This study compared the urinary excretion characteristics of ethyl glucuronide (EtG) with that of ethanol, with focus on the effect of water-induced diuresis. Six healthy volunteers ingested an ethanol dose of 0.5 g/kg (range 25.0-41.5 g) as 5% (v/v) beer in 30 min and the same volume of water after 3 h. Urine collections were made before starting the experiment and at timed intervals over 31.5 h. The concentration of EtG was determined by an LC-MS method (LOQ = 0.1 mg/L). The urine samples collected immediately before starting drinking were all negative for ethanol and EtG, thus confirming that the participants had not recently ingested alcohol. Intake of beer resulted in a marked increase in excreted urine volume and a concomitant drop in creatinine concentration. The concentration of ethanol peaked at a mean value of 17 mmol/L in the 1.5-h urine collection. Except for one subject, EtG was first detectable (range 0.9-5.5 mg/L) at 1 h. Intake of water at 3 h produced another increase in urine volume and a drop in creatinine. The ethanol concentration curve was not influenced by the water diuresis, whereas this caused a distinct drop in the EtG concentration. When EtG was expressed relative to the creatinine value, this ratio was seemingly not affected by the intake of water. The ethanol concentration returned to zero at 6.5 h, whereas EtG was still detectable for up to 22.5-31.5 h, albeit at low levels in the end (< 1 mg/l). Only about 0.02% of the administered dose of ethanol (on a molar basis) was recovered in the urine as EtG. The results demonstrated that EtG remains detectable in the urine for many hours after the ethanol itself has been eliminated. Moreover, it was possible to lower the concentration of EtG by drinking large amounts of water prior to voiding, whereas this strategy did not influence the EtG/creatinine ratio or the concentration of ethanol.

After consumption of alcoholic beverages, the bulk of the ethanol dose (95%-98%) is eliminated in a 2-stage oxidation process mainly in the liver, first to acetaldehyde by alcohol dehydrogenase and then further to acetic acid by aldehyde dehydrogenase. The remainder is excreted unchanged in urine, sweat, and expired air (1 ). In addition, a very small fraction (Ͻ0.1%) of the ingested ethanol undergoes phase II conjugation reactions to produce ethyl glucuronide (EtG) and ethyl sulfate (EtS) (2, 3 ), catalyzed by uridine diphosphate-glucuronosyltransferase or sulfotransferase, respectively. EtG and EtS are eventually excreted in the urine. As both of these nonoxidative direct ethanol metabolites show much longer elimination times than ethanol itself (3 ), the interest in EtG and EtS has focused largely on their use as sensitive and specific biomarkers of recent alcohol intake with clinical and forensic applications (4,5 ). A positive finding of EtG and/or EtS provides a strong indication that the person was recently drinking alcohol, even when the ethanol concentration has returned to 0 or is no longer measurable.Glucuronide and sulfate conjugates of endogenous and exogenous origin are cleaved by ␤-glucuronidase and sulfatase, enzymes that are widely distributed among animals and plants. ␤-Glucuronidase is also present with high activity in most strains of Escherichia coli (6 ). Because this characteristic is rather unique for E. coli compared with other bacterial species, ␤-glucuronidase assays with chromogenic and fluorogenic substrates have been developed for the rapid and specific identification of E. coli in clinical microbiological diagnostics and for testing contamination of food and water (7,8 ). Sulfatase activity has been detected in many different bacteria (9 ), but not in E. coli (10,11 ), or only in very low amounts (12 ).E. coli is the most common bacterium isolated in clinical laboratories and is also the predominant pathogen (ϳ80%) in urinary tract infections (UTIs) (13 ). This study, therefore, evaluated whether the presence of E. coli or other common pathogens in urine specimens, resulting from UTIs or possible contamination during sampling and handling, could give false-negative EtG and EtS results in the detection of recent alcohol consumption because of hydrolysis by bacterial ␤-glucuronidase and sulfatase.Fresh clinical urine specimens (n ϭ 46; stored refrigerated) containing confirmed bacterial growth at a density of 10 3 to Ͼ10 5 colony-forming units (CFU)/mL and with Ͼ80% of samples containing Ͼ10 5 CFU/mL, as identified by culture on standard solid media, were obtained from the microbiology laboratory at the Karolinska University Hospital. Specimens were collected consecutively from the routine pool of infected urine samples and were also selected to include different pathogens. The samples were supplemented with 1 mg/L each of EtG (Medichem Diagnostics) and EtS (TCI); they were then split into 3 tubes (without preservatives), which were placed at Ϫ20, 4, and 22°C. Urine without the addit...

Background: Ethyl glucuronide (EtG) is a minor ethanol metabolite used as a specific marker to document recent alcohol consumption; confirm abstinence in treatment programs, workplaces, and schools; and provide legal proof of drinking. This study examined if bacterial pathogens in urine may enable postsampling synthesis of EtG and ethyl sulfate (EtS) from ethanol, leading to clinical false-positive results. Methods: Urine specimens with confirmed growth of Escherichia coli, Klebsiella pneumoniae, or Enterobacter cloacae were stored at room temperature in the presence of ethanol. Ethanol was either added to the samples or generated by inoculation with the fermenting yeast species Candida albicans and glucose as substrate. EtG and EtS were measured by LC-MS. Results: High concentrations of EtG (24-h range 0.5-17.6 mg/L) were produced during storage in 35% of E. coliinfected urines containing ethanol. In some specimens that were initially EtG positive because of recent alcohol consumption, EtG was also sensitive to degradation by bacterial hydrolysis. In contrast, EtS was completely stable under these conditions. Conclusions: The presence of EtG in urine is not a unique indicator of recent drinking, but might originate from postcollection synthesis if specimens are infected with E. coli and contain ethanol. Given the associated risks for false identification of alcohol consumption and false-negative EtG results due to bacterial degradation, we recommend that measurement of EtG be combined with EtS, or in the future possibly replaced by EtS. © 2007 American Association for Clinical ChemistryEarly recognition of problem drinking or relapse is important to ensure adequate alcohol treatment strategies (1 ). This goal has been hampered by a lack of sufficiently sensitive and specific diagnostic methods. The reliability of self-reporting is limited by denial and underreporting (2 ). The time frame for identifying alcohol use by ethanol testing is usually limited to Ͻ12 h, because of rapid metabolism and excretion (3 ). Research has therefore focused on developing alcohol biomarkers with a longer detection window (4 ).A new laboratory marker for detecting recent alcohol consumption is ethyl glucuronide (EtG) (5 ). EtG and ethyl sulfate (EtS) (6 ) are minor ethanol metabolites formed by uridine diphosphate-glucuronosyltransferase (UGT) and sulfotransferase (SULT), respectively, and excreted in urine for a longer time than ethanol (7-10 ). Positive EtG and/or EtS test results thus provide a strong indication that the person has recently consumed alcohol, even when ethanol is no longer detectable (9 ). LC-MS methods are available for EtG and EtS detection (6, 10 ), as is an enzyme immunoassay for EtG (DRI ® EtG, Microgenics). EtG has been recommended for forensic application (11-13 ) and is used for documentation of abstinence in treatment programs, for alcohol testing in the workplace and schools, and as legal proof of drinking (known as "the 80-h alcohol test"). However, the high diagnostic sensitivity of the EtG test has ...