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CORK Bibliography: Biochemical Markers of Substance Use

135 citations. January 2003 to present

Prepared: March 2009

Neumann T; Helander A; Dahl H; Holzmann T; Neuner B; Weiss-Gerlach E et al. Value of ethyl glucuronide in plasma as a biomarker for recent alcohol consumption in the emergency room. Alcohol and Alcoholism 43(4): 431-435, 2008. (29 refs.)

Aim: This emergency department (ED) study compared the value of plasma ethyl glucuronide (EtG) testing with the information about alcohol consumption obtained using the standard alcohol biomarkers gamma-glutamyltransferase (GGT) and carbohydrate-deficient transferrin (CDT) and the AUDIT questionnaire. Methods: Minimally injured and clinically non-intoxicated male patients (n = 81) admitted to an ED were screened regarding their alcohol consumption, using the computerized AUDIT questionnaire and a paper-and-pencil assessment including the type, amount and time of alcohol intake. Blood samples were collected for determination of ethanol, EtG (LC-MS) and GGT in plasma and %CDT in serum (Axis-Shield %CDT immunoassay). Results: Out of the 81 patients, 23 (28%) were positive (>= 8 points) on the AUDIT questionnaire. Only 3 (4%) showed a detectable ethanol concentration (range 0.01-0.07 g/L) but 31 (38%) showed a detectable EtG (0.16-39.5 mg/L). In four patients, EtG was detectable in plasma for > 48 h after estimated completed elimination of ethanol. EtG was not correlated with the long-term biomarkers %CDT or GGT, or the AUDIT results, but with the time since estimated completed ethanol elimination. Conclusion: EtG testing in blood was found useful in the ED as a way to detect recent drinking, even in cases of a negative ethanol test, and to confirm abstinence from alcohol. This sensitive and specific short-term biomarker provides valuable additional information about individual drinking habits and might also be helpful to identify an alcohol hangover.

Allen JP. Assessment of alcoholic patients: Advances and future challenges. IN: Galanter M, ed. Research on Alcoholism Treatment. Recent Developments in Alcoholism: Volume 16. New York: Kluwer Academic/Plenum Publishers, 2003. pp. 13-24. (25 refs.)

The assessment of alcoholic patients is discussed, with a focus on advances and future challenges. The author notes that more than 100 measures are now available for use in patient screening and diagnosing, as well as in developing individualized treatment plans. Self-report scales are the most commonly used means of screening for alcohol problems. However, screening for alcohol problems has not yet become standard health care practice. Biological tests, such as carbohydrate deficient transferrin (CDT), may aid in the initial recognition of alcohol-related problems and of relapse to drinking by patients in treatment. Treatment planning instruments are designed to assess patient characteristics that should influence the nature and setting of treatment. The author concludes that there is a current need for intense research on existing, promising scales, rather than for construction of new measures. Research is needed for the development of more comprehensive norms; development of abbreviated scales; and determination of the way measures relate to each other. Studies on the effects of gender, ethnicity, age, and collateral psychopathology on test results are also needed. Section headings in this book chapter include: (1) self-report screening measures; (2) biochemical markers; (3) self-report measures of consumption; (4) diagnostic measures; (5) treatment planning instruments; and (6) crosscutting issues.

Allen JP. Use of biomarkers of heavy drinking in health care practice. Military Medicine 168(5): 364-367, 2003. (32 refs.)

Several biochemical tests are available to screen for heavy drinking in patients. Most of these (e.g., liver function tests and macrocytic volume) rise due to adverse effects of alcohol on organs or the organ's functions. Carbohydrate-deficient transferrin, however, seems to elevate because of the direct action of alcohol or one of its metabolites. A second valuable contribution of biomarkers is identification of relapse events in recovering alcoholics that either would not be voluntarily reported or reported only much later. Finally, providing feedback to patients in treatment based on their biomarker values and changes in them during the course of treatment can enhance motivation, a key component in recovery from alcohol problems. The current article offers a clinical rationale for routine use of biomarkers of heavy drinking in health care practice and proposes recommendations on how they might be best used.

Allen JP; Anton R. Biomarkers as aids to identification of relapse in alcoholic patients. IN: Galanter M, ed. Recent Developments in Alcoholism. Volume 16: Research on Alcoholism Treatment. Methodology/Psychosocial Treatment/Selected Treatment Topics/Research Priorities. New York: Kluwer Academic, 2003. pp. 25-38. (53 refs.)

The use of biomarkers as aids to identification of relapse of alcoholic patients is discussed. Specifically, the authors note that recurrent assessment with biochemical measures can provide health care providers with useful information on a patient's drinking status. A return to some level of drinking is common among alcoholic patients during and shortly after treatment. A fundamental goal of continuing care of alcoholic patients is the early identification of initial relapses in an effort to prevent development of full-blown relapses. Future research is needed on degree of possible reactivity of biomarkers, scheduling of follow-up testing with biomarkers, and development of markers that are more accurate in screening of women and adolescents. Section headings in this book chapter include: (1) reasons that relapse should be recognized early; (2) role of biomarkers in relapse recognition; (3) desirable qualities in a biomarker of relapse; (4) biomarkers of relapse; (5) summary of findings on biomarkers of relapse; (6) future research questions; and (7) recommendations to clinicians. Design features of studies involving carbohydrate deficient transferrin as a marker of relapse to drinking and sensitivity and specificity for relapse for carbohydrate deficient transferrin (CDT), gamma glutamyl transpeptidase (GGT), and the combination of CDT and GGT are presented in table format.

Allen JP; Sillanaukee P; Strid N; Litten RZ. Biomarkers of heavy drinking. IN: Allen JP; Wilson VB, eds. Assessing Alcohol Problems: A Guide for Clinicians and Researchers. Second Edition. Rockville MD: National Institute on Alcohol Abuse and Alcoholism, 2003. pp. 37-54. (231 refs.)

While often used as screens for diagnosis of alcohol abuse or dependence, strictly speaking, biomarkers are reflections of physiological reactions to heavy drinking. Self-report screening scales, on the other hand, generally use a diagnosis of alcohol dependence as the criterion against which they are evaluated. Assessment of drinking behavior per se and severity of alcohol dependence are both important, albeit somewhat non-overlapping phenomena. This chapter addresses the following issues: criteria for selection of biomarkers, traditional biomarkers, emerging biomarkers, use of biomarkers in combination, use of biomarkers in alcohol treatment research and clinical practice, and research needs. Although the chapter focuses only on biomarkers, it is, of course, important to recognize that their use is in no way in competition with informed use of other psychometric measures. Rather, clinicians and researchers need to know how to maximize the information value of each class of measures.

Allen JP; Wilson VB, eds. Assessing Alcohol Problems: A Guide for Clinicians and Researchers. Second Edition. Rockville MD: National Institute on Alcohol Abuse and Alcoholism, 2003. (Chapter refs.)

This resource intended for clinicians and researchers is an update of an earlier (1996) volume.There are nine chapters which include a review of the assessment provess along with a quick reference to the available instruments; screening through self-report and use of biomarkers of heavy drinking; the diagnostic process and criteria for diagnosis; assessemnt in terms of consumption, assessment of adults, and assessment among adolescents with attention to both alcohol and other drug use, and as a means of informing treatment planning; and concludes with a discussion of treatment outcome.

Public Domain

Alte D; Ludemann J; Piek M; Adam C; Rose HJ; John U. Distribution and dose response of laboratory markers to alcohol consumption in a general population: Results of the study of health in Pomerania (SHIP). Journal of Studies on Alcohol 64(1): 75-82, 2003. (36 refs.)

Objective: Biomarker distributions must be well known for use as screening tools for hazardous alcohol consumption in general populations. The aim of this study was to investigate characteristics of carbohydrate-deficient transferrin (CDT) and to compare it with gamma-glutamyltransferase (GGT) and erythrocyte mean corpuscular volume (MCV) in regard to distribution in the general population; the dose response relationship between alcohol consumption and biomarkers; and the effect of gender, age, body mass index (BMI) and smoking. Method: In a cross-sectional health survey in northeast Germany (1997-2001), a sample of 7,008 men and women aged 20-79 years was drawn, following stratification by age and gender. Of this sample, 4,3 10 subjects (2,193 women) took part in the study. Alcohol consumption was evaluated by self-reports with a beverage specific quantity-frequency method. Results: Biomarker distributions differ across age and gender. The distribution of drinkers and nondrinking subjects showed considerable overlap. The association of alcohol consumption to laboratory markers is weak in the general population: strongest for GGT, followed by CDT and MCV For CDT and MCV it is weaker in women than in men. We found increasing risk of all three elevated marker values with increasing alcohol consumption and significant effects of age, gender, smoking and body mass index on the alcohol- biomarker dose response curve. Conclusions: When biomarkers are used for screening, all confounding effects have to be taken into account and adjusted normal ranges must be used. CDT shows no overall advantage over GGT. Low correlation of biomarkers with alcohol consumption, their high variability and widely spread ranges in nondrinking subjects limit the usefulness of these markers in general population settings.

Ambrozic J; Bunc M; Osredkar J; Brvar M. S100B protein in benzodiazepine overdose. Emergency Medicine Journal 25(2): 90-92, 2008. (22 refs.)

Background: Severe benzodiazepine overdose can result in coma and respiratory depression that might cause brain hypoxia, necrosis and delayed post-anoxic leucoencephalopathy with permanent neurological sequelae. The aim of this study was to assess the possible role of S100B, a structural protein of astroglial cells, as a biochemical marker of brain injury in acute benzodiazepine overdose. Methods: Serum S100B determination was performed in 38 consecutive patients admitted to the emergency department (ED) in Ljubljana with benzodiazepine overdose. The level of consciousness and respiratory insufficiency on the scene were assessed by responsiveness to a verbal stimulus and pulse oximetry. Blood samples were taken immediately after arrival at the ED and S100B concentrations were measured with a commercial immunoluminometric assay. 20 healthy sex- and age-matched volunteers formed a control group. Results: There were significant differences in S100B levels between the control group and the patients with benzodiazepine overdose according to their responsiveness to a verbal stimulus. Post hoc test results showed that S100B levels in patients with benzodiazepine overdose who were unresponsive to a verbal stimulus were significantly higher than those in patients responsive to a verbal stimulus (median 0.31 vs 0.11 mu g/l; p = 0.001). Both groups of patients with benzodiazepine overdose had significantly higher S100B levels than the control group (median 0.07 mu g/; both p = 0.001). Arterial oxygen saturation of patients with benzodiazepine overdose unresponsive to a verbal stimulus was significantly lower than in patients responsive to a verbal stimulus (median 83% vs 94%; p = 0.001). There was no significant difference in the systolic blood pressure of patients with benzodiazepine overdose responsive or unresponsive to a verbal stimulus. Conclusion: Raised levels of S100B protein are associated with depressed levels of consciousness and respiratory insufficiency in patients with benzodiazepine overdose.

Anttila P; Jarvi K; Latvala J; Romppanen J; Punnonen K; Niemela O. Biomarkers of alcohol consumption in patients classified according to the degree of liver disease severity. Scandinavian Journal of Clinical & Laboratory Investigation 65(2): 141-151, 2005

Objective. In the search for optimal biomarkers of excessive drinking, only a few studies have been conducted to compare the relationships between ethanol consumption, liver status, and various laboratory markers of ethanol-induced diseases. Material and methods. Concentrations of carbohydrate-deficient transferrin (%CDT and CDTect methods), serum sialic acid (SA), gamma-glutamyl transferase (gamma-GT), aspartate aminotransferase (ASAT), mean corpuscular volume (MCV), and a marker of fibrogenesis (PIIINP) were studied in 102 alcoholics with (n=59) or without (n=43) alcoholic liver disease. Controls were 34 healthy volunteers who were either social drinkers or abstainers. Results. Although concentrations of all markers were significantly higher in the alcoholic patients than in the healthy controls, their diagnostic characteristics showed a considerable degree of variation. The %CDT, SA, and MCV showed the strongest correlations with the amount of recent alcohol intake. The presence of liver pathology notably influenced the results of CDTect, GT, ASAT, and PIIINP. In ROC analyses, the highest rates of diagnostic accuracy for detecting hazardous drinking were reached with GT (0.94), CDT (0.86), and SA (0.85), followed by MCV (0.79) and ASAT (0.77). Upon abstinence, the estimated times for normalization varied between 10 days (CDTect) and 25 days (GT). Conclusions. Our data suggest distinct differences in the clinical characteristics of biological markers of ethanol consumption. While the overall accuracy of CDT and GT appear to be highest in the detection of problem drinking, serum SA and PIIINP measurements are of further value when the effects of liver pathology and ethanol drinking need to be differentiated.

Anttila P; Jrvi K; Latvala J; Niemel O. Method-dependent characteristics of carbohydrate-deficient transferrin measurements in the follow-up of alcoholics. Alcohol and Alcoholism 39(1): 59-63, 2004. (25 refs.)

Aims: There are only limited data comparing the diagnostic characteristics of carbohydrate-deficient transferrin (CDT) measurements in assays for excessive alcohol consumption under controlled conditions. Methods: We compared different CDT assays and the conventional laboratory markers of ethanol consumption, -glutamyl transferase (gamma-GT) aspartate aminotransferase (AST) and mean corpuscular volume (MCV) in the assessment and follow-up of 36 alcoholics (31 men, five women, mean age 44 years), who were admitted for detoxification. Detailed interviews to assess the amount of alcohol consumption were carried out for each patient. A hospital follow-up with supervised abstinence for 8 � 4 days (range 5-19 days) was carried out for 17 patients. Controls were 30 apparently healthy individuals (22 men, eight women, mean age 49 years), who had no history of hazardous drinking. Results: At the time of admission, the %CDT method, which excludes the trisialotransferrin isoform from the measurement, yielded elevated values in 69% of the patients, compared to 61% for CDTect. The corresponding sensitivities for gamma-GT, AST and MCV were 61, 56 and 47%, respectively. The self-reported alcohol consumption for a period of 1 month prior to admission showed a stronger correlation with the %CDT results (r = 0.59, P = 0.0003) than with the CDTect results (r = 0.36, P = 0.04), GT (r = 0.40, P = 0.02) or AST (r = 0.35, P = 0.05). During follow-up with supervised abstinence the mean %CDT values were found to show a slower rate to normalization (mean 14 � 4 days) than the CDT values measured with the CDTect method (mean 10 � 5 days) (P < 0.05). Conclusions: The data indicate distinct differences and method-dependent rates of normalization in CDT assays, possibly reflecting different degrees of transferrin desialylation in the alcoholics. The present findings should be considered in studies on alcohol markers for monitoring abstinence.

Bayerlein K; Hillemacher T; Reulbach U; Mugele B; Sperling W; Kornhuber J et al. Alcoholism-associated hyperhomocysteinemia and previous withdrawal seizures. Biological Psychiatry 57(12): 1590-1593, 2005. (21 refs.)

