Friedrich Martin Wurst

Chronic intake of fermented floral nectar by wild treeshrews

For humans alcohol consumption often has devastating consequences. Wild mammals may also be behaviorally and physiologically challenged by alcohol in their food. Here, we provide a detailed account of chronic alcohol intake by mammals as part of a coevolved relationship with a plant. We discovered that seven mammalian species in a West Malaysian rainforest consume alcoholic nectar daily from flower buds of the bertam palm (Eugeissona tristis), which they pollinate. The 3.8% maximum alcohol concentration (mean: 0.6%; median: 0.5%) that we recorded is among the highest ever reported in a natural food. Nectar high in alcohol is facilitated by specialized flower buds that harbor a fermenting yeast community, including several species new to science. Pentailed treeshrews (Ptilocercus lowii) frequently consume alcohol doses from the inflorescences that would intoxicate humans. Yet, the flower-visiting mammals showed no signs of intoxication. Analysis of an alcohol metabolite (ethyl glucuronide) in their hair yielded concentrations higher than those in humans with similarly high alcohol intake. The pentailed treeshrew is considered a living model for extinct mammals representing the stock from which all extinct and living treeshrews and primates radiated. Therefore, we hypothesize that moderate to high alcohol intake was present early on in the evolution of these closely related lineages. It is yet unclear to what extent treeshrews benefit from ingested alcohol per se and how they mitigate the risk of continuous high blood alcohol concentrations.

Phosphatidylethanol as a sensitive and specific biomarker: comparison with gamma-glutamyl transpeptidase, mean corpuscular volume and carbohydrate-deficient transferrin.

Phosphatidylethanol (PEth), a direct ethanol metabolite, is detectable in blood for more than 2 weeks after sustained ethanol intake. Our aim was to assess the usefulness of PEth [comparing sensitivity, specificity and the area under the curve (AUC)] as compared with carbohydrate‐deficient transferrin (CDT), gamma‐glutamyl transpeptidase (GGT) and mean corpuscular volume (MCV), calculating the results from sober patients against those from alcohol‐dependent patients during withdrawal. Fifty‐six alcohol‐dependent patients (ICD‐10 F 10.25) in detoxification, age 43 years, GGT 81 U/l, MCV 96.4 fl, %CDT 4.2, 1400 g ethanol intake in the last 7 days (median), were included in the study. Over the time of 1 year, 52 samples from 35 sober forensic psychiatric addicted in‐patients [age 34 years, GGT 16 U/l, MCV 91 fl, CDT 0.5 (median)] in a closed ward were drawn and used for comparison . PEth was measured in heparinized whole blood with a high‐performance liquid chromatography method. GGT, MCV and %CDT were measured using routine methods. A receiver operating characteristic curve analysis was carried out, with ‘current drinking status’ (sober/drinking) as the state variable and PEth, MCV, GGT and CDT as test variables. The resulting AUC was 0.974 (P < 0.0001, confidence interval 0.932–1.016) for PEth. At a cut‐off of 0.36 µmol/l, the sensitivity was 94.5% and specificity 100%. The AUC for CDT, GGT and MCV were 0.931, 0.894 and 0.883, respectively. A significant Spearman’s rank correlation was found between PEth and GGT (r = 0.739), CDT (r = 0.643), MVC (r = 0.639) and grams of ethanol consumed in the last 7 days (r = 0.802). Our data suggest that PEth has potential to be a sensitive and specific biomarker, having been found in previous studies to indicate longer lasting intake of higher amounts of alcohol.

Measurement of direct ethanol metabolites suggests higher rate of alcohol use among pregnant women than found with the AUDIT—a pilot study in a population-based sample of Swedish women

OBJECTIVES The objective of the study was to investigate whether biomarkers of alcohol consumption would provide additional information to the use of a validated alcohol questionnaire in pregnant women. STUDY DESIGN One hundred three pregnant women were included in the study. The women completed the Alcohol Use Disorders Identification Test (AUDIT) questionnaire, and a urine and hair sample was collected. The urine samples were used for determination of ethyl glucuronide (EtG) and ethyl sulfate and the hair samples for EtG and fatty acid ethyl esters (FAEE). RESULTS Twenty-six women (25.2%) were identified as possible alcohol consumers by the combined use of AUDIT and direct ethanol metabolites. Seven subjects had EtG or FAEE levels in hair highly suspicious of heavy drinking, but only 1 of these were positive according to the AUDIT questionnaire CONCLUSION The combined use of the AUDIT questionnaire and direct ethanol metabolites appear to identify more potential alcohol consumers among pregnant women than does the sole use of the AUDIT questionnaire.

