Studies on the incubation mixtures for the in vitro mutagenesis test with metabolic activation (microsomal assay) = 2. Behaviour of cytochromes P-450, P-420, b5, of NADPH-CYT. C-reductases, and p-nitroanisole demethylase.

Cytochromes P-450, P-420, b5, NADPH-cytochrome c-reductase, aminopyrine and p-nitroanisole demethylase and lipid peroxidation were determined at various times in the incubation mixtures for the in vitro microsomal assay for the mutagenic activity of xenobiotics. No effect was observed on cytochromes b5 and P-420. A decrease in cytochrome P-450 (about 50% in 2 hrs.) and a much faster decrease of aminopyrine demethylase and NADPH cytochrome c-reductase (about 50% in 30 min) was noted with mice microsomes. With S9 liver fraction of rat, p-nitroanisole demethylase activity was much more stable than aminopyrine demethylase activity in the presence of lipid peroxidation, but the decrease was faster and at comparable rates for both activities in the presence of 50 mM styrene. The use of simple colorimetric assay as proves of microsomal monooxygenase activity and the importance of this kind of enzyme studies for a better understanding of the in vitro mutagenesis results are discussed.