroborate C18H22BF4N2O4PS 480 2.73 Anti-hemolytic, Antibacterial activity
22 35.93 Milbemycin B C32H44ClNO7 589 2.57 Insecticidal activity
23 35.93 Carboxylate C17H23NO2 273 2.57 Neuropsychiatric events
24 37.05 Propylamine C13H18FN 207 2.35 Cytotoxicity
25 37.05 Enoic Acid C11H11FO2 194 2.35 Inhibitors of fatty acid and cyclooxygenase
26 37.05 Butenoic acid C9H7BrO3 242 2.35 Antimicrobial, Effective dose to inhibit gastric acid secretion
27 37.25 Pyridine C14H11N3O 253 2.20 Drug metabolism
28 37.70 Methyl Ethyl Pyrazine C7H10N2 122 4.22 Cytoprotective activity
29 37.70 Propenoate C9H11NO3 181 4.22 Antiapoptotic, Cardioprotective effect
30 37.70 Pyrimidine C15H17FN2 244 4.22 Drug metabolism
31 37.70 Glucopyranoside C33H55NO7 577 4.22 Inhibition of Saccharomyces
32 38.21 Dichloro[tri(trimethylsilyl)methyl]Silane C10H28Cl2Si4 330 1.90 Inhibit Cholesterol Biosynthesis, Cytotoxicity
33 38.21 Diazepine C20H17N3O 315 1.90 Anticonvulsant activity
34 38.43 Carboxylate C27H35NO6 469 1.88 Neuropsychiatric events
35 38.43 Naphthalene C27H23N3O5 469 1.88 Inhibition of COX1 
36 38.43 Phenylimidazole C26H24N6OS 468 1.88 Cholesterol lowering effect
37 38.43 Isophthalate C11H8F3IO7S 468 1.88 Cancer drug development
38 38.43 Thiophene C29H24O2S2 468 1.88 Drug metabolism
39 38.82 Quinazoline C19H13Br2N3 441 3.87 Antimicrobial activity
40 39.17 Dihydrofuran C14H12O 196 1.87 Cytotoxicity Antibacterial activity
41 39.92 Tetrahydroisoquinoline C17H20N2O2 284 2.45 Drug metabolism

*Source of the Biological activities of the compound is PubChem database.

Identification of compounds

Using the database of National Institute Standard and Technology (NIST) having more than 62,000 patterns the interpretation of mass spectrum of GC-MS was conducted. The spectrum components of known compounds were compared with unknown components stored in the NIST library. The name, molecular weight and structure of the compounds were ascertained in above said way.

Identification of compounds having biological activity

The obtained list of compounds was checked for having biological properties which are reported in PubChem. All the compounds which are subjected for Bioassay are listed in tables and phytochemicals having anti-inflammatory action is considered as positive hits for our study.

RESULTS AND DISCUSSION

Characterization of secondary metabolites from medicinal plants provides an extensive resource for the isolation and identification of novel therapeutic agents for inflammation. The major non-volatile compounds can be identified by GC-MS analysis. The crude methanolic extract revealed the high peak intensity compound predominant presence of major compounds like pentadecanoic acid, 14-methyl-, methyl esters (RT-14.43) and methyl stearate among other derivatives. Minor compounds such as 10-methyl ester, methyl tetradecanoate, tetradecanoic acid, 12-methyl-methyl ester, 9-hexadecenoic acid, hexadecanoic acid, 14-methyl-methyl ester, 10-octadecenoic acid and heptadecanoic acid were also identified. These compounds are exhibited activities like antioxidant, cancer-preventive, hypercholesterolemic, nematicide, antifungal and antimicrobial. The advantage of GC-MS is its highest accuracy in the identification of derivatized compounds. The list of compounds from different extracts and plant parts are as tabulated in table 1-4 and the respective spectrum is as shown in fig. 1-4.

