Ethyl Glucuronide Concentrations Research Articles

Retrospective Data Analysis Reveals Unusual Metabolism Pattern of Ethanol in Pediatrics as Compared to Adult and Geriatric Populations.

About 95% of consumed ethanol is metabolized by oxidative pathways. Less than 1% is metabolized via nonoxidative pathways: glucuronidation, sulfation, and the formation of fatty acid esters of ethanol. In neonates, the glucuronidation pathway has been reported to be underdeveloped but matures with age. This work compared the test results of patients' random urine samples submitted to our facility for ethyl glucuronide (EtG) and ethyl sulfate (EtS) measurements across pediatric and adult populations. Test results (n = 63 498) from urine samples tested for EtG and EtS by quantitative liquid chromatography-tandem mass spectrometry at our facility were utilized for this study. EtG and EtS concentrations were compared across the age partitions 0 to 17 years (pediatric), 18 to 80 years (adult), and 81 to 100 years (geriatric). Eight pediatric patients from a tertiary academic hospital contributed clinical context via abstracted clinical information. Across the individual age partitions, 60% to 65% of patients had both EtG and EtS present in urine. Approximately 5% to 10% of patients had only EtG, and 25% to 35% had neither metabolite present. The lowest percentages (<1.5%) had EtS present in the absence of EtG. Markedly, no pediatric patients had only EtS present; compared to the adult population, this was statistically significant (Fisher exact test, P = 0.025). From the data presented in this work, EtG is more prevalent relative to EtS in urine samples of patients assessed for ethanol exposure.

B-316 Retrospective Data Analysis Reveals Unusual Metabolism Pattern of Ethanol in Pediatrics as Compared to Adult and Geriatric Populations

Abstract Background Ethanol is utilized in many pharmaceutical formulations and in the treatment of methanol poisoning. It is also consumed recreationally and is the most abused drug globally. About 95% of consumed ethanol is metabolized by alcohol dehydrogenase into acetaldehyde, followed by conversion of the acetaldehyde into acetic acid by acetaldehyde dehydrogenase. Less than one percent is metabolized via nonoxidative pathways, which may include: glucuronidation, sulfation, and the formation of fatty acid esters of ethanol. In neonates the glucuronidation pathway has been reported to be underdeveloped and matures with age. In this work, the patterns and concentrations of phase II nonoxidative metabolites of ethanol, ethyl glucuronide (EtG) and ethyl sulfate (EtS), were assessed in random urine collections of pediatric, adult, and geriatric patients. Methods Test results (n = 63498) from urine samples tested for EtG and EtS by quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) at our facility were utilized for this study. For both analytes, the lower limit of positivity was 100 ng/mL. The age range under consideration was 0–100 years. Data was partitioned into age groups: 0–17 years, 18–80 years and 81–100 years. The EtG and EtS concentrations across the partitions were compared. Results Across all ages, 60%–65% of patients had both EtG and EtS present in urine. Between 5%–10% had EtG present without EtS, and 25%–35% of patients had neither of the ethanol metabolites present. In all age groups there was a low positivity rate of EtS without the presence of EtG (0%–1%). All age partitions showed a similar pattern with the highest percentages of patients showing the presence of both metabolites, and the lowest percentage of patients showing the presence of EtS only. Markedly, there were no pediatric patient samples that had the sulfated metabolite present in the absence of the glucuronidated metabolite; this was statistically significantly different than this proportion in adults (Fisher’s Exact test, P = 0.025). This finding suggests greater glucuronidation activity in the pediatric population. Conclusion From the data presented in this work, EtG is more prevalent relative to EtS in urine samples of patients assessed for ethanol exposure. This work also shows that glucuronidation may be the predominant pathway for ethanol metabolism in pediatric populations. Although the level and frequency of ethanol exposure, as well as the time between exposure and sampling were unknown for the patients studied, the data offers insights into the relative distribution of EtG and EtS in random urine samples for pediatric and adult populations.

