Enantioselective Metabolism and Metabolite Identification of Simeconazole in Rat Liver Microsomes

ABSTRACT

This study investigated the enantioselective metabolism of the chiral fungicide simeconazole in rat liver microsomes. The absolute configurations of the two enantiomers were determined to be S ‐(+)‐simeconazole and R ‐(−)‐simeconazole. The half‐life of S ‐(+)‐simeconazole was 2.7 min, while that of R ‐(−)‐simeconazole reached 25.3 min; the elimination half‐life of R ‐(−)‐simeconazole was approximately 9.4 times longer than that of the S ‐(+)‐enantiomer. Such remarkable enantiomeric discrepancy originates from the markedly higher V _max of S ‐(+)‐simeconazole (1.7836 μM/min/mg) versus R ‐(−)‐simeconazole (0.686 μM/min/mg). Their K _m values were comparable (5.93 μM and 6.80 μM, respectively), suggesting similar substrate binding affinity toward metabolic enzymes. Accordingly, the intrinsic clearance (CL _int ) of S ‐(+)‐simeconazole (300.17 mL/min/mg) was three times that of R ‐(−)‐simeconazole (101.47 mL/min/mg). Chemical inhibition assays verified that CYP3A2 serves as the predominant metabolic isoform for S ‐(+)‐simeconazole, with an inhibition efficiency up to 98%. Molecular docking results demonstrated that S ‐(+)‐simeconazole binds to CYP3A2 with lower binding energy and stronger hydrophobic interactions, whereas R ‐(−)‐simeconazole preferentially interacts with CYP2D2. A total of five Phase I metabolites originating from hydroxylation, demethylation and oxidation reactions were characterized. Toxicity prediction via the TEST software revealed that the primary metabolites exhibit lower toxicity to fathead minnows yet higher toxicity to rats compared with the parent simeconazole. This work offers fundamental experimental evidence for the metabolic behavior and toxicological risk assessment of simeconazole enantiomers.