Abstract Gas chromatographic (GC) and high-performance liquid chromatographic (HPLC) methods were developed for the determination of genaconazole in biological fluids. Both methods involved organic solvent extraction followed by solid-phase extraction on a C 18 column. GC analysis utilized a megabore column (DB-17) with 63Ni electron-capture detection, whereas HPLC analysis utilized separation on a reversed-phase column with a methanol—phosphate buffer mixture as the mobile phase and quantitation by UV absorbance at 208 nm. Both methods yielded good linearity, accuracy and precision. The limit of quantitation (LOQ) was 0.025 μg per ml of serum for GC and 0.2 μg per ml of serum of 0.5 μg per ml of urine for HPLC analysis. Both GC and HPLC methods were used for the determination of serum concentration—time curves of genaconazole in man following oral administration of a 50-mg dose.