Background. Higher homocysteine levels were found in actively drinking alcoholics as well as in early abstinent patients. Furthermore, it has been shown that high homocysteine levels predicted first-onset alcohol withdrawal seizures. The aim of the present study was to determine plasma homocysteine levels in actively drinking alcoholics and patients with early abstinence in order to evaluate whether there is an additional association between elevated plasma homocysteine levels and a history of alcohol withdrawal seizures. Methods: Two groups of patients with an established diagnosis of alcohol dependence were studied, Group A comprised 56 consecutively admitted alcoholics who bad been abstinent from alcohol between 24 to 72 hours before hospitalization. Group B consisted of 144 consecutively recruited alcoholics who were admitted - acutely intoxicated - for withdrawal treatment, Furthermore, groups were divided into two subgroups: patients with and without a history of alcohol withdrawal seizures, Results: Alcoholics of GROUP B with a history of withdrawal seizures had significantly (p <.0001) higher homocysteine levels than actively drinking patients without seizures in their history, 42.0 mu mol/l (SD 26.4) versus 22.5 mu mol/l (SD 11.4). Using a logistic regression analysis, history withdrawal seizures in Group B but not in Group A patients were best predicted by a high homocysteine level at admission (Wald chi(2) = 15.5, p <.0001; odds ratio 1.11, 95% CI 1.05-1.20). Conclusions. Homocysteine levels on admission may be a useful screening method to identify actively drinking patients with a higher risk of alcohol withdrawal seizures.

Bearer CF; Stoler JM; Cook JD; Carpenter SJ. Biomarkers of alcohol use in pregnancy. Alcohol Research & Health 28(1): 38-43, 2004. (27 refs.)

To date, no laboratory test has been widely available to identify and quantify prenatal alcohol use that takes place over a protracted period. Therefore the reliance is upon the use of self-report screening instuments. The authors review biological samples available to detect drinking during pregnancy. These are summarized in a table by maternal samples, fetal samples, and newborn, with the advantages and disadvantages of each. There is also discussion of the use of tests such as liver enzyme levels, GGT or CDT which are known to be associated with longer term and/or high risk alcohol use. However, their sensitivity and specificity in pregnancy has not been established. Some additional potential markers are described (Fatty Acid Ethyl Esters) and proteinomics (the interation of alcohol and protein) which can potentially indicate levels of alcohol use.

Beck O; Helander A. 5-Hydroxytryptophol as a marker for recent alcohol intake. Addiction 98(Supplement 2): 63-72, 2003. (68 refs.)

Aims: To review the mechanism behind the alcohol-induced shift in serotonin metabolism, and the use of urinary 5-hydroxytryptophol (5-HTOL) as a biochemical marker of acute alcohol consumption. Background: The serotonin metabolite 5-HTOL is a normal, minor constituent of urine and is excreted mainly in conjugated form with glucuronic acid. The formation of 5-HTOL increases dramatically after alcohol intake, due to a metabolic interaction, and the elevated urinary excretion remains for some time (>5-15 hours depending on dose) after ethanol has been eliminated. This biochemical effect can be used for detection of recent alcohol intake. Results 5-HTOL is determined by the gas chromatography-mass spectrometry (GC-MS) or liquid chromatography and mass spectrometry (LC-MS) techniques. A new ELISA method for 5-HTOL glucuronide provides a promising clinical assay. The most robust way to use the marker is by measuring the ratio of 5-HTOL to 5-hydroxyindoleacetic acid, because this compensates for urine dilution and dietary intake of serotonin. 5-HTOL is a very sensitive and specific indicator of recent alcohol consumption and, as such, a valuable complement to self-report. In clinical use, 5-HTOL is effective for monitoring lapses into drinking during out-patient treatment and for objective evaluation of treatment efforts. Other applications include detection of high-risk patients in elective surgery, monitoring of disulfiram treatment and a method to rule out artefactual ethanol formation in forensic toxicology. 5-HTOL can also be used as a sensitive reference method for validation of self-report data in clinical alcohol research. Conclusions: An elevated urinary 5-HTOL level can serve as a sensitive and reliable marker for recent alcohol intake with a number of clinical and forensic applications.

Behera D; Uppal R; Majumdar S. Urinary levels of nicotine & cotinine in tobacco users. Indian Journal of Medical Research 118: 129-133, 2003. (28 refs.)

Background & objectives: Of the various biochemical markers used to validate the smoking status of a person, nicotine and continine are considered as good markers for both active and passive smoking. In the present study an attempt was made to estimate urinary levels of nicotine and cotinine in healthy individuals from north India using different types of tobacco to identify and validate the smoking status. Methods: Twenty four hour urine sample of 130 healthy volunteers (smokers=70, passive smokers=20, tobacco chewers=20, non smokers=20) were analyzed by high-pressure liquid chromatography (HPLC) assay. Smokers were divided into different groups, viz., cigarette, bidi and hooka smokers. Results: The mean values of nicotine (ng/ml) and cotinine (ng/ml) in urine were highest in cigarette smokers (nicotine=703.50 +/- 304.34; cotinine=2736.20 +/- 983.29), followed by hooka smokers (nicotine 548.0 +/- 103.47 and cotinine 2379.0 +/- 424.25), and bidi smokers (nicotine=268.53 +/- 97.62, cotinine=562.60 +/- 249.38). There was no correlation of nicotine or cotinine values with smoking index. In passive smokers (nicotine=109.75 +/- 22.33, cotinine=280.75 +/- 86.30) and in nonsmokers, the values were much lower (nicotine=55.00 +/- 13.71, cotinine=7.30 +/- 2.47) compared to smokers. In tobacco chewers, the values for nicotine and cotinine were 447.75 +/- 145.09 and 2178.30 +/- 334.29 respectively.Interpretation & conclusion: All forms of tobacco users had significantly higher values compared to passive smokers and nonusers. Thus, cotinine and nicotine levels in urine may be considered as good indicators to assess the exposure to tobacco in our population.

Benowitz NL; Bernert JT; Caraballo RS; Holiday DB; Wang JT. Optimal serum cotinine levels for distinguishing cigarette smokers and nonsmokers within different racial/ethnic groups in the United States between 1999 and 2004. American Journal of Epidemiology 169(2): 236-248, 2009. (19 refs.)

Cotinine, a metabolite of nicotine, is widely used to distinguish smokers from nonsmokers in epidemiologic studies and smoking-cessation clinical trials. As the magnitude of secondhand smoke exposure declines because of proportionally fewer smokers and more clean-indoor-air regulations, the optimal cotinine cutpoint with which to distinguish smokers from nonsmokers is expected to change. The authors analyzed data on 3,078 smokers and 13,078 nonsmokers from the National Health and Nutrition Examination Survey for 1999-2004. Optimal serum cotinine concentrations for discriminating smokers from nonsmokers were determined using receiver operator characteristic curve analysis. Optimal cotinine cutpoints were 3.08 ng/mL (sensitivity = 96.3%, specificity = 97.4%) and 2.99 ng/mL (sensitivity = 86.5%, specificity = 93.1%) for adults and adolescents, respectively. Among adults, optimal cutpoints differed by race/ethnicity: They were 5.92 ng/mL, 4.85 ng/mL, and 0.84 ng/mL for non-Hispanic blacks, non-Hispanic whites, and Mexican Americans, respectively. Among adolescents, cutpoints were 2.77 ng/mL, 2.95 ng/mL, and 1.18 ng/mL for non-Hispanic blacks, non-Hispanic whites, and Mexican Americans, respectively. Use of the currently accepted cutpoint of 14 ng/mL overestimates the number of nonsmokers in comparison with the proposed new overall cutpoint of 3 ng/mL or the race/ethnicity-specific cutpoints of 1-6 ng/mL.

Berggren U; Berglund K; Eriksson M; Fahlke C; Zachrisson O; Balldin J. Subnormal alpha-2-adrenoceptor-mediated sedation during 6 months of sobriety in male type 1 alcohol-dependent subjects. Alcohol and Alcoholism 38(4): 321-326, 2003. (30 refs.)

Aims: In the present study, alpha(2)-adrenoceptor function was investigated over 6 months of sobriety in eight male alcohol-dependent subjects. Methods: Subjects were investigated with repeated clonidine (CLON, 2 mug/kg body weight intravenously) challenge tests at days 1 and 7, and months 2 and 6 after the end of a period of heavy alcohol intake. CLON-induced sedation was rated at challenge tests. Mental well-being was self-reported before all challenge tests. Three challenge tests were performed at 1-week intervals in six male healthy controls. Results: Sedation was significantly lower after CLON at all time-points for the challenge tests in alcohol-dependent subjects compared with mean values for three challenge tests in controls. Three dimensions of mental well-being were negatively correlated with scores of CLON-induced sedation at month 6. Conclusions: alpha(2)-Adrenoceptor function is subnormal, as assessed by CLON-induced sedation, for at least 6 months after termination of alcohol intake. Whether this subnormal receptor function is pre-existing and possibly genetically determined or is a consequence of long-term alcohol intake must be further investigated, as should this receptor status in alcohol-dependent subjects with longer time-periods of sobriety.

Bergstrom JP; Helander A. Clinical characteristics of carbohydrate-deficient transferrin (%disialotransferrin) measured by HPLC: Sensitivity, specificity, gender effects, and relationship with other alcohol biomarkers. Alcohol and Alcoholism 43(4): 436-441, 2008. (30 refs.)

Aims: The sensitivity and specificity of the relative disialotransferrin amount (%DST), considered the primary single target for measurement of the alcohol biomarker carbohydrate-deficient transferrin (CDT), were compared with the absolute CDT amount determined by the CDTect assay and with GGT and AST. Methods: Serum samples (n = 1387) were collected within the WHO/ISBRA Study on State and Trait Markers of Alcohol Use and Dependence. The subjects had been classified as "nondrinkers" (26%), "light/moderate drinkers" (50%), or "heavy drinkers" (24%) by use of the WHO/ISBRA Interview Schedule. An HPLC candidate reference method for CDT was used to quantify individual transferrin glycoforms. Results: No gender difference in %DST was noted for nondrinkers, but light/moderate and heavy drinking males had significantly higher levels than females. Of the alcohol biomarkers examined, %DST showed the strongest correlation with self-reported alcohol intake, except for female heavy drinkers. The area under the %DST ROC curve for male (0.83) and female (0.82) heavy drinkers was significantly higher compared with CDT by CDTect (0.68) and GGT (0.69). At the 40, 60, or 80 g ethanol/day thresholds, %DST showed lower test sensitivity in women but there was no significant gender difference in overall accuracy according to ROC curve analysis. Conclusions: %DST measured by HPLC showed overall higher sensitivity for "heavy drinking" and better correlation with recent high alcohol intake, compared with the absolute CDT amount, and GGT and AST. The observation that several "light/moderate drinkers" had elevated %DST levels and some also a measurable asialotransferrin indicated misclassification with the WHO/ISBRA Interview Schedule and emphasize the limitations of self-reports of drinking.

Bergstrom JP; Helander A. Influence of alcohol use, ethnicity, age, gender, BMI and smoking on the serum transferrin glycoform pattern: Implications for use of carbohydrate-deficient transferrin (CDT) as alcohol biomarker. Clinica Chimica Acta 388(1/2): 59-67, 2008. (36 refs.)

Background: An alcohol-induced change in the serum transferrin glycoform pattern, carbohydrate-deficient transferrin (CDT), is used as a biomarker for detection and follow-up of heavy alcohol consumption. Besides studying the effects of drinking, this study evaluated any baseline differences in the transferrin pattern in relation to ethnicity, age, gender, body mass index (BMI) and smoking, as these could be confounders causing bias in CDT testing. Methods: The transferrin glycoform pattern was determined in 1387 sera (68% men, 32% women) collected in Australia, Brazil, Canada, Finland and Japan from subjects classified as non-drinkers, light/moderate drinkers, or heavy drinkers by use of the WHO/ISBRA Interview Schedule. The iron-saturated glycoforms were separated by an HPLC candidate reference method, and the relative amounts of individual glycoforms to total transferrin were determined. Results: In non-drinkers, the differences in the serum transferrin glycoform pattern in relation to ethnicity, age, gender and BMI were small and mostly not statistically significant. A higher disialotransferrin level in smokers compared with non-smokers could largely be explained by a higher alcohol intake in smokers. In the drinking subgroups, the main CDT glycoform disialotransferrin showed a positive correlation (r=0.80) with asialotransferrin, and disialo- and asialotransferrin a negative correlation with tetrasialotransferrin, that was dependent on the alcohol consumption level. Conclusions: With respect to CDT testing, the results indicated that adjustment of reference intervals for disialotransferrin and CDT in relation to ethnicity, age, gender, BMI and smoking is not required.

Bisaga A; Laposata M; Xie S; Evans SM. Comparison of serum fatty acid ethyl esters and urinary 5-hydroxytryptophol as biochemical markers of recent ethanol consumption. Alcohol and Alcoholism 40(3): 214-218, 2005. (29 refs.)

Aims: To examine the effects of an acute dose of ethanol on serum fatty acid ethyl esters (FAEEs) concentration and urinary 5-hydroxytryptophol (5-HTOL)/5-hydroxyindole-3-acetic acid (5-HIAA) ratio. Methods: Sixteen (14 male, 2 female) heavy alcohol drinkers were tested in a single, 2-day long session. Six participants received 1.5 g/l of ethanol/l of body water (similar to 0.75 g/kg of body weight, low dose group: LD) and 10 participants received 2.0 g/l of ethanol (similar to 1.0 g/kg of body weight, high dose group: HD) in four divided doses every 20 min. Blood, urine, and breath samples were collected repeatedly over 36 h following the ingestion of ethanol and were analyzed for the presence of FAEE, 5-HTOL/5-HIAA, and ethanol, respectively. Serum gamma-glutamyltransferase (GGT), a marker of chronic ethanol use, was also included. Results: The breath ethanol level peaked similar to 1 h after the last dose, at 95 and 120 mg/dl for the LD and HD groups, respectively. The mean ratio of urinary 5-HTOL/5-HIAA was significantly elevated 5 and 9 h after ethanol administration, but returned to baseline 13 h after ethanol administration. This ratio was twice as high for the HD group compared with the LD group. Serum levels of FAEEs were significantly elevated at 5 h, but not 13 h after ethanol administration. There were no time-dependent changes in serum GGT levels. Conclusions: Measuring the levels of FAEE and 5-HTOL/5-HIAA ratio provides a convenient method to detect recent, particularly binge-type, ethanol use, but these measures may have limited applicability in detecting ethanol use in traditional clinical trial settings.

