Journal of Analytical Toxicology, Vol. 15, November/December 1991 Simultaneous HPLC Analysis of the Hypnotic

Abstract l A new analytical methodology was developed for simultaneous high-performance liquid chromatographic quantitation of nitrazepam, estazolam, flunitrazepam, and trlazolam in plasma. After addition of the internal standard nor-prazepam, biological samples and calibration standards were extracted at basic pH into dlethyl ether-methylene chloride (2:1, v/v). The reconstituted extract was separated on a reversed-phase Nova Pak Ct8 column with acidic buffer (6mM)-acetonitrile-methanol (64:23:13, v/v) as mobile phase. The detection was performed at 242 nm. Within-run and day-to-day precision values were about 2-8%. The method is thus sensitive and reproducible for use in clinical and experimental toxicokinetic studies. High-performance liquid chromatographic (HPLC) method