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Identification of the enzymes catalyzing metabolism of methoxyflurane
Abstract:
The hepatic microsomal metabolism of methoxyflurane in rabbits is markedly stimulated by treatment with phenobarbital. However, the increased rate of metabolism cannot be completely accounted for by the activity of the purified phenobarbital-inducible cytochrome P-450 isozyme 2, even in the presence of cytochrome b5. The discovery of a second hepatic phenobarbital-inducible cytochrome P-450, isozyme 5, led us to undertake experiments to determine in hepatic and pulmonary preparations the portion of microsomal metabolism of methoxyflurane catalyzed by cytochrome P-450 isozymes 2 and 5. We report herein that isozyme 2 accounts for 25% and 29%, respectively, of the O-demethylation of methoxyflurane in hepatic microsomes from untreated and phenobarbital-treated rabbits, and for 25% of the methoxyflurane metabolism in pulmonary microsomes. Results for isozyme 5 indicate that it catalyzes 19% and 27% of methoxyflurane metabolism in control and phenobarbital-induced liver, and 47% of O-demethylation in the lung. In summary, we demonstrate that methoxyflurane O-demethylation in lung, phenobarbital-induced liver, and control liver microsomes is catalyzed by cytochrome P-450 isozymes 2 and 5. Results with purified cytochrome P-450 isozyme 5 are consistent with those obtained using microsomal preparations. Furthermore, metabolism of methoxyflurane by purified isozyme 5 is markedly stimulated by cytochrome b5. A role for cytochrome b5 in cytochrome P-450 isozyme 5-catalyzed metabolism of methoxyflurane was also demonstrated in microsomes. Antibody to isozyme 5 was unable to inhibit methoxyflurane metabolism in the presence of maximally inhibiting concentrations of cytochrome b5 antibody.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Phenobarbital treatment significantly increases methoxyflurane metabolism in rabbits. This study identifies cytochrome P-450 isozymes 2 and 5 as key catalysts in both liver and lung microsomes for this enhanced drug metabolism.
Area of Science:
- Pharmacology
- Biochemistry
- Drug Metabolism
Background:
- Phenobarbital administration is known to induce hepatic microsomal enzymes.
- Previous studies indicated that cytochrome P-450 isozyme 2 alone could not fully explain the phenobarbital-induced increase in methoxyflurane metabolism.
Purpose of the Study:
- To quantify the contribution of cytochrome P-450 isozymes 2 and 5 to methoxyflurane O-demethylation in rabbit hepatic and pulmonary microsomes.
- To investigate the role of cytochrome b5 in the metabolism of methoxyflurane catalyzed by isozyme 5.
Main Methods:
- Enzyme kinetics studies using purified cytochrome P-450 isozymes 2 and 5.
- Incubation of methoxyflurane with hepatic and pulmonary microsomes from untreated and phenobarbital-treated rabbits.
- Use of antibodies against isozyme 5 to assess its role in metabolism.
Main Results:
- Cytochrome P-450 isozyme 2 accounted for 25-29% of hepatic O-demethylation and 25% of pulmonary metabolism.
- Cytochrome P-450 isozyme 5 catalyzed 19-27% of hepatic and 47% of pulmonary methoxyflurane O-demethylation.
- Cytochrome b5 significantly stimulated methoxyflurane metabolism by purified isozyme 5 and in microsomal preparations.
Conclusions:
- Both cytochrome P-450 isozymes 2 and 5 are significant contributors to methoxyflurane O-demethylation in rabbit liver and lung microsomes.
- Cytochrome b5 plays a crucial role in potentiating the activity of isozyme 5 in methoxyflurane metabolism.