High-performance liquid chromatographic determination of midazolam in rat brain.

A high-performance liquid chromatography method for the determination of midazolam in rat brain is described. Midazolam and the internal standard halazepam were extracted with toluene and analyzed isocratically on a reversed-phase column with a mobile phase consisting of methanol, acetonitrile and potassium phosphate buffer. Detection was monitored by ultraviolet absorption at 240 nm. The standard curves were linear over the range of 25-350 ng midazolam per 50 mg brain tissue. The day-to-day coefficient of variation ranged from 1.7 to 6.9%. The limit of quantification was 80 ng/g brain tissue. The method is rapid, simple and reproducible for brain analysis.