Genotype-Environment Interactions Shape the Phytochemical Profile and Antibacterial Activity of Hypericum perforatum L.

St. John’s wort is a medicinal plant of significant pharmaceutical importance, yet its metabolic composition and biological activity vary considerably depending on genetic and environmental factors. Understanding the interplay between genotype and growing conditions is essential for ensuring reproducible therapeutic properties of herbal raw materials. This study aimed to comprehensively evaluate the effects of genotype and growing location on the metabolite profiles and antibacterial/antioxidant activities of H. perforatum. Seven accessions from the VIR collection were grown in two climatically distinct regions of Russia: the temperate zone (Pushkin, Leningrad Region) and a warm region (Maykop, Republic of Adygea). Methanolic extracts were analyzed using LC-MS/MS, yielding 89 annotated metabolites belonging to 8 major compound classes. Antibacterial activity was assessed against Gram-positive (Staphylococcus aureus, Micrococcus luteus) and Gram-negative (Escherichia coli, Pseudomonas aeruginosa) strains, while antioxidant capacity was evaluated using DPPH and TEAC assays. PCA identified cultivation location (36.7%) and genotype (14.8%) as the major sources of metabolic variation, with a significant location × cultivar interaction detected for 70.2% of all analysed features. Naphthodianthrones and procyanidins consistently accumulated in the shoots of H. perforatum plants grown in the temperate region, while flavonoids and acylphloroglucinols exhibited genotype-dependent responses. Extracts from the plants grown in Pushkin showed more pronounced antibacterial activity, particularly against M. luteus and E. coli, and higher antioxidant capacity compared to the Maykop-grown plants, correlating with elevated levels of bioactive compounds. Genotype k-14 displayed the most pronounced metabolic plasticity. The metabolite profiles and bioactivity of H. perforatum are associated with complex genotype-environment interactions. These findings highlight the importance of considering both genetic background and cultivation location for the production of standardized herbal materials with more consistent pharmacological properties.