Estimation of half-life with confidence interval for phosphatidylethanol (PEth) in blood based on two consecutive measurements.

Angiotensin-Converting Enzyme Inhibitors and Risk of Esophageal and Gastric Cancer: A Nested Case-Control Study

Low Alcohol Consumption as a Predictor of Higher CD4+ Cell Count in HIV-Treated Patients

The goal of this preliminary study was to evaluate the relationship between blood phosphatidylethanol (PEth) and recent drinking in patients with liver disease and hypertension. Twenty-one patients with liver disease and 21 patients with essential hypertension were recruited at an academic medical center. Alcohol consumption was estimated using validated self-report methods, and blood PEth was measured by HPLC-MS/MS at a contracted laboratory. Nonparametric comparisons were made between abstainers/light drinkers, moderate drinkers consuming between 1 and 3 drinks per day, and those drinking above this level. Regression methods were used to estimate the effects of liver disease, gender, and age on the relationship between PEth and alcohol use, and to estimate the strength of the linear relationship between PEth and drinking. PEth differed significantly between the three drinking groups (P < 0.001). The relationship between PEth and alcohol did not differ between hypertension and liver disease patients (P = 0.696), nor by gender and age. While there was substantial variability between subjects in the PEth concentration given a similar level of reported drinking, the amount of ethanol consumed was strongly associated with the PEth concentration (P < 0.001). Results support PEth measurement by HPLC-MS/MS as a promising marker of past 1- to 2-week moderate to heavy alcohol consumption in patients with and without liver disease. PEth appears useful for differentiating abstinence or light drinking from moderate to heavy consumption, but may have limited utility for differentiating moderate from heavy alcohol use.

Phosphatidylethanol (PEth) forms in erythrocyte membranes after alcohol consumption, offering a persisting biomarker, that is measurable in whole blood, washed erythrocytes and dried blood spots. For a predominantly erythrocyte-restricted analyte, erythrocyte concentrations seem to have most validity in patients who are anemic through alcoholism or other pathologies, despite preparation increasing assay complexity. Differences in specimen preparation alter PEth concentrations for the same patient, meaning that criteria for interpreting PEth results should relate to specimen type, presenting a barrier to achieving harmonization. We therefore tested whether erythrocyte PEth might be validly calculated by hematocrit correction of a whole blood PEth measurement. PEth testing primarily serves to distinguish drinkers from non-drinkers. In choosing between specimen types, it is important to compare their utility in separating those two groups. We therefore processed 281 blood samples from 17 non-drinkers and 61 drinkers, to prepare matched whole blood and washed erythrocyte specimens. These were assayed by liquid chromatography-tandem mass spectrometry and compared in identifying alcohol consumption. The erythrocyte PEth concentration in the whole blood specimens was also calculated by correcting whole blood concentration by the specimen's hematocrit, as an alternative to prepare washed erythrocytes. The hematocrit-corrected erythrocyte concentrations were included in these comparisons. Predictably, this work found that sensitivity was consistently better at the lower cut-off of 8 µg/L than at 20 µg/L. Sensitivities were also higher for washed erythrocytes than whole blood, explained by the lower erythrocyte mass in the same volume of whole blood. Hematocrit-corrected whole blood PEth concentrations correlated with erythrocyte concentrations, except for the four highest values, which did not influence comparative sensitivity. Specificity was 100% for washed erythrocytes, whole blood and hematocrit-corrected whole blood at either cut-off because non-drinkers had undetectable PEth. We conclude that hematocrit correction of whole blood PEth concentrations theoretically provides an alternative to the preparation of washed erythrocytes.

