Advertisement

Analytical and Bioanalytical Chemistry

, Volume 410, Issue 20, pp 4967–4978 | Cite as

Rapid determination of designer benzodiazepines, benzodiazepines, and Z-hypnotics in whole blood using parallel artificial liquid membrane extraction and UHPLC-MS/MS

  • Linda Vårdal
  • Gladys Wong
  • Åse Marit Leere Øiestad
  • Stig Pedersen-Bjergaard
  • Astrid Gjelstad
  • Elisabeth Leere Øiestad
Research Paper

Abstract

Benzodiazepines (BZD) and Z-hypnotics are frequently analyzed in forensic laboratories, and in 2012, the designer benzodiazepines (DBZD) emerged on the illegal drug scene. DBZD represent a particular challenge demanding new analytical methods. In this work, parallel artificial liquid membrane extraction (PALME) is used for sample preparation of DBZD, BZD, and Z-hypnotics in whole blood prior to UHPLC-MS/MS analysis. PALME of BZD, DBZD, and Z-hypnotics was performed from whole blood samples, and the analytes were extracted across a supported liquid membrane (SLM) and into an acceptor solution of dimethyl sulfoxide and 200 mM formic acid (75:25, v/v). The method was validated according to EMA guidelines. The method was linear throughout the calibration range (R2 > 0.99). Intra- and inter-day accuracy and precision, as well as matrix effects, were within the guideline limit of ± 15%. LOD and LLOQ ranged from 0.10 to 5.0 ng mL−1 and 3.2 to 160 ng mL−1, respectively. Extraction recoveries were reproducible and above 52%. The method was specific, and the analytes were stable in the PALME extracts for 4 and 10 days at 10 and − 20 °C. No carry-over was observed within the calibration range. PALME and UHPLC-MS/MS for the determination of DBZD, BZD, and Z-hypnotics in whole blood are a green and low-cost alternative that provides high sample throughput (96-well format), extensive sample clean-up, good sensitivity, and high reproducibility. The presented method is also the first method incorporating analysis of DBZD, BZD, and Z-hypnotics in whole blood in one efficient analysis.

Graphical abstract

Keywords

Designer benzodiazepines Benzodiazepines Z-hypnotics Parallel artificial liquid membrane extraction LC-MS/MS Whole blood samples 

Notes

Acknowledgements

The Research Council of Norway is acknowledged for financial support through grant no. 231917. Gerrit Middelkoop, Elianne Seeberg, Marit Langødegaard, and Elin Eliassen are gratefully acknowledged for their help with the method comparison.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Supplementary material

216_2018_1147_MOESM1_ESM.pdf (509 kb)
ESM 1 (PDF 141 kb)