Detection of Recent Ethanol Intake With New Markers: Comparison of Fatty Acid Ethyl Esters in Serum and of Ethyl Glucuronide and the Ratio of 5-Hydroxytryptophol to 5-Hydroxyindole Acetic Acid in Urine

BACKGROUND At present, recent ethanol consumption can be routinely detected with certainty only by direct measurement of ethanol concentration in blood or urine. Because ethanol is rapidly eliminated from the circulation, however, the time span for this detection is in the range of hours. Several new markers have been proposed to extend the detection interval, but their characteristics have not yet justified their use in routine clinical practice. We therefore investigated three new markers and compared their kinetics and sensitivities: (1) fatty acid ethyl esters (FAEEs) in serum, (2) ethyl glucuronide (EtG) in urine, and (3) the ratio of 5-hydroxytryptophol to 5-hydroxyindole acetic acid (5-HTOL/5-HIAA) in urine. METHODS Seventeen healthy men participated in a drinking experiment. Blood and urine samples were collected twice daily on three consecutive days and once daily on days 4 and 5. Ethanol concentration was determined by gas chromatography, FAEE levels, by gas chromatography with mass spectrometry, EtG concentration, by liquid chromatography-tandem mass spectrometry, and 5-HTOL/5-HIAA ratio, by high-performance liquid chromatography. RESULTS The peak serum ethanol concentrations of the subjects ranged from 5.4 to 44.7 mmol/liter (mean +/- SD, 30.1 +/- 9.1 mmol/liter). In the case of the serum ethanol determination, 100% sensitivity was reached only immediately after the end of the drinking experiment, and in the case of FAEE levels and 5-HTOL/5-HIAA ratio, it tested for 6.7 hr after the end of the ethanol intake. Thereafter, these latter parameters declined until 15.3 hr (FAEEs) and 29.4 hr (5-HTOL/5-HIAA), subsequently remaining in a stable range until 78.5 hr without further decrease. In contrast, EtG concentration showed 100% sensitivity until 39.3 hr and thereafter decreased, falling to below the limit of quantification of 0.1 mg/liter at 102.5 hr. CONCLUSION After moderate drinking, EtG in the urine proved to be a superior marker of recent ethanol consumption in healthy subjects. This is because EtG is a direct ethanol metabolite, it occurs in the urine only when ethanol has been consumed, and its sensitivity remains at the level of 100% for 39.3 hr.

Ethyl glucuronide discloses recent covert alcohol use not detected by standard testing in forensic psychiatric inpatients

BACKGROUND Considerable lives and money could be saved if one could detect early stages of lapsing/relapsing behavior in addicted persons (e.g., in safety-sensitive workplaces) and could disclose harmful drinking in social drinkers. Due to the serious public health problem of alcohol use and abuse worldwide, markers of alcohol use have been sought. Both ethyl glucuronide (EtG) and phosphatidyl ethanol (PEth) appear to have high sensitivity and specificity and a time frame of detection that may elucidate alcohol use not detected by standard testing. Our aim was to assess their potential for detecting recent covert alcohol use under controlled conditions. METHODS Thirty-five forensic psychiatric inpatients in a closed ward who had committed a substance-related offense ( section sign 64 StGB), were followed for 12 months. The complete time spectrum of possible alcohol consumption was covered by the complementary use of breath and urinary ethanol (hours), urinary EtG (days), %carbohydrate-deficient transferrin (CDT)/PEth (weeks), and gamma-glutamyltranspeptidase (GGT)/mean corpuscular volume (MCV) (weeks-months). RESULTS Fourteen of the 146 urine samples examined were positive for EtG. In all EtG-positive cases, patients reported alcohol consumption of between 40 and 200 g of ethanol 12-60 hr prior to testing. Urinary and breath ethanol were positive in only one case. In the blood samples, PEth was not positive in any case and %CDT did not exceed the reference value. Isoelectric focusing showed no abnormal Tf subtypes. CONCLUSIONS The findings emphasize the diagnostic and therapeutic usefulness, specificity, and sensitivity of EtG as a marker of recent alcohol use. Such a test is needed in numerous settings, including alcohol and drug treatment (to detect lapse/relapse), in safety-sensitive work settings where use is dangerous or in other settings where use may be inappropriate (e.g., such as driving, workplace, pregnancy, or monitoring physicians or other professionals who are in recovery and working), or for testing other groups (such as children or those with medical problems) where alcohol use would be unhealthy or unsafe. The health, social and socioeconomic benefits arising from the future use of these markers is hard to overestimate.

