Markers Of Ethanol Intake Research Articles

N-Acetyltaurine as a novel urinary ethanol marker in a drinking study.

The forensic utility of N-acetyltaurine (NAcT) in urine as a marker for ethanol intake was examined. A HILIC-based liquid chromatography method for the mass spectrometric determination of NAcT, taurine, and creatinine in urine was developed and validated to investigate NAcT formation and elimination in a drinking study. Thereby, eight subjects ingested 0.66 to 0.84g/kg alcohol to reach a blood alcohol concentration (BAC) of 0.8g/kg. Blood and urine were taken every 1.5-2h, during the first 8h. NAcT and taurine levels were measured and corrected for the urine's dilution by normalization to a creatinine concentration of 1mg/mL. For the determination of NAcT and taurine, uncorrected lower limits of quantitation (LLOQs) were at 0.05μg/mL of urine. In the drinking study, NAcT proved to be an endogenous compound, which is present at a range of 1.0 to 2.3μg/mL in urine of alcohol-abstinent subjects. Maximum NAcT concentrations were reached in samples taken 3 to 6h after the start of drinking, whereby an upregulation in N-acetyltaurine could be found for all the subjects. The mean peak concentrations (c̅ max) of 14 ± 2.6μg/mL (range 9-17.5μg/mL) were reached. Within 24h, the NAcT levels declined to endogenous concentrations. The detectability of NAcT was found to be slightly shifted compared to BAC: When BAC was not detectable anymore, NAcT levels were still elevated. After 24h, when ethyl glucuronide (EtG) and ethyl sulfate (EtS) were still detectable, NAcT concentrations showed endogenous levels again. Positive NAcT results can be used as an indicator for recent alcohol consumption. A direct relationship between NAcT and taurine concentrations could not be found. Graphical abstract N-acetyltaurine concentrations for eight subjects during the first 24 h after an alcohol consumption of 0.8 g/kg.

Ethyl arachidonate is the predominant fatty acid ethyl ester in the brains of alcohol-intoxicated subjects at autopsy.

The role of fatty acid ethyl esters (FAEE), the nonoxidative ethanol metabolites, as mediators of alcohol-induced organ damage is increasingly being recognized. FAEE are detectable in the blood and in liver and adipose tissue after ethanol ingestion, and on that basis, FAEE can be used as markers of ethanol intake. In this study, 10 samples of human brain were collected at autopsy at the Massachusetts Medical Examiner's Office and analyzed for FAEE. FAEE were isolated and quantified as mass per gram of wet weight. The blood ethanol level was also obtained in each case along with the other drugs detected in routine postmortem toxicology screening tests. Ethyl arachidonate was the predominant FAEE species in the brain, representing up to 77.4% of total FAEE in the brain. The percent age of ethyl arachidonate of the total FAEE in the brain was significantly higher than what has been found in all other organs and tissues previously analyzed. Linoleate, the precursor of arachidonate, was a poor substrate for FAEE synthesis, as the percentage of ethyl linoleate of the total FAEE content was extremely low. Thus, this reflects preferred incorporation of arachidonate into newly synthesized FAEE in the brain. Since arachidonate is derived from linoleate, which is depleted in FAEE while arachidonate is enriched, the synthesis of FAEE may be linked to the desaturation and elongation of linoleate to arachidonate.

Fatty acid ethyl esters. Ethanol metabolites that reflect ethanol intake.

Fatty acid ethyl esters (FAEEs) are nonoxidative ethanol metabolites that have been implicated as mediators of alcohol-induced organ damage. FAEEs are detectable in the blood after ethanol ingestion, and on that basis represent markers of ethanol intake. FAEEs have also been quantitated in human liver and adipose tissue and have been shown to be postmortem markers of premortem ethanol intake. A substantial difference in FAEE concentration was found in liver and adipose tissue of patients with detectable blood ethanol at the time of autopsy vs those with no detectable blood ethanol, who were either chronic alcoholics or social drinkers. Most currently available diagnostic markers for chronic alcoholism have limited clinical utility. Data in this report demonstrate that the amount or type of FAEEs can be used to differentiate a chronic alcoholic from an episodic heavy drinker (binage drinker) at or near peak blood ethanol concentrations and approximately 24 hours after discontinuation of ethanol. Thus, FAEEs are markers of ethanol intake in blood and tissues and can be useful in distinguishing chronic alcoholics from binge drinkers.

