Abstract
Ethyl glucuronide (EtG) is a metabolite and a specific marker of alcohol consumption that can be detected days after the complete elimination of alcohol after drinking. A rapid, simple, and sensitive LC-ESI-MS/MS method for the determination of urinary ethyl glucuronide was developed and fully validated in accordance with analytical standards, using the C18 column. The whole process including sample preparation and LC-MS/MS lasted 10 min. A comprehensive validation including HorRat, measurement uncertainty, system suitability and intermediate precision calculations among analysts, and a cut-off limit study was performed. The method was applied to real samples and a cutoff limit determination study. The LOD and LOQ (using the IUPAC and Eurachem methods) were determined as 104.21 ng mL−1 and 165.00 ng mL−1. A cut-off limit of ≈ 818 ng mg−1 (normalised to creatinine) was found for urinary EtG. The results showed that the cut-off limits currently in use should be re-considered in further studies and standardised on a global scale. Normalisation to creatinine is important because of the risk of the dilution of urine intentionally or with a change of diet. The concentrations of real samples from subjects who had consumed alcohol were successfully predicted using this method, after zero HS-GC/MS results of urine alcohol concentration.
References
Albermann, M. E., Musshoff, F., & Madea, B. A. (2012). A highperformance liquid chromatographic-tandem mass spectrometric method for the determination of ethyl glucuronide and ethyl sulfate in urine validated according to forensic guidelines. Journal of Chromatographic Science, 50, 51-56. DOI: 10.1093/chromsci/bmr012.10.1093/chromsci/bmr012Suche in Google Scholar
AOAC International (2012). Guidelines for standard method performance requirements (AOAC Official methods of analysis, Appendix F, pp. 16). Rockville, MD, USA: AOAC International.Suche in Google Scholar
Retrieved June 2, 2015, from http://www.eoma.aoac.org/appf.pdf Suche in Google Scholar
Bergström, J., Helander, A., & Jones, A. W. (2003). Ethyl glucuronide concentrations in two successive urinary voids from drinking drivers: relationship to creatinine content and blood and urine ethanol concentrations. Forensic Science International, 133, 86-94. DOI: 10.1016/s0379-0738(03)00053-7.10.1016/S0379-0738(03)00053-7Suche in Google Scholar
Beyer, J., Vo, T. N., Gerostamoulos, D., & Drummer, O. H. (2011). Validated method for the determination of ethlyglucuronide and ethylsulfate in human urine. Analytical and Bioanalytical Chemistry, 400, 189-196. DOI: 10.1007/s00216-011-4667-2.10.1007/s00216-011-4667-2Suche in Google Scholar
Bicker, W., Lämmerhofer, M., Keller, T., Schuhmacher, R., Krska, R., & Lindner, W. (2006). Validated method for the determination of the ethanol consumption markers ethyl glucuronide, ethyl phosphate, and ethyl sulfate in human urine by reversed-phase/weak anion exchange liquid chromatography-tandem mass spectrometry. Analytical Chemistry, 78, 5884-5892. DOI: 10.1021/ac060680+.10.1021/ac060680+Suche in Google Scholar
Costantino, A., DiGregorio, E. J., Korn, W., Spayd, S., & Rieders, F. (2006). The effect of the use of mouthwash on ethylglucuronide concentrations in urine. Journal of Analytical Toxicology, 30, 659-662. DOI: 10.1093/jat/30.9.659.10.1093/jat/30.9.659Suche in Google Scholar
Cross, A. J., Major, J. M., & Sinha, R. (2011). Urinary biomarkers of meat consumption. Cancer Epidemiology, Biomarkers & Prevention, 20, 1107-1111. DOI: 10.1158/1055-9965.epi-11-0048.10.1158/1055-9965.EPI-11-0048Suche in Google Scholar
