To permit the characterization of cefpirome disposition in lactating females, a previously published high-performance liquid chromatographic (HPLC) method for determining the drug in serum was adapted for use with milk and urine. This automated, microanalytical technique requires 50 microliters of biological matrix, which is subjected to an isopropanol extraction. Chromatography was accomplished using a microbore HPLC system, a reversed-phase C18 column and a mobile phase of 0.3% triethylamine in water (pH 5.1). Cefpirome and the internal standard (beta-hydroxypropyltheophylline) were monitored using UV detection at 240 nm and had retention times of 2.84 and 5.05 min, respectively. The method was linear up to 500 mg/l for both matrices and had a limit of detection of 0.6 mg/l. The interday variation (relative standard deviation) at concentrations of 5.0, 50.0 and 500.0 mg/l was consistently less than 5% in both urine and breast milk. The method was found to be free from interference by other commonly administered medications and readily adaptable for use in clinical investigations. The ease of sample preparation, small sample volume requirement, short chromatographic time, apparent lack of interferences, analytical sensitivity and high precision and accuracy make this method ideal for use in pharmacokinetic investigations involving the determination of cefpirome in human milk and urine.