Table 2: Showing the list of compounds having pharmaceutical importance obtained from GC-MS profile of Ethyl acetate stem extract of Trichosanthes lobata

S. No. RT (min) Compound name Molecular
formula		 Mol. weight (g/Mol) Area % Biological activity*
1 13.90 Dimethylbenz Oate C14H20O4 252 0.33 Drug Metabolism and Pathologic Process 
2 13.90 Chloropyridine C11H8ClNO 205 0.33 Antinociceptive Activity
3 13.90 Tetralone C13H16OS2 252 0.33 Parkinson's disease and depression
4 13.90 Dihydroisobenzofuran C13H16O3 220 0.33 Drug-protein interactions at serotonin transporter
5 13.90 Dicarboxylate C15H13ClO6 324 0.33 Agonist for metabotropic glutamate receptors
6 13.90 Pyrazine C12H20N2Si 220 0.33 Antimalarial agent
7 14.43 Acetamide C24H20BrN3O2S2 525 0.29 Radioligand for benzodiazepine receptor in brain
8 14.43 Glaucine C21H25NO4 355 0.29 Antioxidant and Antiviral activity
9 14.43 Quinoline C17H15Br2NO 455 0.29 Drug metabolism
10 19.25 Carboxylate C13H18O5 254 0.37 Neuropsychiatric events in influenza 
11 19.25 Hexanoic acid C19H34N4O6 414 0.37 Inhibit cyclooxygenase activity
12 23.00 Hexadecanoic acid C18H36O2 284 0.44 Promote cancer cell death
13 24.92 Glucopyranose C15H19D3O10 362 0.34 Cytotoxicity
14 24.92 Oxodehydroabietate C21H28O3 328 0.34 Cytotoxicity
15 24.92 Cyclohexanol C13H24O2 212 0.34 MDR-reversal drug lead
16 26.00 Bromobenzene C15H13BrO2S 336 0.44 Anticancer,  Antimicrobial Activity
17 26.00 Cyclopropane C10H12S 164 0.44 Inhibition Assay
18 28.33 Phthalimide C19H16ClNO2S 357 0.38 Hypolipidemic activity
19 28.33 Colchicine C20H23NO5 357 0.38 Binding affinity
20 28.33 Diphenylquinoline C22H17N 295 0.38 Metabolic stability
21 28.77 Haliclonacyclamine C32H58N2 470 0.54 Cytotoxicity
22 29.42 Cyanopyridine C18H20N2 264 0.51 Antimicrobial activity
23 29.42 Ethanone hydrochloride C6H11NOS2 177 0.51 Antiproliferative activity
24 29.42 Oxazoline C21H25NO7 403 0.51 Ovicidal activity
25 29.89 Pyridinedicarboxylic acid C21H27NO5 373 1.92 Inhibitory against Hepatitis B
26 31.74 Stigmastane C29H52 400 0.29 Antimicrobial, Effective dose to inhibit gastric acid secretion
27 31.74 Phenylquinoxaline C15H10ClN3O2 299 0.29 Binding affinity to beta-amyloid in Alzheimer's disease
28 31.74 Glucopyranosiduronic acid C29H56N2O10S 676 0.29 Cytotoxicity
29 32.38

Methyl propyl 6-[3-(propylthiocarbonyl)benzyl]benzene-1-carboxylat

e-3-carboxythioic S-ester

 C23H26O4S2 430 0.72 Analgesic activity
30 32.86 Carbaldehyde C34H35N5O5 593 1.23 Inhibitory drugs for smoking reduction
31 32.86 Oxazolidinone C20H27NO4S 377 1.23 Antimycobacterial activity
32 32.86 Sanguinarine C22H19NO5 377 1.23 Antibacterial activity
33 33.31 Oxazoline C21H25NO7 403 0.27 Ovicidal activity
34 33.31 Tetrahydroisoquinolin-4-ol C19H20N2O6 372 0.27 Potentiating and inhibiting activity
35 33.70 Imidazole C16H10Br2N2O 404 0.46 Kinase Assay
36 33.70 Oxobutanoic acid C20H23FN2O5 390 0.46 Stimulated Gastric Acid Secretion
37 34.23 Phenylindole C16H15NO2 253 1.29  Pro-apoptotic in cancer cells
38 34.23 Azepine C14H14N2OSi 254 1.29 Antioxidant activity
39 34.64 Acetohydrazide C24H30N2O6 442 1.08 Inhibition of topoisomerase 2
40 34.64 Physodic acid C26H30O8 470 1.08 Drug metabolism 
41 39.16 ë(2,2')-Bis(1,3-dithio[4,5-c]selenophene) C10H4S4Se2 412 69.34 Antimicrobial activity
42 39.57 Quinolinecarboxylic acid C10H9NO4 207 3.56 Gastrointestinal disorder
43 39.57 4-Methylthio-3-quinolinesulfonamide C10H10N2O2S2 254 3.56 Antidepressant activity
44 40.24 4H-1-Benzopyran-4-one C28H26O10 522 5.24 Potent anti-estrogens.
45 40.42 N-Benzyl-2-Bromoaniline C13H12BrN 261 1.55 Inhibition of Fibroblast

*Source of the Biological activities of the compound is PubChem database.

