Abstract

Human and ecological systems in the Anthropocene are exposed to complex mixtures of chemicals from industrial, 
agricultural, pharmaceutical, and consumer-product sources. These combined exposures frequently produce effects 
that are not predicted by single-chemical toxicology: interactions can be additive, antagonistic, or synergistic, with 
synergy posing the greatest concern because low-level co-exposures may produce unexpectedly large adverse 
outcomes. This review synthesizes mechanistic knowledge of mixture toxicity with an emphasis on biochemical 
interactions that drive synergy. We examine key interaction modalities - competitive and noncompetitive enzyme 
inhibition, receptor co-activation and cross-talk, redox cycling and oxidative stress amplification, disruption of 
detoxification pathways and transporters, endocrine network interference, and microbiome-mediated 
biotransformation. Methods for assessing mixture effects are evaluated, including experimental designs (fixed-ratio 
and factorial mixtures), dose–response modelling (concentration addition, independent action, response-surface 
methods, isobolograms), and emerging in vitro, in vivo, and in silico tools (high-throughput screening, organoids, 
PBPK-toxicodynamic models, systems toxicology). We discuss biomarkers and multi-omics strategies for detecting 
synergistic biochemical perturbations and highlight illustrative case examples where co-exposures intensify risk 
(e.g., pesticide combinations, metal–organic pollutant interactions, polypharmacy and environmental contaminants). 
Critical challenges are identified: exposure characterization, realistic dose metrics, non-monotonic and low-dose 
effects, interindividual variability, and regulatory frameworks that still largely rely on single-chemical assessment. 
The review concludes with research priorities to improve prediction and prevention of synergistic toxicity, 
advocating integrated experimental–computational pipelines, standardized mixture reference materials, human
relevant models, and policy adaptations that account for cumulative and interaction-driven risks.