Discovery of bioactive compounds from Thai medicinal plants as potential SARS-CoV-2 main protease inhibitors

Thai medicinal plants represent valuable sources of structurally diverse secondary metabolites with potential antiviral relevance. In this study, a phytochemical investigation of three Thai medicinal plants, Phyllanthus emblica, Averrhoa bilimbi, and Helicteres isora, was conducted to identify natural products with inhibitory activity against SARS-CoV-2 main protease (M^pro). The crude MeOH extracts of the three plants showed moderate M^pro inhibition at 100 µg mL^-1, with inhibition values of 61.7 ± 1.5%, 56.9 ± 3.8%, and 53.3 ± 1.7% for P. emblica, A. bilimbi, and H. isora, respectively. Bioactivity-guided fractionation and repeated chromatographic separation led to the isolation of 24 compounds, including nine constituents from P. emblica (P1-P9), six compounds from A. bilimbi (A1-A6), and nine compounds from H. isora (H1-H9). These metabolites comprised phytosterols, lupane-type triterpenoids and triterpenoid esters, lignans, phenolic esters, flavonoids, and related oxygenated derivatives. Among them, A1 was identified as a new phenolic glycoside and named demethoxybilimoside A, while stigmastane-3β,6α-diol 3-O-tetradecanoate (P1) was isolated and characterized as a pure compound. All isolated compounds were evaluated for SARS-CoV-2 M^pro inhibition. High apparent inhibition was observed for (stigmastane-3β,6α-diol 3-O-tetradecanoate) P1, lupeol (P4), kaempferol (A5), and quercetin (A6), with inhibition values of 99.3 ± 2.0%, 88.5 ± 0.9%, 95.8 ± 1.0%, and 99.5 ± 1.1%, respectively. Compound P1 showed an apparent IC₅₀ value of 86.5 ± 3.8 µM and was subsequently selected for molecular docking analysis. These findings expand the phytochemical knowledge of Thai P. emblica, A. bilimbi, and H. isora and identify several compounds for further biochemical evaluation.