Table 3: Showing the list of compounds having pharmaceutical importance obtained from GC-MS profile of Methanol leaf extract of Trichosanthes lobata

S. No. RT (min) Compound name Molecular
formula		 Mol. weight (g/Mol) Area % Biological activity*
1 15.51 Monohydroxamic acid C8H7NO4 181 2.78 Inhibition Of Leukemia Cells 
2 15.51 Piperidine C11H24N2O 200 2.78 Potential cocaine antagonists.
3 15.51 Serine C3H6DNO3 105 2.78 Inhibit, osteoarthritis, Alzheimer's, age-related macular degeneration
4 16.84 Arsine, trimethyl-(CAS) C3H9As 120 1.24 Decreased heart uptake with alkyl substitution
5 16.84 Pyridine C11H9NO2 187 1.24 Hypo-cholesteremic activity
6 19.85 Propanone C24H51NO5Si5 573 1.11 Antagonist activity
7 19.85 Quinazolin-8-one C20H20N2O5 368 1.11 Anti-inflammatory activity
8 19.85 Trimethylsilyl ester C20H40O5Si4 472 1.11 Antiviral activity and Cytotoxicity
9 21.77 Annulene C21H18 270 1.96 Cytotoxicity
10 21.77 Anthraquinone C15H10O3S 270 1.96 Anti-filarial activity, Cytotoxicity
11 21.77 Isoquinolin C28H21NO 387 1.96 Increase of intracellular drug uptake
12 21.77 Azulene C20H14O 270 1.96 Agonist Activity
13 22.03 Hexadecenoic acid C17H32O2 268 0.91 Antibacterial activity Anti-plasmodial activity
14 22.03 Hydroxy palmitate C19H36O4 328 0.91 Antihyperglycemic activity
15 22.87 Benzenedicarboxylic acid C16H22O4 278 1.09 Antitumor activity
16 22.87 phenylpropionate C13H17ClO2 240 1.09 Cellular uptake in human PBMC nucleus
17 22.87 Naphthalene C11H19NO3 213 1.09 Inhibition of HIV1 protease dimerization
18 22.87 Phenylpropan-1-one C17H18O2 254 1.09 Bio-reduction by bacterial ferredoxin
19 23.97 Methacrylate C21H32O3 332 0.86 Antiparasitic activity
20 23.97 Butyl acrylate C21H32O3 332 0.86 Cytotoxicity Antioxidant activity
21 23.97 1,3-Dithiolane C19H30S2 322 0.86 Detoxification activity
22 25.60 Octadecanoic acid C19H38O2 298 1.33 Highest toxicity against pathogens
23 28.25 N-phenylnitrone C16H21NO3 275 1.43 Cytoprotective activity
24 28.25 Carboxylic acid C13H17NO3 235 1.43 Anti-spermatogenic activity, Drug metabolism
25 28.25 Oxoquinoline C12H12N2O4S2 312 1.43 Antimicrobial activity
26 28.25 Methy L Dopamine C15H10Cl3F6N 503 1.43 Binding affinity towards human dopamine D4 receptor
27 28.25 Glucopyranosiduronic acid C29H57NO8Si5 687 1.43 Inhibition of HIV1 protease
28 28.25 Trimethylsilyl ester C29H57NO8Si5 687 1.43 Antiviral activity, Cytotoxicity
29 28.88 Glutamic acid dimethyl
ester		 C7H12DNO4 175 2.16 Inhibitory Activity Against Leukaemia Cells
30 28.88 Dihydropyrrole C11H5F16N 455 2.16 Mitotic arrest of ovarian carcinoma cells