Introduction There is a strong need for a reliable marker of harmful alcohol consumption to identify injured patients that can benefit from alcohol interventions, and blood phosphatidyl ethanol (PEth) has not previously been tested on this population. This study aims to compare the performance of blood PEth concentration, blood alcohol concentration (BAC) and the Alcohol Use Disorders Identification Test Consumption (AUDIT-C) for the screening of alcohol misuse in trauma patients. Methods Prospective cross-sectional study of 238 adult patients presenting in the emergency department with any type of trauma. PEth concentration was determined in whole blood by high-performance liquid chromatography with tandem mass spectrometry. Consent, AUDIT-C score and demographic data were obtained. Results The sample consisted of majority male (67.6%), single (46.2%) and employed (66%) patients. The most common type of trauma was traffic collision (63.9%). The mean age was 41.7 years. We found a significant correlation between PEth levels with AUDIT-C score (Spearman’s r = 0.654; p < .0001). PEth had an area under the ROC curve of 0.885 to detect hazardous alcohol consumption (AUDIT-C score ≥ 6) and PEth ≥23.9 ng/mL cutoff point provided 91.2% of sensitivity and 78.4% of specificity. Twelve patients reported alcohol abstinence, but had quantifiable levels of PEth. Conclusions PEth levels and AUDIT-C score had a moderate correlation in our population. PEth was useful to identify 12 cases of underreporting of alcohol consumption habits. PEth shows promising results, but more research is needed to identify the best screening tool for alcohol misuse in trauma patients.

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. 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. 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. 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.

ABSTRACTBackgroundPhosphatidylethanol 16:0/18:1 (PEth) is a widely used alcohol marker, but how sex, body size and composition and kidney function affect the dose–response relationship between alcohol consumption and PEth is not fully understood.ObjectiveThe aim of the study was to investigate whether sex, age, body size and composition and kidney function influence the estimate of alcohol consumed at a given PEth concentration.MethodsThis is a longitudinal population‐based cohort study including 24 902 participants from the Trøndelag Health Study (HUNT4). Associations were assessed using a regression model.FindingsWhen analysed together, sex and soft lean mass were statistically significant independent predictors of alcohol consumption at a given PEth concentration. To achieve the same PEth concentration as a female, a male with identical body composition could on average consume 21% more alcohol, according to our model. Independent of sex, a person with 10% higher soft lean mass could consume 3.9% more alcohol to achieve the same PEth concentration. When soft lean mass was excluded, height and body weight became significant predictors.ConclusionsSex, body composition and body size may have an impact on the dose–response relationship between alcohol consumption and PEth concentration. Thus, these factors will contribute to the uncertainty when estimating ethanol intake from a given PEth concentration.

To estimate the prevalence of alcohol consumption and binge drinking during pregnancy among the general population in the World Health Organization (WHO) African Region, by country. First, a comprehensive systematic literature search was performed to identify all published and unpublished studies. Then, several meta-analyses, assuming a random-effects model, were conducted to estimate the prevalence of alcohol consumption and binge drinking during pregnancy among the general population for countries in the WHO African Region with two or more studies available. Lastly, for countries with less than two studies or no known data predictions were obtained using regression modelling. The estimated prevalence of alcohol consumption during pregnancy among the general population ranged from 2.2% (95% confidence interval [CI]: 1.6-2.8%; Equatorial Guinea) to 12.6% (95% CI: 9.9-15.4%; Cameroon) in Central Africa, 3.4% (95% CI: 2.6-4.3%; Seychelles) to 20.5% (95% CI: 16.4-24.7%; Uganda) in Eastern Africa, 5.7% (95% CI: 4.4-7.1%; Botswana) to 14.2% (95% CI: 11.1-17.3%; Namibia) in Southern Africa, 6.6% (95% CI: 5.0-8.3%; Mauritania) to 14.8% (95% CI: 11.6-17.9%; Sierra Leone) in Western Africa, and 4.3% (95% CI: 3.2-5.3%; Algeria) in Northern Africa. The high prevalence of alcohol consumption and binge drinking during pregnancy in some African countries calls for educational campaigns, screening and targeted interventions for women of childbearing age.

Direct alcohol biomarkers are of growing interest for the assessment of alcohol consumption, with particular interest in phosphatidylethanol (PEth) in recent years. PEth is only formed when alcohol is present in the body. However, there is no statement about how much the PEth concentration increases after single moderate alcohol consumption. This study was conducted to determine the increase in PEth concentrations after a single drinking event. Additionally, a new volumetric sampling device (volumetric dried blood spot cards (DBSV)) was evaluated, which was designed to simplify further sampling processes and to allow for easy self-sampling. Dried blood samples from 31 volunteers were collected before and after single alcohol consumption with a mean maximum breath alcohol concentration of 0.4 mg/L (range: 0.30-0.55 mg/L). PEth concentrations were determined after automated extraction by liquid chromatography--tandem mass spectrometry. PEth 16:0/18:1 and PEth 16:0/18:2 concentrations increased to an average of 45 ng/mL each in patients starting below 20 ng/mL (range: 25.0-57.0 ng/mL for PEth 16:0/18:1; range 26.8-62.3 ng/mL for PEth 16:0/18:2). PEth concentrations in patients starting above 20 ng/mL increased by a mean of 30 ng/mL (range: 6.2-71.3 ng/mL for PEth 16:0/18:1; range 8.8-65.3 ng/mL for PEth 16:0/18:2). In addition, the comparison of the new sampling device DBSV with a standard filter paper card (with volumetrically applied 20 µL of blood samples) yielded a close agreement for the determined PEth concentrations in 24 forensic samples and three external controls. Therefore, the sampling device DBSV proved to be suitable for the determination of PEth concentrations in blood.

