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A simple and sensitive high-performance liquid chromatography method was developed for the simultaneous quantitative determination of fluvoxamine and its two metabolites, fluvoxamino alcohol and fluvoxamino acid, in human liver microsomes. Chromatographic separation was achieved with a Grand-pak C4-5 column using a mobile phase at pH 2.5 of 0.5% KH2PO4-acetonitrile ( 75: 25, v/v). Analysis involved a solid-phase extraction with an Oasis HLB cartridge, which gave high extraction recovery (> 92.8%) with good selectivity. The lower limit of quantification of this assay was 78.6 nM for fluvoxamine and fluvoxamino acid, and 82.2 nM for fluvoxamino alcohol, respectively. The coefficient of variation of intra- and interday assays was less than 5.8% and accuracy was within 5.3% for all analytes (concentration range 78.6 nM - 2.36 mu M for fluvoxamine and fluvoxamino acid, and 82.2 nM 2.47 mu M for fluvoxamino alcohol, respectively). This method is applicable for accurate and simultaneous determination of oxidative metabolites of fluvoxamine by human liver microsomes.