Borucki K; Kunstmann S; Dierkes J; Westphal S; Diekmann S; Bogerts B et al. In heavy drinkers fatty acid ethyl esters in the serum are increased for 44 hr after ethanol consumption. Alcoholism: Clinical and Experimental Research 28(7): 1102-1106, 2004. (17 refs.)

Background: Fatty acid ethyl esters (FAEEs) have been proposed as a marker of ethanol consumption because they can be detected for up to 24 hr after a moderate intake of ethanol, even though blood ethanol remains increased for only 8 hr. Therefore, this study investigated whether FAEEs can be found during a time period exceeding 24 hr in a group of patients who were hospitalized for ethanol detoxification. A second aim was to study the distribution of FAEEs between lipoproteins during that time. Methods: Serum samples of 12 patients with acute ethanol intoxication were assayed for FAEEs. Blood samples were drawn 8.2, 20.2, 32.2, and 44.2 hr after hospitalization. FAEEs were quantified by gas chromatography-mass spectrometry. Results: Ethanol was no longer detectable after 20.2 hr from hospitalization, whereas FAEEs were still found after 32.2 and 44.2 hr. These late FAEEs were significantly higher than the FAEEs in 15 different healthy men who had abstained from ethanol for 4.5 days (p < 0.001 and p = 0.001). FAEEs were associated mainly with lipid-free serum but tended to accumulate in very-low-density lipoprotein in patients with moderate hypertriglyceridemia. Conclusions: In heavy drinkers, the FAEEs were increased after ethanol consumption for at least 44 hr. It remains to be studied whether they originate from a single ethanol intake or, in addition, from a slow release out of body storage compartments.

Brown TG; Gianoulakis C; Tremblay J; Nadeau L; Dongier M; Kin NMKNY et al. Salivary cortisol: A predictor of convictions for driving under the influence of alcohol? Alcohol and Alcoholism 40(5): 474-481, 2005. (57 refs.)

Aims: To examine the relationship between salivary cortisol and frequency of past driving under the influence of alcohol (DUI) convictions. Methods: A total of 104 males with previous DUI convictions (from one to eight) and mean age of 44.7 years were assessed on measures characterizing repeat DUI offenders, including sociodemographic information, alcohol use behaviours, biological indices of the organic consequences of chronic abuse, negative consequences of excessive drinking, past DUI conviction history, impulse control, and antisocial behaviour tendencies. Saliva samples were taken approximately every 30 min over a 6 h period during an exhaustive multidimensional assessment protocol, and were then assayed to obtain cortisol responses. Results: Blunted cortisol response, typically observed in alcoholics and in high-risk non-alcoholics, was associated with increased number of past DUI convictions. This association was particularly pronounced in multiple DUI offenders, and was stronger than, and independent of, other measures of alcohol use severity and chronicity commonly used for DUI assessment. Conclusions: Cortisol response may be useful in understanding the mediators underlying repeat DUI offending and the frequent failure of intervention efforts in curbing DUI behaviour.

Campuzano JC; Hernandez-Avila M; Jaakkola MS; Ponce EL; Morales PK; Bautista P et al. Determinants of salivary cotinine levels among current smokers in Mexico. Nicotine & Tobacco Research 6(6): 997-1008, 2004. (38 refs.)

The present study describes salivary cotinine levels and their relationship to cigarettes smoked per day in Mexican smokers. Using a sampling strategy based on the number of cigarettes per day, we recruited 1,222 smokers from Mexico City and the state of Morelos in Mexico during 1999. Smoking behaviors and other factors known to affect nicotine intake and cotinine level were identified in an interview using a standardized questionnaire. Salivary cotinine was measured by capillary gas chromatography with nitrogen-phosphorus detection. We used generalized additive models to describe the relationship between salivary cotinine levels and variables of interest. The mean age of the population was 39.7 years (SD = 15.6 years), with a mean cotinine level of 194.7 ng/ml (SD = 134.8; range = 10.1-767). Participants smoked a mean of 15.5 cigarettes per day (SD = 11.3). Salivary cotinine and cigarettes smoked per day were positively related, although the association was not linear. flattening above 20 cigarettes per day. After adjusting for cigarettes per day, we found that significant predictors of cotinine levels included age, body mass index, cigarette producer, and smoking behavior variables. These results may have, implications for dosing with nicotine medications to aid smoking cessation in Mexican smokers and suggest that whether the cigarette is labeled light or regular has no relationship to nicotine dose from smoking cigarettes.

Caprara D. Fatty acid ethyl esters and hair analysis: Biomarkers for alcohol consumption and their possible application in the diagnosis of FASD. Journal of FAS International 1(1): e7, 2003

This is a critical review of three recent published articles that deal with biomarkers for alcohol consumption: Analysis of fatty acid ethyl esters in hair as possible markers of chronically elevated alcohol consumption by headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC- MS) by Pragst, Auwaerter, Sporkert, Spiegel in "Forensic Science International" (2001); Fatty acid ethyl esters in hair as markers of alcohol consumption. Segmental hair analysis of alcoholics, social drinkers, and teetotalers by Auwarter, Sporkert, Hartwig, Pragst, Vater and Diefenbacherin in "Clinical Chemistry" (2001); and Fatty acid ethyl esters: A novel biologic marker for heavy in utero ethanol exposure: a case report. by Klein, Karaskov and Koren in "Therapeutic Drug Monitoring" (1999).

Ceccanti M; Mancinelli R; Sasso GF; Allen JP; Binetti R; Mellini A et al. Erythrocyte thiamine (Th) esters: A major factor of the alcohol withdrawal syndrome or a candidate marker for alcoholism itself? Alcohol and Alcoholism 40(4): 283-290, 2005. (63 refs.)

Aims: Thiamine (Th) deficiency is a major problem in alcoholics. In this study, the relationship of alcohol withdrawal syndrome (AWS) to Th and its esters, as well as the diagnostic power of Th and its esters were investigated. Patients and methods: Th and its esters were assessed in a series of chronic alcoholics (and in controls) using an improved method. Results: No association was found between AWS severity and Th and its esters, while the diagnostic power of thiamine diphosphate (TDP) and Th was very high. TDP was the most significant among the parameters under study, confirming that erythrocyte TDP is a suitable marker of alcoholism: TDP sensitivity across subjects was 84.1%, specificity 85.4%, positive predictive value 82.4%, and negative predictive value 88.0%.

Chan D; Knie B; Boskovic R; Koren G. Placental handling of fatty acid ethyl esters: Perfusion and subcellular studies. Journal of Pharmacology and Experimental Therapeutics 310(1): 75-82, 2004. (32 refs.)

The measurement of fatty acid ethyl esters ( FAEE) in neonatal meconium is a novel test to confirm prenatal ethanol exposure. The origin of FAEE in the maternal-placental-fetal unit is not known. The objectives of this study were to investigate whether FAEE are transferred and metabolized by the human placenta. Isolated placental cotyledons were perfused with a mixture of four FAEE ( palmitic, stearic, oleic, and linoleic acid ethyl esters) commonly detected in the meconium of neonates exposed to ethanol in utero, and the transfer of FAEE to the fetal unit was investigated in the absence and presence of albumin. The metabolic degradation of FAEE by human placental microsomes was subsequently determined. FAEE disappeared from the maternal circulation but remained undetectable in the fetal unit following perfusions. The addition of albumin had no effect on FAEE transfer. The unrecoverable fraction of individual FAEE in the perfusion system accounted for >50% of the initial amount used, suggesting significant metabolic degradation. Subcellular studies documented the enzymatic degradation of FAEE by placental microsomes (mean K-m, 35-95 muM; V-max, 0.6-1.8 nmol/min/mg for individual FAEE). FAEE at levels found in alcoholics that are originated from the mother are not transferred to the fetus because they are taken up and degraded extensively by the human placenta. Hence, FAEE detected in neonatal matrices are likely produced by the fetus from ethanol that has been transferred to and metabolized by the fetus, rendering FAEE a powerful direct biomarker reflective of true fetal exposure to ethanol in utero.

Chen J; Conigrave KM; Macaskill P; Whitfield JB; Irwig L. Combining carbohydrate-deficient transferrin and gamma-glutamyltransferace to increase diagnostic accuracy for problem drinking. Alcohol and Alcoholism 38(6): 574-582, 2003. (20 refs.)

Aim: To examine methods for combining quantitative results for serum carbohydrate deficient transferrin (CDT), gamma-glutamyltransferase (GGT) and/or aspartate aminotransferase (AST), and refining these by inclusion of patient characteristics. Methods: Data from 1684 subjects, recruited from the general population, abstainer groups and alcohol treatment centres (participants in the five nations WHO/ISBRA study of biological markers of alcohol use), were used to develop clinical rules for combining results of GGT, AST and CDT. The algorithm derived by Sillanaukee and Olsson was tested, and compared with new algorithms derived by logistic regression and discriminant analysis. Diagnostic accuracy was assessed by area underneath the receiver operator characteristic curve. Effects of adding gender and clinical information to the algorithm were estimated. Results: The predictive ability of combination rules derived from the two studies and by two different statistical techniques was remarkably consistent. For men, combining lnCDT and lnGGT provided the best accuracy for detecting daily consumption of 60 g ethanol or more in the past 30 days. For women, GGT alone provided the best accuracy for that consumption level. Clinical variables added significantly to the diagnostic accuracy of the models for both men and women, and conversely the test results modified the probability of problem drinking as assessed from clinical data alone. A graphic method was produced to help clinicians estimate probabilities for consumption of 60 g or more per day. Conclusions: Combining biochemical markers enhances detection of problem drinking in men but not in women. Information on clinical variables increases the ability to correctly detect problem drinking.

Cluver JC; Miller PM; Anton RF. Case studies of the utility of serum carbohydrate-deficient transferrin (%CDT) in the clinical management of alcoholics. Journal of Addiction Medicine 1(1): 44-47, 2007. (17 refs.)

A laboratory test that measures the percentage of carbohydrate-deficient transferrin (%CDT) has been approved by the FDA for the detection of heavy alcohol use. This biomarker has unique properties and has proven to be a useful tool in multiple clinical settings. It can be used for screening and monitoring purposes in patients with suspected or known alcohol use disorders. We review the use of this test in the management of 5 patients seen in the mental health and substance abuse clinics at a Department of Veterans Affairs Medical Center.

Czermak C; Lehofer M; Wagner EM; Prietl B; Lemonis L; Rohrhofer A et al. Reduced dopamine D4 receptor mRNA expression in lymphocytes of long-term abstinent alcohol and heroin addicts. Addiction 99(2): 251-257, 2004. (44 refs.)

Aim: It has been repeatedly suggested that dopamine receptor expression in peripheral blood lymphocytes reflects, to some extent, brain status. The aim of the present study was to investigate dopamine receptor expression in peripheral blood lymphocytes of long-term abstinent alcohol and heroin addicts against the background of the hypothesis, that a persisting dysfunction of the dopaminergic system contributes a biological cause to the chronic character of addiction. Design: Dopamine D3 and D4 receptor mRNA expression in peripheral blood lymphocytes was measured by real-time polymerase chain reaction (PCR) in 19 alcohol addicts, abstinent for 6.2 � 4.7 months (mean � SD), and 20 heroin addicts, abstinent for 6.7 � 3.7 months (mean � SD), and compared to a control group of 29 age- and sex-matched individuals with no life-time history of substance abuse. Findings One-way anova showed significant differences in D4 mRNA expression between the groups (P = 0.005): both groups of addicts showed an approximately 50% reduction in D4 receptor mRNA expression in peripheral blood lymphocytes (PBL) compared to controls. No differences were found for D3 mRNA expression between the groups. Conclusion The results of the present study indicate a withdrawal-persisting dopaminergic imbalance in abstinent addicts as measured by a suggested peripheral marker.

Daeppen JB; Anex F; Leutwyler J; Secretan F; Gammeter R; Besson J; Darioli R; Chossis I. Role of high normal gamma-glutamyltransferase level in identifying heavy alcohol use in young men. Alcohol 32(2): 157-161, 2004. (16 refs.)

The objective of the current study was to determine the predictive value of high normal gamma-glutamyltransferase (GGT) level as an indication of heavy drinking in young men. In a sample of 577 men attending a one-day army recruitment process mandatory for all Swiss men at age 19 years, GGT level was evaluated as the dependent variable for each of eight dichotomous classifications of individuals on the basis of meeting cut-off criteria for five indexes of alcohol use, two indexes of alcohol-related problems, and one index of body mass. The sensitivity, specificity, and predictive values of GGT level in identifying subjects as either heavy drinkers or being overweight were determined. Compared with findings for their counterparts, GGT level was higher in subjects reporting consumption of more than 14 drinks per week (20.5 +/- 7.81 vs. 18.9 +/- 7.60, P < .05), in those reporting being drunk at least once during the past 30 days (20.3 +/- 7.80 vs. 18.3 +/- 7.43, P < .001), and in individuals with body mass indexes greater than or equal to25 kg/m(2) (25.8 +/- 10.84 vs. 18.3 +/- 6.59, P < .001). At a GGT level cut-off of 20 U/l, the sensitivity, specificity, and positive and negative predictive values of either being a heavy drinker or overweight were 48.2%, 70.2%, 67.7%, and 51.2%, respectively. Exclusion of subjects with body mass indexes of :25 kg/m(2) revealed similar results. High normal GGT level in young men is indicative of heavy alcohol use or being overweight; when present, subjects should be screened further for potential concomitant drinking problems.

Dai X; Thavundayil J; Gianoulakis C. Differences in the peripheral levels of beta-endorphin in response to alcohol and stress as a function of alcohol dependence and family history of alcoholism (review). Alcoholism: Clinical and Experimental Research 29(11): 1965-1975, 2005. (115 refs.)

