Gender Differences in Microsomal Metabolic Activation of Hepatotoxic Clivorine in Rat

Abstract

The gender differences in the in vitro microsomal metabolic activation of hepatotoxic clivorine, a representative naturally occurring hepatotoxic otonecine type pyrrolizidine alkaloid, in Sprague−Dawley rats and their relation to the gender differences in susceptibility to clivorine intoxication were reported in the present study. Clivorine-induced liver damage in the male rat via metabolic activation to form the reactive pyrrolic ester followed by covalent binding to liver tissue constituents has been reported previously by our research group. The present study demonstrated, for the first time, that cytochromes P450 3A1 and 3A2, which are constitutively expressed in the male rat, might play a significant role in the metabolic activation of clivorine in the rat. Thus, in the male rat, the metabolic activation by liver microsomes to form the reactive pyrrolic ester was found as the only direct metabolic pathway of clivorine followed by subsequent formation of the toxic tissue-bound pyrroles leading to hepatotoxicity. In the case of the female rat, a less significant metabolic activation was observed, whereas the formations of two novel nonpyrrolic metabolites were determined as the predominant biotransformations. None of the four cDNA-expressed rat enzymes (cytochrome P450 2C12, 2E1, 3A1, 3A2) tested could catalyze the formation of these two new metabolites. Furthermore, the female rat (LD₅₀ = 114 ± 9 mg/kg, ip) was found to be significantly less susceptible to clivorine intoxication than the male rat (LD₅₀ = 91 ± 3 mg/kg, ip). Therefore, the results suggested that a significantly lower metabolic activation due to the lack of cytochrome P450 3A1 and P450 3A2 activities mainly accounted for the smaller susceptibility of the female rat to clivorine intoxication.