Synergistic and Antagonistic Contributions of Essential Oil Components to Their Bioactivity Against Tribolium: In Vitro and In Silico Approaches

The complex chemical composition of essential oils (EOs) and the interactions among their constituents highlight the need for approaches that consider their overall chemical profiles. In this study the toxic and repellent activities of 13 EOs from pre-domesticated aromatic plants were evaluated against the stored-product pest and model organism Tribolium confusum. All EOs exhibited repellent activity, whereas toxicity varied considerably. An unbiased composition-based hierarchical clustering grouped EOs according to chemical similarity and guided the selection of carvacrol, p-cymene, eucalyptol (1,8-cineole), linalool, linalyl acetate, γ-terpinene and thymol for interaction studies where 12 constituent combinations were evaluated. Bioassays demonstrated synergistic, additive and antagonistic interactions, with synergism strongly dependent on the natural relative proportions of constituents. Molecular docking against odorant-binding proteins (OBPs), cytochrome P450 4BR3 (CYP450 4BR3), acetylcholinesterase (AChE), the GABA receptor (GABAr), and octopamine/tyramine receptors (OATAr) supported a sequential and cooperative multi-targeted mode of action. Thymol and carvacrol displayed the strongest interaction profiles across protein families, whereas linalool and linalyl acetate suggested alternative molecular mechanisms. Overall, these findings support a shift from abundance-based approaches to an interaction-based framework integrating EO composition, natural constituent ratios and multi-target mechanisms for the rational development of effective and sustainable formulations for stored-product pest management.