Fluorometric assay of rat brain N-methyltransferase with 4-methylnicotinamide.

A fluorometric method for the assay of rat brain N-methyltransferase (NMT) has been developed using rat brain 9000 x g supernatant fluid as an enzyme preparation. The method is based on the enzyme reaction at pH 8.6 using 4-methylnicotinamide (4-MN) as the methyl acceptor substrate in the presence of cofactor S-adenosyl-L-methionine (AdoMet), and the fluorometric determination of the product 1,4-dimethylnicotinamide by means of its reaction with 4-methoxybenzaldehyde. The apparent Km values for 4-MN and AdoMet were 0.47 mM and 19 microM, respectively. The method was successfully applied to the studies of age- and sex-related differences of brain NMT activity in rats. The enzyme activity decreased gradually between 3 and 14 weeks of age whereas there were no differences between males and females in the levels of NMT activity.