Defense Date
2024
Document Type
Directed Research Project
First Advisor
Michelle Peace
Abstract
The emergence of designer benzodiazepines (DBZDs) on the illicit market has raised concerns regarding the detection of them within forensic casework. In an attempt to get ahead of this ever evolving market, the analytical validation of a qualitative method for detecting four DBZDs—4-chloro-deschloroalprazolam, deschloroetizolam, desalkylgidazepam, and nifoxipam—was performed. This was done across multiple biological matrices using a liquid-liquid extraction method and analysis using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Following ANSI/ASB Standard 036 guidelines, the study examines endogenous and exogenous interferences, carryover, sample stability, ion suppression/enhancement, and limits of detection (LOD) within blank blood, antemortem blood, postmortem blood, blank urine, and liver homogenate.
Interference studies confirmed no signal overlaps from endogenous or commonly encountered exogenous substances, indicating a reliable analytical method free from background noise within these matrices. Minimal carryover was observed, which was primarily at elevated analyte concentrations in postmortem blood, meeting acceptable criteria and suggesting rare carryover occurrences within practical forensic applications. Stability analysis revealed consistent results for deschloroetizolam, desalkylgidazepam, and 4-chloro-deschloroalprazolam over seven days; however, nifoxipam demonstrated pronounced instability, especially in urine, likely impacting its forensic reliability in delayed analysis scenarios. Ion suppression and enhancement effects varied by matrix, with liver homogenate and urine displaying suppression/enhancement for specific analytes. These matrix-specific interferences prompted reassessment of LODs to ensure analytical sensitivity, with deschloroetizolam validated at a 0.0025 mg/L detection threshold and desalkylgidazepam at 0.01 mg/L across matrices. However, 4-chloro-deschloroalprazolam and nifoxipam did not consistently meet LOD criteria, suggesting a need for methodological refinement to improve signal reliability, especially at low concentrations. Structural similarity to alprazolam posed detection challenges for 4-chloro-deschloroalprazolam, while nifoxipam’s instability and signal inconsistency complicated reliable detection.
The findings emphasize the method's potential for forensic analysis of DBZDs, though specific analytes such as nifoxipam require further investigation to resolve instability and signal variability. Suggested future directions include quantitative validation for robust detection of deschloroetizolam and desalkylgidazepam and enhanced extraction and storage protocols to address stability challenges with nifoxipam and 4-chloro-deschloroalprazolam. These refinements could improve the reliability of DBZD detection in forensic toxicology, providing critical tools for addressing the increased presence of DBZDs in illicit substance analysis.
Rights
© The Author(s)
Is Part Of
VCU Master of Science in Forensic Science Directed Research Projects
Date of Submission
12-14-2024