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Salting-out-assisted liquid–liquid extraction as a suitable approach for determination of methoxetamine in large sets of tissue samples

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Abstract

A new designer drug, a dissociative anesthetic, and a putative N-methyl-d-aspartate receptor antagonist, methoxetamine (MXE) noted by the EU Early Warning System has been already identified as a cause of several fatalities worldwide. The primary objective of this work was to develop a suitable sample preparation method allowing for isolation of MXE and its main metabolites in high yields from rat brain, liver, and lungs. For the purpose of the project, MXE and five metabolites were synthesized in-house, specifically O-desmethyl-normethoxetamine, O-desmethylmethoxetamine, dihydro-O-desmethylmethoxetamine, normethoxetamine, and dihydromethoxetamine. A sample preparation procedure consisted in the homogenization of the tissue applying salting-out-assisted liquid–liquid extraction (SALLE). A subsequent liquid chromatography-mass spectrometry (LC-MS) analysis was based on reversed-phased chromatography hyphenated with a triple quad MS system in a positive electrospray mode. Multiple reaction monitoring (MRM) was used for qualification and quantification of the analytes. The quantification was based on the application of an isotopically labeled internal standard, normethoxetamine-d3. The matrix-matched calibrations were prepared for each type of matrix with regression coefficients 0.9943–1.0000. The calibration curves were linear in the concentration range of 2.5–250 ng g−1. Limits of quantification (LOQs) were estimated as 2.5 and 5 ng g−1, respectively. Recovery (80–117 %) and matrix effect (94–110 %) at 100 ng g−1 and intra- and inter-day accuracy and precision at low (2.5 ng g−1), middle (25 ng g−1), and upper (250 ng g−1) concentration levels for all the analytes in all three types of tissues were also determined. The developed analytical method was applied to a set of real samples gathered in toxicological trials on rats and MXE, and its metabolites were determined successfully.

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Acknowledgments

This study was funded by a specific university research grant (project MSMT No 20/2015) and by the Ministry of Interior of the Czech Republic (project VG20122015075, New synthetic drugs (NSD)—creation of toxicological database, development and validation of detection methods including fast immunochemical tests, behavioral pharmacology, pharmacokinetics and biotransformation in rats, epidemiology).

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Authors and Affiliations

  1. Forensic Laboratory of Biologically Active Substances, University of Chemistry and Technology Prague, Technická 5, 166 28, Prague 6, Dejvice, Czech Republic

    Katerina Hajkova, Bronislav Jurasek, Petra Miksatkova & Martin Kuchar

  2. Department of Analytical Chemistry, University of Chemistry and Technology Prague, Technická 5, 166 28, Prague 6, Dejvice, Czech Republic

    Katerina Hajkova & David Sykora

  3. National Institute of Mental Health, Topolová 748, 250 67, Klecany, Czech Republic

    Katerina Hajkova, Tomas Palenicek, Petra Miksatkova & Martin Kuchar

Authors
  1. Katerina Hajkova
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  2. Bronislav Jurasek
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  3. David Sykora
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  4. Tomas Palenicek
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  5. Petra Miksatkova
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  6. Martin Kuchar
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Correspondence to David Sykora.

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Hajkova, K., Jurasek, B., Sykora, D. et al. Salting-out-assisted liquid–liquid extraction as a suitable approach for determination of methoxetamine in large sets of tissue samples. Anal Bioanal Chem 408, 1171–1181 (2016). https://doi.org/10.1007/s00216-015-9221-1

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  • DOI: https://doi.org/10.1007/s00216-015-9221-1

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