Ethyl glucuronide and ethyl sulfate in the zebrafish after ethanol exposure

Ethanol exposure during pregnancy is an important problem and is the cause of fetal alcohol syndrome (FAS) and fetal alcohol spectrum disorder (FASD). The etiology of FAS and FASD can be elucidated using animal models. Recently, a novel model, the zebrafish (Danio rerio), has garnered the interest of researchers. This study confirmed the negative influence of ethyl alcohol (0.5 %, 1.5 %, and 2.5 % v/v) on the development of zebrafish embryos. The observed malformations included pericardial and yolk sac edema, increased body curvature, tail edema, and a decreased embryo hatching rate. The differences in body length, body width, and heart rate were statistically significant. Due to the similarities in the quantity and function of ethanol biotransformation enzymes between zebrafish and mammals, this study investigated the nonoxidative metabolites of ethanol – ethyl glucuronide (EtG) and ethyl sulfate (EtS) – in zebrafish following ethanol exposure. This research confirmed that EtG and EtS concentrations can be measured in zebrafish embryos, and the levels of these metabolites appear to be associated with the ethyl alcohol concentration in the medium.

Formation of phosphatidylethanol and ethylglucuronide after low to moderate alcohol consumption in volunteers with a previous three-week alcohol abstinence.

Phosphatidylethanol (PEth) is only formed when ethanol is present in blood. This direct alcohol marker has been widely discussed, including the minimum amount of ethanol being necessary to form as much PEth as to exceed the threshold of 20ng/mL in previously PEth negative subjects. In order to corroborate hitherto existing results, a drinking study including 18 participants after a 3-week alcohol abstinence was performed. They consumed a pre-calculated amount of ethanol to reach a blood alcohol concentration (BAC) of at least 0.6g/kg. Blood was drawn before and periodically seven times after alcohol administration on day 1. Blood and urine were also collected the next morning. Dried blood spots (DBS) were prepared immediately from collected venous blood. BAC was determined by head space gas chromatography and the concentrations of both PEth (16:0/18:1, 16:0/18:2 and five additional homologues) and ethyl glucuronide (EtG) were analysed using liquid chromatography-tandem mass spectrometry. Out of 18, 5 participants had concentrations of PEth 16:0/18:1 above the threshold of 20ng/mL, and 11 out of the 18 subjects had concentrations between 10 and 20ng/mL. In addition, four persons had PEth 16:0/18:2 concentrations above 20ng/mL the following morning. All test subjects tested positive for EtG in DBS (≥ 3ng/mL) and urine (≥100ng/mL) upon 20-21h after alcohol administration. By combining both a lower cutoff of 10ng/mL and the homologue PEth 16:0/18:2, the sensitivity to detect a single alcohol intake after a 3-week abstinence increases to 72.2%.

Assessing maternal alcohol consumption in pregnancy: comparison of confidential postnatal maternal interview and measurement of alcohol biomarkers in meconium

ObjectiveKnowledge of alcohol consumption in pregnancy is important for early identification of children with fetal alcohol spectrum disorder. We investigated whether alcohol biomarkers fatty acid ethyl esters (FAEEs) and ethyl...

EtG Quantification in Hair and Different Reference Cut-Offs in Relation to Various Pathologies: A Scoping Review.

Ethyl glucuronide (EtG) is a non-volatile, non-oxidative, hydrophilic, and stable ethanol phase II metabolite. EtG is produced through ethanol glucuronidation by UDP-glucuronosyltransferase (UGT), a phase II enzyme. EtG can be extracted from different biological matrices, including keratin ones, such as hair or nails. The purpose of this scoping review is to describe the relationship between EtG levels in hair and some of the most common and frequent pathological conditions and verify whether different reference cut-offs in relation to various pathologies have been identified in the scientific literature. In fact, in-depth knowledge of the influence of pathologies, such as diabetes mellitus, hepatic and renal dysfunction, on EtG production and its storage in keratin matrices would allow a more appropriate interpretation of obtained data and rule out false positives or false negatives. This scoping review is based on bibliographic research carried out on PubMed regarding the quantification of EtG in hair of subjects affected by different pathological conditions. According to the scientific literature, the main and most common pathologies that can affect the concentration of EtG in hair are liver and kidney diseases and diabetes. The EtG quantification analytical data should be interpreted carefully as they may have a great impact in both forensic and clinical contexts.

