Skip to main content

Advertisement

SpringerLink
Log in

Determination of ethyl glucuronide and ethyl sulfate from dried blood spots

  • Original Article
  • Published:
International Journal of Legal Medicine Aims and scope Submit manuscript

Abstract

Background

Ethyl glucuronide (EtG) and ethyl sulfate (EtS) are non-oxidative minor metabolites of ethanol. They are detectable in various body fluids shortly after initial consumption of ethanol and have a longer detection time frame than the parent compound. They are regarded highly sensitive and specific markers of recent alcohol uptake. This study evaluates the determination of EtG and EtS from dried blood spots (DBS), a simple and cost-effective sampling method that would shorten the time gap between offense and blood sampling and lead to a better reflectance of the actual impairment.

Methods

For method validation, EtG and EtS standard and quality control samples were prepared in fresh human heparinized blood and spotted on DBS cards, then extracted and measured by an LC-ESI-MS/MS method. Additionally, 76 heparinized blood samples from traffic offense cases were analyzed for EtG and EtS as whole blood and as DBS specimens. The results from these measurements were then compared by calculating the respective mean values, by a matched-paired t test, by a Wilcoxon test, and by Bland–Altman and Mountain plots.

Results and discussion

Calibrations for EtG and EtS in DBS were linear over the studied calibration range. The precision and accuracy of the method met the requirements of the validation guidelines that were employed in the study. The stability of the biomarkers stored as DBS was demonstrated under different storage conditions. The t test showed no significant difference between whole blood and DBS in the determination of EtG and EtS. In addition, the Bland–Altman analysis and Mountain plot confirmed that the concentration differences that were measured in DBS specimens were not relevant.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  1. Schmitt G, Aderjan R, Keller T, Wu M (1995) Ethyl glucuronide: an unusual ethanol metabolite in humans. Synthesis, analytical data, and determination in serum and urine. J Anal Toxicol 19(2):91–94

    Article  PubMed  CAS  Google Scholar 

  2. Skipper GE, Weinmann W, Thierauf A, Schaefer P, Wiesbeck G, Allen JP, Miller M, Wurst FM (2004) Ethyl glucuronide: a biomarker to identify alcohol use by health professionals recovering from substance use disorders. Alcohol Alcohol 39(5):445–449

    Article  PubMed  CAS  Google Scholar 

  3. Wurst FM, Skipper GE, Weinmann W (2003) Ethyl glucuronide—the direct ethanol metabolite on the threshold from science to routine use. Addiction 98(Suppl 2):51–61

    Article  PubMed  Google Scholar 

  4. Litten RZ, Bradley AM, Moss HB (2010) Alcohol biomarkers in applied settings: recent advances and future research opportunities. Alcohol Clin Exp Res 34(6):955–967

    Article  PubMed  CAS  Google Scholar 

  5. Substance Abuse and Mental Health Services Administration (SAMHSA) (2012) The role of biomarkers in the treatment of alcohol use disorders. SAMHSA Advisory 11 (2)

  6. Schmitt G, Droenner P, Skopp G, Aderjan R (1997) Ethyl glucuronide concentration in serum of human volunteers, teetotalers, and suspected drinking drivers. J Forensic Sci 42(6):1099–1102

    PubMed  CAS  Google Scholar 

  7. Halter CC, Dresen S, Auwaerter V, Wurst FM, Weinmann W (2008) Kinetics in serum and urinary excretion of ethyl sulfate and ethyl glucuronide after medium dose ethanol intake. Int J Legal Med 122(2):123–128

    Article  PubMed  Google Scholar 

  8. Hoiseth G, Bernard JP, Karinen R, Johnsen L, Helander A, Christophersen AS, Morland J (2007) A pharmacokinetic study of ethyl glucuronide in blood and urine: applications to forensic toxicology. Forensic Sci Int 172(2–3):119–124

    Article  PubMed  Google Scholar 

  9. Helander A, Bottcher M, Fehr C, Dahmen N, Beck O (2009) Detection times for urinary ethyl glucuronide and ethyl sulfate in heavy drinkers during alcohol detoxification. Alcohol Alcohol 44(1):55–61

    Article  PubMed  CAS  Google Scholar 

  10. Pragst F, Balikova MA (2006) State of the art in hair analysis for detection of drug and alcohol abuse. Clin Chim Acta 370(1–2):17–49

