Highly sensitive and specific high-performance liquid chromatographic analysis of 7-hydroxywarfarin, a marker for human cytochrome P-4502C9 activity

The formation of 7-hydroxywarfarin in incubations of (S)-warfarin with human liver microsomes reflects their cytochrome P-4502C9 activity. This paper describes a rapid high-performance liquid chromatographic method for the determination of 7-hydroxywarfarin with high sensitivity, selectivity, and a simple sample clean-up procedure. Separation was achieved with a C₁₈ reversed-phase column and quantification by fluorometric detection. The method employs an internal standard resulting in good accuracy and precision. The limit of detection is 150 fmol for 7-hydroxywarfarin.