Fatty acid ethyl esters: toxic non-oxidative metabolites of ethanol and markers of ethanol intake.

This report describes the biochemistry of fatty acid ethyl esters (FAEE), non-oxidative metabolites of ethanol, and their clinical significance. We review information regarding the enzymes responsible for FAEE synthesis and degradation, and the mechanisms involved with the intracellular and extracellular transport of FAEE and FAEE-mediated cytotoxicity. Also a summary of reports on the emerging clinical significance and diagnostic utility of FAEE as short and long-term markers of alcohol consumption, and the methodological aspects of FAEE assessment is included.

Fatty acid ethyl esters: recent observations

Fatty acid ethyl esters (FAEE), esterification products of fatty acids and ethanol, have been shown to be mediators of ethanol-induced cell injury and their presence in the blood and tissues is a marker of ethanol intake. Recently, it has been shown that FAEE are produced within seconds of infusion of ethanol into the heart, when using a protocol similar to that used for myocardial ablation. This raises the possibility that the mechanism for the death of myocytes in cardiac ablation involves the generation of toxic FAEE. It has also been recently demonstrated that chronic alcoholics have a high concentration of a specific FAEE species—ethyl oleate. The use of the serum ethyl oleate concentration may be helpful in differentiating binge drinkers from chronic alcoholics.

Pathogenesis of Alcoholic Liver Disease–Recent Advances

The article summarizes the proceedings of a symposium on recent advances in research on the pathogenesis of alcoholic liver disease at the 2001 RSA meeting in Montreal, Canada. The chairs were Amin A. Nanji and Samuel W. French. The presentations were (1) Role of inflammatory mediators in alcoholic liver injury by Amin A. Nanji, (2) Role of endotoxin, lipopolysaccharide binding protein, CD14 and Toll receptors in alcoholic liver injury by Grace Su, (3) Fatty acid ethyl esters: toxicity, metabolism and markers of ethanol intake by Michael Laposata, and (4) Cyclic changes in gene expression when rats are fed alcohol at a constant rate by Samuel W. French.

Liver and Adipose Tissue Fatty Acid Ethyl Esters Obtained at Autopsy Are Postmortem Markers for Premortem Ethanol Intake

Fatty acid ethyl esters (FAEEs) are nonoxidative ethanol metabolites that have been implicated as mediators of alcohol-induced organ damage. FAEEs are detectable in the blood after ethanol ingestion, and on that basis have been proposed as markers of ethanol intake. Because blood is not always available at autopsy, in this study we quantified FAEEs in human liver and adipose tissue as potential postmortem markers of premortem ethanol intake. Twenty-four sets of samples were collected at the Massachusetts State Medical Examiner's Office, and 7 sets of samples were obtained from the Pathology Department of Massachusetts General Hospital. Samples of liver and adipose tissue were collected at autopsy, and FAEEs were isolated and quantified from these organs as mass per gram of wet weight. Postmortem analysis of blood involved assessment for ethanol and other drugs. The study shows a substantial difference in FAEE concentrations in liver and adipose tissue of patients with detectable blood ethanol at the time of autopsy vs those with no detectable blood ethanol, who were either chronic alcoholics or social drinkers. In addition, a specific FAEE, ethyl arachidonate, was found at concentrations >200 pmol/g almost exclusively in the liver and adipose tissue of individuals with detectable blood ethanol at the time of death, providing an additional FAEE-related marker for prior ethanol intake. The mass of FAEEs in liver and adipose tissue and the presence of ethyl arachidonate can serve as postmortem markers of premortem ethanol intake when no blood sample can be obtained.

