Integrated chemical and biological characterization of Hypericum perforatum extract using LC-MS/MS and in vitro functional assays.

Hypericum perforatum L. (St. John's wort) is a medicinal plant known for its diverse secondary metabolites and pharmacological potential. This study provides a comprehensive evaluation of the phytochemical composition and bioactivities of methanolic extracts from wild H. perforatum. LC-MS/MS analysis identified 36 compounds, including phenolic acids (chlorogenic acid, protocatechuic acid) and flavonoids (rutin, isoquercitrin, quercetin), with the first-time detection of genkwanin, vicenin-2, schaftoside, and afzelin, to the best of our knowledge. The extract demonstrated significant antioxidant activity, with a total phenolic content of 203.04 mg GAE/g DW and strong radical scavenging activity in DPPH (544.78 mg TE/g DW) and ABTS (312.15 mg TE/g DW) assays. In vitro assays showed marked cytotoxicity across multiple cancer cell lines, with IC50 values as low as 3.57 µg/mL in A549 cells, indicating stronger activity compared to cisplatin. Flow cytometric analysis revealed substantial apoptosis induction (up to 53.67% in A549 cells) and G0/G1 cell cycle arrest (76.37% in A549). The extract also exhibited moderate antibacterial activity against E. coli and S. aureus, but no activity against P. aeruginosa or Candida species. These results position H. perforatum as a rich source of bioactive compounds with antioxidant, anticancer, and antimicrobial potential, and suggest further research into its therapeutic applications.