Dependence of hydrogen peroxide formation in rat liver microsomes on the molecular structure of cytochrome P-450 substrates: A study with barbiturates and β-adrenoceptor antagonists

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Abstract
In the present study, the molecular structure of xenobiotics has been successfully linked to their effect on the oxidase activity of cytochrome P-450, determined as microsomal hydrogen peroxide formation. A homologous series of 5-alkyl-5-ethyl barbiturates and a heterologous series of β-adrenoceptor antagonists was used. The logarithm of the rate of microsomal hydrogen peroxide formation could be correlated with the logarithm of the apparent partition (n-octanol/buffer) coefficient for the barbiturate derivatives according to a parabolic function. The statistics of the correlation improved considerably by applying a bilinear model in order to fit the data. This probably indicates that both transport of the substrate to cytochrome P-450 and interaction with hydrophobic substrate binding sites of cytochrome P-450 are involved in the modulating effect of substrates on the oxidase function of cytochrome P-450. With the series of β-adrenoceptor antagonists no clear-cut structure activity relationship with regard to the oxidase activity was apparent at first sight. However, when the inhibitory effect of the β-antagonists on the ‘cytochrome P-450 metabolic intermediate (MI) complex’ formation that occurs during the microsomal biotransformation of 33 μM tofenacine was studied a relationship with the lipophilicity could be demonstrated. It is known that MI complex formation occurs with specific subforms of cytochrome P-450. By using this inhibitory activity of the β-adrenoceptor antagonists, the interaction of the compounds becomes restricted to these specific subforms of cytochrome P-450. Li both the oxidase activity as well as the MI complex formation phénobarbital induced cytochrome P-450 is involved. Accordingly a relationship could be established between the percentage of inhibition of microsomal hydrogen peroxide formation and the percentage of inhibition of MI complexation. A distinction between β-antagonists that interact through a ligand or a substrate binding with cytochrome P-450 improved the correlation. Remarkably, d,l-propranolol enhanced the microsomal hydrogen peroxide formation. Both enantiomers of propranolol increased the oxidase activity of cytochrome P-450 to the same extent. The present study demonstrates that prediction of the oxidase activity of cytochrome P-450 is possible provided certain precautions with regard to the specificity of interactions of substrates with cytochrome P-450 are taken into account.