FAST DETERMINATION OF LOW-LEVEL CYTOCHROME-P-450 1A1 ACTIVITY BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH FLUORESCENCE OR VISIBLE ABSORBENCY DETECTION

A method to determine the activity of the cytochrome P-450 1A1 enzyme, by measuring 7-ethoxyresorufin-O-deethylase (EROD) activity using hig h-performance Liquid chromatography (HPLC) with fluorescence or with v isible absorbance detection of resorufin, is described. The lowest qua ntifiable activity (0.2 pmol/mg min) is obtained by incubation of 0.3 mg of human duodenal microsomal proteins using HPLC fluorescence detec tion. Using HPLC with visible absorbance detection, sensivity was ten times lower. However, the equipment for this last method is available in most laboratories. The use of both HPLC assays allows determination of the low EROD activity level in samples of small size, such as two or three human duodenal biopsies obtained by routine endoscopy. These methods will be a useful tool to study the role of drug intestinal met abolism by cytochrome P-450 1A1.