The Synthesis and Secretion of Fatty Acid Ethyl Esters From HepG2 Cells Are Stimulated by Lipoproteins and Albumin

Background: Fatty acid ethyl esters (FAEEs) are nonoxidative metabolites of ethanol produced by the esterification of fatty acids and ethanol. FAEEs have been implicated as mediators of ethanol-induced organ damage in vivo and in vitro. They are detectable in the blood and in many organs after ethanol ingestion, and on this basis they are useful markers of ethanol intake in living patients as well as subjects at autopsy. FAEEs found in human plasma after ethanol ingestion bind to lipoproteins and albumin. Methods: In this study, we used a hepatoblastoma cell model (HepG2) to determine if lipoproteins or albumin stimulates the synthesis and/or secretion of FAEEs from HepG2 cells. Because FAEEs have been shown to decrease HepG2 cellular proliferation and protein synthesis, their removal from cells potentially could reestablish normal cell activity. HepG2 cells were incubated with 100 mM ethanol and 6 nM 3H oleic acid to generate 3H-FAEEs within the cells. Dose response and time course studies were performed by using low density lipoproteins, high density lipoproteins, and albumin as FAEE acceptors. Results: The results indicate that FAEEs are extracted efficiently by each of these FAEE carriers and that FAEE synthesis also is stimulated by the addition of FAEE carriers to the extracellular medium. Conclusion: These observations indicate that lipoproteins and albumin can extract ethyl esters from HepG2 cells and thereby may limit alcohol-induced liver damage.

The Synthesis and Secretion of Fatty Acid Ethyl Esters From HepG2 Cells Are Stimulated by Lipoproteins and Albumin

Fatty acid ethyl esters (FAEEs) are nonoxidative metabolites of ethanol produced by the esterification of fatty acids and ethanol. FAEEs have been implicated as mediators of ethanol-induced organ damage in vivo and in vitro. They are detectable in the blood and in many organs after ethanol ingestion, and on this basis they are useful markers of ethanol intake in living patients as well as subjects at autopsy. FAEEs found in human plasma after ethanol ingestion bind to lipoproteins and albumin. In this study, we used a hepatoblastoma cell model (HepG2) to determine if lipoproteins or albumin stimulates the synthesis and/or secretion of FAEEs from HepG2 cells. Because FAEEs have been shown to decrease HepG2 cellular proliferation and protein synthesis, their removal from cells potentially could reestablish normal cell activity. HepG2 cells were incubated with 100 mM ethanol and 6 nM 3H oleic acid to generate 3H-FAEEs within the cells. Dose response and time course studies were performed by using low density lipoproteins, high density lipoproteins, and albumin as FAEE acceptors. The results indicate that FAEEs are extracted efficiently by each of these FAEE carriers and that FAEE synthesis also is stimulated by the addition of FAEE carriers to the extracellular medium. These observations indicate that lipoproteins and albumin can extract ethyl esters from HepG2 cells and thereby may limit alcohol-induced liver damage.

Effect of Specimen Anticoagulant and Storage on Measurement of Serum and Plasma Fatty Acid Ethyl Ester Concentrations.

Fatty acid ethyl esters (FAEEs) are cytotoxic nonoxidative metabolites of ethanol that are produced by esterification of alcohol and fatty acids (1)(2)(3). After ethanol intake, FAEEs are found mainly in the liver, pancreas, heart, and brain, which are the major organs adversely affected by ethanol intake (4)(5). FAEEs are detectable in the blood for up to 24 h after consumption of ethanol to at least 600 mg/L (6)(7)(8), making the presence of FAEEs in blood a useful marker for ethanol intake. This investigation presents the results of studies on the effects of collection tube, storage time, and storage temperature on FAEE concentrations in blood, with the goal of creating a reliable clinical assay for serum and plasma FAEE quantification. Four volunteers participated in this study. Each subject was given a weight-adjusted amount of 100-proof vodka mixed with fruit juice in a 1:3 ratio (8). The vodka-juice beverage was divided into nine equal aliquots, which were administered every 10 min over a 90-min time period. After the last aliquot was drunk, blood samples were collected in four different 5-mL Vacutainer Tubes from each subject to assess the influence of the collection tube …

Comparison of the formation of proteins modified by direct and indirect ethanol metabolites in the liver and blood of rats fed the Lieber-DeCarli liquid diet.