Background: Evidence indicates that both genetic and environmental factors, such as stress, may play an important role for the development of alcoholism, while beta-endorphin may be implicated in the control of alcohol consumption. The objective of the present studies was to test the hypothesis that there are differences in the response of the pituitary beta-endorphin system to stress as a function of family history of alcoholism and alcohol dependence. Methods: The response of the pituitary beta-endorphin to a placebo or an alcohol (0.50g ethanol/kg) drink and to a stress task performed 30 min following ingestion of either the placebo or the alcohol drink was measured in social and heavy drinkers with [high risk (HR)] and without flow risk (LR)] a family history of alcoholism. Thus, each subject participated in 4 experimental sessions given on different days in a randomized order. Four groups of subjects were investigated: 1) low risk nonalcoholics (LRNA); 2) high risk nonalcoholics; (HRNA), 3) low risk alcoholics (LRA); and 4) high risk alcoholics (HRA). Plasma beta-endorphin was estimated prior to and for 3.5 hr post-stress. Changes in the concentration of plasma beta-endorphin following ingestion of either the placebo or alcohol drink without performance of the stress task served as controls to compare the stress-induced changes. Results: Basal plasma beta-endorphin levels were higher in LRNA than LRA, HRNA and HRA participants, while basal plasma beta-endorphin levels were higher in LRA than those in HRNA and HRA participants. Furthermore, there was no significant difference in the plasma beta-endorphin levels between HRNA and HRA participants. Stress, induced a significant increase in plasma beta-endorphin concentration in all four groups of participants. However, the stress-induced increase in plasma beta-endorphin levels was more pronounced in LRNA than HRNA, LRA and HRA participants. Thus, alcohol dependence decreased the basal plasma beta-endorphin levels of participants without, but not of those with, a family history of alcoholism. Alcohol prior to stress attenuated the stress-induced increase in plasma beta-endorphin levels of all four groups of participants. Conclusions: The present data indicates that there are differences in both, the basal plasma beta-endorphin levels as well as the response of the pituitary beta-endorphin to stress as a function of family history of alcoholism and alcohol dependence. Thus, in HR individuals a dysfunction in the activity of the pituitary beta-endorphin system predates the development of alcoholism, while in LR individuals it develops following alcohol dependence. Furthermore, alcohol dependence did not alter the alcohol-induced attenuation of beta-endorphin response to stress.

Das SK; Dhanya L; Vasudevan DM. Biomarkers of alcoholism: an updated review. (review). Scandinavian Journal of Clinical & Laboratory Investigation 68(2): 81-92, 2008. (141 refs.)

Alcoholic beverages, and the problems they engender, have been familiar in human societies since the beginning of recorded history. Among a variety of blood tests used to aid the diagnosis of alcohol consumption and related disorders, laboratory tests are particularly useful in settings where cooperativeness is suspected or when a history is not available. Biochemical and haematological tests, such as gamma-glutamyltransferase activity, aspartate aminotransferase activity and erythrocyte mean corpuscular volume, are established markers of alcohol intake. Carbohydrate-deficient transferrin is the only test approved by the FDA for the identification of heavy alcohol use. Total serum sialic acid and sialic acid index of Apolipoprotein J have the potential to be included in a combination of measurements providing an accurate, more exact, assessment of alcohol consumption in a variety of clinical and research settings. Several other markers with considerable potential for measuring recent alcohol intake include beta-hexosaminidase, acetaldehyde adducts and the urinary ratio of serotonin metabolites, 5-hydroxytryptophol and 5-hydroxyindoleacetic acid. These markers provide hope for more sensitive and specific aids to diagnosis and improved monitoring of alcohol intake.

Derauf C; Katz AR; Easa D. Agreement between maternal self-reported ethanol intake and tobacco use during pregnancy and meconium assays for fatty acid ethyl esters and cotinine. American Journal of Epidemiology 158(7): 705-709, 2003. (37 refs.)

Reliance on self-reported use of tobacco and intake of ethanol during pregnancy is associated with a high probability of error. Use of biological markers, or biomarkers, potentially offers a more valid method to assess exposure. Although cotinine is an established biomarker for tobacco use, there is no established biomarker for in utero ethanol exposure. Recent reports suggest that fatty acid ethyl esters (FAEE) could serve this purpose. To assess agreement between maternal self-reported tobacco use and ethanol intake during pregnancy and detection of metabolites associated with tobacco use (cotinine) and ethanol intake (FAEE), the authors studied maternal histories and meconium samples obtained in November-December 1999 from 436 consecutive mother-infant pairs at a large urban regional perinatal center in Honolulu, Hawaii. Cohen's kappa coefficient and 95% confidence intervals were calculated. Moderate agreement was found between reported tobacco use during the third trimester and detected cotinine level (kappa = 0.53, 95% confidence interval: 0.39, 0.68); however, there was no agreement between reported ethanol intake during the third trimester and detected FAEE (kappa = -0.02, 95% confidence interval: -0.04, 0.00). No mother reporting ethanol intake during the third trimester had detectable FAEE. Findings support the need for additional refinement and validation of the use of FAEE as a biomarker for maternal ethanol intake.

Dhar P. Measuring tobacco smoke exposure: quantifying nicotine/cotinine concentration in biological samples by colorimetry, chromatography and immunoassay methods. (review). Journal of Pharmaceutical and Biomedical Analysis 35(1): 155-168, 2004. (186 refs.)

Procedures to assess tobacco smoke exposure are reviewed and biomarkers used for determining the smoking status of an individual are compared. Methods used to extract these biomarkers from saliva, urine, and blood and the advantages and disadvantages of the assays are discussed. Finally, the procedures used to measure the levels of cortisol, a stress hormone speculated to be linked to nicotine metabolism, are discussed.

do Amaral RA; Malbergier A. Effectiveness of the CAGE questionnaire, gamma-glutamyltransferase and mean corpuscular volume of red blood cells as markers for alcohol-related problems in the workplace. Addictive Behaviors 33(6): 772-781, 2008. (28 refs.)

Objective: To evaluate the usefulness of gamma-glutamyltransferase (GGT) and mean corpuscular volume (MCV), as well as that of the CAGE questionnaire, in workplace screening for alcohol abuse/dependence. Methods: A total of 183 male employees were submitted to structured interviews (Structured Clinical Interview for DSM-IV 2.0 and CAGE questionnaire). Blood samples were collected. Diagnostic accuracy and odds ratio were determined for the CAGE, GGT and MCV. Results: The CAGE questionnaire presented the best sensitivity for alcohol dependence (91%; specificity, 87.8%) and for alcohol abuse (87.5%, specificity, 80.9%), which increased when the questionnaire was used in combination with GGT (sensitivity, 100% and 87.5%, respectively; specificity, 68% and 61.5, respectively). CAGE positive results and/or alterations in GGT were less likely to occur among employees not presenting alcohol abuse/ dependence than among those presenting such abuse (OR for CAGE = 13, p < 0.05; OR for CAGE-GGT = 11, p < 0.05) or dependence (OR for CAGE = 76, p < 0.0 1; OR for GGT = 5, p < 0.0 1). Employees not presenting alcohol abuse/dependence were also several times more likely to present negative CAGE or GGT results. Conclusions: The use short, simple questionnaires, combined with that of low-cost biochemical markers, such as GGT, can serve as an initial screening for alcohol-related problems, especially for employees in hazardous occupations. The data provided can serve to corroborate clinical findings.

Dolman JM; Hawkes ND. Combining the AUDIT Questionnaire and biochemical markers to assess alcohol use and risk of alcohol withdrawal in medical inpatients. Alcohol and Alcoholism 40(6): 515-519, 2005. (24 refs.)

Aims: Alcohol consumption is often under-reported in patients admitted to general hospitals with acute illness. For alcohol-dependent individuals hospital admission results in an enforced period of abstinence with potential alcohol withdrawal symptoms, and possible life threatening complications. Early detection of alcohol use is therefore beneficial to patients and health services. The purpose of this study was to investigate the performance of the alcohol use disorders identification test (AUDIT) questionnaire in the acute medical setting, and the effect of combining routine biological markers-glutamyltransferase, alanine aminotransferase, aspartate aminotransferase, and mean corpuscular volume (MCV) on its performance in the early identification of in-patients with alcohol use disorders and at risk of developing symptoms of alcohol withdrawal. Methods: Prospective study in consecutive patients admitted to an acute medical admissions ward. All patients were screened using the AUDIT questionnaire and routine blood tests. Patients were then monitored for symptoms of withdrawal using clinical institute withdrawal assessment for alcohol (CIWA-Ar). Results: Of the 874 patients screened using the AUDIT, 98 (11%) screened positive of whom 17 (2% of the 874) experienced clinically significant alcohol withdrawal symptoms, when using serial CIWA-Ar. The AUDIT and serial CIWA-Ar detected all patients who went on to manifest acute withdrawal symptoms. There was no loss of sensitivity at an AUDIT cut-off of 13 or more compared with the lower cut-off of 8 or more. A positive predictive value of 17.3% for an AUDIT score of 8 or more in the detection of withdrawal, increased to 47.1% when found in combination with at least two abnormal biological markers whilst maintaining a sensitivity of 94.1% and specificity of 97.9%. Conclusion: These findings confirm that AUDIT is a useful alcohol screen in general medical settings and that its ability to correctly predict which patients will experience alcohol withdrawal is increased when used in combination with biological markers.

Domino EF; Kadoya C; Matsuoka S; Ni LS; Fedewa KS. Comparative American and Japanese tobacco smoke uptake parameters after overnight tobacco deprivation. (review). Progress in Neuro-Psychopharmacology & Biological Psychiatry 27(6): 973-984, 2003. (62 refs.)

American and Japanese overnight deprived tobacco smokers were compared with respect to expired CO, plasma nicotine and cotinine, and red cell carboxyhemoglobin. The participants were 5 1 of 59 American and 55 of 86 Japanese cigarette smokers of mixed gender who met similar strict criteria. Female and male American tobacco smokers were similar in mean age, number of cigarettes smoked per day, machine-rated nicotine and tar yield per cigarette and per 24 h plasma cotinine, calculated previous 24 h nicotine dose, and exhaled CO. Only mean plasma nicotine levels were significantly higher in American females. American and Japanese female smokers had similar tobacco uptake parameters. American and Japanese male smokers differed; the latter had higher plasma nicotine and lower cotinine levels as well as calculated 24 h dose of nicotine and lower exhaled CO. Japanese females and males were similar in all tobacco smoke uptake parameters. When the two racial groups were compared, irrespective of gender, the only statistically significant differences were lower mean exhaled CO levels and percent COHb in the Japanese. It is concluded that Japanese males inhale cigarettes in moderation compared to Americans. The results are discussed in relation to known ethnic, social, and genetic differences in CYP2A6 gene polymorphism.

Ebbert JO; Dale LC; Nirelli LM; Schroeder DR; Moyer TP; Hurt RD. Cotinine as a biomarker of systemic nicotine exposure in spit tobacco users. Addictive Behaviors 29(2): 349-355, 2004. (14 refs.)

Unlike cigarette smokers, spit tobacco (ST) users absorb a significant amount of nicotine through the gastrointestinal tract while swallowing tobacco juice. The majority of the absorbed nicotine is rapidly converted to cotinine during first-pass hepatic metabolism. This process potentially compromises the utility of cotinine as a biomarker for systemic nicotine exposure in ST users. To investigate this question, we correlated nicotine and cotinine concentrations with clinical measures of ST use in 68 daily ST users enrolled in a nonnicotine pharmacologic intervention trial. We found that a higher frequency of swallowing tobacco juice (P=.007) was an independent predictor of higher serum cotinine concentrations. Serum nicotine concentrations, on the other hand, were not correlated with a higher frequency of swallowing. In the absence of a reliable way to measure frequency of swallowing, we conclude that cotinine should not be used for guiding clinical decisions that depend upon a precise quantification of systemic nicotine exposure, such as tailored nicotine replacement therapy.

Fidler JA; Jarvis MJ; Mindell J; West R. Nicotine intake in cigarette smokers in England: Distribution and demographic correlates. Cancer Epidemiology, Biomarkers & Prevention 17(12): 3331-3336, 2008. (31 refs.)

Documenting smoke intake by objective biochemical markers is important for quantification of exposure to toxins. The aim of this report is to show the most definitive distribution of the nicotine metabolite, cotinine, yet available from English smokers in the period before implementation of the legislation banning smoking in indoor public areas. A total of 6,423 cigarette smokers, ages 1.6 years and above, taking part in the Health Survey for England between 1998 and 2003 provided a saliva cotinine value. Data on cigarette consumption, age, gender, social class, deprivation (as indicated by manual or nonmanual occupation, car ownership, and housing tenure), marital status, and region were collected. Cotinine concentrations showed no overall trend over the study period. The mean value was 289.15 ng/mL (SD 174.43); excluding those who had not smoked in the past 24 hours, the mean value was 302.08 ng/mL (SD 168.78). A higher cotinine concentration was associated with being middle-aged (peaking at 40 years), being male, being single, greater economic deprivation, and not living in London. After adjusting for cigarette consumption, higher values were associated with middle age, being male, and greater deprivation. This study provides the most complete picture yet of the smoke intake of cigarette smokers in England. The fact that cotinine peaks at around the age of 40 years raises the possibility that nicotine intake continues to increase decades after a person starts to smoke. Greater nicotine intake in more deprived smokers could explain why they find it harder to stop smoking.

Gimenez TJC; Adame ML. The influence of salivary activity in evaluating adolescent tobacco consumption by determining the level of thiocyanate in saliva. Addictive Behaviors 28(1): 81-89, 2003. (33 refs.)

The thiocyanate (SNC) concentration in saliva is a biochemical measure frequently used as an objective indicator of tobacco consumption. However, variations in the volume of saliva can influence SNC levels, making it more difficult to ascertain the relation between tobacco consumption and SNC concentration. The goal of this study was to determine the extent to which salivary activity can influence SNC concentration in saliva among adolescents. We measured the SNC concentration in saliva and the salivary activity (using cotton dental rolls) of 592 students aged 14-17 years. The results of a stepwise regression analysis did not show any significant effect of the salivary activity on SCN levels.

Golka K; Sondermann R; Reich SE; Wiese A. Carbohydrate-deficient transferrin (CDT) as a biomarker in persons suspected of alcohol abuse. Toxicology Letters 151(1): 235-241, 2004. (11 refs.)

The coherence of carbohydrate-deficient transferrin (CDT) as a biomarker of alcohol abuse was investigated with 15 conventional laboratory parameters, with the self-reported medical history and with clinical findings, all previously reported to be associated with chronic alcohol intake. In total, 100 male persons who were at least suspected of abusing alcohol were assessed. Medical history, clinical picture and physical examination were taken, and laboratory parameters regarding blood count, liver enzymes, serum lipids, iron balance, Ig A and uric acid were determined. These data were correlated with the CDT values, the daily ethanol intakes reported, and several findings from medical history and clinical examination. The mean CDT level (mean +/- S.D.) of the entire group was 29.4 +/- 19.7 U/l. Eighty-one patients admitted a daily ethanol intake of 60 g or more. The ratio ASPALT (de Ritis ratio) appeared as the best conventional parameter correlated with both CDT and ethanol intake. Mean corpuscular volume (MCV), serum iron, AST and red blood cell count also correlated significantly with CDT. CDT, AST and ferritin correlated significantly with the reported daily ethanol intake. It is concluded that CDT provides a reliable estimate of long-term alcohol intake.

Graham AW; Schultz TK; Mayo-Smith MF; Ries RK; Wilford BB, eds. Principles of Addiction Medicine. Chevy Chase MD: American Society of Addiction Medicine, 2003. (Chapter refs.)

