MSACL 2016 US Abstract

Quantitation of Antidepressant Drugs and their Metabolites by High-Resolution Mass Spectrometry Utilizing PRM and Full-MS Scan Modes for Forensic Research

Anastasia Kalli (Presenter)
Thermo Fisher Scientific

Authorship: Anastasia Kalli, Marta Kozak
Thermo Fisher Scientific, 355 River Oaks Parkway, San Jose, CA

Short Abstract Quantitative analysis of antidepressant drugs is performed in forensic laboratories to support criminal cases. We have performed quantitative analysis of antidepressants drugs and their metabolites in human plasma on a Q ExactiveTM Orbitrap mass spectrometer utilizing high mass-accuracy high-resolution mass spectrometry. Quantitation was performed with both full MS scan followed by MS/MS for confirmation and PRM scan modes. The LOQs obtained with both full-MS and PRM scan modes were well below the values required by forensic labs for all compounds examined. Method specificity, linearity and precision were comparable between the two scan modes.

Long Abstract

Introduction

In forensic toxicology laboratories, quantitative analysis of antidepressant drugs and their metabolites is performed in cases such as driving under the influence, violent crime and unknown cause of death. Here we present a quantitative method for the analysis of antidepressant drugs on the Thermo ScientificTM Q ExactiveTM Orbitrap mass spectrometer utilizing high-resolution, high-mass accuracy mass spectrometry. We have utilized full-MS and PRM scan modes for the quantitative analysis of venlafaxine, O-desmethylvenlafaxine, doxepin, desmethyldoxepin, clomipramine, N-desmethylclomipramine, imipramine and desipramine in human plasma. The specificity of full-MS followed by MS/MS for confirmation was compared to that of the PRM scan mode. Linearity, method precision and reproducibility, LOD and LOQ obtained by each scan mode will be presented.

Methods

The plasma calibration standards were prepared by spiking human plasma with aqueous drug standard (venlafaxine, O-desmethylvenlafaxine, doxepin, desmethyldoxepin, clomipramine, N-desmethylclomipramine, imipramine and desipramine). After the addition of internal standards,protein precipitation was achieved by the addition of ACN. The concentration of each calibration standard ranged from 1 ng/ml to 1000 ng/ml. Ten microliters were injected for analysis. Chromatographic separation was performed with a Thermo ScientificTM DionexTM UltimateTM 3000 RS LC on a Thermo ScientificTM Accucore PFP column (50 × 2.1 mm, 2.6 µ particle size). Mobile phase A was 10 mM ammonium formate in water and mobile phase B was 10 mM ammonium formate in 80:20 MeOH: ACN. Mass spectrometric analysis was performed on a Thermo ScientificTM Q ExactiveTM Orbitrap mass spectrometer operated in full-MS and PRM scan modes. Full MS scans were acquired at a resolution of 70,000, whereas, PRM scans were acquired at 35,000 resolution. Analysis time, including column equilibration, was 5 min.

Results

Data processing was performed with TraceFinderTM 3.3. Quantitation of all analytes was based on high mass accuracy measurements. Mass error of < 3 ppm was obtained with external calibration for both the PRM and full MS scan modes. Quantitation in PRM scan mode was based on the most abundant fragment ion of each analyte in collected MS/MS spectrum. The LOQs obtained with both full-MS and PRM scan modes were well below the values required by forensic laboratories for all compounds examined. Linearity and specificity were comparable between PRM and full MS scan mode followed by MS/MS for confirmation.

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