Environmental pyridinium disinfectants impair cortisol metabolism via 11β-hydroxysteroid dehydrogenase 2 inhibition: A cross-species risk evaluation.

11β-Hydroxysteroid dehydrogenase type 2 (11β-HSD2), a key enzyme in glucocorticoid inactivation, is expressed in human and rat placentas and kidneys. However, the inhibitory effects of alkyl (C1-C20) pyridinium quaternary ammonium disinfectants (PYRs) on 11β-HSD2 remain unexplored. This study evaluated PYRs through inhibitory potency assays, mechanistic investigations, structure-activity relationship (SAR)/3D-QSAR modeling, and molecular docking using human and rat 11β-HSD2 enzymes. Human 11β-HSD2 inhibition by PYRs followed a distinct V-shaped trend: potency increased from C12 to C16 (IC50: C12 = 132.81 μM, C14 = 19.62 μM, C16 = 1.96 μM) but declined beyond C16 (C18 = 14.45 μM, C20 = 100.76 μM). Shorter-chain PYRs (C1-C8) were inactive at ≤100 μM. Rat 11β-HSD2 exhibited a similar inhibition pattern. SPR analysis revealed that C16 displayed the highest binding affinity (KD = 1.98 μM) with human 11β-HSD2. Mechanistic analysis revealed mixed/noncompetitive inhibition, with PYRs binding both the free enzyme and enzyme-substrate complex. In BeWo cells, C12-C20 PYRs inhibited cortisol metabolism at 10-100 μM, whereas C16 was effective at ≥1 μM. Molecular docking indicated PYRs interact with the NAD + binding site in both species. SAR analysis identified lipophilicity, steric bulk, and flexibility as critical inhibitory determinants, with higher values enhancing potency. A 3D-QSAR pharmacophore model confirmed hydrophobic interactions as essential for inhibition. Notably, species-specific binding differences suggest caution when extrapolating rodent data to humans. These findings highlight the endocrine-disrupting potential of PYRs, demonstrating their structure- and species-dependent interference with cortisol metabolism, with implications for chemical safety assessment.