PH effects on the N-demethylation and formation of the cytochrome P-450 iron II nitrosoalkane complex for erythromycin derivatives

Abstract

The effects of pH on access to the cytochrome P-450 active site, N-demethylation and formation of the cytochrome P-450 Fe(II)-RNO metabolite complex for a series of erythromycin derivatives were examined. Studies were performed with dexamethasone-treated rat liver microsomes containing large amounts of cytochrome P-450 3A isozymes. In addition to factors such as hydrophobicity or hindrance around the dimethyl-amino function, the ionisation state of the N(CH 3) 2 group played an important role in the recognition and metabolism of the substrate by cytochrome P-450. Esterification of the desosamine in the β position of the N(CH 3) 2 group leads to lower p K a values for the R-N + H(CH 3) 2 ag [R-N (CH 3) 2] + H + equilibrium. At physiological pH, the amine group is mainly in the unprotonated form. Consequently, easier access to the protein active site and significant formation of cytochrome P-450 Fe(II)-RNO metabolite complex are observed for these derivatives. These results led us to interpret the formation of cytochrome P-450 Fe(II)-RNO metabolite complex as a series of multiple steps equilibria depending on the ionisation state of the N(CH 3) 2 group, the partition coefficient of the substrate between the microsomal layer and the aqueous media and a series of metabolic reactions leading partially to the final inhibitory nitrosoalkane-cytochrome P-450 Fe(II) complex.