A high-performance liquid chromatography-UV methodology (lambda=230 nm) was developed and validated for the simultaneous determination of vincristine and doxorubicin in pharmaceutical preparations used in oncology. The chromatography was carried out on a C18 column using acetonitrile 90% in water-potassium hydrogenphosphate buffer 50 mM, pH 3.2+/-0.1 (32:68, v/v) as mobile phase at a flow rate of 1.5 mL/min. The method proved to be specific, exact, and accurate, in accordance with the ICH standards, presenting linearity in the 1-5 microg/mL and 5-100 microg/mL intervals, and detection (0.19x0.51 microg/mL) and quantification (0.63x1.7 microg/mL) limits for vincristine and doxorubicin, respectively.