High-Throughput Quantitative LC-MS/MS Analysis of Benzodiazepines in Human Urine

We hereby present a fast, high-throughput, and clinical LC-MS/MS assay for the simultaneous analysis of benzodiazepines in human urine. The assay is used as both a confirmations and semi-quantitative assay for the abovementioned drugs of abuse following immunoassay urine drug screens. Urine levels are reported in the range of 25 ng/mL–500 ng/mL for each of the 22 analytes, based on a six-level calibration and using a subset (10) of stable isotopically labeled analogues as internal standards. The urine sample is clarified, diluted ten times in internal standard reagent, and thereafter injected into the LC-MS/MS instrument. Reversed-phase liquid chromatography is used to separate the mixture, and the TSQ Endura triple-quadrupole (QqQ) MS instrument performs detection via positive-mode electrospray ionization multiple reaction monitoring.