Eurachem (1998). The fitness for purpose of analytical methods: A laboratory guide to method validation and related topics (Eurachem guide). Leoben, Austria: Eurachem. Retrieved June 2, 2015, from http://eurachem.org/images/stories/ Guides/pdf/valid.pdf Suche in Google Scholar
Favretto, D., Nalesso, A., Frison, G., Viel, G., Traldi, P., & Ferrara, S. D. (2010). A novel and an effective analytical approach for the LC-MS determination of ethyl glucuronide and ethyl sulfate in urine. International Journal of Legal Medicine, 124, 161-164. DOI: 10.1007/s00414-009-0376-1.10.1007/s00414-009-0376-1Suche in Google Scholar
Ghante, M. R., Pannu, H. K., Loni, A., & Shivsharan, T. (2012). Development and validation of a RP- HPLC method for simultaneous estimation of metronidazole and norfloxacin in bulk and tablet dosage form. International Journal of Pharmacy and Pharmaceutical Sciences, 4(Suppl4), 241-245.Suche in Google Scholar
Hegstad, S., Helland, A., Hagemann, C., Michelsen, L., & Spigset, O. (2013). EtG/EtS in urine from sexual assault victims determined by UPLC-MS-MS. Journal of Analytical Toxicology, 37, 227-232. DOI: 10.1093/jat/bkt008.10.1093/jat/bkt008Suche in Google Scholar
Janda, I., & Alt, J. (2001). Improvement of ethyl glucuronide determination in human urine and serum samples by solidphase extraction. Journal of Chromatography B, 758, 229-234. DOI: 10.1016/s0378-4347(01)00186-4.10.1016/S0378-4347(01)00186-4Suche in Google Scholar
Jurado, C., Soriano, T., Giménez, M. P., & Menéndez, M. (2004). Diagnosis of chronic alcohol consumption: Hair analysis for ethyl-glucuronide. Forensic Science International, 145, 161-166. DOI: 10.1016/j.forsciint.2004.04.031.10.1016/j.forsciint.2004.04.031Suche in Google Scholar PubMed
Keten, A., Tumer, A. R., & Balseven-Odabasi, A. (2009). Measurement of ethyl glucuronide in vitreous humor with liquid chromatography-mass spectrometry. Forensic Science International, 193, 101-105. DOI: 10.1016/j.forsciint.2009.09.018.10.1016/j.forsciint.2009.09.018Suche in Google Scholar PubMed
Kummer, N., Wille, S., Di Fazio, V., Lambert, W., & Samyn, N. (2013). A fully validated method for the quantification of ethyl glucuronide and ethyl sulphate in urine by UPLC- ESI-MS/MS applied in a prospective alcohol self-monitoring study. Journal of Chromatography B, 929, 149-154. DOI: 10.1016/j.jchromb.2013.04.011.10.1016/j.jchromb.2013.04.011Suche in Google Scholar PubMed
Lykken, G. I., Jacob, R. A., Munoz, J. M., & Sandstead, H. H. (1980). A mathematical model of creatine metabolism in normal males-comparison between theory and experiment. The American Journal of Clinical Nutrition, 33, 2674-2685.10.1093/ajcn/33.12.2674Suche in Google Scholar PubMed
Mocak, J., Bond, A. M., Mitchell, S., & Scollary, G. (1997). A statistical overview of standard (IUPAC and ACS) and new procedures for determining the limits of detection and quantification: Application to voltammetric and stripping techniques (Technical Report), Pure and Applied Chemistry, 69, 297-328. DOI: 10.1351/pac199769020297.10.1351/pac199769020297Suche in Google Scholar
Morini, L., Politi, L., Zucchella, A., & Polettini, A. (2007). Ethyl glucuronide and ethyl sulphate determination in serum by liquid chromatography-electrospray tandem mass spectrometry. Clinica Chimica Acta, 376, 213-219. DOI: 10.1016/j.cca.2006.08.024.10.1016/j.cca.2006.08.024Suche in Google Scholar PubMed
Musshoff, F., Albermann, E., & Madea, B. (2010). Ethyl glucuronide and ethyl sulfate in urine after consumption of various beverages and foods-misleading results? International Journal of Legal Medicine, 124, 623-630. DOI: 10.1007/s00414-010-0511-z.10.1007/s00414-010-0511-zSuche in Google Scholar PubMed
