Effect of hydrolysed Trifolium repens L. extract on menopause induced obesity and depressive symptoms: an in-vitro and in-silico approach

This study aimed to evaluate acid-hydrolysis effects on phytoestrogen content and the efficacy of phytoestrogen-rich extract in alleviating post-menopausal symptoms (PMS). Bioactive phytoestrogens were quantified using HPTLC, and unhydrolyzed and hydrolysed extracts were assessed for oestrogenic, anti-proliferative, MAO-A inhibition, ROS quenching, and PL inhibition potential through in-vitro models. In-silico analysis of quantified phytoestrogens against PMS-related proteins was conducted. Hydrolysis increased phytoestrogen content significantly, particularly biochanin-A (56.48-fold), daidzein (22.2-fold), genistein (10.77-fold), and formononetin (5.26-fold). The hydrolysed extract demonstrated improved efficacy in in-vitro models, including reduced IC50 in oestrogenic activity, decreased cell proliferation, increased MAO-A inhibition, and de-creased IC50 for lipase. High binding energies of phytoestrogens against ERβ, MAO-A, and PL supported their potential in managing post-menopausal symptoms. The study scientifically validated T. repens for women's health at menopause, highlighting acid-hydrolysis for enhancing therapeutic activity. T. repens could be explored commercially as an alternative to T. pratense.