Development of a dried blood spot screening test using gas chromatography coupled to high-resolution mass spectrometry: application in authentic postmortem samples

This study aims to develop a toxicological screening analysis using gas chromatography coupled with high-resolution mass spectrometry (GC-HRMS) on postmortem dried blood spots (DBSs) and apply it to authentic cases. Twenty-five microliters of blood was deposited and dried on a paper card. Compounds of interest were desorbed, extracted, and acetylated, then injected into the GC-HRMS. The limits of detection (LOD) and of identification (LOI) were determined for 22 of the compounds that were most frequently detected in postmortem blood samples in the laboratory in 2022. Stability on DBS was studied at three temperatures (-20°C, +4°C, and +20°C) over 15 days. The method was then applied to 102 postmortem blood samples. Results were compared to the two conventional screening methods implemented in the laboratory: liquid chromatography coupled with diode-array detection and mass spectrometry (LC-DAD-MS) and GC-MS. Selectivity was demonstrated by the analysis of 10 negative postmortem blood samples. All LODs were between <10.0 and 20.0 ng/mL. LOIs were within the therapeutic concentration range for each compound or at a value not leading to acute intoxication (narcotics). Overall, compounds remained stable over the 15 days at all test temperatures, except for midazolam and tramadol and its metabolites. A comparison of screenings of 102 postmortem samples resulted in 239 identifications, corresponding to 74 compounds, across 70 positive cases. In 32 cases, no compound was identified. Compounds of 57% and 60%, respectively, were detected by LC-DAD-MS and GC-MS screenings, while DBS-GC-HRMS identified 81%. Application of the method to a hundred authentic cases demonstrated its ability to meet the constraints of low sample volume, sensitivity, and ease of preservation for urgent cases or cases with limited blood availability.