This volume is a comprehensive text on addictions. It is organized into 14 major sections, each of which has multiple chapters. There are over 200 contributors. The sections deal with the following themes: basic science and core concepts; pharmcology; diagnosis, assessment and early intervention; overview of addiction treatment; management of intoxication and withdrawal; pharmacologic interventions; behavioral interventions; 12-step programs and other recovery-oriented interventions; alcohol and drug problems in the workplace; medical disorders and complications of addiction; co-occurring addictive and psychiatric disorders; pain and addiction; and children and adolescents. There are also six appendices.

Gray TR; Kelly T; LaGasse LL; Smith LM; Derauf C; Haning W et al. Novel biomarkers of prenatal methamphetamine exposure in human meconium. Therapeutic Drug Monitoring 31(1): 70-75, 2009. (40 refs.)

Meconium analysis can detect fetal exposure to drugs taken by the mother during pregnancy. Methamphetamine (MAMP) and amphetamine (AMP) have previously been observed in meconium of MAMP-exposed neonates; the presence of other metabolites has not been investigated. Detection of such analytes may lead to more sensitive identification and thus improved medical treatment of affected infants. Forty-three MAMP-positive meconium specimens were analyzed for newly identified MAMP biomarkers, p-hydroxymethamphetamine, p-hydroxyamphetamine, and norephedrine. Due to MAMP adulteration in illicit ecstasy and to simultaneously monitor 3,4-methylenedioxymethamphetaniine and MAMP prenatal exposure, 3,4-methylenedioxymethamphetamine, its metabolites, and related sympathomimetic amines were assayed. MAMP AM-P, and unconjugated p-hydroxyrnethamphetamine were the most prevalent and abundant analytes present in meconium; however, unconjugated p-hydroxyamphetarnine and norephedrine also were identified. It is possible that one of these additional analytes could be important for predicting toxicity or maternal or neonatal outcome measures in fetuses exposed to MAMP at specific gestational ages or with different metabolic capabilities. Although these new biomarkers were present in lower concentrations than MAMP and AMP in the meconium of previously confirmed specimens, additional research will determine if inclusion of these analytes can increase identification of MAMP-exposed neonates. Novel methamphetamine biomarker concentrations were characterized in meconium of infants exposed in utero to MAMP

Harasymiw J; Forster I; Bean P. Using the EDAC (TM) test to monitor abstinence and relapses during outpatient treatment. Journal of Addictive Diseases 24(3): 101-113, 2005. (17 refs.)

The main objective of this study is to show the performance of the EDAC (TM) test in monitoring alcohol consumption during outpatient treatment. The EDAC (TM) is a new approach that uses routine laboratory tests to identify binge drinking as well as chronic drinking. The overall diagnostic performance of the EDAC (TM) fluctuates around 80 to 90% for both specificity and sensitivity. Close to two thousand subjects have been tested by the EDAC (TM) since the early eighties; 300 of these were patients undergoing treatment at different institutions across the U.S. This article selected five case studies to represent examples of classical drinking behaviors encountered in most outpatient clinics. The first four cases illustrate the use of the EDAC (TM) alone and the last case represents the use of the EDAC (TM) combined with CDT. These five case studies illustrate the use of the EDAC (TM) to detect relapse episodes, to monitor abstinence during outpatient treatment and to recognize a slip early enough to prevent more severe drinking. The use of biomarkers to monitor drinking behavior in alcohol dependent patients is gaining popularity because they provide objective information on a patient's drinking status when used as an adjunct to self-report.

Hardin E; Adinoff B. Family history of alcoholism does not influence adrenocortical hyporesponsiveness in abstinent alcohol-dependent men. American Journal of Drug and Alcohol Abuse 34(2): 151-160, 2008. (32 refs.)

Background: Early abstinence in alcohol-dependent subjects is marked by adrenocortical hyporesponsivity. However, it is uncertain whether the blunted response is primarily attributable to a genetic vulnerability or to the chronic abuse of alcohol. In the present study, the authors investigated the influence of a family history (FH) of alcoholism upon suppressed glucocorticoid reactivity. Methods: Twenty-two abstinent alcohol-dependent and 14 control men were studied. The cortisol response was assessed in 11 patients following oCRH infusion (.4 ug/kg) and in a separate group of 11 patients following cosyntropin infusion (.01 ug/kg) preceded by high-dose intravenous dexamethasone (8 mg). FH, as determined by self-report, was assessed using two different methods: history of parental alcoholism and number of alcohol-dependent first- and second-degree relatives. Results: Neither a parental history or familial loading of alcoholism had a significant effect upon glucocorticoid responsivity in abstinent alcohol-dependent men. Conclusions: Adrenocorticol responsiveness in recently abstinent alcohol-dependent men does not appear to reflect a preexisting biologic vulnerability to alcoholism.

Hartwig S; Auwarter V; Pragst F. Fatty acid ethyl esters in scalp, pubic, axillary, beard and body hair as markers for alcohol misuse. Alcohol and Alcoholism 38(2): 163-167, 2003. (23 refs.)

Aims: This study examined the possibility of using hair samples other than scalp hair for analysis of fatty acid ethyl esters (FAEE) as markers for alcohol misuse. Methods: Samples of scalp and pubic hair and, if available, axillary, beard and body hair of one teetotaller, five moderate social drinkers and 22 fatalities were analysed for FAEE by head-space solid-phase microextraction and gas chromatography-mass spectrometry. The sum of the concentrations of ethyl myristate, ethyl palmitate, ethyl oleate and ethyl stearate (C-FAEE) in the hair samples was compared with information about the drinking behaviour of the individuals. Results: Although there were large differences in the analyte concentrations in hair from different sites in the same individual, cases of chronic excessive alcohol consumption were characterized by C-FAEE > 1.0 ng/mg in almost all samples. Conclusion: FAEE concentrations in hair other than scalp hair can be used as markers of chronic alcohol misuse.

Hastie CE; Haw S; Pell JP. Impact of smoking cessation and lifetime exposure on C-reactive protein. Nicotine & Tobacco Research 10(4): 637-642, 2008. (35 refs.)

C-reactive protein (CRP) levels predict coronary heart disease (CHD) risk. Levels are raised among smokers, but the effect of smoking cessation is unclear. Exposure to secondhand smoke (SHS) may be a confounder. Lifetime smoking exposure may have a dose effect on CRP among smokers, but it is unclear if this persists after cessation. We analyzed cross-sectional data on 4,072 adults recruited to a Scotland-wide population health survey who did not have CHD and were not on nicotine replacement therapy. CRP fell with time from cessation but was still raised up to 5 years after adjustment for case-mix (p <.001). SHS exposure was greater among ex-smokers than neversmokers (median cotinine 0.5 ng/ml vs. 0.4 ng/ml, p <.001) but did not explain the difference. Among smokers, there was a dose relationship between pack years and CRP on both univariate, F(4,1279)=31.841, p <.001, and multivariate, F(4,1085)=3.499, p=.008, analysis. Among ex-smokers there was also a dose relationship between pack-years and CRP, F(4,751)=14.108, p <.001, which was independent of time from cessation and case-mix, F(4,466) = 3.744, p =.005. That CRP does not fall to normal levels immediately and that lifetime smoking exposure continues to impact on CRP levels post cessation suggest that CRP is not raised as a direct effect of cigarette smoke but rather via a secondary mechanism, such as tissue damage causing an inflammatory stimulus. Our results reinforce the need to encourage smoking cessation as early as possible.

Hatsukami DK; Giovino GA; Eissenberg T; Clark PI; Lawrence D; Leischow S. Methods to assess potential reduced exposure products. (review). Nicotine & Tobacco Research 7(6): 827-844, 2005. (108 refs.)

The availability of tobacco products purported to reduce toxin exposure or potentially reduce health risks necessitates the development of methods and identification of biomarkers that can be used to assess these products. These assessments occur on multiple levels and stages, from identifying constituents in the tobacco products and smoke, to human exposure and health effects trials, to postmarketing surveillance. A conference of multidisciplinary experts was convened to present and discuss methods and biomarkers to assess these products and to consider the infrastructure necessary to facilitate the evaluation process. Although no currently available set of measures was thought to be sufficient for determining the relative health risk of potential reduced exposure products, this paper provides a blueprint for future research toward this end.

Hatsukami DK; Kotlyar A; Allen S; Jensen J; Li S; Le C et al. Effects of cigarette reduction on cardiovascular risk factors and subjective measures. Chest 128(4): 2528-2537, 2005. (29 refs.)

Study objectives: To assess the effect of continued smoking and smoking reduction on cardiovascular biomarkers (eg, WBC count, cholesterol concentrations, BP, heart rate). Design, setting, and participants: This study, conducted at the University of Minnesota, randomized smokers interested in significantly reducing cigarette use but not quitting to either start 12 weeks of smoking reduction immediately (n = 102), assisted by nicotine replacement therapy, or to a 6-week wait list (n = 49). Those starting smoking reduction were required to reduce smoking by 25% for 2 weeks, 50% for 2 weeks, and 75% during the final 2 weeks. After 6 weeks, the subjects were asked to maintain a 50% reduction or quit. Nicotine gum and, if necessary, nicotine patch were used to achieve reduction goals. The wait list group (n = 49) smoked ad libitum for 6 weeks and then reduced smoking as previously described. Measurements and results: Cardiovascular biomarkers (eg, WBC count, cholesterol concentrations, BP, heart rate) were assessed at several time points after enrollment. During ad libitum smoking, cardiovascular biomarkers remained relatively stable with correlation coefficients across the various time measurements, ranging from 0.44 to 1.00 (p < 0.01 for all measures). Among successful nonabstinent reducers (64 of 151 subjects), significant improvements were found in many biomarkers (eg, hemoglobin, hematocrit, RBC and WBC counts, lipids, BP, heart rate, respiratory symptoms, all p < 0.0167). Conclusions: These results show the availability of reliable and dose-sensitive biomarkers and that reduction in smoking can lead to significant but only modest changes in cardiovascular risk factors in healthy smokers. It is not known whether the reductions in cardiovascular risk factors observed after smoking reduction are also associated with reduced disease risk. Additional research is necessary to address this issue.

Hecht SS. Carcinogen derived biomarkers: Applications in studies of human exposure to secondhand tobacco smoke. (review). Tobacco Control 13(Supplement 1): 48-56, 2004. (103 refs.)

Objective: To review the literature on carcinogen derived biomarkers of exposure to secondhand tobacco smoke (SHS). These biomarkers are specifically related to known carcinogens in tobacco smoke and include urinary metabolites, DNA adducts, and blood protein adducts. Method: Published reviews and the current literature were searched for relevant articles. Results: The most consistently elevated biomarker in people exposed to SHS was 4-(methylnitrosamino) (3-pyridyl)-1-butanol (NNAL) and its glucuronides (NNAL-Gluc), urinary metabolites of the tobacco I specific lung carcinogen 4-(methylnitrosamino)-1 -(3-pyridyl)- 1 -butanone (NNK). The tobacco specificity of this biomarker as well as its clear relation to an established lung carcinogen are particularly appropriate for its application in studies of SHS exposure. Conclusion: The results of the available carcinogen derived biomarker studies provide biochemical data which support the conclusion, based on epidemiologic investigations, that SHS causes lung cancer in nonsmokers.

Hecht SS; Carmella SG; Le KA; Murphy SE; Li YS; Le C; Jensen J; Hatsukami DK. Effects of reduced cigarette smoking on levels of 1-hydroxypyrene in urine. Cancer Epidemiology, Biomarkers & Prevention 13(5): 834-842, 2004. (30 refs.)

We investigated the effects of smoking fewer cigarettes/day (CPD) on urinary levels of 1-hydroxypyrene (1-HOP), a biomarker of carcinogenic polycyclic aromatic hydrocarbon (PAH) uptake. We randomly assigned 151 smokers to either a reduction group or a waitlist group. In the reduction group, we measured urinary 1-HOP at two baseline intervals. Then, the smokers were expected to reduce their CPD by 25% in weeks 0-2, 50% in weeks 2-4, and 75% in weeks 4-6 and to maintain reduced smoking through week 26. In the waitlist group, four baseline measurements were taken and then the smokers joined the reduction group. Urinary 1-HOP was quantified at weeks 4, 6, 8, 12, and 26 after baseline. Statistically significant reductions in urinary 1-HOP were observed at most time points examined in groups of smokers who reduced to different extents. Reductions in the waitlist group were also generally significantly greater than baseline levels. The reductions in 1-HOP were usually modest (ranging from 14% to 35% in all groups and time points examined), which partially reflects the fact that there are sources of pyrene exposure other than cigarette smoke. Thus, cessation of smoking would only be expected to result in partial reductions of 1-HOP. The observed reductions in 1-HOP were not fully consistent with reductions in CPD probably due to uncontrolled dietary factors. Collectively, the results demonstrate that some smokers can achieve substantial reductions in 1-HOP, reflecting reduced uptake of polycyclic aromatic hydrocarbons, by reducing CPD, but there was not a consistent relationship between these parameters.

Helander A; Bottcher M; Fehr C; Dahmen N; Beck O. Detection times for urinary ethyl glucuronide and ethyl sulfate in heavy drinkers during alcohol detoxification. Alcohol and Alcoholism 44(1): 55-61, 2009. (41 refs.)

Aims: Ethyl glucuronide (EtG) and ethyl sulfate (EtS) are conjugated ethanol metabolites formed in low amounts after alcohol consumption. Compared with ethanol, EtG and EtS are excreted in urine for a prolonged time, making them useful as sensitive alcohol biomarkers. This study determined the detection times for EtG and EtS in alcoholic patients undergoing alcohol detoxification. Methods: Alcohol-dependent patients (n = 32) with an initial alcohol concentration >= 1 g/L based on breath testing were followed during detoxification. Urine samples for determination of EtG, EtS, ethanol and creatinine were collected on admission to the hospital and thereafter once daily for several days. EtG and EtS measurements were performed by liquid chromatography-mass spectrometry (LC-MS) and EtG also using an immunochemical assay (DRI-EtG EIA, ThermoFisher/Microgenics). Results: The detection time for urinary EtG was weakly correlated (r = 0.434, P = 0.013) with the initial alcohol concentration (range 1.0-3.4 g/L). For EtG, the individual time range until return to below the applied cut-off limit (< 0.5 mg/L) was similar to 40-130 h (median 78) with a similar time course observed for EtS. After correction for urine dilution, the time until an EtG/creatinine ratio < 0.5 mg/g was similar to 40- 90 h (median 65). The detection times after an estimated zero ethanol concentration were similar to 30-110 h (median 66) for EtG and similar to 30- 70 h (median 56) for EtG/creatinine. The EtG results by LC-MS and the immunoassay were in good agreement. Conclusions: During alcohol detoxification, EtG and EtS remained detectable in urine for several days. The detection times showed wide inter-individual variations, also after adjusting values for urine dilution and to the estimated times for a completed ethanol elimination.

