Characterization of known and unknown AhR-active substances in freshwater fish from the Gapcheon River, South Korea: Application of effect-directed analysis and nontarget screening.

This study investigated the distribution and composition of persistent toxic substances in freshwater fish collected from the Gapcheon River, Daejeon, South Korea. To identify key toxicants responsible for biological effects, effect-directed analysis was applied, focusing on aryl hydrocarbon receptor (AhR) agonists. The target species included crucian carp (S1, Carassius carassius), common carp (S2, Cyprinus carpio), Far Eastern catfish (S3, Silurus asotus), barbel steed (S4, Hemibarbus labeo), and skygager (S5, Erythroculter erythropterus). Trophic positions (TP), estimated from δN values of amino acids, were 2.2 for S1 and S2, 3.0 for S3, and 3.2 for S4 and S5. Concentrations of mercury (Hg), polychlorinated biphenyls, and polycyclic aromatic hydrocarbons (PAHs) exhibited species- and organ-specific patterns, generally increasing with TP. The H4IIE-luc bioassay indicated elevated AhR-mediated potencies in mid-polar fractions (F2) from S1 muscle and egg, S3 fillet, and S5 liver. Target analysis of 21 known AhR agonists, including 7 traditional and 14 emerging PAHs, accounted for 20-94 % of total AhR activity, with benzo[b]anthracene emerging as the dominant contributor (mean = 44 %). Nontarget screening using GC-QTOFMS on F2 identified 19 AhR agonist candidates, including compounds potentially associated with crude oil and vehicle emissions, combustion byproducts, dye precursors, industrial materials, silicone-based compounds, and natural products. In silico modeling predicted AhR binding potential for all candidates, with triphenylmethane and 1,1,3,3-tetraphenyl-1,3-dimethyldisiloxane showing the highest overall toxic potential. These findings highlight the diversity and ecological relevance of AhR-active contaminants in freshwater fish and their implications for contaminant transfer in aquatic food webs.