A novel reversed-phase high-performance liquid chromatographic (HPLC) method has been established for the determination of a newly developed anti-cancer agent desoxyepothilone B (dEpo B) in nude mice plasma. The sample preparation involved deproteination of 200μl of plasma sample first, followed by liquid–liquid extraction of the resultant supernatant with chloroform. The compound taxol was used as the internal standard. Chromatographic separations were carried out on a 250mm×4.6mm Zorbax SB-phenyl column with acetonitrile–0.25% orthophosphoric acid (50/50, v/v) as mobile phase and UV detection at 250nm. For dEpo B and taxol at the concentration level of 10μg/ml in nude mice plasma, the absolute extraction recoveries were 85.3 and 87.2%, respectively. The linear quantification range of the method was 0.1–100μg/ml in nude mice plasma with linear correlation coefficients greater than 0.999. The within-day and between-day relative standard deviations (R.S.D.s) for dEpo B at 0.5, 2.5 and 10μg/ml levels in nude mice plasma fell in the range of 2.8–4.8 and 1.5–4.6%, and the within-day and between-day recoveries were in the range of 96.5–101.7 and 97.7–101.2%, respectively.