Krstic_2026_Int.J.Mol.Sci_27_

Alchemilla alpina L. (Rosaceae), belongs to a genus well recognized in traditional medicine for treating gynecological disorders and hormonal imbalance; however, the specific bioactivity of A. alpina itself remains poorly characterized. This study aimed to elucidate the phenolic composition and the biological potential of the methanolic (MeOH) extract of A. alpina. LC-MS/MS analysis identified 39 phenolic compounds, with rutin, catechin, kaempferol-3-O-glucoside, and caffeic acid being the dominant constituents. The extract exhibited high total phenolic and flavonoid contents, consistent with strong antioxidant capacities. It demonstrated notable alpha-glucosidase and acetylcholinesterase inhibitory activities, indicating its potential relevance for metabolic and neurodegenerative disorders. The extract effectively reduced AAPH-induced ROS levels in MRC-5 fibroblasts, indicating cytoprotective and antioxidative effects. The cytotoxicity toward cervical cancer cells HeLa and ovarian cancer cells A2780 was moderate and concentration dependent. A yeast-based fluorescent screen revealed a strong and selective binding affinity toward estrogen receptor alpha (ERalpha) and selective inhibition of human recombinant AKR1C3 (59.5%), without affecting AKR1C4. Additionally, high COX-1/COX-2 inhibition (>70%) supported its anti-inflammatory potential. Collectively, these findings provide the first integrated evidence of A. alpina's phenolic richness and multifunctional bioactivity, scientifically supporting its potential in managing hormone-dependent and oxidative stress-related disorders.