Unraveling Antagonistic Interactions in Complex Mixtures of Benzotriazole UV Stabilizers: A High-Throughput NAMs...

Deconvoluting the contribution of individual components and assessing the combined toxicity of chemical mixtures remain formidable challenges in environmental risk assessment. To address this, we developed an economical and high-throughput New Approach Methodology (NAM) framework that integrates leave-one-out method with dose-dependent yeast functional genomic analysis (DYFGA). We applied this framework to investigate environmentally prevalent benzotriazole ultraviolet stabilizers (BUVSs: UV-326, UV-327, UV-328, UV-329, UV-P, and UV-PS). Results demonstrated that UV-PS contributed most significantly to the mixture's antagonistic effect, as evidenced by the lowest point-of-departure (POD) similarity between the "Leave_UV-PS" and the "All" mixture, followed by UV-P and UV-328. The joint toxicity of the six BUVSs exhibited antagonistic interactions, indicated by a higher POD value for the "All" mixture compared to others. This antagonistic interaction was consistently observed in binary and ternary mixtures of the main antagonism contributors (UV-P, UV-PS, and UV-328) and was validated by conventional cytotoxicity assays. Mechanistic insights from the genomic analysis suggest these effects are potentially mediated through disruption of the cell cycle and DNA replication. These findings not only enhance understanding of the complex toxicology of BUVS mixtures but also present a robust methodological framework for evaluating the health risks posed by more intricate chemical mixtures in real-world settings.