Probabilistic methods are gaining in importance in aerospace engineering due to their ability to describe the behavior of the system in the presence of input value variance. A frequently employed probabilistic method is the Monte Carlo Simulation (MCS). There, a sample of random representative realizations is evaluated deterministically and their results are afterwards analyzed with statistical methods. Possible statistical results are mean, standard deviation, quantile values and correlation coefficients. Since the sample is generated randomly, the result of a MCS will differ for each repetition. Therefore, it can be regarded as a random variable. Confidence Intervals (CIs) are commonly used to quantify this variance. To gain the true CI, many repetitions of the MCS have to be conducted, which is not desirable due to limitations in time and computational power. Hence, analytical formulations or bootstrapping is used to estimate the CI. In order to reduce the variance of the result of a MCS, sampling techniques with variance reduction properties like Latin Hypercube Sampling (LHS) are commonly used. But the known methods to determine the CI do not consider this variance reduction and tend to overestimate it instead. Furthermore, it is difficult to predict the change of the CI size with increasing size of the sample. In the present work, new methods to calculate the CI are introduced. They allow a more precise CI estimation when LHS is used for a MCS. For this purpose, the system is approximated by means of a meta model. The distribution of the result value is now approximated by repeating the MCS many times. The time consuming deterministic calculations of a MCS are thus replaced with an evaluation on the meta model. These so called virtual MCS can therefore be performed in a short amount of time. The estimated distribution of the result value can be used to estimate the CI. It is, however, not sufficient to use only the meta model. The error ε, defined as the difference between the true value y and the approximated value y, must be considered as well. The generated meta model can also be used to predict the size of the CI at different sample sizes. The suggested methods were applied to two test cases. The first test case examines a structural mechanics application of a bending beam, which features low computational cost. This allows to show that the predicted sizes of the CI are sufficiently precise. The second test case covers the aerodynamic application. Therefore, an aerodynamic Computational Fluid Dynamics (CFD) analysis accounting for geometrical variations of NASA’s Rotor 37 is conducted. For this, the blade is parametrized with the in-house tool Blade2Parameter. For different sample sizes, blades are generated using this parametrization. Their geometrical variance is based on experience values. CFD calculations for these blades are performed with the commercial software NUMECA. Afterwards, the CIs for result values of interest like mechanical efficiency are evaluated with the presented methods. The suggested methods predict a narrower and thus less conservative CI.

A prospective study of the association between phosphatidylethanol concentration and risk of first-ever venous thromboembolism.

BackgroundAlcohol consumption during pregnancy is associated with major birth defects and developmental disabilities. Questionnaires concerning alcohol consumption during pregnancy underestimate alcohol use while the use of a reliable and objective biomarker for alcohol consumption enables more accurate screening. Phosphatidylethanol can detect low levels of alcohol consumption in the previous two weeks. In this study we aimed to biochemically assess the prevalence of alcohol consumption during early pregnancy using phosphatidylethanol in blood and compare this with self-reported alcohol consumption.MethodsTo evaluate biochemically assessed prevalence of alcohol consumption during early pregnancy using phosphatidylethanol levels, we conducted a prospective, cross-sectional, single center study in the largest tertiary hospital of the Netherlands. All adult pregnant women who were under the care of the obstetric department of the Erasmus MC and who underwent routine blood testing at a gestational age of less than 15 weeks were eligible. No specified informed consent was needed.ResultsThe study was conducted between September 2016 and October 2017. In total, we received 1,002 residual samples of 992 women. After applying in- and exclusion criteria we analyzed 684 samples. Mean gestational age of all included women was 10.3 weeks (SD 1.9). Of these women, 36 (5.3 %) tested positive for phosphatidylethanol, indicating alcohol consumption in the previous two weeks. Of women with a positive phosphatidylethanol test, 89 % (n = 32) did not express alcohol consumption to their obstetric care provider.ConclusionsOne in nineteen women consumed alcohol during early pregnancy with a high percentage not reporting this use to their obstetric care provider. Questioning alcohol consumption by an obstetric care provider did not successfully identify (hazardous) alcohol consumption. Routine screening with phosphatidylethanol in maternal blood can be of added value to identify women who consume alcohol during pregnancy.