31 29.73 Azetidinone C24H25NO6S 455 2.16 Reduction of liver cholesteryl esters
32 29.73 Quinoxalinone C27H25N3O4 455 2.39 Cytotoxicity
33 29.73 Methylphenol C26H35ClN4O 454 2.39 Antimicrobial activity
34 29.73 Argentatin C33H54O4 514 2.39 Inhibitory towards leukemic cell line
35 29.73 Isopropylamide C32H41NO 455 2.39 Inhibitor of beta-galactosidases.
36 31.32 Montiporyne C15H20O 216 5.25 Cytotoxicity
37 31.32 Chlorophenol C12H13ClO 208 5.25 Anti-trypanosomal activity
38 31.32 Dimethylindole C10H10N2O3 206 5.25 Growth inhibitory activity against pancreatic cancer cells.
39 31.32 Benzimidazole C11H13N3S 219 3.65 Reduced blood pressure 
40 31.32 Cyclopropane C20H18N2 286 3.65 Potential targets for anti-tuberculous drugs
41 32.20 Aminocyclohexane C25H26BrN2O 512 0.82 Antidiuretic activity
42 32.20 Dimethoxy indole C33H27N3O3 513 0.82 Antitumor activity
43 32.20 Methylpyrazole C20H20Br2N2 510 0.82 Drug metabolism in human liver, osteoporosis
44 32.93 Dimethylcoumarin C11H10O3 190 0.91 Inhibition of monoamine oxidases
45 33.12 Lupeol C30H50O 426 6.50 Reduce blood glucose, Cytotoxicity
46 33.12 Sclareol C20H36O2 308 6.50 Cytotoxicity, Induction of apoptosis
47 33.12 Farnesyl bromide C15H25Br 284 6.50 Anti-plasmodial activity
48 33.12 Geranylgeraniol C20H34O 290 6.50 Cytotoxicity
49 33.42 Illudinic acid C15H18O4 262 7.32 Antimicrobial activity, antitumor activity
50 34.36 Ilicic Acid C15H24O4 268 2.12 Anti-inflammatory activity
51 35.05 Dicarboxylic acid
dimethyl ester		 C27H32O5 436 10.51 Antagonist activity
52 35.05 Phthalazin-1(2H)-one C20H19N5OS 377 10.51 Antibacterial activity
53 35.89 Quinazolinone C27H22N4O3 450 2.01 Treatment for neurodegenerative disorders
54 35.89 Dimethoxybenzene C17H29BrO3Si 388 2.01 Cross allergenicity
55 35.89 Scopadulane C24H38O4 390 2.01 Cytotoxicity, Antiviral activity
56 36.13 Methylcrinasiadine C15H11NO3 253 2.31 Cytotoxic alkaloid
57 36.13 Methylpyrrole C13H19NO4 253 2.31 Antitumor Activity
58 36.84 Undecanolide C12H20O3 212 1.57 Antimicrobial activity, Cell cycle arrest in HepG2 cells
59 36.84 Galactitol C18H26O12 434 1.57 Antihyperglycemic agent
60 37.56 Aminodiphenylmethane C22H21NO2 331 0.98 Antimicrobial activity, Antiviral activity
61 37.8 Propargyl ether C14H20O2S 252 1.21 Antinociceptive tests
62 37.8 Phthalic acid C22H26O4 354 1.21 Insecticidal activity
63 38.41 Porphine C36H42N4O4 594 10.21 Cytotoxicity
64 38.78 Glucopyranosiduronic acid C27H52N2O10S 648 12.27 Cytotoxic activity, Anti-HIV-1 protease
65 38.78 Hydroxypregnenolone C23H36O3 360 12.27 Steroid-producing neurons
66 40.33 Azaanthracene C34H24N6 516 1.91 Activity against L1210 leukaemia cells

