Liquid chromatographic assay and pharmacokinetics of halazepam and its metabolite in humans.

A reversed-phase high-performance liquid chromatographic method is described for simultaneous quantification of halazepam and its major active metabolite, nordiazepam, in plasma. The method uses a solid-phase extraction procedure to prepare plasma samples. After extraction, the methanolic extract is evaporated, and the residue is then reconstituted in a small volume of mobile phase (a 40:60, v/v, mixture of 0.02 M phosphate buffer, pH 4.0, and methanol) and chromatographed. The total chromatography time for a single sample is approximately 10 min. A sensitivity of 1 ng/mL for halazepam and nordiazepam is attained when 1 mL of plasma is extracted. Analytical recovery of halazepam and nordiazepam added to the plasma ranged from 89 to 96%. The maximum within-day and day-to-day coefficients of variation for each compound at the concentration range of 2 to 100 ng/mL were 8.7 and 10.3%, respectively. Suitability of the method was assessed in a preliminary pharmacokinetic study in which three subjects were given a single 20-mg oral dose of halazepam following an overnight fast. It appeared from our kinetic analysis that halazepam is a drug with a fairly rapid absorption phase that is followed by a slow elimination phase. Mean oral plasma clearance of halazepam was 24 L/h. The mean apparent elimination half-life of nordiazepam (45.22 h) is considerably longer than that of halazepam (21.15 h).