Politi, L., Morini, L., Groppi, A., Poloni, V., Pozzi, F., & Polettini, A. (2005). Direct determination of the ethanol metabolites ethyl glucuronide and ethyl sulfate in urine by liquid chromatography/electrospray tandem mass spectrometry. Rapid Communications in Mass Spectrometry, 19, 1321-1331. DOI: 10.1002/rcm.1932.10.1002/rcm.1932Suche in Google Scholar PubMed
Robinson, T. M., Sewell, D. A., Casey, A., Steenge, G., & Greenhaff, P. L. (2000). Dietary creatine supplementation does not affect some haematological indices, or indices of muscle damage and hepatic and renal function. British Journal of Sports Medicine, 34, 284-288. DOI: 10.1136/bjsm.34.4.284.10.1136/bjsm.34.4.284Suche in Google Scholar PubMed PubMed Central
Schloegl, H., Rost, T., Schmidt,W.,Wurst, F. M., &Weinmann, W. (2006). Distribution of ethyl glucuronide in rib bone marrow, other tissues and body liquids as proof of alcohol consumption before death, Forensic Science International, 156, 213-218. DOI: 10.1016/j.forsciint.2005.03.024.10.1016/j.forsciint.2005.03.024Suche in Google Scholar PubMed
Skipper, G. E.,Weinmann, W., Thierauf, A., Schaefer, P.,Wiesbeck, G. Allen, J. P., Miller, M., & Wurst, F. M. (2004). Ethyl glucuronide: a biomarker to identify alcohol use by health professionals recovering from substance use disorders. Alcohol and Alcoholism, 39, 445-449. DOI: 10.1093/alcalc/ agh078.Suche in Google Scholar
Stephanson, N., Dahl, H., Helander, A., & Beck, O. (2002). Direct quantification of ethyl glucuronide in clinical urine samples by liquid chromatography-mass spectrometry. Therapeutic Drug Monitoring, 24, 645-651.10.1097/00007691-200210000-00011Suche in Google Scholar PubMed
Thierauf, A., Serr, A., Halter, C. C., Al-Ahmad, A., Rana, S., & Weinmann, W. (2008). Influence of preservatives on the stability of ethyl glucuronide and ethyl sulphate in urine. Forensic Science International, 182, 41-45. DOI: 10.1016/j.forsciint.2008.09.011.10.1016/j.forsciint.2008.09.011Suche in Google Scholar PubMed
Thierauf, A., Halter, C. C., Rana, S., Auwaerter, V., Wohlfarth, A., Wurst, F. M., & Weinmann, W. (2009). Urine tested positive for ethyl glucuronide after trace amounts of ethanol. Addiction, 104, 2007-2012. DOI: 10.1111/j.1360-0443.2009.02722.x.10.1111/j.1360-0443.2009.02722.xSuche in Google Scholar PubMed
Thierauf, A., Wohlfarth, A., Auwärter, V., Große Perdekamp, M., Wurst, F. M., & Weinmann, W. (2010). Urine tested positive for ethyl glucuronide and ethyl sulfate after the consumption of yeast and sugar. Forensic Science International, 202, e45-e47. DOI: 10.1016/j.forsciint.2010.06.028.10.1016/j.forsciint.2010.06.028Suche in Google Scholar PubMed
Weinmann, W., Schaefer, P., Thierauf, A., Schreiber, A., & Wurst, F. M. (2004). Confirmatory analysis of ethylglucuronide in urine by liquid-chromatography/electrospray ionization/ tandem mass spectrometry according to forensic guidelines. Journal of the American Society for Mass Spectrometry, 15, 188-193. DOI: 10.1016/j.jasms.2003.10.010.10.1016/j.jasms.2003.10.010Suche in Google Scholar PubMed
Wurst, F. M., Kempter, C., Metzger, J., Seidl, S., & Alt, A. (2000). Ethyl glucuronide: a marker of recent alcohol consumption with clinical and forensic implications. Alcohol, 20, 111-116. DOI: 10.1016/s0741-8329(99)00076-2. Wurst, F. M., Wiesbeck, G. A., Metzger, J. W., Weinmann, W., & Graf, M. (2004). On sensitivity, specificity, and the influence of various parameters on ethyl glucuronide levels in urine-results from the WHO/ISBRA study. Alcoholism: Clinical & Experimental Research, 28, 1220-1228. DOI: 10.1097/01.alc.0000134230.21414.11 10.1097/01.ALC.0000134230.21414.11Suche in Google Scholar
