Determination of delavirdine in very small volumes of plasma by high-performance liquid chromatography with fluorescence detection

Delavirdine is a newly developed anti-HIV-1 drug for AIDS therapy. This study describes a sensitive high-performance liquid chromatographic method for the determination of delavirdine in 50 μl of plasma. Samples were deproteinized with 150 μl of a solution of internal standard (cisapride 10 μg/ml) in acetonitrile. An aliquot of the supernatant was injected onto the column. HPLC separation was achieved on a C18 column with the mobile phase of acetonitrile-50 mM sodium dihydrogen phosphate (60:40, v/v) at a flow-rate of 1 ml/min. The eluants were measured by fluorescence detection with excitation at 295 nm and emission filtration at 425 nm. The retention time was about 5.3 min for delavirdine and 6.5 min for cisapride. The specificity was demonstrated, as there were no interferences from plasma samples of different batches in the regions of peak interest. Calibration curves were linear from 25 to 25000 ng/ml. The limit of quantitation was 25 ng/ml. The within- and between-day precision (C.V.) was 9.3%, or less, and the accuracy was within 9.2% of the nominal concentration. The small sample volume needed is especially advantageous for the application both in pharmacokinetic studies in HIV-infected adults and pediatric patients, and in small animals, where limited samples are available.