A highly sensitive method to quantify triazolam and its metabolites with liquid chromatography--mass spectrometry

We established a highly sensitive method to determine triazolam and its major metabolites, alpha-hydroxytriazolam and 4-hydroxytriazolam, in human plasma and urine with a liquid chromatography-mass spectrometry system which incorporates an atmospheric chemical ionization interface. A plasma sample and a urine sample after solvent extraction were injected into an ODS column of reversed phase with a mobile phase in a linear solvent gradient of initially 50 mM ammonium acetate (pH 4.0) 50%:methanol 50% and 15 min later methanol 100%; quantification limits of as low as 20 pg/mL at an SNR of 3 were obtained for each compound. With diazepam as an internal standard, recovery yields of 84, 81, and 77% were obtained for triazolam, alpha-hydroxytriazolam and 4-hydroxytriazolam, respectively.