Estimating the time of last drinking from blood ethyl glucuronide and ethyl sulphate concentrations

The determination of length of time from the last drinking is critical for cases like drunk driving, sexual assault victims, and also postmortem suspected poisoning cases. The study was aimed to established a method of estimating the time of last drinking through the pharmacokinetic study of conjugation metabolites of alcohol in blood after a single oral dose. Twenty-six volunteers (14 males) consumed alcohol with food at a fixed dose of 0.72 g/kg after fasting for 12 h. Five milliliters of blood were collected 120 h after the start of drinking, and all samples were analyzed with headspace-gas chromatography and high-performance liquid chromatography–tandem mass spectrometry. The time point of last drinking was estimated through the relationship between the concentration ratio of ethyl glucuronide to ethyl sulphate and the length of time after drinking. Pharmacokinetic parameters were analyzed by a pharmacokinetic software DAS according to the non-compartment model. A good correlation model was obtained from the relationship between concentration ratio of ethyl glucuronide to ethyl sulphate in blood and the time of alcohol use, and the margin of error was mostly lower than 10%. The time of maximum concentration, maximum concentration, and elimination half-life of ethyl glucuronide in blood were 4.12 ± 1.07 h, 0.31 ± 0.11 mg/L and 2.56 ± 0.89 h; the time of maximum concentration, maximum concentration, and elimination half-life of ethyl sulphate in blood were 3.02 ± 0.70 h, 0.17 ± 0.04 mg/L, and 2.04 ± 0.76 h. The study established a potential method to estimate the length of time after a moderate oral dose, and provided pharmacokinetic parameters of ethyl glucuronide and ethyl sulphate in Chinese population.

Assessment of prenatal exposure to alcohol and gestational consumption by EtG testing in neonatal meconium and maternal hair and neonatal: First Italian national survey

Use of alcohol during pregnancy and subsequent fetal exposure can cause multiple perinatal disorders (prematurity at birth, withdrawal syndromes, tremors, hyperreflexia) and impaired physical and mental development in the later stages of life, known as fetal alcohol spectrum disorders (FADS) including as major expression of impairment fetal alcohol syndrome (FAS). Gestational drinking is usually, worldwide reported by questionnaires filled during antenatal visits, but international studies demonstrated their inconsistency for under-reporting or misreporting of toxic habits. Information concerning the true prevalence of alcohol consumption during pregnancy and consequent prenatal exposure to this teratogen is essentially lacking in Italy. To assess objective gestational consumption to alcohol and fetal exposure to it, we measured one of the most specific ethyl alcohol metabolites, ethylglucuronide (EtG), in maternal hair and neonatal meconium involving delivering mothers and newborns along the whole National Italian territory. Non-paired twenty delivering women and twenty newborns per city from hospitals of the National Health system located in the North, Centre and South Italy (including islands) were enrolled in the project. Maternal hair, 8 cm as maximum length, was collected at the end of pregnancy and analyzed as entire strand and neonatal meconium was collected within the first 24 hours after birth. Ethylglucuronide was quantified in both biological matrices through a validated gas chromatography–tandem mass spectrometry methodology. Hair EtG concentrations below 5 pg/mg hair were attributed to abstinent women, those > 5 pg/mg and below 30 pg/mg were attributed to women consuming “some” alcohol during pregnancy and ≥ 30 pg/mg were attributed to excessive drinkers. Furthermore, according to previous studies from our research group, maternal hair EtG above 11 pg/mg was shown to correspond with high probability to a neonatal meconium with EtG > 30 ng/g. Hence, it was considered as a “cut-off” matching to fetal exposure to alcohol. Up to now, 781 maternal hair and 642 meconium samples have been analyzed. Only one woman out of 781(0.1%) presented chronic excessive ethanol consumption with EtG concentration > 30 pg/mg, while the 8.2% of maternal hair presented EtG concentrations > 5 pg/mg with 1.4% > 11 pg/mg. Four newborns (0.6%) resulted prenatally exposed to ethanol with meconium EtG concentration > 30 ng/g. The highest percentage of women involved in the study (mean age + SD 34 + 5.1 years) were Italian (88%) and employed (69%). With regard to educational level, 44% had a university degree and 39% high-school education level. The four exposed newborns did not present at birth any sign of evident disability or malformation. These preliminary findings, obtained in separate cohorts of pregnant and delivering mothers and newborns showed that, currently, a negligible amount of Italian women drink during pregnancy and that a minute percentage of newborns are prenatally exposed to alcohol. The policies applied by health Italian women have acquired awareness about risks associated with drinking during pregnancy.