References

  1. 1.
    Persona K, Madej K, Knihnicki P, Piekoszewski W. Analytical methodologies for the determination of benzodiazepines in biological samples. J Pharm Biomed Anal. 2015;113:239–64.CrossRefPubMedGoogle Scholar
  2. 2.
    Manchester KR, Lomas EC, Waters L, Dempsey FC, Maskell PD. The emergence of new psychoactive substance (NPS) benzodiazepines: a review. Drug Test Anal. 2017Google Scholar
  3. 3.
    Bogstrand ST, Gjerde H. Which drugs are associated with highest risk for being arrested for driving under the influence? A case–control study. Forensic Sci Int. 2014;240:21–8.CrossRefPubMedGoogle Scholar
  4. 4.
    Magrini L, Cappiello A, Famiglini G, Palma P. Microextraction by packed sorbent (MEPS)-UHPLC-UV: a simple and efficient method for the determination of five benzodiazepines in an alcoholic beverage. J Pharm Biomed Anal. 2016;125:48–53.CrossRefPubMedGoogle Scholar
  5. 5.
    Švidrnoch M, Boráňová B, Tomková J, Ondra P, Maier V. Simultaneous determination of designer benzodiazepines in human serum using non-aqueous capillary electrophoresis–tandem mass spectrometry with successive multiple ionic–polymer layer coated capillary. Talanta. 2018;176:69–76.CrossRefPubMedGoogle Scholar
  6. 6.
    Moosmann B, Hutter M, Huppertz LM, Ferlaino S, Redlingshöfer L, Auwärter V. Characterization of the designer benzodiazepine pyrazolam and its detectability in human serum and urine. Forensic Toxicol. 2013;31(2):263–71.CrossRefGoogle Scholar
  7. 7.
    EMCDDA. European Drug Report 2017: Trends and Developments. 2017. http://www.emcdda.europa.eu/publications/edr/trends-developments/2017. Accessed 05.12.17.
  8. 8.
    Bergstrand MP, Helander A, Beck O. Development and application of a multi-component LC–MS/MS method for determination of designer benzodiazepines in urine. J Chromatogr B. 2016;1035:104–10.CrossRefGoogle Scholar
  9. 9.
    Høiseth G, Tuv SS, Karinen R. Blood concentrations of new designer benzodiazepines in forensic cases. Forensic Sci Int. 2016;268:35–8.CrossRefPubMedGoogle Scholar
  10. 10.
    Middelkoop G. Oslo University Hospital, personal communication 2017.Google Scholar
  11. 11.
    Sauve E, Langødegård M, Ekeberg D, Øiestad ÅM. Determination of benzodiazepines in ante-mortem and post-mortem whole blood by solid-supported liquid–liquid extraction and UPLC–MS/MS. J Chromatogr B. 2012;883:177–88.CrossRefGoogle Scholar
  12. 12.
    Valen A, Leere Øiestad ÅM, Strand DH, Skari R, Berg T. Determination of 21 drugs in oral fluid using fully automated supported liquid extraction and UHPLC-MS/MS. Drug Test Anal. 2017;9(5):808–23.CrossRefPubMedGoogle Scholar
  13. 13.
    Mullett WM, Pawliszyn J. Direct determination of benzodiazepines in biological fluids by restricted-access solid-phase microextraction. Anal Chem. 2002;74(5):1081–7.CrossRefPubMedGoogle Scholar
  14. 14.
    de Bairros AV, de Almeida RM, Pantaleão L, Barcellos T, e Silva SM, Yonamine M. Determination of low levels of benzodiazepines and their metabolites in urine by hollow-fiber liquid-phase microextraction (LPME) and gas chromatography–mass spectrometry (GC–MS). J Chromatogr B. 2015;975:24–33.CrossRefGoogle Scholar
  15. 15.
    De Boeck M, Dehaen W, Tytgat J, Cuypers E. Ionic liquid-based liquid–liquid microextraction for benzodiazepine analysis in postmortem blood samples. J Forensic Sci. 2018;Google Scholar
  16. 16.
    De Boeck M, Missotten S, Dehaen W, Tytgat J, Cuypers E. Development and validation of a fast ionic liquid-based dispersive liquid–liquid microextraction procedure combined with LC–MS/MS analysis for the quantification of benzodiazepines and benzodiazepine-like hypnotics in whole blood. Forensic Sci Int. 2017;274:44–54.CrossRefPubMedGoogle Scholar
  17. 17.
    Fisichella M, Odoardi S, Strano-Rossi S. High-throughput dispersive liquid/liquid microextraction (DLLME) method for the rapid determination of drugs of abuse, benzodiazepines and other psychotropic medications in blood samples by liquid chromatography–tandem mass spectrometry (LC-MS/MS) and application to forensic cases. Microchem J. 2015;123:33–41.CrossRefGoogle Scholar
  18. 18.
    Gjelstad A, Rasmussen KE, Parmer MP, Pedersen-Bjergaard S. Parallel artificial liquid membrane extraction: micro-scale liquid–liquid–liquid extraction in the 96-well format. Bioanalysis. 2013;5(11):1377–85.CrossRefPubMedGoogle Scholar
