Development and Validation of an LC-MS/MS Method for the Simultaneous Determination of Alprazolam, Bromazepam, Clonazepam, Diazepam and Flunitrazpam in Human Urine and Its Application to Samples from Suspected Drug Abusers
Year: 2025
Type: article
Abstract: A simple and reliable method was developed using LC-MS/MS to quantify alprazolam, bromazepam, clonazepam, diazepam, and flunitrazepam in clinical samples. This method was validated for the simultaneous determination of alprazolam, bromazepam, clonazepam, diazepam, and flunitrazepam. It was applied to human urine samples collected from people suspected of drug abuse in the Kuwaiti region. Formic acid in water and acetonitrile was used in mobile phase with a gradient mode of elution using C18 reverse-phase column. The instrument was operated in a positive mode with an electrospray ionization source using multiple reaction monitoring. For sample extraction, the liquid-liquid extraction technique was used. The method was validated for limit of detection, limit of quantitation, selectivity, linearity, accuracy, and precision. The concentration for limit of quantitation was 6.0 ng/mL, the linearity ranged from 2.0 to 300 ng/mL for each of the analytes, and the r2 values were ≥0.99. The accuracy was found to be within a range of 80–120% and precision had a %RSD of ≤15% for each of the analytes. The method was applied to 48 urine samples collected from those suspected of drug abuse by the Toxicology Department of the General Department of Criminal Evidence, Kuwait, and alprazolam, bromazepam, clonazepam, diazepam and flunitrazepam were identified commonly in the samples. The overall drug positivity rate obtained considering 48 samples was 93.75%. Based on these results and successful determination of alprazolam, bromazepam, clonazepam, diazepam and flunitrazepam in human urine samples from those suspected of drug abuse, this method is deemed to be suitable for its routine analysis.
Source: Molecules
Institutions: University of Strathclyde, Kuwait University
Language: English
Subfield: Toxicology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,258
OpenAlex ID: W4413440500