Dried blood spot (DBS) for ethyl glucuronide (EtG) and ethyl sulfate (EtS) determination in blood with a new design of “volumetric” DBS filter paper card

Determination of EtG in blood can be performed from capillary blood with high sensitivity and detection limits of down to 1 ng/mL for EtG–and in addition to phosphatidyl ethanol (PEth) determination, EtG concentrations can be used for interpretation of consumption habits of patients in maintenance therapy. Sampling of capillary blood is a crucial preanalytical step–and different methods have been suggested for collection of a defined volume (VAMS, Capitainer, HemaXis, and capillaries for collection of samples). We used and validated a new volumetric capillary blood sampling card (DBSv cards, Protzek, Lörrach/Germany) which can be used either for self-sampling of dried blood spots (up to 4 times 20 microliter of blood) on filter paper with convenient handling by the patients or also in laboratory work. In contrast to standard DBS cards with printed circles on a filter paper, a newly designed card with a paper comb with four teeth per card, which allows a volumetric collection of 20 microlitres of blood per tooth, was used. Venous blood was pipetted on the teeth of DBSv cards, dried, and one paper tooth was ripped off and extracted by adding 490 microliter of methanol and 10 microliter of ISTD (400 ng/ml D5-EtG and 400 ng/ml D5-EtS). The samples were then vortexed for 5 minutes and centrifuged for 10 minutes. After transferring them into vials, the solutions were evaporated under a stream of nitrogen and reconstituted in 80 microliter of a water/0.1% formic acid. The samples were shaken and 10 μL were injected onto the LC column. A previously published LC-MS/MS method was adapted for a QTrap 5500 triple-quadrupole ion trap mass spectrometer (Sciex, Toronto, Canada) with an Acquity C18 HSS T3 column (Waters, Wexford, Ireland) and detection of EtG, D5-EtG, EtS and D5-EtS was performed according to forensic guidelines using one precursor and two product ions. Lower limits of quantification were 10 ng/mL for EtG and 20 ng/mL for EtS–with linear ranges of 10 to 5000 ng/mL for EtG and (50 to 2500 ng/mL for EtS. Traces of EtS were found in HPLC grade solvent, and a solvent selection between products of different manufacturers had to be performed. One brand methanol showed the lowest baseline level of EtS, which was in the range of 20 ng/mL. Thus, for determination of EtS, a lower limit of quantitation of 50 ng/mL was defined, whereas EtG could be quantified at concentrations above 10 ng/mL. A validation with spiked blood samples was performed. Samples from forensic cases and from drinking studies, where DBS were collected for the analysis of alcohol biomarkers (PEth and EtG/EtS) have been analyzed. The dried blood spot sample collection in laboratory or by self-sampling (of capillary blood) can be used to detect EtG and EtS at very low levels. The results show the potential of DBS analysis for alcohol markers EtG and EtS–which can be used for abstinence monitoring by very low limits of detection, especially for EtG, or for the detection of a recent alcohol consumption, when ethanol has already been eliminated from the body. A contamination of pure HPLC-grade solvent methanol was found for EtS determination, this has to be kept in mind, and needs further optimization for detection of EtS at lower concentrations in biological samples.

Blood simultaneous determination of PEth 16:0/18:1, PEth 16:0/20:4 and ethylglucuronide using DBS: Application to a clinical study of patients hospitalized for withdrawal

