A New Strategy for Efficient Retrospective Data Analyses for Designer Benzodiazepines in Large LC-HRMS Datasets.

Area of Science:

• Forensic Chemistry
• Analytical Chemistry
• Mass Spectrometry

Background:

• The increasing prevalence of new psychoactive substances (NPSs) presents analytical challenges for forensic laboratories.
• Retrospective data analysis (RDA) offers a way to identify previously missed analytes but is often time-consuming and labor-intensive.
• Developing efficient and scalable workflows is crucial for monitoring NPS abuse.

Purpose of the Study:

• To establish an efficient and scalable retrospective data analysis (RDA) workflow for detecting designer benzodiazepines (DBZDs) in whole blood samples.
• To apply this workflow to previously analyzed driving-under-the-influence-of-drugs (DUID) cases.
• To optimize the workflow for groups of NPSs, particularly benzodiazepines.

Main Methods:

• Developed an RDA workflow utilizing ultrahigh-performance liquid chromatography-quadrupole time-of-flight-mass spectrometry (UHPLC-QTOF-MS) data.
• Used a training set of confirmed benzodiazepine hits (analyzed by UHPLC-MS/MS) to establish true/false positive criteria and set filters (retention time, count, mass error).
• Applied the validated workflow to 13,514 UHPLC-QTOF-MS data files from DUID cases (2014-2020) to screen for 47 targeted DBZDs.

Main Results:

• The RDA workflow successfully identified 16 designer and uncommon benzodiazepines (DBZDs) in the analyzed DUID cases.
• A total of 47 confirmed positive findings and 43 tentative positive findings were reported.
• The method demonstrated high efficiency with only nine false-positive hits and the capability to screen previously acquired DUID data files for emerging DBZDs in under one minute.

Conclusions:

• The developed RDA workflow is an efficient, scalable, and accurate method for identifying DBZDs in large forensic datasets.
• This technological advancement significantly improves the monitoring of NPS abuse by enabling rapid screening of historical data.
• The workflow can be easily adapted for other groups of NPSs, offering a valuable tool for forensic laboratories worldwide.