A high-performance liquid chromatographic method with precolumn fluorescence derivatization using 2-(5-chlorocarbonyl-2-oxazolyl)-5, 6-methylenedioxybenzofuran is described for the quantification of 2', 3'-dideoxyinosine, a therapeutic drug for acquired immunodeficiency syndrome, and 2', 3'-dideoxyadenosine, an anti-human-immunodeficiency-viral agent, in rat plasma. The dideoxyribonucleosides and 3'-deoxythymidine (internal standard) in rat plasma (0.1 ml) are cleaned up by a solid-phase extraction technique using an octadecyl silica (ODS) cartridge, Toyopak ODS M, and the dideoxyribonucleosides in the eluate are reacted with the reagent to produce the corresponding fluorescent esters. The esters are separated by chromatography on a reversed phase column, TSKgel ODS-80T_M. The detection limits (signal-to-noise raito = 3) for the dideoxyribonucleosides are 1.3-5.4 pmol on column. Plasma concentrations of 2', 3'-dideoxyinosine after intra-jugular-vainous administration to rat can be monitored by this method.