LAUSR

Fipronil has been frequently detected in waterways worldwide at concentrations threatening aquatic organisms, yet metabolic behavior of fipronil enantiomers in aquatic organisms is largely unknown, which is of significance in enantioselective toxicity evaluation. Herein, we quantitatively identified the specific cytochrome P450 (CYPs) isozymes involved in metabolizing fipronil enantiomers in tilapia by combining in vitro metabolic kinetic assays and molecular docking. Inhibition studies suggested that CYP1A enzyme was the main isoform catalyzing metabolism of fipronil, and CYP3A limitedly contributed to the metabolism in fish liver S9. Both dissipation rate constant (k e ) and maximum metabolic velocity (V max ) of R-(-)-fipronil were greater than that of S-(+)-fipronil in tilapia liver S9, suggesting that tilapia selectively metabolized R-(-)-fipronil. The CYP1A1 isozyme exhibited the highest binding capacity to R-(-)-fipronil and S-(+)-fipronil (binding energy: -9.39 and -9.17 kcal/mol, respectively), followed by CYP1A2 (-7.30 and -6.94 kcal/mol, respectively) and CYP3A4 (-7.16 and -6.91 kcal/mol, respectively). The results of in vitro metabolic assays and molecular docking were consistent, i.e., CYP1A, specifically CYP1A1 exhibited a higher metabolic capacity to fipronil than CYP3A, and the fish liver S9 selectively metabolized R-(-)-fipronil. The present study provided insights in understanding enantioselective metabolic behavior and toxicological implications of the in vitro metabolic kinetics of fipronil in fish. This article is protected by copyright. All rights reserved.