Development and validation of rapid and sensitive HPLC method for the quantitative determination of doxorubicin in human plasma

An isocratic reversed-phase high-performance liquid chromatographic method with ultraviolet detection at 254 nm has been developed for the determination of doxorubicin in human plasma. Plasma samples were extracted by a selective one-step liquid–liquid extraction using dichloromethane. Doxorubicin and the internal standard epirubicin were separated using a column packed with C18 material, using a mobile phase consisting of water:acetonitrile (75:25v/v). The calibration graph for doxorubicin was linear in the range 0.2–10 μg/mL, with a correlation coefficient, R2 = 0.9986. Lower limit of quantitation was 0.2 μg/mL, using 1 mL plasma samples. The extraction recovery ranged from 98.5–101.1%, and the recovery rate was consistent for drug and internal standard examined at each level. The interday and intraday precision values ranged between 0.5–2%. Validation data showed that the assay for doxorubicin is sensitive, selective, accurate, and reproducible. The assay has been used in population pharmacokinetics study.