E THYLGLUCURONIDE (E T G) ASSESSED BY TWO METHODS FOLLOWING FRAGRANCE AND HAND SANITIZER EXPOSURE IN MEN AND WOMEN Mollie Starkie, Pharm.D. Candidate Mercer University College of Pharmacy and Health Sciences

BACKGROUND

EtG analysis Enzyme multiplied immunoassay technique (EMIT) Semi-quantitative laboratory screen for urinary EtG Liquid chromatography-mass spectrometry-mass spectrometry (LC/MS/MS) Confirmatory test for urinary EtG Positive EtG concentrations: range of ng/mL

BACKGROUND Cologne and perfume in US contains 78-83% alcohol by volume. Hand sanitizer can contain 62% alcohol by volume. Exposure to such products would increase ethanol exposure.

OBJECTIVES Primary objective: Evaluate whether positive urine EtG results were detected following: 1) fragrance exposure alone 2) combined fragrance plus hand sanitizer exposure in men and women when evaluated with a screen (EMIT analysis) and confirmatory test (LC/MS/MS analysis) Secondary objectives: Comparison of EMIT analysis to LC/MS/MS analysis Detection of gender differences between the two analyses

HYPOTHESIS Combined fragrance (cologne or perfume) and hand sanitizer exposure is more likely to produce positive EtG screen and confirmation results than the fragrance exposure alone.

METHODS

Days 1-7: Abstinence from alcohol consumption & use of cologne, perfume, or hand sanitizer Day 8: Begin BID fragrance application behind ear Begin collection of first morning urine void Day 13: Hand sanitizer applied every 15 minutes for 8 hours Urine samples collected at end of 8 hour period & 4 hours later Day 18: End fragrance application Day 21: End collection of urine

Urine Sample Tube A Refrigerated EMIT analysis every 3-4 days Tube B Frozen LC/MS/MS analysis at the end of study URINE SAMPLE DIVISION

DEFINITIONS POSITIVE RESULT : EtG concentration > 100 ng/mL FALSE POSITIVE : EMIT positive screen result not confirmed by LC/MS/MS confirmation analysis FALSE NEGATIVE : Negative EMIT screen but positive LC/MS/MS confirmation result

STATISTICS Mixed model repeated measures Assess difference between male and female positive results Mixed model Chi Square Comparison of probability of having EtG concentrations >100 ng/mL by gender Pearson r coefficient and Kappa agreement Evaluated correlation and association of EMIT and LC/MS/MS analyses

RESULTS

4 males & 4 females provided 128 urine samples Range for EtG values: EMIT: ng/mL LC/MS/MS: ng/mL Mean scores of EMIT and LC/MS/MS EtG positive = > 500 ng/mL, 100% agreement EtG positive = >100 ng/mL, 37% agreement (r = 0.42, p<0.001) Females less likely to test EMIT positive

RESULTS * On the morning of hand sanitizer application, 1 female subject provided first morning sample after hand sanitizer exposure.

CONCLUSIONS Study participants abstained from alcohol beverages during the study. Positive EtG concentrations > 100 ng/mL were found following exposure of fragrance alone and fragrance and hand sanitizer when evaluated with both analyses. False positive results occurred more frequently in males. In laboratory screens the biomarker EtG detects ethanol exposure from sources other than oral alcoholic intake and should not be used as a sole method to determine covert alcohol intake with positive levels defined as > 100 ng/mL. Further research should be done to determine if false positives and false negatives results would occur with concentrations > 500 ng/mL.

REFERENCES 1. Bean P. State of the art contemporary biomarkers of alcohol consumption. MLO Med Lab Obs. 2005;37:10-2, 14, 16-7; quiz Wurst FM, Vogel R, Jachau K, et al. Ethyl glucuronide discloses recent covert alcohol use not detected by standard testing in forensic psychiatric inpatients. Alcohol Clin Exp Res. 2003;27: Wurst FM, Metzger J, WHO/ISBRA Study on State and Trait Markers ofAlcohol Use and Dependence Investigators. The ethanol conjugate ethyl glucuronide is a useful marker of recent alcohol consumption. Alcohol Clin Exp Res. 2002;26: Wurst FM, Kempter C, Seidl S, Alt A. Ethyl glucuronide--a marker of alcohol consumption and a relapse marker with clinical and forensic implications. Alcohol Alcohol. 1999;34: Schmitt G, Droenner P, Skopp G, Aderjan R. Ethyl glucuronide concentration in serum of human volunteers, teetotalers, and suspected drinking drivers. J Forensic Sci. 1997;42: Scott-Ham M, Burton FC. A study of blood and urine alcohol concentrations in cases of alleged drug-facilitated sexual assault in the united kingdom over a 3-year period. J Clin Forensic Med. 2006;13: Skipper GE, Weinmann W, Thierauf A, et al. Ethyl glucuronide: A biomarker to identify alcohol use by health professionals recovering from substance use disorders. Alcohol Alcohol. 2004;39: Dahl H, Stephanson N, Beck O, Helander A. Comparison of urinary excretion characteristics of ethanol and ethyl glucuronide. J Anal Toxicol. 2002;26:

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