Hietala J; Puukka K; Koivisto H; Antillia P; Niemela O. Serum gamma-glutamyl transferase in alcoholics, moderate drinkers and abstainers: Effect on GT reference intervals at population level. Alcohol and Alcoholism 40(6): 511-514, 2005. (21 refs.)

Aims: To clarify in the association between amount of ethanol consumption and serum gamma-glutamyl transferase (GT) levels. Methods: GT values were measured from 195 individuals with a wide variety of well-documented ethanol consumption assessed by detailed personal interviews using a time-line follow-back technique. These included 103 heavy drinkers (90 men, 13 women) and 92 healthy volunteers (54 men, 38 women) who were either abstainers (n = 30) or moderate drinkers (n = 62). For comparisons, data were collected from GT measurements for establishing GT reference intervals from 2485 healthy volunteers including 1156 abstainers and 1329 moderate drinkers. Results: GT values in the individuals whose mean ethanol consumption exceeded 40 g of ethanol per day were significantly higher than those in the moderate drinkers with a mean consumption of 1-40 g/day (P < 0.001) or in abstainers (P < 0.001). The GT values in the group of moderate drinkers also exceeded those of the abstainers (P < 0.001). The upper normal GT limits obtained from the data from abstainers were markedly lower (men 45 U/l, women 35 U/l) than those obtained from the population of moderate drinkers (men 66 U/l, women 40 U/l). Conclusions: Serum GT concentrations may respond to relatively low levels of ethanol consumption, which should be considered when defining GT reference intervals. The continuous increase in alcohol consumption at population level may lead to increased GT cut-off limits and hamper the detection of alcohol problems and liver affection in their early phase.

Hillemacher T; Weinland C; Heberlein A; Groschl M; Schanze A; Frieling H et al. Increased levels of adiponectin and resistin in alcohol dependence-possible link to craving. Drug and Alcohol Dependence 99(1-3): 333-337, 2009. (26 refs.)

Recent studies suggested a role of appetite regulating peptides like leptin and ghrelin in alcohol dependence and particularly in the neurobiology of alcohol craving. Aim of the present study was to investigate alterations of the adipocytokines adiponectin and resistin in alcohol-dependent patients. We analyzed a sample of 88 patients at admission for alcohol detoxification and after I week of withdrawal treatment in comparison to 89 healthy controls. Adiponectin and resistin serum levels were measured using commercial ELISA kits. The extent of alcohol craving was obtained using the obsessive Compulsive Drinking Scale (OCDS). Adiponectin and resistin serum levels were significantly elevated in patients with alcohol dependence at both dates (admission and after I week of treatment) compared to healthy controls. Adiponectin decreased significantly during the course of withdrawal (T = 3.44, p = 0.001) while resistin serum levels showed a slight increase (T = -1.83, p = 0.071). In a multivariate approach the extent of alcohol craving was significantly associated with adiponectin but not with resistin serum levels in male patients (Beta = -0.255, p = 0.025). Results for female patients were not significant. Our findings provide first evidence for an alteration of file adipocytokines adiponectin and resistin during alcohol withdrawal. Furthermore, adiponectin may be involved in the neurobiology of alcohol craving, possibly via its effects on the hypothalamic circuits.

Hock B; Schwarz M; Domke I; Grunert VP; Wuertemberger M; Schiemann U et al. Validity of carbohydrate-deficient transferrin (%CDT), gamma-glutamyltransferase (gamma-GT) and mean corpuscular erythrocyte volume (MCV) as biomarkers for chronic alcohol abuse: A study in patients with alcohol dependence and liver disorders of non-alcoholic and alcoholic origin. Addiction 100(10): 1477-1486, 2005. (42 refs.)

Aim: To test the clinical performance of carbohydrate-deficient transferrin (%CDT), gamma-glutamyltransferase (gamma-GT) and mean corpuscular erythrocyte volume (MCV) as biomarkers for alcoholism with a special focus on patients suffering from liver diseases. Design: Well-characterized collectives of alcohol-dependent patients with current consumption (ALC patients, n = 101), and relevant control groups (115 social drinkers, 46 patients with unspecifically increased gamma-GT, 51 hepatitis patients and 20/31 patients with non-alcohol/alcohol-dependent liver cirrhosis) were included into the study. The Positive Alcohol Use Disorders Test (AUDIT) score, International Classification of Diseases version 10 (ICD-10)/Diagnostic and Statistical Manual version IV (DSM-IV) criteria and blood drawn within 4 days of last drinking were inclusion criteria for subjects with regular heavy drinking. %CDT was determined using an automated assay which recently had been completely modified. Findings: Median AUDIT scores of patients without/with regular heavy drinking were 1-3/27. The following medians/95th percentiles were obtained for %CDT: social drinkers 2.2/3.0, patients with unspecifically increased gamma-GT 2.1/3.0, hepatitis 2.0/4.4, non-alcohol-dependent liver cirrhosis 2.4/4.8, alcohol-dependent liver cirrhosis 3.0/5.9, ALC patients 3.9/14.9. Differences between patients without and with alcohol abuse were highly significant (P < 0.001). No differences in CDT values were found between males and females. There was no correlation between %CDT values, gamma-GT, MCV and the amount of alcohol consumed in ALC patients; 3.0%CDT (95th percentile social drinkers) is proposed as cut-off for the test used (Tina-quant((R))%CDT 2nd-generation). At this cut-off, the sensitivity for ALC patients was 73.3%, whereas gamma-GT/MCV had a sensitivity of 71.3%/64.4%. Multivariate analysis performed at 95% specificity resulted in an improvement of the sensitivity by combining %CDT with gamma-GT (83.2%). A further enhancement of the sensitivity to 88.1% was obtained by combination of %CDT, gamma-GT and MCV. The diagnostic specificity of %CDT calculated at the cut-off of 3% was 93.5% in patients with unspecifically increased gamma-GT, 88.2% in hepatitis patients and 70.0% in patients with non-alcohol-dependent liver cirrhosis. %CDT was more specific in these patient collectives than MCV, and especially more than gamma-GT (specificity in hepatitis 52.9%, and 35.0% in non-alcohol-dependent liver cirrhosis). Conclusion: %CDT is of high diagnostic value to support diagnosis of alcohol-use disorders. The specificity of this marker in patient groups with liver disorders is superior to the biomarkers gamma-GT and MCV.

Hoiseth G; Bernard JP; Stephanson N; Normann PT; Christophersen AS; Morland J et al. Comparison between the urinary alcohol markers EtG, EtS, and GTOL/5-HIAA in a controlled drinking experiment. Alcohol and Alcoholism 43(2): 187-191, 2008. (30 refs.)

Aim: Urinary ethyl glucuronide (EtG), ethyl sulfate (EtS), and the ratio between 5-hydroxytryptophol-glucuronide and 5-hydroxyindole-3-acetic acid (GTOL/5-HIAA) are all suggested as biomarkers for recent alcohol ingestion with longer detection times than measurement of ethanol itself. The aim of this controlled study was to compare the sensitivities and detection times of EtG, EtS, and GTOL/5-HIAA, after a single ingestion of ethanol. Methods: 0.5 g ethanol/kg body weight was ingested by 10 healthy male volunteers in a fasted state. Ethanol, EtG, EtS, and GTOL/HIAA levels were measured in urine samples collected during a 4550 h period. The total amount of ethanol excreted as EtG and EtS was also determined. Results: Urinary EtG, EtS, and GTOL/5-HIAA showed 100% sensitivity as biomarkers for recent drinking. Compared to ethanol testing in urine, the detection times for GTOL/5-HIAA were 5 h longer and for EtG and EtS 25 h longer. The maximum EtG concentrations were higher than for EtS in all subjects, and a higher fraction of the ethanol dose was excreted as EtG (median 0.019%) compared with EtS (median 0.011%). Conclusions: This study is the first controlled experiment comparing the time-courses for ethanol, EtG, EtS, and GTOL/5-HIAA in urine. In cases where surveillance of alcohol relapse is needed, measurements of urinary EtG and EtS are sensitive and specific alternatives to ethanol testing. The GTOL/5-HIAA ratio is equally sensitive but with a much shorter window of detection.

Imbert-Bismut F; Naveau S; Morra R; Munteanu M; Ratziu V; Abella A et al. The diagnostic value of combining carbohydrate-deficient transferrin, fibrosis, and steatosis biomarkers for the prediction of excessive alcohol consumption. European Journal of Gastroenterology and Hepatology 21(1): 18-27, 2009. (25 refs.)

Background and aim: The validity of biomarkers of excessive alcohol drinking (EAD) (30g/day or more), such as carbohydrate-deficient transferrin (CDT%), is confounded by liver disease severity. The aim was to improve the accuracy of the percentage of CDT by taking into account the presence of fibrosis and steatosis, estimated using biomarkers FibroTest and SteatoTest. Methods Three hundred and twenty consecutive patients, 97 with alcoholic liver disease (ALD), and 223 non-ALD, were included. In ALD, 58% had advanced fibrosis and 58% had steatosis; in non-ALD, 25% had advanced fibrosis and 25% had steatosis. Results The mean percentage of CDT was lower in ALD with advanced fibrosis [2.4 (SE=0.2)] versus without [4.1 (0.3) P<0.0001], and lower in ALD with steatosis versus without (2.4 vs. 3.9; P=0.0007). Among non-ALD, there was no difference in the percentage of CDT according to fibrosis or steatosis. gamma-glutamyl-transpeptidase was higher in patients with advanced fibrosis or with steatosis both in ALD and non-ALD. Aspartate aminotransferase/alanine aminotransferase (AST/ALT) was higher in ALD patients with fibrosis versus without (2.5 vs. 1.3 P<0.0001) but not in non-ALD 0.01 vs. 0.98). AST/ALT was higher in ALD patients with steatosis versus without (2.2 vs. 1.6 P=0.04) and the inverse was observed in non-ALD (0.6 vs. 1.1 P<0.0001). In the entire population the percentage of CDT, gamma-glutamyl-transpeptidase, AST/ALT was associated with EAD, the area under the receiver operating characteristic curve =0.89 (95% Cl: 0.84-0.93), 0.93 (0.89-0.93) and 0.77 (0.71-0.82). An algorithm combining the percentage of CDT, FibroTest and SteatoTest permitted to obtain area under the receiver operating characteristic curve=0.92 versus 0.88 for the percentage of CDT (P=0.004) with 874% of patients classified correctly. Conclusion: Biomarkers of EAD are confounded by fibrosis and steatosis. Accuracy of the percentage of CDT is significantly increased when combined with biomarkers of fibrosis and steatosis.

Javors MA; Johnson BA. Current status of carbohydrate deficient transferrin, total serum sialic acid, sialic acid index of apolipoprotein J and serum beta-hexosaminidase as markers for alcohol consumption. Addiction 98(Supplement 2): 45-50, 2003. (13 refs.)

Aims: The purpose of this paper is to present a brief review of the literature and to summarize the current status of four biochemical markers for alcohol consumption, carbohydrate deficient transferrin (CDT), total serum sialic acid (TSA), sialic acid index of apolipoprotein J (SIJ) and serum -hexosaminidase (-HEX). Findings: Of these markers, CDT has been the most widely studied, is currently thought to be the most accurate predictor of alcohol consumption, is most readily available and is the only test approved by the FDA for the identification of heavy alcohol use. TSA and SIJ have the potential to be useful markers, but have only recently been discovered, are not readily available and have not yet been studied comprehensively. Finally, the relationship between serum -HEX and heavy alcohol consumption has been studied for about 20 years, but the test is not readily available and has not been widely accepted or used as a marker for heavy alcohol consumption. Conclusions: These markers have the potential to be included in a combination of measurements to provide an accurate, more exact assessment of alcohol consumption in a variety of clinical and research settings.

Johansen D; Andersen PK; Jensen MK; Schnohr P; Gronbaek M. Nonlinear relation between alcohol intake and high-density lipoprotein cholesterol level: Results from the Copenhagen City Heart Study. Alcoholism: Clinical and Experimental Research 27(8): 1305-1309, 2003. (26 refs.)

Background: It has been suggested that the level of high-density lipoprotein cholesterol (HDL-C) in the blood can be used as a marker of recent alcohol intake. However, before using HDL-C as a predictor of alcoholism, the relation between alcohol intake and HDL-C in the entire range of consumption must be explored. Most studies model the relation between alcohol intake and HDL-C linearly, although a threshold effect is expected. The objective of this study was to evaluate the shape of the relation between intake of alcohol and HDL-C and to determine whether there are differential effects of beer, wine, and spirits on HDL-C and whether they remain after adjusting for total alcohol. Method: The relation between alcohol intake and HDL-C was investigated by means of generalized additive models using data from the Copenhagen City Heart Study. Results: A nonlinear effect of alcohol improved the model fit significantly, and the nonlinearity of alcohol was highly significant in both men and women. The relation was concave: HDL-C was stable in men and women who drank more than approximately 35 and 20 drinks per week, respectively. We found a significant nonlinear term of wine on HDL-C in men after adjustment for total alcohol intake. Conclusions: There was a concave relation between alcohol intake and HDL-C, indicating a threshold effect of alcohol on HDL-C. The association between wine and HDL-C in men after adjusting for total alcohol intake may be due to residual lifestyle confounding.

Joseph AM; Hecht SS; Murphy SE; Carmella SG; Le CT; Zhang Y et al. Relationships between cigarette consumption and biomarkers of tobacco toxin exposure. Cancer Epidemiology, Biomarkers & Prevention 14(12): 2963-2968, 2005. (22 refs.)

Epidemiologic studies show a dose-response relationship between cigarettes per day and health outcomes such as heart and lung disease, and health outcomes are related to some biomarkers of tobacco exposure. The objective of this study was to examine the relationships between cigarettes per day and levels of selected biomarkers of tobacco toxin exposure: carbon monoxide (CO), metabolites of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and polycyclic aromatic hydrocarbons [total 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and 1-hydroxypyrene (1-HOP), respectively], and total cotinine (cotinine plus cotinine-N-glucuronide). We did a cross-sectional analysis of merged data from (a) two clinical trials and (b) two cohorts of light smokers (total n = 400). The mean age of participants was 50.4 years and the range of cigarette consumption was 1 to 100/d; however, few subjects smoked >45 cigarettes/d (n = 12). Results show that levels of the biomarkers CO, total NNAL, and total cotinine increase with an increase in the number of cigarettes smoked per day, but not in a linear fashion. 1-HOP is a less discriminating biomarker as levels are relatively stable regardless of the number of cigarettes smoked per day. There is considerable variability in toxin measurement, especially at high levels of smoking. There was a significant correlation between cigarettes per day and total NNAL, 1-HOP, total cotinine, and CO. Total NNAL was highly significantly correlated with total cotinine and CO and also significantly correlated with 1-HOP. These findings suggest that the number of cigarettes smoked per day is not necessarily a reliable measure of toxin exposure and may underestimate tobacco toxin exposure at low levels of smoking or overestimate exposure at high levels of smoking.

