Quaternary ammonium-linked glucuronidation of 1-substituted imidazoles by liver microsomes: interspecies differences and structure-metabolism relationships.

N-Glucuronidation at an aromatic tertiary amine of 5-membered polyaza ring systems was investigated for a model series of eight 1-substituted imidazoles in liver microsomes from five species. The major objectives were to investigate substrate specificities of the series in human microsomes and interspecies variation for the prototype molecule, 1-phenylimidazole. The formed quaternary ammonium-linked metabolites were characterized by positive ion electrospray mass spectrometry. The incubation conditions for the N-glucuronidation of 1-substituted imidazoles were optimized; where for membrane disrupting agents, alamethicin was more effective than the detergents examined. The need to optimize alamethicin concentration was indicated by 4-fold interspecies variation in optimal concentration and by a change in effect from removal of glucuronidation latency to inhibition on increasing concentration. For the four species with quantifiable N-glucuronidation of 1-phenylimidazole, there were 8- and 18-fold variations in the determined apparent K(m) (range, 0.63 to 4.8 mM) and V(max) (range, 0.08 to 1.4 nmol/min/mg of protein) values, respectively. The apparent clearance values (V(max)/K(m)) were in the following order: human congruent with guinea pig congruent with rabbit > rat congruent with dog (no metabolite detected). Monophasic kinetics were observed for the N-glucuronidation of seven substrates by human liver microsomes, which suggests that one enzyme is involved in each metabolic catalysis. No N-glucuronidation was observed for the substrate containing the para-phenyl substituent with the largest electron withdrawing effect, 1-(4-nitrophenyl)imidazole. Linear correlation analyses between apparent microsomal kinetics and substrate physicochemical parameters revealed significant correlations between K(m) and lipophilicity (pi(para) or log P values) and between V(max)/K(m) and both electronic properties (sigma(para) value) and pKa.