The identification and quantification of biomarkers in clinical blood samples play a pivotal role in personalized medicine, enabling disease monitoring and therapeutic assessment. Protein phosphorylation, a key post-translational modification, reflects cellular signaling in immune or tumor cells. To exploit this, we developed a platform detecting phosphorylation states of key signaling proteins from clinical blood samples via flow cytometry. Our approach analyzes phosphorylated STAT5 (pSTAT5), ERK1/2 (pERK1/2), and p38 (p-p38), vital components in immune regulation and cancer progression. A major challenge in the analysis of phosphorylated proteins is the rapid degradation or change in phosphorylation state ex vivo, which requires immediate stabilization of samples after collection. To solve this problem, we use Smart Tubes, which enable rapid fixation of clinical blood samples and preserve the phosphorylation state upon collection, ensuring signal integrity and minimizing variability. Sample collection with Smart Tubes requires no technical equipment and can be easily implemented and standardized at any clinical trial site. Our test system employs flow cytometry, enabling the analysis of linage-specific markers (e.g., T/B cells, monocytes) and phospho-specific antibodies to quantitatively detect pSTAT5, pERK1/2, and p-p38 in peripheral blood mononuclear cell (PBMCs) subpopulations. Validation experiments with specific inhibitors demonstrate sensitivity and specificity in detecting these phosphorylated targets, even in heterogeneous cell populations within blood samples. The method allows for its application across a range of clinical conditions, from cancer immunotherapy monitoring to autoimmune disease characterization. By integrating rapid sample fixation with advanced analytical techniques, this platform addresses key challenges in analyzing phosphorylated biomarkers in clinical samples. The use of flow cytometry as the central analytical tool enables single-cell resolution, allowing detailed investigation of individual cell populations within complex samples by differential marker expression. This precision makes it possible to differentiate and study specific subpopulations of immune cells or circulating tumor cells, within a heterogeneous sample. The ability to preserve and accurately measure phosphorylation states directly in patient-derived samples provides invaluable insights into real-time cellular signaling and facilitates the optimization of therapeutic strategies targeting dysregulated signaling pathways. In summary, our system can detect phosphorylated proteins in clinical samples, with Smart Tubes ensuring sample stability and reliable biomarker analysis, advancing translational research and precision medicine. Citation Format: Valerie Oberhardt, Philipp Metzger, Carla N. Castro, Holger Weber. Detection of phosphorylated biomarkers in clinical blood samples: a flow cytometry platform utilizing smart tube fixation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4616.

Validation of a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for erythrocyte phosphatidylethanol revealing critical considerations for its use as a clinical biomarker

The present paper aims at a systematic review of the current knowledge on phosphatidylethanol (PEth) in blood as a direct marker of chronic alcohol use and abuse. In March 2012, the search through “MeSH” and “free-text” protocols in the databases Medline/PubMed, SCOPUS, Web of Science, and Ovid/Embase, combining the terms phosphatidylethanol and alcohol, provided 444 records, 58 of which fulfilled the inclusion criteria and were used to summarize the current evidence on the formation, distribution and degradation of PEth in human blood: (1), the presence and distribution of different PEth molecular species (2), the most diffused analytical methods devoted to PEth identification and quantization (3), the clinical efficiency of total PEth quantification as a marker of chronic excessive drinking (4), and the potential utility of this marker for identifying binge drinking behaviors (5). Twelve papers were included in the meta-analysis and the mean (M) and 95% confidence interval (CI) of total PEth concentrations in social drinkers (DAI ≤ 60 g/die; M = 0.288 μM; CI 0.208–0.367 μM) and heavy drinkers (DAI > 60 g/die; M = 3.897 μM; CI 2.404–5.391 μM) were calculated. The present analysis demonstrates a good clinical efficiency of PEth for detecting chronic heavy drinking.