In Vitro Bioassay Evidence for Chemical Mixture Propagation from the Environment to Humans.

Complex mixtures of organic chemicals extracted from representative but not directly related environmental samples (wastewater, surface water, fish), food items (drinking water, fish, milk) and human blood were tested in 22 in vitro bioassays targeting pathways associated with neurodevelopmental and reproductive health. Extraction methods were optimized to extract common chemicals across matrices capturing both persistent and nonpersistent, neutral and charged organic chemicals─albeit with some bias toward more hydrophilic chemicals over highly hydrophobic chemicals. Most bioassay end points─except genotoxicity─were responsive, with strongest effects observed higher up the food chain in fish and humans. Experimental mixture effects of 24 chemicals quantified in these extracts conformed to the mixture prediction model of concentration addition in the six most responsive bioassays, namely neurite outgrowth inhibition, mitochondrial membrane potential inhibition, transthyretin protein binding, sodium-iodide symporter inhibition and androgen receptor antagonism. Designed mixtures explained little of total bioactivity, indicating that many of the thousands of unannotated molecular features detected by nontarget analysis contribute to mixture effects. Preliminary effect-based trigger (EBT) values defined for water and food by extrapolation from safe levels of individual chemicals indicate no immediate health risks at these average contamination levels. The high complexity and multivalent bioactivity of these mixtures on neurodevelopmental and reproductive pathways necessitate further toxicological scrutiny.