In vitro relative potencies of polycyclic aromatic hydrocarbons and selected hydroxylated, oxygenated, and nitrated derivatives to activate aryl hydrocarbon receptor in HepG2 and PLHC-1 cell lines

Environmental monitoring of polycyclic aromatic hydrocarbons (PAHs) and their derivatives (e.g., oxy- and nitro-PAHs) remains challenging due to their chemical diversity and the limitations of targeted chemical analyses, which capture only a subset of compounds and cannot address mixture effects. Aryl hydrocarbon receptor (AhR)-based in vitro bioassays have therefore emerged as key complementary tools that integrate the biological activity of all AhR-active compounds in a sample. Linking chemical and biological data requires investigating the AhR activity of individual compounds and determining bioassay-specific relative effect potencies (REPs), which remain unavailable for many environmentally relevant PAH derivatives. This study aims to comprehensively characterize the in vitro AhR-mediated activity for 64 individual compounds, including 17 parent PAHs, 6 hydroxy-PAHs, 21 oxy-PAHs, 16 nitro-PAHs, and 4 p-phenylenediamine (PPD) quinones, and derive their REPs using two AhR-responsive cell lines (human HepG2 and fish PLHC-1). Out of the 64 compounds tested, 27 induced AhR activity in at least one cell assay, with compound-, cell line-, and exposure time-dependent differences being observed. Medium to high-molecular-weight parent PAHs (4-6 rings) and a few derivatives showed the highest potencies, whereas other compounds were weakly active or inactive. PLHC-1 cells were more sensitive to parent PAHs, HepG2 cells to oxy-PAHs, and nitro-PAHs elicited broadly comparable responses using both cell lines. By expanding AhR activity data for numerous previously uncharacterized PAH derivatives and emerging contaminants, this study provides a harmonized reference dataset to support mass balance, structure-activity analyses, and enhance effect-based hazard evaluation of complex PAC mixtures.