A high-performance liquid chromatographic assay for the determination of labetalol, a novel antihypertensive agent, in human plasma was developed. Reversed-phase separation of labetalol and the internal standard was accomplished on a 150 X 4.1 mm column commercially packed with a spherical (8-12 micron particle size) macroporous co-polymer (PRP-1). Unlike silica-based columns, the unique properties of PRP-1 permit operation at pH extremes. Based on this advantage, a mobile phase which was sufficiently basic (pH 9.5) to optimize the fluorescent yield of analyte and provide the necessary specificity was selected. Detector response (peak area ratio) was linear from 4 to 500 ng/nl. Following a simple extraction procedure, samples were automatically injected and analyzed using micro-processor-controlled equipment. No interferences were observed in the extracts obtained from drug-free plasma which were processed under the conditions described for unchanged drug. The limit of quantitation using 0.5 ml of plasma was validated to 4 ng/ml. The inter-assay precision (coefficient of variation) was less than 4.6% at all concentrations evaluated from 4 to 300 ng/ml. This method is suitable for the routine quantitation of labetalol or its RR isomer (dilevalol) in plasma (0-24 h) following the administration of therapeutically effective doses to man.