A microassay for the detection of low levels of cytochrome P450 O-deethylation activities with alkoxyresorufin substrates.

A microfluorometric method for the detection of low levels of cytochrome P450 was developed to increase the sensitivity of the assay, since a low level of CYP450 associated enzymatic activities was expected in human placenta tissues and small samples of placenta (approximately 10 g) could be easily collected, stored and processed. The dual fluorescence assay of Kennedy et al. [1], which was developed to simultaneously quantitate microsomal proteins and ethoxyresorufin-O-deethylase (EROD) activity was adapted for 96 wells microtiter plates. Placental microsomes samples were analyzed. For samples obtained from non-smoking mothers from the general southern Quebec population, results ranged from less than 1-3.3 pmol/mg protein.min. Samples collected from smoking mothers showed activity levels ranging from 30-69 pmol/mg protein.min. These results showed the suitability of the microassay for measuring low level of CYP450 activity in tissues such as placenta.