This study aims to present a LC-MS/MS method for simultaneous determination of phosphatidyléthanol (PEth)16:0/18:1, PEth16:0/20:4 and ethylglucuronide (EtG) using dried blood spots (DBS) and to report preliminary results of PETHOXICO, a monocentric non-interventional non-randomized cross-sectional clinical study (ClinicalTrials.gov NCT04557631 ) in a population of patients hospitalized for withdrawal. Blood samples can alternatively consist in 10 μL from usual blood sampling tubes (e.g. fluoride preserved) transferred on DBS card, or direct DBS sampling using Volumetric Absorptive MicroSampling devices (VAMS; Hemaxis™, Neoteryx™). For PETHOXICO, PEth16:0/18:1, PEth16:0/20:4 and EtG were analyzed simultaneously together with urinary EtG and blood alcohol concentration (BAC), determined using immunoassay and GC-FID, respectively. The fully validated LC-MS/MS method exhibits a LLOQ of 2 μg/L for the 3 analytes. A total of 37 chronic and excessive ethanol consumers (ethanol per week consumptions ranging from 560 to 1750 g) were included in PETHOXICO [sex ratio 11/26 (F/M); 30 to 67 YO with a median at 46 YO]. At D0, the mean concentration of PEth16:0/18:1 was 1180 μg/L (range: 53–3889 μg/L) with 95% of participants above the 210 μg/L threshold. PEth16:0/20:4-to-PEth16:0/18:1 mean ratio was 0.46 with PEth16:0/20:4 concentrations ranging from 9 to 2353 μg/L. The mean BAC was 1060 mg/L (range: undetected–3540 mg/L) and for 78% of participants, urinary EtG was above the 500 μg/L threshold. At D7, no BAC nor EtG (urine and blood) were found as expected. The mean concentration of PEth16:0/18:1 dropped to 527 μg/L (range: 16–1655 μg/L) with still 78% of participants above the 210 μg/L threshold. PEth16:0/20:4 concentrations dropped more significantly (range: undetected–923 μg/L) with a mean PEth16:0/20:4-to-PEth16:0/18:1 ratio of 0.26. This decreased ratio (0.46 at D0 and 0.26 at D7) is coherent with the expectation of a shortened detection window for PEth16:0/20:4. Blood elimination half-life of PEths were calculated with the assumption of a single elimination phase: mean half-life were 6.9 days (range: 2.2–18.6 days) and 4.6 days (range: 1.3–9.3 days) for PEth16:0/18:1 and PEth16:0/20:4, respectively. PEth16:0/18:1 observed half-life is consistent with literature data (around 7 days with a significant inter-individual variability). Observed PEth16:0/20:4 half-life is higher than the very scarce data from literature (2 days). In this study, the use of an analytical method combining 3 direct metabolites of ethanol allows, in one run, to discriminate a very recent consumption (< 48 h) from a recent consumption (< 2 weeks) and an older consumption (2–4 weeks). PEth16:0/18:1 sensitivity to detect chronic and excessive alcohol consumers was 95% with a 210 μg/L threshold. However, these preliminary results need yet to be confronted to detailed clinical data.

Ethyl glucuronide and alcohol abstinence: A correlation study in hair and fingernails to establish a cut-off value in fingernails for teetotalers

IntroductionExcessive use of alcohol increases the risk to be involved in a road traffic accident. According to many legislations, certain maximal blood-alcohol-concentrations (BAC) have to be respected to be allowed to drive on public roads. Acute alcohol intoxication is evaluated by blood analysis or analysis of the exhaled alveolar air. In many cases, evaluation of the alcohol consumption during the past months can be useful. In this light, ethyl glucuronide (EtG), a direct alcohol biomarker which can be found in keratinized matrices (hair, nails) is valuable and can be used for the long-term follow-up of alcohol consumption. Research aimTo compare the EtG concentration in hair and fingernails from teetotalers, and to propose a cut-off value for EtG in fingernails for alcohol abstinence. Material and methodsPaired samples of hair and nails were collected from participants, with a minimum age of 18 years. They all stated alcohol abstinence for at least 6 months. Participants were asked to complete a questionnaire about age, gender and the use of hair care products and nail polish. Analysis of EtG in the nail and hair samples were conducted following a validated analytical method. ResultsFrom 126 participants a hair and nail sample were collected. Of this group, 15 participants were finally not included in the study because of insufficient amount of hair or nails collected. There were more female participants (65%) and the average age of participants was 39 years. The EtG concentration in hair was below the limit of detection of 2.1 pg/mg in all but 4 samples (2.1, 2.1, 2.9, and 3.5 pg/mg). The EtG concentration in nails was below the limit of detection in 97 of the 111 samples. The concentrations in nails ranged between 2.3 and 23 pg/mg. Discussion and conclusionIn a population of 111 teetotalers, the 97.5% percentile of EtG concentrations in fingernails is 7.6 pg/mg. The highest EtG concentration observed was 23 pg/mg. These results suggest that the cut-off value for alcohol abstinence may be lower than the previous suggested 59 pg/mg and 37 pg/mg.

Monitoring alcohol-use-disorder medication compliance by LC-MS/MS determination of urinary ethyl glucuronide, acamprosate, naltrexone, and 6β-naltrexol using zirconia-based hybrid solid-phase extraction

