Phase I and phase II biotransformation of methoxychlor and its demethylated metabolites in channel catfish (Ictalurus punctatus) liver and intestine.

摘要

Methoxychlor (MXC) is a chlorinated pesticide that is relatively persistent in the environment. Its demethylated metabolites interact with estrogen receptors and have been associated with endocrine disruption in a variety of species. [14C]-MXC biotransformation was examined in channel catfish (Ictalurus punctatus), a freshwater species commonly found in the southern United States. Hepatic microsomes formed monohydroxymethoxychlor (mono-OH-MXC) and bishydroxymethoxychlor (bis-OH-MXC), as assessed by co-migration with authentic standards. Hepatic formation of mono-OH-MXC showed a K m of 3.28 +/- 0.78 muM (mean +/- S.D., n = 4) and V max of 99.0 +/- 17.4 pmoles/min/mg protein for fish pretreated with MXC, 2 mg/kg i.p., for 6 days. These values did not differ significantly from those in control fish, but were significantly lower (p0.05) than the kinetic parameters for catfish treated with 3-methylcholanthrene (3-MC), 10 mg/kg i.p., for 4 days. The 3-MC-treated fish produced significantly more of the secondary metabolite, bis-OH-MXC, than controls. Intestinal microsomes formed mono-OH-MXC at lower rates than liver. Methoxychlor pretreatment significantly reduced intestinal metabolite formation while 3-MC-treatment significantly raised mono-OH-MXC production. Ketoconazole, clotrimazole, and alpha-naphthoflavone all decreased the production of mono-OH-MXC in liver microsomes, while alpha-naphthoflavone stimulated bis-OH-MXC formation. The results suggested that CYP1 and other CYP isoforms were responsible for MXC demethylation in catfish.; Slow phase II metabolism of mono-OH-MXC or bis-OH-MXC may delay their excretion and contribute to MXC toxicity. In hepatic microsomes for both mono-OH-MXC and bis-OH-MXC, the Vmax was significantly (p0.05) higher in 3-MC-treated fish than in controls. Microsomes from MXC-treated fish had a lower apparent Km value. Product was formed in microsomes from MXC-treated fish without added substrate, suggesting the lower apparent K m was an artifact due to residues of mono-OH-MXC or bis-OH-MXC. Analysis of these microsomes revealed residues of mono-OH-MXC and bis-OH-MXC, but not parent MXC. Intestinal microsomes from control and 3-MC-treated fish glucuronidated mono-OH-MXC and bis-OH-MXC more rapidly than liver. The results suggested that although mono-OH-MXC and bis-OH-MXC were readily glucuronidated in catfish liver and intestine, the Km values for glucuronidation were 50-fold higher than for CYP-dependent formation of the mono-OH-MXC and bis-OH-MXC. This suggests that glucuronidation may be inefficient at environmental exposure concentrations.