Urinary Tract Infection: A Risk Factor for False-Negative Urinary Ethyl Glucuronide but Not Ethyl Sulfate in the Detection of Recent Alcohol Consumption

Abstract

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...

A study of ethyl glucuronide in post-mortem blood as a marker of ante-mortem ingestion of alcohol

“…Both our study (paper II) and others have documented that EtG is unstable in the presence of bacteria, both in post mortem material (115,119) and in clinical urine samples (146,147 Weinmann, personal information). We found that refrigeration and the presence of preservatives made EtG stable (paper II), results that are also supported by other publications (146,148).…”

“…Both our study (paper II) and others have documented that EtG is unstable in the presence of bacteria, both in post mortem material (115,119) and in clinical urine samples (146,147 Weinmann, personal information). We found that refrigeration and the presence of preservatives made EtG stable (paper II), results that are also supported by other publications (146,148). It is also well documented that EtS is stable in the presence of bacteria, both in clinical urine samples and post mortem material (119,146,147), although a very recent publication showed some instability under extreme conditions, also of EtS (149).…”

“…Nine of these ethanol positive cases were negative for EtG in both blood and urine. We cannot totally exclude the possibility that they were false negatives caused by degradation in both media, but the analysis of EtS could answer this question, as recent research has found superior stability of EtS (119,(146)(147)(148). The analyses of EtS showed that total loss of EtG also in urine probably did not occur, because we found that none of the nine urine samples negative for EtG contained EtS.…”

Vitreous Humor: A Short Review on Post-mortem Applications

“…Another study has also shown that EtG levels were concordant with alcohol intake habits [15]. However, it has been suggested that due to false negative results during putrefaction, it has to be interpreted carefully and it was suggested to use both EtG and EtS to be more accurate [104,105]. However, as EtG degradation and also possible artifactual formation both require microbial activity, VH is expected to be less prone to false positive and false negative EtG results [106][107].…”

Ethyl glucuronide

“…In addition, samples can be taken into containers with fluoride preservatives and frozen to prevent bacterial growth. 22 Other preservatives, e.g. boric acid, have not been investigated with respect to EtG.…”