Junghanns K; Backhaus J; Tietz U; Lange W; Bernzen J; Wetterling T; Rink L; Driessen M. Impaired serum cortisol stress response is a predictor of early relapse. Alcohol and Alcoholism 38(2): 189-193, 2003. (24 refs.)

Aims: to investigate a possible association of cortisol stress response during early abstention with relapse. Methods: Thirty-six alcohol-dependent males, half of them with a comorbid anxiety disorder, and 15 healthy controls were exposed to a standardized psychosocial stress test. Thirty-one of the patients were assessed for relapse 6 weeks after discharge. Results: The relapsers showed almost no cortisol responses in the stress test. Comorbid anxiety disorder influenced neither stress response nor relapse. Conclusions: During early abstention from alcohol, reduced stress-responsivity of the hypothalamo-pituitary-adrenocortical axis seems to be connected to early relapse.

Kalivas PW; LaLumiere RT; Knackstedt L; Shen HW. Glutamate transmission in addiction. (review). Neuropharmacology 56(Supplement 1): 169-173, 2009. (79 refs.)

Cortico-striatal glutamate transmission has been implicated in both the initiation and expression of addiction related behaviors, such as locomotor sensitization and drug-seeking. While glutamate transmission onto dopamine cells in the ventral tegmental area undergoes transient plasticity important for establishing addiction-related behaviors, glutamatergic plasticity in the nucleus accumbens is critical for the expression of these behaviors. This information points to the value of exploring pharmacotherapeutic manipulation of glutamate plasticity in treating drug addiction.

Kapaki E; Paraskevas GP; Theotoka I; Liappas I. Neurochemical profile of elderly alcoholic patients with cognitive decline compared with patients with other dementias. International Journal on Disability and Human Development 6(2): 215-223, 2007. (58 refs.)

Alcoholism may result in impaired cognition and dementia. The increased risk of dementia in older individuals interferes with the differential diagnosis, especially when an elderly patient with a long history of alcohol abuse is the case. The aim of the present study was to evaluate the diagnostic value of the putative cerebrospinal fluid (CSF) biomarkers tau, beta-amyloid 1-42 (A beta 42) and their ratio in differentiating alcohol related dementia (ARD) from others of vascular or degenerative aetiology. Double-sandwich ELISAs (Innotest htau antigen and beta-Amyloid (142), Innogenetics) were used to quantify the above markers in a total of 151 patients and 82 controls. Patient groups comprised: 24 ARD, 17 vascular dementia, I I dementia with Lewy bodies, 23 frontotemporal dementia and 76 Alzheimer's disease (AD) patients. Tau protein succeessfully differentiated ARD from AD with 88% specificity and 86% sensitivity. A beta 42 alone had a specificity of 86% and a sensitivity of 70%, while tau/A beta 42 ratio was better than tau alone with corresponding values 100% and 91% respectively. For the discrimination of ARD from other dementias the diagnostic value of the above markers is substantially lower. In conclusion, the combined use of CSF tau and A beta 42 seems to be a useful tool in the differential diagnosis of ARD from AD, while in other primary dementias only a positive result may be useful.

Kaphalia BS; Cai P; Khan MF; Okorodudu AO; Ansari GAS. Fatty acid ethyl esters: Markers of alcohol abuse and alcoholism. Alcohol 34(2/3): 151-158, 2004. (34 refs.)

Chronic alcoholism, which is associated with hepatic, pancreatic, and myocardial diseases, is one of the major health problems in the United States with high morbidity and mortality. Many individuals who abuse alcohol chronically die even before reaching the clinical stage of the disease. Reliable biomarkers of the diseases induced by chronic alcohol abuse, as well as for alcoholism, currently are not available. In the current study, we measured plasma concentrations of fatty acid ethyl esters [(FAEEs), nonoxidative metabolites of ethanol] in 39 patients with a detectable concentration of alcohol in their blood samples. In turn, we determined the relation of FAEE concentrations with blood alcohol concentration (BAC). Of 39 patients in whom we evaluated this relation, only five had a history of chronic alcohol abuse, and six had a history of acute alcohol abuse. Patients' age ranged from 25 to 71 years. Within this age range, greater concentrations of FAEEs were found in the plasma samples obtained from patients in the 41- to 50-year age group. There were no sex-related differences in BAC, nor in FAEE concentrations. Thirteen patients had a BAC greater than 300 mg%. For 11 patients, the BAC ranged between 200 and 299 mg%, and, for 12 patients, the BAC ranged between 100 and 199 mg%. In comparison with Findings: for patients with a BAC that ranged between 100 and 299 mg%, the FAEE concentrations were approximately twofold higher in patients with a BAC greater than 300 mg%. Ethyl palmitate and ethyl oleate were the main FAEEs detected in most patients. In general, FAEE concentrations increased with increasing BAC. However, in comparison with patients with a history of acute alcohol abuse, a greater increase in total FAEE concentrations was observed in patients with a history of chronic alcohol abuse (4,250 ng/ml and 15,086 ng/ml, respectively). Fatty acid ethyl esters were either detected in trace amounts or not detectable in the plasma of control subjects with no known alcohol ingestion. These results support our hypothesis that nonoxidative metabolism of ethanol to FAEEs is an important pathway of ethanol disposition during chronic alcohol abuse, and that FAEE concentrations can be a more reliable biomarker of chronic alcohol abuse than a history of acute alcohol abuse.

Kim I; Darwin WD; Huestis MA. Simultaneous determination of nicotine, cotinine, norcotinine, and trans-3 '-hydroxycotinine in human oral fluid using solid phase extraction and gas chromatography-mass spectrometry. Journal of Chromatography B 814(2): 233-240, 2005. (31 refs.)

Nicotine is rapidly and extensively metabolized in humans. We present an analytical method to simultaneously quantify nicotine, cotinine, norcotinine. and trans-3'-hydroxycotinine in human oral fluid. Solid phase extraction (SPE) and GC/MS/EI with selected ion monitoring (SIM) were utilized. Linearity ranged from 5 to 1000 ng/mL of oral fluid; correlation coefficients for calibration curves were >0.99. Recoveries were 90-115% nicotine, 76-117% cotinine, 88-101% norcotinine, and 67-77% trans-3'-hydroxycotinine. Intra-assay precision and accuracy ranged from 1.6 to 5.7% and 1.6 to 17.8%, respectively. Inter-assay precision and accuracy ranged from 4.3 to 10.2% and 0 to 12.8%, respectively. Suitable precision and accuracy were achieved for the simultaneous determination of nicotine and three metabolites in the oral fluid of smokers. This assay is applicable to pharmacokinetic studies of nicotine, cotinine, and trans-3'-hydroxycotinine from tobacco smokers and can be utilized for routine monitoring of tobacco smoke exposure. 3-Hydroxycotinine requires additional investigation to determine its usefulness as a biomarker for tobacco smoke exposure.

Kim I; Wtsadik A; Choo RE; Jones HE; Huestis MA. Usefulness of salivary trans-3 '-hydroxycotinine concentration and trans-3 '-hydroxycotinine/cotinine ratio as biomarkers of cigarette smoke in pregnant women. Journal of Analytical Toxicology 29(7): 689-695, 2005. (32 refs.)

Nicotine is rapidly and extensively metabolized in humans and negatively impacts the developing fetus. The concentrations of nicotine, cotinine, trans-3'-hydroxycotinine (hydroxycotinine), and norcotinine in pregnant smokers' oral fluid were evaluated to determine usefulness as biomarkers of cigarette smoking. Sixteen participants were divided into two groups: eight light smokers (LS) who smoked < or =10 cigarettes/day and eight heavy smokers (HS) who smoked > or =20 cigarettes/day. Oral fluid specimens (n=415) were collected throughout pregnancy and analyzed with solid-phase extraction followed by gas chromatography-mass spectrometry-electron impact selected ion monitoring. Median concentrations of nicotine, cotinine, and hydroxycotinine in oral fluid of LS ranged from 241.1 to 622.0, 80.6 to 387.5, and 14.4 to 117.7 ng/mL and for HS 146.5-1372.2, 66.0-245.8, and 38.3-184.4 ng/mL, respectively. Salivary cotinine and hydroxycotinine concentrations were significantly correlated in LS (r = 0.55, p < 0.01) and HS (r = 0.74, p < 0.01). Ratios of hydroxycotinine/cotinine in oral fluid from pregnant women averaged 0.30 +/- 0.18 (range, 0.07-1.05) for LS and 0.68 +/- 0.25 (range, 0.29-1.83) for HS. Based on these preliminary data, the best ratio to differentiate light from heavy pregnant smokers was 0.41. Salivary hydroxycotinine and cotinine concentrations are both good biomarkers of cigarette smoking. Determining the hydroxycotinine/cotinine ratio may differentiate light from heavy tobacco use and help predict increased fetal tobacco exposure.

Kip MJ; Spies CD; Neumann T; Nachbar Y; Alling C; Aradottir S et al. The usefulness of direct ethanol metabolites in assessing alcohol intake in nonintoxicated male patients in an emergency room setting. Alcoholism: Clinical and Experimental Research 32(7): 1284-1291, 2008. (43 refs.)

Background: A major part of medical pathology in internal medicine is associated with chronic alcoholism. The aim of the current study was to investigate whether screening for Alcohol Use Disorders (AUD) can be improved through determination of direct ethanol metabolites compared to traditional biological state markers, the Alcohol Use Disorders Identification Test (AUDIT) and additional self-reports beyond the detection time period of a positive blood alcohol concentration (BAC). Methods: A total of 74 blood alcohol negative male patients who presented at the emergency room with either thoracic or gastrointestinal complaints were included. Phosphatidylethanol (PEth) was determined in whole blood, and ethyl glucuronide (EtG) in serum and urine samples. Traditional biological state markers [carbohydrate deficient transferrin (%CDT), gamma glutamyl transpeptidase (GGT), mean corpuscular volume (MCV)] were determined. The AUDIT was obtained and furthermore, all patients completed an additional self-report of alcohol consumption. Patients were divided into two (2) groups: AUDIT scores < 8 and AUDIT scores >= 8. Results: After assessment of the AUDIT, patients were allocated to one of the following groups: patients with AUDIT scores < 8 (n = 52) and with AUDIT scores >= 8 (n = 22). Twenty-five percent of the patients with AUDIT scores below the cut-off (n = 13/52) were tested positive for both PEth and UEtG. Of the patients who declared to be sober during the past 12 months, 38.5% were tested positive for PEth and UEtG. PEth discriminated similarly as %CDT for AUDIT scores >= 8 (AUC: 0.672; 95%CI 0.524 to 0.821). Self-reports of alcohol consumption were unreliable. Conclusion: Determination of direct ethanol metabolites such as PEth and UEtG provides additional evidence in screening for AUD in an ER setting. Determination of PEth might be considered complementary with or alternatively to %CDT.

Klein J; Blanchette P; Koren G. Assessing nicotine metabolism in pregnancy: A novel approach using hair analysis. Forensic Science International 145(2-3): 191-194, 2004. (32 refs.)

Nicotine and its metabolite cotinine are important biochemical markers to determine active 4 passive exposure to cigarette smoke. The amount of nicotine and cotinine in hair provides a cumulative index of tobacco exposure. This study uses segmental hair analysis to assess changes in nicotine metabolism in a cohort of pregnant women who smoked steadily during the whole gestational period according to their self-report. Maternal hair was collected at the time of delivery and sectioned into segments representing the three trimesters of pregnancy. The concentrations of nicotine and cotinine in each section of hair were measured by radioimmunoassay. Hair concentration of nicotine decreased during pregnancy without any reported reduction in smoking, while the cotinine concentration remained constant. The nicotine-cotinine ratio in these consistent smokers was: 1st trimester 20.6, 2nd trimester 19.9, and 3rd trimester 13.3. These findings are consistent with results from other studies showing increased nicotine metabolism during pregnancy. Our data suggest that the results of segmental hair analysis should be carefully examined in pregnancy. Both nicotine and cotinine levels should be evaluated in order to confirm any significant change in maternal tobacco exposure during pregnancy.

Klous MG; Rook EJ; Hillebrand MJX; van den Brink W; van Ree JM; Beijnen JH. Deuterodiacetylmorphine as a marker for use of illicit heroin by addicts in a heroin-assisted treatment program. Journal of Analytical Toxicology 29(6): 564-569, 2005. (17 refs.)

Delta-9-tetrahydrocannabinol (THC) is frequently found in the blood of drivers suspected of driving under the influence of cannabis or involved in traffic crashes. the present study used a double-blind crossover design to compare the effects of medium (16.5 mg THC) and high does (45.7 mg) of hemp milk decoctions or of a medium dose of dronabinol (20 mg synthetic THC, Marinol) on several skills required for safe driving. forensic interpretation of cannabinoids blood concentrations were attempted using the models proposed by Daldrug and Huestis and coworkers. First, the time concentration-profiles of THC, 11-dydroxy-THCin whole blood were determined by chromatography-mass- spectrometry-negative ion chemical ionization. compared to smoking studies, relatively low concentrations were measured in blood. the highest mean THC concentration (8.4 ng/mL) was achieved 1 h after ingestion of the strongest decoction. Mean maximum ``-OH-THC reached its highest concentration (66.2 ng/mL) 2.5-5.5 hours after intake. Individual blood levels showed considerable intersubject variability. the willingness to drive was influenced by the importance of the requested task. Under significant cannabinoids influence, the participants refused to drive when they were asked whether they would glee to accomplish several unimportant tasks (e.g., driving a friend to a party). Most of the participants reported a significant feeling of intoxication and did not appreciate the effects, notably those felt after drinking the strongest decoction. Road sign and tracking testing revealed obvious and statistically significant differences between placebo and treatments. A marked impairment was detected after ingestion of the strongest decoction. ACIF value, which relies on the molar ration of maina active to inactive cannabinoids, greater than 10 was found to correlate with a strong feling of intoxication. It also matched with a significant decrease in the willingness to drive, and it matched also with a significant impairment in tracking performances. The mathematic model II proposed by Huestis et al. provided at best a rough estimate of the time of oral administration with 27% of actual values being out of range of the 95% confidence interval. The sum of THC and 11-OH-THC blood concentrations provided a better estimate of impairment than THC alone. This controlled clinical study points out the negative influence on fitness to drive after medium or high dose oral THC or dronabinol.

Koch H; Meerkerk GJ; Zaat JOM; Ham MF; Scholten RJPM; Assendelft WJJ. Accuracy of carbohydrate-deficient transferrin in the detection of excessive alcohol consumption: A systematic review. (review). Alcohol and Alcoholism 39(2): 75-85, 2004. (58 refs.)

