997 Article Info Article history Received 1 August 2023 Revised 16 September 2023 Accepted 17 September 2023 Published Online 30 December 2023 Keywords Antioxidant studies HR-LCMS In silico docking studies Hepatoprotective Evaluation of the phytochemistry of Gymnostachyum febrifugum Benth. extract and its hepatoprotective activity using in vitro and in silico methods Lekha Mathew*  , R. Suresh* and A. Anil Babu** *Department of Pharmacy, Annamalai University, Chidambaram-608 002, Tamil Nadu, India **Department of Pharmacy, Westfort College of Pharmacy, Pottore-680581, Thrisur, Kerala Abstract The plant, Gymnostachyum febrifugum Benth. (Nelumucchala) of the Acanthaceae family is a herb, significant to traditional medicine practitioners of the South Western India. The dried and powdered plant was subjected to successive Soxhlet extraction using petroleum ether, ethyl acetate, and ethanol. The ethyl acetate extract was chosen for further studies and its antioxidant capacity was evaluated using four different methods; FRAP, DPPH, ABTS, and total antioxidant studies. The extract was further subjected to HR-LCMS, for its chemical characterization. Licoricesaponins A3, AFN911, Austalide A, and Caryoptin were a few of the compounds that were identified in abundance in the extract. These compounds were evaluated for their capacity to bind to various receptors involved in hepatic disorders using the molecular docking approach. In silico studies were carried out using the Schrodinger suite. The docking study was done on four receptors PPAR (PDB-2ZNN), PPAR (PDB-2ATH) , 5-LOX (PDB-6N2W), and TGF- receptor I(PDB- 6B8Y) . The compound AFN911 exhibited an excellent binding score and stability with all four receptors. Licoricesaponins A3, Austalide A, and Caryoptin exhibited good docking scores and binding stability with the receptor 5-LOX. Annals of Phytomedicine 12(2): 997-1004, 2023 Annals of Phytomedicine: An International Journal http://www.ukaazpublications.com/publications/index.php Print ISSN : 2278-9839 Online ISSN : 2393-9885 DOI: http://dx.doi.org/10.54085/ap.2023.12.2.119 Original Article : Open Access Copyright © 2023 Ukaaz Publications. All rights reserved. Email: ukaaz@yahoo.com; Website: www.ukaazpublications.com Corresponding author: Ms. Lekha Mathew Department of Pharmacy, Annamalai University, Annamalai Nagar, Chidambaram-608 002, Tamil Nadu, India E-mail: lekhamathewphd@gmail.com Tel.: +91-8943221189 1. Introduction G. febrifugum, is a small scrapigerous, stemless herb, indigenous to the South Western Ghats of India (Silpa and Thomas, 2021). Based on the literature review, the decoction prepared from the root of the plant was used by traditional medicine practitioners, for treating various diseases like purpureal fever, metrorrhagia, and indigestion (Vijayalakshmi and Haridasan, 2017). Despite its significant usage in folklore medicine, the plant has not been extensively studied for its ethnopharmacology. The purpose of the study was to investigate the phytochemistry of the plant, G. febrifugum and evaluate its potential in the management of various liver disorders. As per the 2023 updates, 4% of all deaths worldwide are due to liver diseases. The current scenario of liver diseases is alarming and the awareness that, not much effective treatment is still available is shocking. Liver disorders ranging from chronic viral hepatitis, genetic/autoimmune/ drug-induced liver diseases, alcoholic liver diseases (ALD), fatty liver disorders progressing to non-alcoholic fatty liver diseases/ nonalcoholic steatohepatitis (NAFLD/NASH), cirrhosis, hepato- cellular carcinoma (HCC), accounts for 2 million deaths per year worldwide (Devarbhavi et al., 2023 ; Bari et al., 2022). The preliminary phytochemical evaluation of the plant was carried out by Vijayalakshmi and Haridasan (2017), and the plant was found to be rich in phenolic compounds. Arunachalam and Parimelazhagan (2011), reported the antioxidant and antimicrobial potential of the methanolic extract of the plant root. Some journals have reported various traditional uses of this plant extract, but no scientific evidence has been produced for its reliability. Hence, the study was oriented towards understanding the phytochemistry of the plant and evaluating its therapeutic potential by in vitro and in silico methods. 2. Materials and Methods 2.1 Plant collection and extraction The fresh, whole plant was collected from its natural habitat, Kakkayam, Kozhikode, Kerala. The plant was identified by Dr. V. B. Sreekumar, Dept. of Botany, KFRI (Kerala Forest Research Institute), Peechi, Thrissur, and a specimen of the authenticated sample (Voucher No. 18126) was deposited in the herbarium. The whole plant samples were rigorously washed under running water to remove soil and other debris sticking to the roots and leaves. The samples were shade-dried for 10 days, powdered, and sieved through a 45-mesh sieve. 2000 g of the powdered plant material was subjected to Soxhlet extraction using 5000 ml of petroleum ether followed by ethyl acetate and ethanol. All the extracts obtained were concentrated at a temperature below 40°C, air dried, and stored at 2-8°C. The qualitative and quantitative preliminary phytochemical evaluation of ethyl acetate extract was found to contain flavonoids, total phenolics, saponins, steroids, and tannins (Hamid et al., 2020). 2.2 Antioxidant study The antioxidant capacity of a plant extract is a preliminary parameter to monitor its therapeutic potency. The capacity of the ethyl acetate extract to scavenge free radicals was evaluated using different in vitro assay methods.