On sensitivity, specificity, and the influence of various parameters on ethyl glucuronide levels in urine-results from the WHO/ISBRA Study

BACKGROUND Ethyl glucuronide (EtG), a direct ethanol metabolite, seems to meet the need for a sensitive and specific marker for monitoring recent alcohol consumption in different settings. Our aim was to study sensitivity, specificity, and the influence of various parameters on EtG levels in urine. PATIENTS AND METHODS Urine samples for a total of 453 patients (373 male, 80 female) were statistically analyzed. The mean age was 37.1 years (median 36, SD 12.59), body mass index was 24.7, total ethanol consumed last month was 1817.66 g (each median), and 80 patients reported cannabis use within the last 30 days. Determination of EtG was performed with a liquid chromatography-tandem mass spectrometry method with deuterium-labeled EtG as internal standard. RESULTS For EtG in urine, a good correlation was found with other state markers and days of sobriety. In a regression analysis, age, gender, marijuana use, kidney disease, and total grams of ethanol consumed last month were the variables that significantly influenced EtG levels in contrast to race, smoking, body mass index, cirrhosis of liver, age began drinking regularly, packs of cigarettes smoked last month, and total body water. Furthermore, in a receiver operating characteristic curve analysis to distinguish between nondrinkers and individuals sober > 4 days versus individuals drinking in the recent 4 days, area under the curve was 0.834. At a cutoff of 0.145 mg/liter, sensitivity was 83.5% and specificity 68.3%. A receiver operating characteristic curve was calculated for lifetime alcohol abuse or dependence against those who had never been abusers or dependent. In this case, subjects were either never dependent or lifetime dependent, but those currently dependent were excluded. The resulting area under the curve was 0.694. At a cutoff of 0.145 mg/liter, sensitivity was 73.8% and specificity 60.3%. For those with a self-reported sobriety of less than 24 hr, the area under the curve was 0.899, sensitivity was 90.8%, and specificity was 76.5% at a cutoff of 0.435 mg/liter when we calculated nondrinkers and light drinkers against heavy drinkers and drinkers needing treatment. Cannabis-using patients showed significant differences with regard to almost all state markers when compared with nonconsuming subjects. CONCLUSIONS Age, gender, marijuana use, kidney disease, and total grams of ethanol consumed last month should be taken into consideration when interpreting results of EtG in urine. Sensitivity and specificity seem promising. Cannabis use can be regarded as an indicator for other serious mental problems in alcohol-using subjects.

Phosphatidylethanol in Human Organs and Blood: A Study on Autopsy Material and Influences by Storage Conditions.

OBJECTIVE Phosphatidylethanol (PEth) is an abnormal phospholipid that is formed and accumulated in mammalian cells that have been exposed to ethanol. PEth has been proposed as a marker of ethanol abuse. This study was conducted to investigate the concentration of PEth in blood and organs obtained during the autopsy of alcoholics. In addition, we performed experiments on rat tissues and human blood to evaluate the effect of various storage conditions on PEth concentrations. METHODS Human tissues and blood from alcoholics and controls were obtained at autopsy and frozen at -20 degrees C until extraction. Blood from healthy donors was incubated with ethanol for 24 hr and thereafter either extracted directly or stored at room temperature, stored at 4 degrees C, frozen at -20 degrees C, or frozen in liquid nitrogen and stored at -80 degrees C before extraction. Rats were given intraperitoneal injections of ethanol and then killed, either while still intoxicated or when sober. Rat organs were homogenized and extracted directly, after a period of storage, and/or after freezing at -20 degrees C. PEth concentration was analyzed using HPLC and verified by mass spectrometry. RESULTS In all rat organs studied, PEth was formed during freezing at -20 degrees C with ethanol present. PEth concentrations of 9 to 205 mumol/liter were observed in the blood obtained at autopsy. The highest value was found in the case with the highest blood alcohol concentration (114 mmol/liter) at the time of death. In the experiments on human blood stored with ethanol present, PEth concentrations were not affected after 72 hr at 4 degrees C or after freezing in liquid nitrogen and storage at -80 degrees C for up to 144 hr but were slightly elevated after 24 hr at room temperature and at -20 degrees C. PEth was found in all organs obtained from the cadavers of alcoholics. Storage of organs at 4 degrees C for 24 hr with ethanol present had no effect on the PEth concentration. The PEth concentration was unaffected when no ethanol was present at the time of freezing. CONCLUSIONS The rat experiments indicated that the very high PEth concentrations found in the organs of the alcoholics were probably largely formed while the organs were frozen at -20 degrees C. Our data suggest that tissue material from bodies that were exposed to ethanol must be stored properly to obtain reliable results from subsequent analysis for PEth. Tissue should not be frozen at -20 degrees C but instead stored refrigerated until extraction, preferably within hours of autopsy, or frozen in liquid nitrogen and stored at -80 degrees C. Blood samples that contain ethanol can be stored refrigerated for up to 72 hr or frozen in liquid nitrogen and stored at -80 degrees C without affecting PEth levels.

