Hepatic Aminopyrine N-Demethylase System: Further Studies of Assay Procedure

Rat hepatic aminopyrine N-demethylase activity was measured by detecting the amount of formaldehyde produced from aminopyrine. Some optimal conditions for the N-demethylation were determined using both isolated microsomes and whole homogenates, and the standard assay method is described. Formaldehyde production from the substrate by microsomal enzyme system was linear to the amount of microsomes added during 3 min reaction time, whereas long-time incubation caused a decrease in the apparent activity of aminopyrine N-demethylation. The N-demethylase activity observed in normal rat liver homogenate was quite similar to that in microsomes when the activity was expressed on the basis of cytochrome P-450 as molecular activity. Pretreatment of animals with typical inducers, phenobarbital and 3-methylcholanthrene, resulted in alteration of the aminopyrine N-demethylase system, which was detectable in both microsomes and whole homogenates.