It has been proposed that proteins modified by ethanol metabolites, such as acetaldehyde (AcH) or alpha-hydroxyethyl radicals (HER) may be an important step in the aetiology of alcoholic liver disease. Furthermore, it has also been suggested that these modified proteins may act as a marker of ethanol intake. In this study, we have measured the generation of various types of modified proteins in the liver and blood of ethanol-fed rats. Multiple types of protein modification were observed in the livers of the ethanol-fed rats. In each case, the level of modification increased over the first 6 weeks of ethanol feeding, but reached a plateau by 10 weeks. In contrast to the liver, elevated levels of proteins modified by malondialdehyde were not seen in the plasma of ethanol-fed animals, whereas elevated levels of modification due to AcH and HER were observed. In haemolysates from these animals, only modification due to AcH was seen. Further investigation of the modification of plasma proteins showed that albumin, a protein produced in the liver, carried all the types of modification investigated, whereas immunoglobulin G, a protein derived from an extra-hepatic source, only carried modifications due to acetaldehyde. This study demonstrates for the first time that modification of plasma proteins by ethanol metabolites can occur at both intra- and extra-hepatic sites.

Assessment of ethanol intake. Current tests and new assays on the horizon.

The number of tests used for the detection of ethanol ingestion is increasing. The field is rapidly moving beyond ethanol alone as a marker of ethanol intake. The combined measurement of carbohydrate deficient transferrin, FAEEs, 5-HTOL/5-HIAA, acetaldehyde adducts, and phosphatidylethanol may one day be used to approximate the time and amount of ethanol ingestion. The ultimate configuration of a panel of tests for monitoring ethanol intake awaits the results of studies that identify the clinical usefulness of each marker.

Increased gamma-glutamyltransferase in hypertriglyceridaemia: the value of carbohydrate-deficient transferrin measurement.

Excess alcohol consumption is a common cause of hypertriglyceridaemia.'.2 It has, therefore, been recommended that hypertriglyceridaemic patients be screened for elevated serum y glutamyltransferase activity (yGT), a marker of ethanol intake.2,3 It has also been suggested that hypertriglyceridaemia itself may be associated with increased yGT.435 This, nevertheless, is controversial, first, because subjects often under-report their alcohol intakeh and, secondly, because of the limited sensitivity and specificity of routine laboratory markers of alcohol intake, including Y C T . ~ Measurement of carbohydrate-deficient transferrin (CDT) has provided a sensitive and specific marker of chronic elevated alcohol ingestion.h Daily intake in excess of 60g of alcohol for longer than 2 weeks results in elevated CDT levels. To assess whether elevated yGT activity in hypertriglyceridaemic patients was due to undisclosed excess ethanol ingestion, we measured CDT in hypertriglyceridaemic patients with and without raised yCT activity who reported an alcohol intake of less than 4 units (40g) per day.

Fatty acid ethyl esters in the blood as markers for ethanol intake

Fatty acid ethyl esters in the blood as markers for ethanol intake

Validated fluorimetric HPLC analysis of acetaldehyde in hemoglobin fractions separated by cation exchange chromatography: three new peaks associated with acetaldehyde.

