Abstract
This study has investigated the interactions of a number of methyl-enedioxyphenyl compounds with hepatic cytochrome P-450. The administration of isosafrole to rats resuited in the formation of an in vivo isosafrole metabolite-cytochrome P-450 complex which caused inhibition of the hepatic monooxygenase activities. In the oxidized state this in vivo metabolite complex readily dissociated in vitro following the addition of certain lipophilic Type I binding compounds, many of which were substrates for the monooxygenase. The time-dependent dissociation of the in vivo metabolite complex resulted in an apparent increase in the cytochrome P-450-mediated monooxygenase activities. The enzyme activities which increased following displacement of the complex were those generally associated with induction by 3-methylcholanthrene. Induction by isosafrole resulted in an increase in the monooxygenase activities of hepatic microsomes from both C57BL/10 and DBA/2 mice, strains which are respectively "responsive" and "non-responsive" to induction by polycyclic aromatic hydrocarbons. The monooxygenase activities which showed the largest increases in activity following dissociation of the isosafrole metabolite-cytochrome P-450 complex were those which were generally increased by 3-methylcholanthrene induction. The induction profile of the hepatic microsomes from both C57BL/10 and DBA/2 mice was compared by sodium dodecyl sulphate polyacrylamide gel electrophoresis. Isosafrole induced a pattern of proteins similar to those induced by both phenobarbitone and 3-methylcholanthrene. Spectral investigations in isolated hepatocytes demonstrated Type I and Type II binding spectra but not Type RI spectra. The addition of methylenedioxyphenyl compounds to a suspension of isolated hepatocytes in vitro resulted in the formation of absorption maxima at 427 nm and 460 nm although the 460 nm peak was small. However, the rate of formation of the 427 nm and 455 nm absorption maxima during oxidative metabolism of isosafrole in hepatic microsomes was shown to be pH-dependent, with formation of the 427 nm maximum favoured at low pH whilst a higher pH favoured formation of the 455 nm maximum. Isosafrole-related material was also found to be apparently irreversibly bound to the cellular components of isolated rat hepatocytes incubated 14 with [o - [14]C] isosafrole in vitro. This binding was increased by prior induction of the hepatic monooxygenase with phenobarbitone, 3-methylcholanthrene and isosafrole.