A simplified, rapid high-performance liquid chromatographic procedure has been developed for the measurement of toloxatone in rabbit plasma and cerebrospinal fluid. The method involves a single-step extraction of the alkalinized sample with diethyl ether and analysis of the evaporated extract on a C8 column. Detection was performed by ultraviolet absorbance monitored at 240 nm. The overall run-time of the assay was 8 min at a flow-rate of 1 ml min-1. The sensitivity limit of toloxatone was 70 ng ml-1 at a signal-to-noise ratio of 3:1 in rabbit cerebrospinal fluid and plasma. The assay has been used to define plasma toloxatone concentration-time profiles and to quantitate cerebrospinal fluid toloxatone levels after a single intravenous injection in rabbits.