Stable hemoglobin-acetaldehyde adducts present in hemoglobin fractions separated by polyaspartic acid cation exchange chromatography were quantified by fluorimetric HPLC. The fluorescent species eluted from the HPLC was confirmed by mass spectrometry to be consistent with the expected product from reaction of acetaldehyde, 1,3-cyclohexanedione (CHD), and ammonium ion. Hemolysate (2.2 mM hemoglobin) was incubated in equivalent volumes of either phosphate-buffered saline or 5 mM acetaldehyde at 37 degrees C for 30 min and washed three times with H2O to remove free acetaldehyde and labile adducts before the injection of 14.7 mg hemoglobin onto the cation exchange column. Fluorimetric HPLC analysis of hemolysate samples either with or without in vitro reaction with acetaldehyde revealed that most acetaldehyde resides in the hemoglobin A0 fraction. The reaction with acetaldehyde in vitro resulted in a significant increase in fast-eluting minor hemoglobin species on cation exchange chromatography concomitant with increased acetaldehyde in the HbA1a+b, HbA1c, and HbA1-AcH fractions. We report three new cation exchange chromatographic peaks after reaction with acetaldehyde: HbA1-AcH-3, HbA1c-1, and HbA0-1. Each new peak was found to associate with a significant quantity of CHD-reactive acetaldehyde. These experiments provide additional evidence that stable adducts form between acetaldehyde and hemoglobin and that these adducts occur in multiple hemoglobin species separated by cation exchange chromatography. Further characterization and structural assignment of these species are warranted in view of their potential utility as markers for ethanol intake.

Comparative evaluation of the clinical utility of three markers of ethanol intake: the effect of gender.

We evaluated three markers of ethanol intake [whole blood associated acetaldehyde (WBAA), serum beta-hexosaminidase, and gamma-glutamyl transpeptidase (GGT)] in four groups of subjects: teetotalers (n = 104), random insurance applicants or "normals" (n = 1,010), subjects enrolling in an alcohol treatment program or "alcoholics" (n = 31), and subjects attending outpatient drug/alcohol treatment follow-up clinics (n = 128). Significant differences (p < 0.004 for each assay and each comparison) were found in the mean values between teetotalers and normals and normals and alcoholics. Male teetotalers and normals had significantly (p < 0.002) higher levels of WBAA than females of the same group. Male normals had significantly higher levels of GGT than females (p < 0.001). GGT increased with age in the normal population into the fifth decade and decreased thereafter. WBAA was the most sensitive assay with 97% of alcoholics having values above the 99th percentile for the teetotaler population (vs. 66% for serum beta-hexosaminidase and 70% for GGT). None of the alcoholic subjects had values for all three assays below the 99th percentile for teetotalers compared with 21% of those in follow-up and 72% of normals. We conclude that WBAA appears to be the best of the three markers studied and that measurement of multiple markers for ethanol use appears clinically useful and incremental.

A Comparative Blinded Study in Miniature Swine of Whole Blood‐, Hemoglobin‐, Platelet‐, Plasma‐, and Lymphocyte‐Associated Acetaldehyde as Markers for Ethanol Intake

Blood samples were obtained from miniature swine maintained on 0, 2, or 6 g/kg/24 hr ethanol for 8 months (N = 6 in each group). Samples from drinking pigs were taken after 8 hr of ethanol abstinence and all were coded and sent for "blinded" analysis. A fluorigenic high performance liquid chromatographic assay was used to quantify whole blood-associated acetaldehyde, hemoglobin-associated acetaldehyde, plasma-associated acetaldehyde, platelet-associated acetaldehyde, and lymphocyte-associated acetaldehyde. Detectable levels of acetaldehyde were found in each sample in both drinking and nondrinking pigs. Analysis of whole blood-associated acetaldehyde was most discriminatory in distinguishing nondrinking from drinking pigs (mean 21.4 +/- 1.0 microM for nondrinkers vs. 24.6 +/- 1.5 SD for the group consuming 2 g/kg ethanol, p = 0.001). Measurements of hemoglobin-associated acetaldehyde normalized to protein concentration (250 +/- 47 nmoles/g vs. 203 +/- 33 SD, p less than 0.05 drinking vs. nondrinking pigs) and platelet-associated acetaldehyde (0.46 0.34 vs. 0.15 +/- 0.16 nmoles/3 x 10(8) platelets, p = 0.05 drinking vs. nondrinking pigs) were also useful in discriminating drinking from nondrinking animals. Analysis of plasma-associated acetaldehyde and lymphocyte-associated acetaldehyde were not useful as markers of ethanol consumption.(ABSTRACT TRUNCATED AT 250 WORDS)