Date of Award

1982

Degree Type

Dissertation

Degree Name

Doctor of Philosophy in Pharmaceutical Sciences

Department

Interdepartmental Program

First Advisor

George C. Fuller

Abstract

Pentachloroanisole (PCA), an environmental degradation product of the biocide pentachlorophenol (PCP), has been detected in the food chain. The metabolic fate of PCA was examined in miniature pig hepatic microsomes, in vitro. The compound was shown to be a substrate for a cytochrome P450-dependent demethylation reaction, which results in the regeneration of the parent compound, PCP. A disproportionately large increase in PCA demethylase activity (PCADM) following pretreatment with phenobarbital suggests that the compound is preferentially metabolized by specific-inducible form(s) of cytochrome P450.

A comparison of the effects of PCA and purified PCP on the hepatic MFO system of miniature pigs was conducted at various stages of postnatal development. Phenobarbital was utilized as a positive control for induction. PCA, PCP and phenobarbital (10 mg/kg/day X 4 days, P.O.) were administered to piglets at 1, 4, and 8 weeks of age and the levels of cytochromes P450 and b5, and the activities of NADPH-Cytochrome c reductase, aniline hydroxylase (ANOH), p-nitroanisole demethylase (NADM), and PCA demethylase were determined. In one week old piglets, PCA produced significant increases in all parameters measured, with the greatest effect (300% of control) on its own in vitro metabolism. The pleiotropic response evoked by PCA was similar to that of phenobarbital, but of lesser magnitude. PCP produced small increases in only P450 and nitroanisole demethylase. The qualitative differences in the induction patterns produced by PCA and PCP suggests that the two compounds exert different effect on MFO. By eight weeks of age, the magnitude of induction by PCA was diminished. Furthermore, although specific activities for ANOH, NADM, and PCADM in phenobarbital treated pigs were similar at 1 and 8 weeks of age, examination of catalytic activity profiles suggested an age dependent decrease in the induction of specific forms of cytochrome P450. On further investigation, Eadie-Hofstee plots from kinetic experiments with ANOH and PCADM exhibited biphasic patterns suggestive of multiple forms of P450 catalyzing the same reaction. By integrating the effects of age and treatment on the various kinetic species for each substrate, a minimum of four forms of cytochrome P450 are suggested to exist in miniature pig hepatic microsomes. Of the four forms, two are inducible by phenobarbital and one of these forms appears to display age-dependency in the magnitude of induction. These data indicate that MFO induction by exogenous chemicals varies qualitatively as well as quantitatively with age.

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