    Article  PubMed  CAS  Google Scholar 

  11. Pragst F, Rothe M, Moench B, Hastedt M, Herre S, Simmert D (2010) Combined use of fatty acid ethyl esters and ethyl glucuronide in hair for diagnosis of alcohol abuse: interpretation and advantages. Forensic Sci Int 196(1–3):101–110

    Article  PubMed  CAS  Google Scholar 

  12. Morini L, Colucci M, Ruberto MG, Groppi A (2012) Determination of ethyl glucuronide in nails by liquid chromatography tandem mass spectrometry as a potential new biomarker for chronic alcohol abuse and binge drinking behavior. Anal Bioanal Chem 402(5):1865–1870

    Article  PubMed  CAS  Google Scholar 

  13. Guthrie R, Susi A (1963) A simple phenylalanine method for detecting phenylketonuria in large populations of newborn infants. Pediatrics 32:338–343

    PubMed  CAS  Google Scholar 

  14. De Jesus VR, Mei JV, Bell CJ, Hannon WH (2010) Improving and assuring newborn screening laboratory quality worldwide: 30-year experience at the Centers for Disease Control and Prevention. Semin Perinatol 34(2):125–133

    Article  PubMed  Google Scholar 

  15. Edelbroek PM, van der Heijden J, Stolk LM (2009) Dried blood spot methods in therapeutic drug monitoring: methods, assays, and pitfalls. Ther Drug Monit 31(3):327–336

    Article  PubMed  Google Scholar 

  16. van der Heijden J, de Beer Y, Hoogtanders K, Christiaans M, de Jong GJ, Neef C, Stolk L (2009) Therapeutic drug monitoring of everolimus using the dried blood spot method in combination with liquid chromatography–mass spectrometry. J Pharm Biomed Anal 50(4):664–670

    Article  PubMed  Google Scholar 

  17. Garcia Boy R, Henseler J, Mattern R, Skopp G (2008) Determination of morphine and 6-acetylmorphine in blood with use of dried blood spots. Ther Drug Monit 30(6):733–739

    Article  PubMed  CAS  Google Scholar 

  18. Jantos R, Veldstra JL, Mattern R, Brookhuis KA, Skopp G (2011) Analysis of 3,4-methylenedioxymetamphetamine: whole blood versus dried blood spots. J Anal Toxicol 35(5):269–273

    Article  PubMed  CAS  Google Scholar 

  19. Ingels AS, De Paepe P, Anseeuw K, Van Sassenbroeck D, Neels H, Lambert W, Stove C (2011) Dried blood spot punches for confirmation of suspected gamma-hydroxybutyric acid intoxications: validation of an optimized GC-MS procedure. Bioanalysis 3(20):2271–2281

    Article  PubMed  CAS  Google Scholar 

  20. Mei JV, Alexander JR, Adam BW, Hannon WH (2001) Use of filter paper for the collection and analysis of human whole blood specimens. J Nutr 131(5):1631S–1636S

    PubMed  CAS  Google Scholar 

  21. McDade TW, Williams S, Snodgrass JJ (2007) What a drop can do: dried blood spots as a minimally invasive method for integrating biomarkers into population-based research. Demography 44(4):899–925

    Article  PubMed  Google Scholar 

  22. Li W, Tse FL (2010) Dried blood spot sampling in combination with LC-MS/MS for quantitative analysis of small molecules. Biomed Chromatogr 24(1):49–65

    Article  PubMed  Google Scholar 

  23. Kissinger PT (2011) Thinking about dried blood spots for pharmacokinetic assays and therapeutic drug monitoring. Bioanalysis 3(20):2263–2266

    Article  PubMed  CAS  Google Scholar 

  24. O'Broin SD, Kelleher BP, Gunter E (1995) Evaluation of factors influencing precision in the analysis of samples taken from blood spots on filter paper. Clin Lab Haematol 17(2):185–188

    PubMed  Google Scholar 

  25. Holub M, Tuschl K, Ratschmann R, Strnadova KA, Muhl A, Heinze G, Sperl W, Bodamer OA (2006) Influence of hematocrit and localisation of punch in dried blood spots on levels of amino acids and acylcarnitines measured by tandem mass spectrometry. Clin Chim Acta 373(1–2):27–31