Institute of Chemistry, Slovak Academy of Sciences
Artikel in diesem Heft
- Catalysis in glycerol: a survey of recent advances
- A rapid LC-MS/MS method for determination of urinary EtG and application to a cut-off limit study
- Validated chiral chromatographic methods for clopidogrel bisulphate and its related substances in bulk drug and pharmaceutical dosage forms
- Effect of SO2 on SCR activity of MnOx/PG catalysts at low temperature
- Preparation of Pd/Al2O3@silicalite-1 core–shell beads and their application to hydrogenation reactions
- Effect of rapid batch decompression on hydrolysate quality after hydrothermal pretreatment of wheat straw
- Anthocyanins profile, total phenolics and antioxidant activity of two Romanian red grape varieties: Feteascǎ neagrǎ and Bǎbeascǎ neagrǎ (Vitis vinifera)
- Polyphenols, radical scavenger activity, short-chain organic acids and heavy metals of several plants extracts from “Bucharest Delta”
- Reaction of anhydrous zinc chloride with 2,3-thiophenedicarbaldehyde bis(semicarbazone) (2,3BSTCH2) and bis(thiosemicarbazone) (2,3BTSTCH2): Crystal structure of {[C6H5N2S]+[ZnCl3(C6H4N2S)]−} complex
- Synthesis and evaluation of a novel hydrophobically associating polymer based on acrylamide for enhanced oil recovery
- Synthesis of cardanol-based photo-active SET-LRP initiator and its application to preparation of UV-cured resin
- Preparation, characterization and ion adsorption properties of functionalized polystyrene modified with 1,4-phenylene diisocyanate and diethylenetriamine
- Synthesis of lanthanide-based SBA-15 mesoporous hybrids by a novel route
- Comparative ESI FT-MS and MALDI-TOF structural analyses of representative human N-linked glycans
Artikel in diesem Heft
- Catalysis in glycerol: a survey of recent advances
- A rapid LC-MS/MS method for determination of urinary EtG and application to a cut-off limit study
- Validated chiral chromatographic methods for clopidogrel bisulphate and its related substances in bulk drug and pharmaceutical dosage forms
- Effect of SO2 on SCR activity of MnOx/PG catalysts at low temperature
- Preparation of Pd/Al2O3@silicalite-1 core–shell beads and their application to hydrogenation reactions
- Effect of rapid batch decompression on hydrolysate quality after hydrothermal pretreatment of wheat straw
- Anthocyanins profile, total phenolics and antioxidant activity of two Romanian red grape varieties: Feteascǎ neagrǎ and Bǎbeascǎ neagrǎ (Vitis vinifera)
- Polyphenols, radical scavenger activity, short-chain organic acids and heavy metals of several plants extracts from “Bucharest Delta”
- Reaction of anhydrous zinc chloride with 2,3-thiophenedicarbaldehyde bis(semicarbazone) (2,3BSTCH2) and bis(thiosemicarbazone) (2,3BTSTCH2): Crystal structure of {[C6H5N2S]+[ZnCl3(C6H4N2S)]−} complex
- Synthesis and evaluation of a novel hydrophobically associating polymer based on acrylamide for enhanced oil recovery
- Synthesis of cardanol-based photo-active SET-LRP initiator and its application to preparation of UV-cured resin
- Preparation, characterization and ion adsorption properties of functionalized polystyrene modified with 1,4-phenylene diisocyanate and diethylenetriamine
- Synthesis of lanthanide-based SBA-15 mesoporous hybrids by a novel route
- Comparative ESI FT-MS and MALDI-TOF structural analyses of representative human N-linked glycans