Cytochrome P450 Enzyme-Mediated Enantioselective Biotransformation of Chiral Fungicide Tebuconazole in Earthworm

Tebuconazole is widely used in agricultural practices, leading to elevated residue levels in agricultural soils. However, its biotransformation and the ecotoxicological effects of the corresponding transformation products (TPs) remain insufficiently explored. To fill this research gap, the TPs of tebuconazole in an earthworm-soil system were identified by UHPLC-QTOF/MS combined with UHPLC-QLiT/MS. Six chiral TPs were tentatively identified, with four TPs detected in both earthworms and soil, while two TPs were found exclusively in earthworm. Significant enantioselectivity was observed for tebuconazole and five TPs in earthworms, involving cytochrome P450 enzyme-mediated hydroxylation and dechlorination. <i>In vitro</i> metabolism experiments using earthworm microsomes revealed that CYP1A2, CYP2J2, and CYP2E1 were involved in the hydroxylation pathway of tebuconazole. Molecular docking results confirmed that <i>S</i>-(+)-tebuconazole produced more hydroxylated transformation products than <i>R</i>-(-)-tebuconazole due to its lower binding energy with these enzymes. Predictions from the ECOSAR model indicated that hydroxylation was the most significant transformation pathway for reducing the toxicity of tebuconazole. These findings provide valuable insights into the environmental fate and risk assessment of tebuconazole at the enantiomeric level.