Ketoconazole is a new antifungal drug which has increasing clinical application due to its wide spectrum of activity and oral availability. Since substituted imidazoles may be potent inhibitors of cytochrome-mediated drug oxidation, we have examined the effect of ketoconazole on aminopyrine and caffeine metabolism in the rat. The 14C breath test was used to determine the elimination rate of aminopyrine and caffeine in vivo after single and chronic (7 day) treatment with ketoconazole. Acutely, ketoconazole (50 mg/kg) impaired markedly both aminopyrine (56% inhibition) and caffeine demethylation (83% inhibition). Inhibition of aminopyrine and caffeine demethylation as studied with the breath tests was related to ketoconazole dose. The decreased rate of aminopyrine demethylation measured by breath test correlated with decreased aminopyrine clearance from plasma after iv dosage. Chronic treatment with ketoconazole further increased the inhibition of aminopyrine demethylation while chronic treatment was associated with less inhibition of caffeine demethylation than found with a single dose. Breath tests repeated up to 72 hr after ketoconazole treatment revealed differences in aminopyrine and caffeine demethylation. At 24 hr after ketoconazole, aminopyrine demethylation was still inhibited at 78% of controls whereas caffeine demethylation was enhanced to 146% of control. Binding of ketoconazole to microsomal P-450 was determined by spectral analysis. A type II difference spectrum was found with absorbance maximum at 430 nm and minimum at 390 nm. Scatchard analysis revealed a biphasic interaction with estimated dissociation constants of 0.6 and 3.69 microM. Aminopyrine N-demethylation in vitro was markedly impaired with an I50 for ketoconazole of 27 microM and Ki of 8.5 microM.(ABSTRACT TRUNCATED AT 250 WORDS)