A method for the determination of a bisheteroarylpiperazine, non-nucleoside HIV-1 reverse transcriptase inhibitor, delavirdine, and its N-desisopropyl metabolite in human plasma, is described. Samples were deproteinized by addition of two parts of a solution of internal standard in acetonitrile (1 microgram/ml) to one part plasma. The supernatant was diluted with 10 mM phosphate buffer, pH 6.0, and injected onto the HPLC system. Fluorescence of the eluent was monitored with excitation at 302 nm and emission at 425 nm. Quantitation of delavirdine and its metabolite was achieved by comparing the peak-height ratio of each component relative to the internal standard to a through-the-origin linear regression curve determined from fortified plasma calibration standards. The assay was linear over the concentration range 0.02-17 microM for both delavirdine and its metabolite. The precision of the method, as expressed by the mean C.V. of the back-calculated, non-zero, standard concentrations, was +/- 4.4% for delavirdine and +/- 4.3% for the metabolite. The assay has been validated and utilized to analyze samples from human and animal pharmacokinetic studies.