    Article  PubMed  CAS  Google Scholar 

  26. Abu-Rabie P, Denniff P, Spooner N, Brynjolffssen J, Galluzzo P, Sanders G (2011) Method of applying internal standard to dried matrix spot samples for use in quantitative bioanalysis. Anal Chem 83(22):8779–8786

    Article  PubMed  CAS  Google Scholar 

  27. Jones J, Jones M, Plate C, Lewis D (2011) The detection of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanol in human dried blood spots. Analytical Methods 3(5):1101–1106

    Article  CAS  Google Scholar 

  28. Faller A, Richter B, Kluge M, Koenig P, Seitz HK, Thierauf A, Gnann H, Winkler M, Mattern R, Skopp G (2011) LC-MS/MS analysis of phosphatidylethanol in dried blood spots versus conventional blood specimens. Anal Bioanal Chem 401(4):1163–1166

    Article  PubMed  CAS  Google Scholar 

  29. Hernandez Redondo A, Korber C, Konig S, Langin A, Al-Ahmad A, Weinmann W (2012) Inhibition of bacterial degradation of EtG by collection as dried urine spots (DUS). Anal Bioanal Chem 402(7):2417–2424

    Article  CAS  Google Scholar 

  30. German Society of Toxicological and Forensic Chemistry (GTFCh) (2009) Anforderungen an die Validierung von Analysenmethoden. T + K 76:185–208

  31. Dresen S, Weinmann W, Wurst FM (2004) Forensic confirmatory analysis of ethyl sulfate—a new marker for alcohol consumption—by liquid-chromatography/electrospray ionization/tandem mass spectrometry. J Am Soc Mass Spectrom 15(11):1644–1648

    Article  PubMed  CAS  Google Scholar 

  32. Bland JM, Altman DG (1986) Statistical methods for assessing agreement between two methods of clinical measurement. Lancet 1(8476):307–310

    Article  PubMed  CAS  Google Scholar 

  33. Mantha S, Roizen MF, Fleisher LA, Thisted R, Foss J (2000) Comparing methods of clinical measurement: reporting standards for Bland and Altman analysis. Anesth Analg 90(3):593–602

    Article  PubMed  CAS  Google Scholar 

  34. Krouwer JS, Monti KL (1995) A simple, graphical method to evaluate laboratory assays. Eur J Clin Chem Clin Biochem 33(8):525–527

    PubMed  CAS  Google Scholar 

  35. Ingels AS, Lambert WE, Stove CP (2010) Determination of gamma-hydroxybutyric acid in dried blood spots using a simple GC-MS method with direct “on spot” derivatization. Anal Bioanal Chem 398(5):2173–2182

    Article  PubMed  CAS  Google Scholar 

  36. Suyagh MF, Iheagwaram G, Kole PL, Millership J, Collier P, Halliday H, McElnay JC (2010) Development and validation of a dried blood spot-HPLC assay for the determination of metronidazole in neonatal whole blood samples. Anal Bioanal Chem 397(2):687–693

    Article  PubMed  CAS  Google Scholar 

  37. Keevil BG (2011) The analysis of dried blood spot samples using liquid chromatography tandem mass spectrometry. Clin Biochem 44(1):110–118

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The authors would like to thank Korbinian Seitz (University of Bern) for language editing and the Swiss National Foundation for financial support.

Author information

Authors and Affiliations

  1. Institute of Forensic Medicine, University of Bern, Bern, Switzerland

    Ana Hernández Redondo, Alexandra Schroeck, Beat Kneubuehl & Wolfgang Weinmann

  2. Institut für Rechtsmedizin, Medizinische Fakultät, Universität Bern, Bühlstrasse 20, 3012, Bern, Switzerland

    Ana Hernández Redondo, Alexandra Schroeck, Beat Kneubuehl & Wolfgang Weinmann

Authors
  1. Ana Hernández Redondo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Alexandra Schroeck
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Beat Kneubuehl
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Wolfgang Weinmann
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Wolfgang Weinmann.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hernández Redondo, A., Schroeck, A., Kneubuehl, B. et al. Determination of ethyl glucuronide and ethyl sulfate from dried blood spots. Int J Legal Med 127, 769–775 (2013). https://doi.org/10.1007/s00414-012-0815-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00414-012-0815-2

Keywords

Search

Navigation