*Source of the Biological activities of the compound is PubChem database.

Table 4: Showing the list of compounds having pharmaceutical importance obtained from GC-MS profile of Methanol stem extract of Trichosanthes lobata

S. No. RT (min) Compound name Molecular
formula		 Mol. weight (g/Mol) Area % Biological activity*
1 15.51 Dihydroisocoumarin C13H16O2 204 2.73 Aromatase inhibitor
2 16.84 Tert-butyldimethylsilyl ether C14H28O5Si 304 3.19 Antiviral activity, Cytotoxicity
3 19.44 Iminoisoquinoline C11H12N2 172 1.32 Inhibitors of nitric oxide synthase
4 19.44 Hydroxypropanoate C11H22O3 202 1.32 Antioxidant activity
5 21.79 Hexadecanoic acid C17H34O2 270 3.12 Anticancer drug target
6 21.79 Pentadecanoic acid C17H34O2 270 3.12 Inhibition of COX1 and prostaglandin biosynthesis
7 24.76 Oxaspiro C14H20O3 236 1.26 Acts against cancer and inflammation
8 25.72 Trimethylsilyl ester C20H40O5Si4 404 2.22 Anti-HIV activity
9 26.80 D-glucopyranose C8H19ClO6 246 1.73 Cytotoxicity
10 26.80 Octadecanoic acid C20H40O2 312 1.73 Antimicrobial agent
11 27.94 Sulfoximine C23H31NO4S2 449 1.21 Cytotoxicity
12 29.45 Benzeneacetic acid C10H12O3 180 1.60 Anti-inflammatory activity 
13 29.92 Cyanoacetate C20H15N3O5 377 1.63 Cytotoxicity, Antimicrobial activity
14 29.92 Phenylmethanone C27H23NO 377 1.63 Inhibitory Activity on diabetes mellitus
15 30.36 Ethanamine C9H13N 135 2.78 Antifungal activity
16 30.36 2-(Chlorovinyl)phenyl sulfide C8H7ClS 170 2.78 Antimalarial design
17 30.36 Dihydrobenzothiophene C15H14S 226 2.78 Potential treatment of Parkinson's disease
18 30.36 Phenylpropan-1-one C17H18O2 254 1.09 Bioreduction by bacterial ferredoxin
19 31.44 Tetraol C27H44O4 432 20.55 Cytotoxicity 
20 31.44 ç-Sitosterol C29H50O 414 20.55 Antiproliferative activity
21 32.20 Cycloheptatrien C10H12O2 164 2.44 Neuroprotective activity
22 32.52 Isopropylidene C39H50O4Si2 638 3.17 Antitumor and antiviral activity
23 33.12 D-Glucopyranosiduronic acid C29H56N2O10S 676 2.42 Cytotoxicity
24 33.12 Oxo-acetamide C24H23N3O3 401 2.42 Highly potent histone deacetylase inhibitors
25 33.79 Azabicyclo C14H22N2 218 1.79 Muscarinic receptor antagonists
26 33.79 Quinazolinone C19H23N3O2 325 1.79 Neurodegenerative disorder
27 34.17 Glucopyranosiduronic acid C29H57NO8Si5 687 1.43 Inhibition of HIV1 protease
28 34.17 Porphycene C31H36N4O4 528 1.24 Induction of apoptosis in human HeLa 
29 34.17 Spongiadioxin C13H6Br4O3 526 1.24 Cytotoxicity
30 34.48 Oxazolidinone C12H15NO5 253 1.42 Antibacterial activity 
31 34.48 Cinnamate C19H18O3S 326 1.42 Depigmenting activity
32 34.48 Dimethyluracil C15H22N2O6 326 1.42 Antibacterial activity
33 36.46 Propylpyrimidine C28H28N2O3 440 16.55 Binding affinity towards Corticotropin
34 37.46 Androstane C21H34N2O2 346 2.16  Inhibition of reduction of dihydrotestosterone
35 38.66 Trifluoromethanesulfonate C9H9F3O5S 286 1.44 Antineoplastic activity 
36 38.66 Benzenecarboximidamide C15H12F3N3O3 339 1.44 Antithrombotic
37 38.92 Norsesterterpene C24H38O2 358 1.62 Cytotoxicity
38 38.92 Methyl pentyl ester C20H27ClO4 366 1.62 Antiviral agent
39 39.72 Hydroxycodeinone C18H19NO4 313 3.97 Antagonistic Activity 
40 39.72 Pyrimidine C22H22N2 314 3.97 Drug metabolism
41 39.72 Phenanthroline C19H11N3O2 313 3.97 Cytotoxicity
42 39.72 Indolizine C21H15NO2 313 3.97 Inhibition of human EGFR autophosphorylation
43 39.72 Nitroanthracene C19H23NO3 313 3.97 Cancer therapy
44 40.02 Norbornadiene C14H13N 195 1.82 Analgesic activity, Psychotomimetic activity
45 40.27 Phenanthrene C28H20Br2 514 1.65 Antiviral activity, Drug metabolism
46 40.27 Gomisin F C28H34O9 514 1.65 Cytotoxicity, Antiviral activity

*Source of the Biological activities of the compound is PubChem database.