  19. 19.
    Ask KS, Bardakci T, Parmer MP, Halvorsen TG, Øiestad EL, Pedersen-Bjergaard S, et al. Parallel artificial liquid membrane extraction as an efficient tool for removal of phospholipids from human plasma. J Pharm Biomed Anal. 2016;129:229–36.CrossRefPubMedGoogle Scholar
  20. 20.
    Øiestad EL, Johansen U, Øiestad ÅML, Christophersen AS. Drug screening of whole blood by ultra-performance liquid chromatography-tandem mass spectrometry. J Anal Toxicol. 2011;35(5):280–93.CrossRefPubMedGoogle Scholar
  21. 21.
    Drummer OH. Requirements for bioanalytical procedures in postmortem toxicology. Anal Bioanal Chem. 2007;388(7):1495–503.CrossRefPubMedGoogle Scholar
  22. 22.
    Sanches LR, Seulin SC, Leyton V, Bismara Paranhos BAP, Pasqualucci CA, Munoz DR, et al. Determination of opiates in whole blood and vitreous humor: a study of the matrix effect and an experimental design to optimize conditions for the enzymatic hydrolysis of glucuronides. J Anal Toxicol. 2012;36(3):162–70.CrossRefPubMedGoogle Scholar
  23. 23.
    Berg T, Karlsen M, Øiestad ÅML, Johansen JE, Liu H, Strand DH. Evaluation of 13C-and 2H-labeled internal standards for the determination of amphetamines in biological samples, by reversed-phase ultra-high performance liquid chromatography–tandem mass spectrometry. J Chromatogr A. 2014;1344:83–90.CrossRefPubMedGoogle Scholar
  24. 24.
    EMA. Guideline on bioanalytical method validation. 2011. http://www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2011/08/WC500109686.pdf. Accessed 26.04.18.
  25. 25.
    GTFCH. Requirements for the validation of Anal Methods (Appendix B). 2009. https://www.gtfch.org/cms/images/stories/files/Appendix%20B%20GTFCh%2020090601.pdf. Accessed 26.04.18.
  26. 26.
    Matuszewski B, Constanzer M, Chavez-Eng C. Strategies for the assessment of matrix effect in quantitative bioanalytical methods based on HPLC−MS/MS. Anal Chem. 2003;75(13):3019–30.CrossRefPubMedGoogle Scholar
  27. 27.
    Eliassen E, Kristoffersen L. Quantitative determination of zopiclone and zolpidem in whole blood by liquid–liquid extraction and UHPLC-MS/MS. J Chromatogr B. 2014;971:72–80.CrossRefGoogle Scholar
  28. 28.
    Strand DH, Langødegård M, Gaare KI, Amundsen I, Nilsen M, Kristoffersen L. Determination of twelve commonly found compounds in DUI cases in whole blood using fully automated supported liquid extraction and UHPLC-MS/MS. SOFT-TIAFT, Boca Raton, Florida, USA2018.Google Scholar
  29. 29.
    Bland JM, Altman D. Statistical methods for assessing agreement between two methods of clinical measurement. Lancet. 1986;327(8476):307–10.CrossRefGoogle Scholar
  30. 30.
    Giavarina D. Understanding Bland Altman analysis. Biochem Med. 2015;25(2):141–51.CrossRefGoogle Scholar
  31. 31.
    Vårdal L, Askildsen H-M, Gjelstad A, Øiestad EL, Edvardsen HME, Pedersen-Bjergaard S. Parallel artificial liquid membrane extraction of new psychoactive substances in plasma and whole blood. J Chromatogr B. 2017;1048:77–84.CrossRefGoogle Scholar
  32. 32.
    Verplaetse R, Cuypers E, Tytgat J. The evaluation of the applicability of a high pH mobile phase in ultrahigh performance liquid chromatography tandem mass spectrometry analysis of benzodiazepines and benzodiazepine-like hypnotics in urine and blood. J Chromatogr A. 2012;1249:147–54.CrossRefPubMedGoogle Scholar
  33. 33.
    Simonsen KW, Hermansson S, Steentoft A, Linnet K. A validated method for simultaneous screening and quantification of twenty-three benzodiazepines and metabolites plus zopiclone and zaleplone in whole blood by liquid-liquid extraction and ultra-performance liquid chromatography-tandem mass spectrometry. J Anal Toxicol. 2010;34(6):332–41.CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Linda Vårdal
    • 1
  • Gladys Wong
    • 1
  • Åse Marit Leere Øiestad
    • 2
  • Stig Pedersen-Bjergaard
    • 1
    • 3
  • Astrid Gjelstad
    • 1
  • Elisabeth Leere Øiestad
    • 1
    • 2
  1. 1.School of PharmacyUniversity of OsloOsloNorway
  2. 2.Department of Forensic SciencesOslo University HospitalOsloNorway
  3. 3.Faculty of Health and Medical Sciences, School of Pharmaceutical SciencesUniversity of CopenhagenCopenhagenDenmark

Personalised recommendations