Phosphatidylethanol: normalization during detoxification, gender aspects and correlation with other biomarkers and self-reports.

Phosphatidylethanol (PEth) is a direct ethanol metabolite, and has recently attracted attention as biomarker of ethanol intake. The aims of the current study are: (1) to characterize the normalization time of PEth in larger samples than previously conducted; (2) to elucidate potential gender differences; and (3) to report the correlation of PEth with other biomarkers and self‐reported alcohol consumption. Fifty‐seven alcohol‐dependent patients (ICD 10 F 10.25; 9 females, 48 males) entering medical detoxification at three study sites were enrolled. The study sample was comprised of 48 males and 9 females, with mean age 43.5. Mean gamma glutamyl transpeptidase (GGT) was 209.61 U/l, average mean corpuscular volume (MCV) was 97.35 fl, mean carbohydrate deficient transferrin (%CDT) was 8.68, and mean total ethanol intake in the last 7 days was 1653 g. PEth was measured in heparinized whole blood with a high‐pressure liquid chromatography method, while GGT, MCV and %CDT were measured using routine methods. PEth levels at day 1 of detoxification ranged between 0.63 and 26.95 µmol/l (6.22 mean, 4.70 median, SD 4.97). There were no false negatives at day 1. Sensitivities for the other biomarkers were 40.4% for MCV, 73.1% for GGT and 69.2% for %CDT, respectively. No gender differences were found for PEth levels at any time point. Our data suggest that PEth is (1) a suitable intermediate term marker of ethanol intake in both sexes; and (2) sensitivity is extraordinary high in alcohol dependent patients. The results add further evidence to the data that suggest that PEth has potential as a candidate for a sensitive and specific biomarker, which reflects longer‐lasting intake of higher amounts of alcohol and seemingly has the above mentioned certain advantages over traditional biomarkers.

Ethyl glucuronide-a biological marker for recent alcohol consumption.

Ethyl glucuronide (EtG) is a non‐volatile, water‐soluble metabolite of ethanol, with a high storage stability. It can be detected in body fluids, tissues, sweat and hair for an extended time period after the elimination of ethanol from the body. EtG closes the gap between short‐term markers for alcohol consumption such as ethanol or methanol and long‐term markers for alcohol misuse such as GGT, MCV and CDT. Due to its specific time‐frame of detection and its high sensitivity and specificity, EtG is a promising marker for alcohol consumption and for relapse control that enables the therapist to intervene at an early stage of relapsing behaviour. The aim of this review is to give an overview of analytical techniques for the detection of EtG, its clinical use and remaining questions.

Phosphatidylethanol (PEth) as a Biomarker of Alcohol Consumption in HIV-Positive Patients in Sub-Saharan Africa

BACKGROUND Alcohol is heavily consumed in sub-Saharan Africa and affects HIV transmission and treatment and is difficult to measure. Our goal was to examine the test characteristics of a direct metabolite of alcohol consumption, phosphatidylethanol (PEth). METHODS Persons infected with HIV were recruited from a large HIV clinic in southwestern Uganda. We conducted surveys and breath alcohol concentration (BRAC) testing at 21 daily home or drinking establishment visits, and blood was collected on day 21 (n = 77). PEth in whole blood was compared with prior 7-, 14-, and 21-day alcohol consumption. RESULTS (i) The receiver operator characteristic area under the curve (ROC-AUC) was highest for PEth versus any consumption over the prior 21 days (0.92; 95% confidence interval [CI]: 0.86 to 0.97). The sensitivity for any detectable PEth was 88.0% (95% CI: 76.0 to 95.6) and the specificity was 88.5% (95% CI: 69.8 to 97.6). (ii) The ROC-AUC of PEth versus any 21-day alcohol consumption did not vary with age, body mass index, CD4 cell count, hepatitis B virus infection, and antiretroviral therapy status, but was higher for men compared with women (p = 0.03). (iii) PEth measurements were correlated with several measures of alcohol consumption, including number of drinking days in the prior 21 days (Spearman r = 0.74, p < 0.001) and BRAC (r = 0.75, p < 0.001). CONCLUSIONS The data add support to the body of evidence for PEth as a useful marker of alcohol consumption with high ROC-AUC, sensitivity, and specificity. Future studies should further address the period and level of alcohol consumption for which PEth is detectable.