The measurement of nitric oxide synthase in rats' brain

An enzyme reaction system which contains L-Arg, NADPH, FAD, FMN, BH4, Ca2+ etc, has been studied to determine the activity of nitric oxide synthase (NOS) in rats' brain. After being incubated at 37 degrees C for 80 min in dark with samples, the sulfanilic acid and N-(l-naphthyl)-ethylenediammonium was added to the reaction system to measure the concentration of produced nitrite in system. It was found that the optimum pH for the enzyme reaction was 7.4, K-m was 0.1 mmol/L. There was a relationship between the amounts of sample and nitrite concentration in the system. This method is simple and convenient. Intra-assay CV value is 3.69% and inter-assay CV value is 5.16%. The specific activity of nitric oxide synthase in ten rats' brains is (39.61 +/- 7.64) nmol/(min . g).