Improved Determination of Drug Metabolism by Perifusion of Recombinant V79 Cells Carrying Human CYP3A4.

Recombinant V79Mz cell lines stably co-expressing human CYP3A4 and human NADPH-cytochrome P450 oxidoreductase were used to compare testosterone metabolism under conventional stationary cultivation and under perifusion. In contrast to stationary culture, steady-state conditions for the production of 6beta-hydoxytestosterone were established in the perifusion system even when different concentrations of testosterone were infused sequentially. As a consequence of these favourable conditions, threefold higher maximal metabolic rates could be measured and kinetic constants could be determined more reproducibly than in stationary culture. The results indicate that CYP3A4 shows an autocatalytic reaction kinetic probably due to two binding sites for testosterone.