Antibacterial activity of Cupressaceae essential oils from woody plant resources: Chemical diversity, seasonal variation, and component interactions

Cupressaceae essential oils (CEOs) derived from woody plant resources and forestry by-products were systematically investigated for their chemical diversity and antibacterial activity. CEOs from 11 species representing seven genera were examined to assess the roles of species identity and harvest season in shaping chemical composition and bioactivity. GC–MS analysis revealed four dominant chemotypes, together with pronounced seasonal shifts in key constituents, highlighting harvest timing as a critical determinant of CEO quality. Antibacterial assays showed that most CEOs were more effective against Escherichia coli than Staphylococcus aureus , with spring-harvested oils displaying generally enhanced activity. CEOs from Cupressus obtusa (Siebold & Zucc.) Endl. and Thuja occidentalis L. exhibited minimum inhibitory concentrations (MICs) as low as 2 mg/mL against E. coli , whereas higher concentrations (8–16 mg/mL) were required for S. aureus . Scanning electron microscopy combined with membrane leakage assays indicated that antibacterial activity was primarily associated with membrane disruption, accompanied by the loss of intracellular proteins, nucleic acids, and ATP. Integrative analysis linking chemical profiles with antibacterial performance identified α-thujone as a key contributor to activity; however, its individual efficacy could not fully account for the potency of whole oils. Synergistic effects were therefore explored. Checkerboard assays demonstrated strong synergy between α-thujone and terpinen-4-ol, reducing the MIC to 0.5 mg/mL with a fractional inhibitory concentration index of 0.19. In a model essential oil system, supplementation with α-thujone further enhanced the antibacterial activity of a terpinen-4-ol–containing lavender essential oil, decreasing its MIC from 2 to 1 mg/mL. Overall, these findings demonstrate that the antibacterial performance of CEOs is governed by species-specific chemistry, seasonal variation, and synergistic interactions among components, providing a mechanistic and practical framework for the valorization of woody plant–derived essential oils as natural antibacterial agents.