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Abstract: Objectives: The aim of this study was to develop a simultaneous determination method for voriconazole (VRCZ) and its N-oxide (VNO) in human plasma and to apply it to clinical samples. Design and methods: Plasma specimens were deproteinized with acetonitrile and then injected into an HPLC-UV system. VRCZ and VNO were separated on an ODS column filled with 2.3-μm particles using an isocratic mixture of acetonitrile/methanol/phosphate buffer 25/10/65 (v/v/v). Plasma concentrations of VRCZ and VNO in 25 patients were determined. Results: Both VRCZ and VNO were linear (r >0.999) over the concentration ranges of 0.1–8.0mg/L. The run time for quantification of the analytes was 4min. Interindividual variation in the plasma concentration of VRCZ that ranged from less than 0.1 to 6.96mg/L was observed. VNO concentrations weakly depended on VRCZ dosage. Conclusions: The present method can contribute to the optimization of antifungal therapy using VRCZ. [Copyright &y& Elsevier]