Fig. 1: Showing the GC-MS spectrum of Trichosanthes lobata leaf extracted from Ethyl acetate

Fig. 2: Showing the GC-MS spectrum of Trichosanthes lobata stem extracted from Ethyl acetate

Fig. 3: Showing the GC-MS spectrum of Trichosanthes lobata leaf extracted from Methanol

Fig. 4: Showing the GC-MS spectrum of Trichosanthes lobata stem extracted from Methanol

The GC-MS profile of Ethyl acetate leaf extract of Trichosanthes lobata showed the presence of 41 bioactive compounds, whereas its stem extract showed 45 compounds which are reported for various bioassays in PubChem. The GC-MS profile of methanol leaf extract of T. lobata showed 66 compounds which are biologically active and its respective stem extract contained 46 compounds having bioassay reports in PubChem. Overall, a large number of phytochemical compounds with good area percentage were found in methanolic extract, which was followed by ethyl acetate. We could also note that the number of active compounds was found more in leaf than in stem extracts. We were also able to find out potent anti-inflammatory compounds such as Octanoic acid, Dodecanoic acid, Octadecane, Tetramethyl, Enoic acid and Napthalene from ethyl acetate leaf extracts; Hexanoic acid and Methyl propyl from ethyl acetate stem extracts. Similarly, anti-inflammatory compounds such as Methylpyrazole, Quinazolin-8-one and Ilicic acid were present in the methanol leaf extract, whereas Pentadecanoic acid, Oxaspiro, Benzeneacetic acid and Norbornadiene were present in the methanol stem extract. With this we conclude that a greater number of bioactive agents against inflammation is found in extracts of leaf generated using ethyl acetate.

Many studies have proven that methanol has higher activities than that of extracts obtained from different solvents. In this case also even though methanolic extract has shown the presence of more number compounds, ethyl acetate leaf extracts stand to be effective against treating inflammation. Based on these examinations the anti-inflammatory activity of ethyl acetate and methanol leaf extracts may be attributed towards the presence of very active agents. The thorough quantitative analysis of phytochemicals using GC-MS studies reveals the presence of all components such as flavonoids, sterols, fatty acid and esters in the sample [5]. GC-MS analysis is also helpful in the determination of the chemical composition, area and molecular weight of the samples [6, 7].

In addition to anti-inflammatory activity we could notice that the extracts contain compounds which are responsible for treating other ailments too. It is reported that T. lobata, T. dioica [8, 9] T. cucumerina [10, 11] and T. kirilowii are said to contain carbohydrates, glycosides, flavonoids, tannins, proteins, steroids and saponins. T. lobata is used for malarial fever and liver disorders [12]. The highlighted compounds had been noted earlier for their anti-inflammatory potential. Active polyphenols, flavones, phenolic terpenes, fatty acids, sterols, amide, esters, alkaloids, flavonoids, lactones and carotenoids such as Orientin, isoorientin, isovitexin, vitexin, chlorogenic acid, catechin, palmitic, stearic, vanillic acids, sitotesnone, vinyl guaiacol, o-Tolylaldehyde, epicatechin, procyanidins, protocatechuic acid, oleanolic acids, Pomolic acid, α-amyrin, β-amyrin and derivatives of stigmasterol, oleic, linoleic, linolenic, limonene, lupeol, phytol, vellaral, cucuminoids, selinene, piperitol, camphor, quercetin, kaempferol, benzoazolinone, tocopherol, eugenol, myricetin, thymol, lutein, carotene, etc. have been proven for their anti-inflammatory potential in varied plant species [13-15]. More number of compounds present in the list are attributed towards cytotoxic activity and hence Trichosanthes lobata can be a potent anti-cancer agent also.

CONCLUSION

The analytical characterization of ethyl acetate and methanol extract of Trichosanthes lobata leaf and stem witness the presence of active metabolites. GC-MS reveals that the plant and its extract has valued agents which fulfil the pharmaceutical need. From the present study, it is confirmed that Trichosanthes lobata ethyl acetate and methanol leaf extract can be used as potent anti-inflammatory drug. Further, the fractions containing active compounds should be isolated from the extract and has to be examined through in vivo experiments. This will confirm their mechanism of action as novel therapeutic agent against inflammation. This research article also emphasizes varied pharmacological properties of Trichosanthes lobata in treating various disorders like cancer, neurological disorder, aging, against bacterial infections and its future prospects for improved usage in.

FUNDING

Nil

AUTHORS CONTRIBUTIONS

Both the authors of the research article have sufficiently participated in the intellectual content, conception, design, analysis, interpretation of data and writing the manuscript.

CONFLICT OF INTERESTS

There remains no conflict of interest.

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