Oxantrazole is an anthrapyrazole analogue developed as an anthracycline-like agent with potentially reduced cardiotoxicity. A reversed-phase high-performance liquid chromatographic assay was developed using a C2 column and mobile solvent system of dimethylformamide-acetonitrile-0.2 M ammonium acetate, pH 4.5 (20:5:75, v/v/v) at a flow-rate of 1 ml/min. Drug and internal standard were detected by ultraviolet absorbance at 514 nm. Isolation of drug and internal standard was afforded by elution from C18 disposable isolation columns with a mixture of methanol-glacial acetic acid-0.02 M sodium acetate, pH 4.0 (12:1:3, v/v/v). The assay was linear (r2 greater than 0.99) over concentrations of 0.025-2.5 micrograms/ml and the limit of detection was 10 ng/ml plasma. Oxantrazole was unstable in neutral and particularly alkaline aqueous solutions. Utilizing this assay, plasma pharmacokinetics were determined following intravenous infusion of oxantrazole to beagle dogs. Plasma elimination was rapid with elimination phase half-life values less than 45 min.