Alcohol use disorder (AUD) is a major risk factor for numerous social concerns worldwide. In the Republic of Korea, the court imposes compulsory medication treatment on criminals diagnosed with AUD who are on probation. The purpose of the treatment is to reduce the number of repeat offenses committed by AUD-afflicted criminals. In this study, a method employing liquid chromatography–tandem mass spectrometry (LC–MS/MS) to determine ethyl glucuronide (EtG), acamprosate (ACAM), naltrexone (NTX), and 6β-naltrexol (6BNT) in urine was developed and validated to monitor the medication compliance and alcohol use of probationers diagnosed with AUD. A zirconia-based hybrid solid–phase extraction technique was introduced to increase the sensitivity toward target analytes having significantly differing pKa values while minimizing the matrix effect of the urine sample. The pretreated urine samples were analyzed by LC–MS/MS performed in a polarity-switching electrospray ionization mode via a three-period multiple-reaction monitoring method. All the target analytes were separated and detected within 6.5 min with an Xselect HSS T3 column and gradient elution using water containing 0.02% formic acid and methanol as the mobile phase. The lower limits of quantification of EtG, ACAM, NTX, and 6BNT were 10.0, 4.0, 0.4, and 0.2 ng mL−1, respectively. The determination coefficients of each calibration curve were greater than 0.9989. The intra-day accuracy ranged from −5.5–5.3% and the precision was ≤ 5.7%, whereas the inter-day accuracy ranged from −5.3–4.6% and the precision was ≤ 4.7%. The recovery, matrix effect, and process efficiency were 99.7–107.9%, 80.7–101.8%, and 80.5–108.1%, respectively. The developed method was successfully applied to analyze 107 urine samples obtained from probationers undergoing medication treatment for AUD and to monitor the probationers’ medication compliance and alcohol use. This study also determined the urinary concentrations of EtG, ACAM, NTX, and 6BNT as well as the metabolic ratio of NTX following repeated oral administration of AUD medications.

DETERMINAÇÃO DE ETIL-GLICURONÍDEO E ETIL-SULFATO EM SANGUE POR UPLC-MS/MS: APLICAÇÃO CLÍNICA EM VÍTIMAS DE TRAUMA

DETERMINATION OF ETHYL-GLUCURONIDE AND ETHYL-SULFATE IN BLOOD BY UPLC-MS/MS: CLINICAL APPLICATION IN TRAUMA VICTIMS. We developed a UPLC-MS/MS method for the quantification of the ethanol metabolites ethyl-glucuronide (EtG) and ethyl-sulfate (EtS) in blood samples. Sample preparation was a protein precipitation with acetonitrile 0.1% formic acid containing internal standards (IS) 0.5 µg mL-1 EtG-D5 and EtS-D5. Chromatographic separation was performed in a fluoro-phenyl column, mobile phases were 0.1% formic acid in water (A) and acetonitrile (B) in gradient elution from 5% B to 70% B, with 10 min run time. Ionization was performed with electrospray source at 550 °C in negative mode, quantification mass transitions of m/z EtG 221- 75, EtS 125- 80.1, EtS-D5 130- 79.9, and EtG-D5 226- 75.1. The method was linear from 0.1 to 18 µg mL-1 for EtG and 0.02 to 6 µg mL-1 for EtS, specific, with no interfering peaks during the blank samples analyses. The method was precise and accurate, the matrix effect was compensated with the IS (-12% a + 6.1%). EtG and EtS were analyzed in 28 blood samples from trauma patients with positive alcoholaemia. The concentrations of EtG and EtS ranged from 0.25 to 14.11 µg mL-1 and 0.13 to 5.53 µg mL-1, with high correlation with ethanol concentrations, r=0.739 and r=0.817 (p &lt;0,001), respectively

Phosphatidylethanol, ethyl glucuronide and ethanol in blood as complementary biomarkers for alcohol consumption

Alcohol biomarkers can monitor both recent and long-term drinking and provide information about drinking habits as a complement to self-reporting. Ethyl glucuronide (EtG) and phosphatidylethanol (PEth) are the most sensitive available biomarkers for this purpose. The present study aimed to collect data on both PEth and EtG in the same blood sample, in addition to ethanol, in order to evaluate the combined use of these biomarkers. Venous EDTA blood samples (n = 1149) sent to the laboratory as part of a clinical routine service for measuring PEth were investigated. PEth and EtG concentrations were analyzed using liquid chromatography–mass spectrometry methods and ethanol with an enzymatic method. Of the 1149 samples, 95 were positive for ethanol (range 0.11–3.12 g/L), 454 for EtG (1.0–9739 ng/mL), 635 for PEth (0.014–6.0 µmol/L), 534 for PEth ≥ 0.050 µmol/L, and 315 for PEth ≥ 0.30 µmol/L. EtG and PEth concentrations seemed largely independent as the coefficient of determination (r2) between PEth and EtG concentrations was 0.15. However, when the EtG concentrations were evaluated for different subgroups depending on ethanol or PEth concentrations a statistically significant difference between successive higher concentrations was observed. EtG and PEth are independent measures of recent alcohol drinking reflecting different time windows. Their combined measurement in the same blood sample is possible and will provide valuable information regarding recent alcohol consumption as a complement to self-reporting.

