GC-NCD-QTOF Dual Detector in Quantitative Screening of 38 Stimulant-Type New Psychoactive Substances in Blood without Using Reference Standards Samuel Mesihää (Presenter) University of Helsinki, Department of Forensic Med >> POSTER (PDF) Presenter Bio: Samuel Mesihää received his MSc degree in Biochemistry (Protein Science) from the University of Oulu (Finland). In his master's thesis, he applied high resolution mass spectrometry in tumor marker discovery from urine samples. Currently, he is a PhD student at the Department of Forensic Medicine, at the University of Helsinki. He is interested in screening methods in the clinical mass spectrometry and in analysis of the drugs of abuse.

Authors: Samuel Mesihää (1), Ilpo Rasanen (2), Anna Pelander (2), Ilkka Ojanperä (1,2)
(1) University of Helsinki, Department of Forensic Medicine, Helsinki, Finland, (2) National Institute for Health and Welfare, Forensic Toxicology Unit, Helsinki, Finland

Analysis of the new psychoactive substances (NPS) is a challenge to the clinical toxicology laboratories because it is difficult to obtain an authentic reference standard in short period of time, while these compounds are still being consumed.

An analytical platform was developed for the simultaneous identification and quantification of NPS by using a GC system that directs the gas flow to quadrupole time-of-flight mass analyzer (QTOF) and nitrogen chemiluminescence detector (NCD). In this system, the drug screening is based on the detection of the accurate masses of the protonated molecular ion (MH+) produced in the atmospheric pressure chemical ionization source. Quantification is based on the equimolar response of drugs to nitrogen.

In this work, GC-NCD-QTOF detector used to measure 38 stimulant-type NPS spiked in the blank blood samples.

Introduction

Constantly transforming illicit NPS drug market is characterized by unpredicted pharmacological effects that in some cases may result in severe intoxications. Assessment of NPS toxicity and abuse patterns in population has been difficult to carry out mainly due to limited availability of authentic reference standards in clinical toxicology laboratories.

Methods

Introduction of high resolution mass spectrometer instruments has enabled a tentative screening of NPS in absence of reference standards based on the high mass resolution and mass accuracy. However, quantification still remains inaccurate.

For identification, a high resolution mass spectrometer with atmospheric pressure chemical ionization (APCI) source was used to produce protonated molecular ions (MH+) with high accurate mass. Confidence of the analysis can be improved if MS/MS data is available.

The universal quantification of NPS was done with the nitrogen chemiluminescence (NCD) which exhibits N-equimolar response to nitrogen. It is estimated that >90% of the drugs contain nitrogen and therefore any containing secondary standard can be used to quantify NPS. Separation with gas chromatography (GC) is more suitable than with liquid chromatography (LC) because LC-NCD exhibits poor sensitivity.

Blank blood samples containing 0.05, 0.25 and 1.25 mg/l of NPS were prepared by liquid-liquid extraction with butyl chloride at basic pH and 5 µl was injected to the GC-system. All primary and secondary amines were acylated with trifluoroacetic anhydride in order to improve their analysis in GC. Amphetamine, MDMA andmethylenedioxypyrovalerone (MDPV) were used as external calibrants to quantify primary amines, secondary and tertiary amines, respectively.

Results

A data independent acquisition method was developed for screening. Tentative identification was successful in GC-APCI-QTOF using an in-house NPS database gathered previously with electrospray ionization (ESI) LC-QTOF instrument. Even the acylated compounds produced the same qualifiers ions in similar ion ratios as those that were measured without acylation in LC-QTOF.

The precision of the NCD quantification was more better than 30% for most compounds. The limit of quantification blood was 0.05 mg/l.

Conclusions & Discussion

The developed GC-NCD-QTOF platform is a dual detector system that is optimized for simultaneous identification and quantification of multiple drugs without using reference standards. Identification parameters can be matched with more commonly used LC-ESI by choosing an APCI as an ionization source, which consequently improves data transferability in the forensic collaboration work. The quantification without reference standards by GC-NCD was proven to be comparable conventional mass spectrometric quantification that uses reference standards.

In addition, both identification and quantification can be done in retrospect in cases where the structure of NPS has been initially escaped the screening. Our next phase is to apply the developed method in clinical studies related to NPS abuse in selected Finnish population.