A new HPLC assay was developed to study dextromethorphan O-demethylation to dextrorphan in vitro using human liver microsomes to investigate the activity of the polymorphic monooxygenase cytochrome P450 2D6 (CYP 2D6). The separation of dextromethorphan and its main metabolite dextrorphan was performed on a polymeric C18 reversed-phase column with UV-detection using levallorphan as an internal standard. Liver samples from ten subjects were screened for dextrorphan formation whereby three groups with different abilities to metabolize dextromethorphan could be found. Seven microsomal preparations from extensive metabolizers showed an average dextrorphan formation rate of 298 +/- 68 pmol/mg protein.min, one sample was classified to belong to an intermediate dextromethorphan metabolizer (79 pmol/mg protein.min), whereas two samples of poor metabolizers exhibited significantly lower rates of dextromethorphan metabolism with values of 11 and 27 pmol/mg protein.min, respectively. This assay permits not only a fast in vitro screening for cytochrome P450 2D6 monooxygenase activity but is also an excellent tool to determine potential drug-drug interactions with this important metabolizing enzyme.