Driving license regranting in DUI subjects: Road accident variables and predictive factors of substance use disorder.

A comparative case study (2017-2020) was conducted to identify demographic, social, medico-legal, and toxicological variables associated with non-fatal accidents in driving under the influence (DUI) subjects. A second aim was to identify the factors predictive of substance use disorders among subjects. Drivers charged with alcohol DUI (blood alcohol concentration (BAC) > 0.5) and/or psychoactive substance DUI were included; cases included those involved in an accident while intoxicated, and the comparison group included DUI offenders negative for road accident involvement. Significance was determined by chi-square and Mann-Whitney tests. To prevent confounding effects, a multivariate binary logistic regression analysis was performed. Our sample encompassed 882 subjects (381 in the case group and 501 in the comparison group). Parameters such as psychoactive substances and BAC at the time of the road crash/DUI and the day of the week, when subjects were involved in the road accident or found DUI, resulted in significant differences (p < 0.01) between groups. The model's independent variables of BAC > 1.5 g/L (p = 0.013), BAC > 2.5 g/L (p < 0.001), and concurrent alcohol and psychoactive substance use (p < 0.001) were independent risk factors for an accident. Smoking >20 cigarettes/day was an independent risk factor for unfitness to drive (p < 0.01). Unfitness to drive was based primarily on ethyl glucuronide levels >30 pg/mg. Our results suggest a detailed assessment of DUI subjects with variables associated with accidents (BAC > 1.5 g/L and concurrent intake of psychoactive substances). Hair analysis, including ethylglucuronide (EtG) concentration, should be always performed. Based on our results, nicotine use should be investigated in cases of driving license regranting.

Validation of hair ethyl glucuronide using transdermal monitoring and self-reported alcohol use in women of childbearing potential.

AimThe present study aimed to evaluate the validity of hair ethyl glucuronide concentrations compared with transdermal alcohol concentration and self‐reported alcohol use.MethodsThis trial included 25 adolescent and young adult females, aged 16‐24, who reported at least one heavy drinking episode (≥4 drinks) in the two weeks prior to baseline. All participants were asked to wear an alcohol biosensor over a one‐month prospective study. Detailed self‐report of drinking behaviors was assessed weekly. Estimates of blood alcohol concentration were computed from self‐report data using the National Highway and Transportation Safety Administration equation. Transdermal alcohol concentration and estimated blood alcohol concentration data were categorized into at‐risk (>0.05 g/dL alcohol) and high‐risk (>0.08 g/dL alcohol) drinking events. Hair ethyl glucuronide concentration, total number of drinking events, moderate (>0.05 g/dL) and high level (>0.08 g/dL) of transdermal alcohol concentration, and estimated blood alcohol concentration drinking events were analyzed with Spearman's rank correlation test for validity comparisons.ResultsNo significant correlations were found between hair ethyl glucuronide values and total number, and moderate or high levels of detected drinking events by estimated blood alcohol concentration or transdermal alcohol concentration. Total number of drinking events detected and number of drinking events >0.08 g/dL using estimated blood alcohol concentration and transdermal alcohol concentration methods were significantly correlated with each other (respectively, R = .33, P < .05; R = .42, P < .05).ConclusionOur findings indicate that, due to the number of false negatives, hair ethyl glucuronide concentrations should be used with caution for monitoring abstinence from alcohol use.

Clinical usefulness of a urine dipstick to detect ethyl glucuronide (EtG): A quantitative clinical study in healthy young female volunteers.

The metabolite of ethanol, ethyl glucuronide (EtG), reflects alcohol intake longer than ethanol and is used as a biomarker in clinical settings to detect alcohol use. We aimed to assess the clinical usefulness in a low-to-moderate alcohol intake setting and validate a new urine EtG dipstick. A three-way, open, cross-over trial was conducted. Data were collected from January to June 2019. Among 12 healthy female volunteers, we quantified urine EtG and used a dipstick following intake of either one, two or four units of alcohol. Main outcomes were concentrations of EtG in urine and serum, and creatinine and ethanol in serum. EtG in urine was determined dichotomously by dipsticks at two different thresholds and by mass spectrometry used as gold standard. EtG in urine was quantifiable up to 24hours after alcohol intake. In some individual cases, EtG was quantifiable up to 72hours at low concentrations. The dipstick detected EtG in urine up to 24hours. At thresholds of 1000 and 1500ng/mL, the dipsticks had a specificity of 100% (both), while sensitivity was 84% and 69%, respectively. The sensitivity of the dipsticks was insufficient to support a screening purpose in this setting of low-to-moderate alcohol intake.

Ethyl Glucuronide and Ethyl Sulphate in Urine: Caution in their use as markers of recent alcohol use.

To clarify the role of the ethanol metabolites, ethyl glucuronide (EtG) and ethyl sulfate (EtS), in monitoring alcohol consumption. We recruited 7 female and 17 male volunteers who were instructed to consume a quantity of beer (containing 48 gm ethanol) with food in one session. We examined urinary excretion of EtG and EtS over time and looked for correlations between the concentrations of the metabolites EtG and EtS. EtG concentrations in urine varied between 0.026 and 430.372μg/ml with average values between 11.85μg/ml (SD 19.75), 30min after alcohol intake, and 100.39μg/ml (SD 101.34), 4.5h after alcohol intake. EtS urinary concentration ranged from 0.006 to 101.432μg/ml with average values between 4.77μg/ml (SD 5.42), 30min after alcohol intake, and 30.14μg/ml (SD 27.20), 4.5h after alcohol intake. Spearman's test showed that urinary EtG and EtS correlated significantly at several time points. The great interindividual variability in their excretion suggests caution in the use of urinary measurement of these metabolites in forensic investigations.

Ethyl glucuronide and ethanol concentrations in femoral blood, urine and vitreous humor from 117 autopsy cases

BackgroundVitreous humor (VH) is a specimen of great value in forensic investigations and is being used for evaluating possible post-mortem formation of ethanol. Ethyl glucuronide (EtG) is an ethanol metabolite that has found interest for the same purpose. Both compounds can be measured in VH and because of differences in rate of distribution and elimination they may offer complementary information. MethodsVH, femoral blood (FB) and urine were collected from 117 autopsy cases for forensic investigation. Ethanol was measured with headspace gas chromatography, while EtG was measured with liquid chromatography – tandem mass spectrometry. Results and conclusionEthanol was detected in all matrices in 39 cases, while EtG was present in 62 cases. The VH-FB and the VH-urine ethanol concentrations in the 39 cases were statistically correlated (p < 0.00001). In one case with an ethanol concentration of 0.11 g/L in FB, no ethanol was detected in VH and urine, and no EtG in any specimen, indicating a possible post-mortem formation. EtG was present in VH in more cases than in FB and urine. The correlation between the EtG concentrations in VH and FB was statistically significant (p < 0.0003) as was the case also for VH and urine (p < 0.001). The combined information on ethanol and EtG concentrations in the three matrices can be used to interpret alcohol drinking habit before death. This study confirms the value of using VH as a specimen in forensic investigations regarding recent exposure to ethanol. EtG can be used not only for investigating post-mortem ethanol formation but also for estimating recent alcohol drinking.

Does Eumelanin Oxidation Play a Role on the Photostability of Ethyl Glucuronide in Hair Exposed to Simulated Solar Radiation?

The action of solar radiation on the concentration of the ethanol metabolite ethyl glucuronide (EtG) in 40 hair samples of nonabstinent subjects was investigated. Hair samples of different colors were analyzed before and after irradiation with artificial sunlight under a light dose corresponding to 3 months of sun exposure. After irradiation, an increase of EtG concentration was detected in 55% of the samples ranging from 5% to 141%. In 16 cases, a concentration reduction ranging from -2% to -74% was observed. The measure of the level of pyrrole-2,3,5-tricarboxylic acid (PTCA), a marker of eumelanin oxidation, demonstrated the largest increase in oxidation in light brown hair where the greatest degradation of EtG was observed after irradiation. However, the rise of PTCA in all hair tested was accompanied by increase in EtG concentration in 8/10 samples and by decrease in 2/10, suggesting no correlation between the two markers. To verify if hair structure was modified by light, scanning electron microscopy (SEM) analysis was performed on irradiated hair of different colors and compared with the dark samples. SEM revealed modification of hair structure in all samples showing partial shaft exfoliation and reduction of hair thickness under the treatment with solar radiation.