Aims: Excessive alcohol consumption is a common problem in society and medical practice. There is a need for a diagnostic tool with both high sensitivity and specificity for the detection of excessive alcohol consumption in unselected medical populations. Therefore, we evaluated the diagnostic accuracy of carbohydrate-deficient transferrin (CDT) in the detection of excessive alcohol consumption. Methods: Computerised literature searches in Medline, Embase and Current Contents databases (01/1966-06/2003) and reference checking. Articles on the detection of excessive alcohol consumption reporting CDT levels and self-reported alcohol consumption as a reference test were selected (n = 101). Studies concerning treatment, relapse detection and traffic offenders were excluded. Quality assessment and data-extraction was done by two reviewers independently. Only studies scoring positive on core validity criteria by Lijmer were eligible for quantitative analysis (n = 29). Results: Only two CDT-assays (CDTect and CDTriTIA) were evaluated in more than two high validity studies fulfilling the criteria for inclusion in the statistical analysis. Sensitivity of CDTect (14 data points) ranged from 20 to 85%, whereas specificity ranged from 77 to 95%. A summary ROC curve was computed for CDTect. Sensitivity of CDTTriTIA (4 data points) ranged from 10 to 67%, and specificity ranged from 90 to 100%. No summary measure could be computed for CDTTriTIA. The heterogeneity of results could not be explained clinically. Conclusions: The validity of CDT as a diagnostic tool is still questionable. If the higher values for sensitivity that some studies report can be confirmed by others it is a useful diagnostic tool in unselected populations. However, more methodologically sound, comparable studies need to be performed before firm conclusions can be drawn.

Korthuis PT; Zephyrin LC; Fleishman JA; Saha S; Josephs JS; McGrath MM; Hellinger J; Gebo KA. Health-related quality of life in HIV-infected patients: The role of substance use. AIDS 22(11): 859-867, 2008. (48 refs.)

HIV infection and substance use disorders are chronic diseases with complex contributions to health-related quality of life (HRQOL). We conducted a cross-sectional survey of 951 HIV-infected adults receiving care at 14 HIV Research Network sites in 2003 to estimate associations between HRQOL and specific substance use among HIV-infected patients. HRQOL was assessed by multi-item measures of physical and role functioning, general health, pain, energy, positive affect, anxiety, and depression. Mental and physical summary scales were developed by factor analysis. We used linear regression to estimate adjusted associations between HRQOL and current illicit use of marijuana, analgesics, heroin, amphetamines, cocaine, sedatives, inhalants, hazardous/binge alcohol, and drug use severity. Current illicit drug use was reported by 37% of subjects. Mental HRQOL was reduced for current users [adjusted beta coefficient -9.66, 95% confidence interval [(CI]) -13.4, -5.94] but not former users compared with never users. Amphetamines and sedatives were associated with large decreases in mental (amphetamines: beta = -22.8 [95% CI -33.5, -12.0], sedatives: beta = -18.6 [95% CI -26.2, -11.0]), and physical HRQOL (amphetamines: beta = -11.5 [95% CI -22.6, -0.43], sedatives: beta = -13.2 [95% CI -21.0, -5.36]). All illicit drugs were associated with decreased mental HRQOL: marijuana (beta = -7.72 [95% CI -12.0, -3.48]), non-prescription analgesics (beta = -13.4 [95% CI -20.8, -6.07]), cocaine (beta = -10.5 [95% CI -16.4, -4.67]), and inhalants (beta = -14.0 [95% CI -24.1, -3.83]). Facilitating sobriety for patients with attention to specific illicit drugs represents an important avenue for elevating HRQOL in patients living with HIV.

Kravos M; Malesic I. Kinetics and isoforms of serum glutamate dehydrogenase in alcoholics. Alcohol and Alcoholism 43(3): 281-286, 2008. (29 refs.)

Aims: The goal of this paper was to determine glutamate dehydrogenase's (GLDH) increased activity and rapid decrease in alcoholics according to last consumption of alcohol as well as to confirm that quick normalisation cannot be a sign of hepatocyte necrosis and that GLDH from rough endoplasmic reticulum exists in the serum of alcoholics. Methods: GLDH activity was assessed in 238 alcoholics admitted to hospital. A blood sample was taken from every subject three times: on admission to hospital, after 24 hours and after 7 days. We established our own reference activities for GLDH in serum, i.e., up to 124.0 nkat/l in males and 64.5 nkat/l in females. Results: Alcoholics were ascertained to have statistically significantly higher mean GLDH serum activities (men 351.53 nkat/L, women 296.08 nkat/L); the higher the level, the less elapsed time there was after the latest alcohol intake. There was an increased GLDH activity in 65.5% of alcoholics; furthermore, the percentage rose sharply to 72.2% with those who had last consumed alcohol within 48 hours. In the serum of alcoholics, it was found that, on average, it was 32.4% thermo-stable and 67.6% thermo-labile GLDH, which means that almost one third of GLDH serum originates from rough endoplasmic reticulum and rest from mitochondria. This is a completely new finding. Conclusions: A statistically significant fast decrease of GLDH serum activity after a break in alcohol consumption has been confirmed. It is estimated that increased GLDH activity in the sera of alcohol dependents and its fast decrease after total abstinence has been restored are specific for alcohol addiction.

Leitch DN; Harkawat R; Askew J; Masel P; Hendrick DJ. Relation of expired carbon monoxide to smoking history, lapsed time, TLCO measurement and passive smoking. Respiratory Medicine 99(1): 32-38, 2005. (17 refs.)

We quantified the influence of lapsed time, measurement of gas-transfer factor (TLCO), and passive smoking on expired carbon monoxide (CO) levels, and then evaluated the accuracy of smoking histories against expired CO measurements in patients newly attending 'occupational' compared with 'general' chest clinics. Expired CO levels had an estimated average rate of decline of 3.4ppm/h in the presumed absence of further smoking, though individual rates depended necessarily on the initial levels (2.1, 3.9, 5.7 and 7.5 ppm/h, respectively, when the initial levels were 10, 20, 30 and 40 ppm). TLCO measurement was associated with a median increase in expired CO of 4.0 ppm, but passive exposure to tobacco smoke in nonsmokers had negligible effect. Expired CO levels indicative of current smoking (> 8 ppm) were noted much more commonly in the current cigarette smokers (88%) than those who claimed to be current non-smokers (6.0%), but without significant difference between the non-smokers attending the occupational and general clinics (6.6% vs 5.3%). We conclude that the lapse of 1 h and the measurement of TACO exert mild but important influences on the expired CO level, but that passive smoking does not. 'Occupational' and 'general' patients give similarly false declarations of current non-smoking when presenting initially for clinical evaluation.

Litten RZ; Fertig J. Self-report and biochemical measures of alcohol consumption. Introduction to Special Supplement. Addiction 98(Supplement 2): iii-iv, 2003. (13 refs.)

Accurate measures of alcohol use are important in several realms -- for clinical assessment and screening of alcohol related problems as well as medical conditions aggravated or caused by alcohol. It is also important in the public health realm, such as biomarkers that can be used to monitor individuals with alcohol problems in occupations requiring abstinence to include medical personnel, airline and maritime pilots, air traffic controllers and railway engineers, as well as monitoring individuals convicted of alcohol-related traffic offenses as part of their probation requirement. Several features in these measures, that they be able to distinguish problematic drinking from light to moderate or abstinence. In addition, it must have a reasonable window of assessment, that is, provide a measure for a reasonable amount of time after drinking; be easy to use; acceptable to practitioners and patients; and easily incorporated in diverse settings. This Supplement describes progress in the development of such measures. A brief description and history of each measure and their strengths, weaknesses and appropriate applications are considered. The first several articles focus on self-report measures. The remainder consider biochemical measures of drinking.

Lo Muzio L; Falaschini S; Rappelli G; Bambini F; Baldoni A; Procaccini M et al. Saliva as a diagnostic matrix for drug abuse. International Journal of Immunopathology and Pharmacology 18(3): 567-573, 2005. (22 refs.)

Scientific interest in saliva as a diagnostic matrix has greatly increased over the last decade. The Triage (R) screening test (Biosite Diagnostics), a rapid immunological test used to detect recreational drugs in the urine, was used to compare two biological matrixes: a non-conventional one, saliva, and a traditional one, urine. Twenty-one drug abusers collected one urine and one saliva specimen, both of which were tested with the Triage kit. Data were validated by gas-chromatography-mass-spectrometry (GC-MS). Results were positive for methadone in 9 saliva and 14 urine specimens, for opiates in 2 and 10, respectively, and for barbiturates in 2 specimens. Saliva specimens were negative for cannabis, THC, benzodiazepines and tricyclic antidepressants, although the GC-MS analysis revealed low concentrations of these drugs in the saliva. The study demonstrates the possibility of using saliva as a diagnostic matrix to test for drug-taking; however, the Triage kit must be improved before being used with saliva.

Ma XY; Coles CD; Lynch ME; LaConte SM; Zurkiya O; Wang DL et al. Evaluation of corpus callosum anisotropy in young adults with fetal alcohol syndrome according to diffusion tensor imaging. Alcoholism: Clinical and Experimental Research 29(7): 1214-1222, 2005. (40 refs.)

Background: Fetal alcohol syndrome (FAS) and associated disorders resulting from maternal alcohol use during gestation are among the most common developmental disorders. However, they are rarely diagnosed and not fully understood in terms of their behavioral and neurocognitive phenotype. Prenatal exposure leads to alterations in facial morphology, growth, and neurocognition. The nature and extent of teratogenic effects on the brain and the relationship between such effects and observed behaviors remain in debate because there are no established markers for the neurological effects of exposure. In this study, we examined the impact of prenatal alcohol exposure on white-matter integrity in the corpus callosum by using diffusion tensor imaging (DTI) and herein describe the relationship between such effects and observed physical and behavioral outcomes. Methods: DTI was used to evaluate diffusion anisotropy in the genu and splenium of corpus callosum in 16 low-income, primarily African-American volunteers. Volunteers were recruited from a cohort of young adults who had received neuropsychological evaluations during adolescence. Nine had been prenatally exposed to alcohol and had characteristics of FAS, and seven were nonexposed controls. Results: Significant difference in the means for diffusion fractional anisotropy (t = 2.26, df = 9, p < 0.002) and apparent diffusion coefficient (t = 2.14, df = 14, p < 0.008) were observed in the corpus callosum of alcohol-exposed youth compared with nonexposed youth. No significant differences were found in intracranial volume between these groups. Conclusions: Our results illustrate that DTI can be used in evaluating the integrity of corpus callosurn in alcohol-exposed individuals. If future studies support these findings, diffusion anisotropy, represented by fractional anisotropy, has the potential to be used as a clinical marker in the diagnosis of FAS.

Mannino DM; Mulinare J; Ford ES; Schwartz J. Tobacco smoke exposure and decreased serum and red blood cell folate levels: Data from the Third National Health and Nutrition Examination Survey. Nicotine & Tobacco Research 5(3): 357-362, 2003. (34 refs.)

The aim of this cross-sectional study was to determine the effects of smoke exposure on serum and red blood cell folate levels. Data collected as part of the Third National Health and Nutrition Examination Survey were analyzed. Serum and red blood cell folate levels were measured in active smokers and nonsmokers with high, moderate, and low exposure to environmental tobacco smoke. After adjusting for dietary intake of folate and other covariates, we found that both smokers and nonsmokers with high smoke exposure had lower red blood cell folate levels than did nonsmokers with low smoke exposure (-86 nmol/l [95% confidence interval, CI, -101 to -71 nmol/l] for smokers; -50 nmol/l [95% CI -69 to -31 nmol/l] for nonsmokers with high smoke exposure, compared with nonsmokers with low smoke exposure). Similarly, after adjustment of dietary intake of folate and other covariates, the log serum folate level also was decreased (-0.29 log nmol/l [95% CI -0.33 to -0.25 log nmol/l] for smokers; -0.16 log nmol/l [95% CI -0.20 to -0.12 log nmol/l] for nonsmokers with high smoke exposure, compared with nonsmokers with low smoke exposure). Tobacco smoke exposure is associated with decreased folate levels, which may be a mechanism for some of the health effects of active and passive smoking.

Mayfield RD; Harris RA. Gene expression profiling in blood: new diagnostics in alcoholism and addiction? Neuropsychopharmacology 34(1): 250-251, 2009. (5 refs.)

Miller PM; Anton RF. Biochemical alcohol screening in primary health care. Addictive Behaviors 29(7 (Special Issue)): 1427-1437, 2004. (48 refs.)

Alcohol biomarkers such as carbohydrate-deficient transferrin (CDT) and gamma-glutamyltransferase (GGT) have significant potential for enhancing the quality of medical treatment in primary health care settings. Recent studies demonstrate that these laboratory tests can help the general practitioner in several ways. First, CDT and GGT can detect current heavy drinking in primary care patients with a fair degree of sensitivity (60% to 70%), with CDT being more specific (90%). When combined with self-report tests, they can provide a clinically useful alcohol screening battery. Second, elevated CDT and GGT levels have been correlated with specific alcohol-sensitive diseases (e.g., hypertension) and, as such, can serve as risk indicators for those diseases. Third, alcohol biomarkers have proven to be useful in monitoring the effectiveness of brief alcohol interventions with medical patients. Unfortunately, preliminary findings indicate that physicians have little knowledge of current biomarker research as applied to primary health care. Translational studies are needed on methods to facilitate knowledge and use of alcohol biomarkers by general practitioners.

Miller PM; Ornstein SM; Nietert PJ; Anton RF. Self-report and biomarker alcohol screening by primary care physicians: The need to translate research into guidelines and practice. Alcohol and Alcoholism 39(4): 325-328, 2004. (16 refs.)

Aims: To assess knowledge and use of alcohol self-report and biomarker screening by physicians. Methods: Forty-eight primary care physicians were surveyed. Results: Knowledge of MCV and GGT was as good as that for non-biomarker screening tools (CAGE, AUDIT) although use was significantly less. Knowledge and use of carbohydrate-deficient transferrin (CDT) was extremely low. Conclusions: Little translation of alcohol biomarker research into guidelines for primary care medicine has occurred. Most physicians report they would utilize these tests more frequently with additional knowledge about availability and use.

Miller PM; Spies C; Neumann T; Javors MA; Hoyumpa AM; Roache J et al. Alcohol biomarker screening in medical and surgical settings. Alcoholism: Clinical and Experimental Research 30(2): 185-193, 2006. (55 refs.)

This article highlights the proceedings of a symposium presented at the 28th Annual Meeting or the Research Society oil Alcoholism in Santa Barbara, CA, Oil June 28 2005, organized and chaired by Peter Miller. The presentations included (1) Screening for Alcohol Use Disorders in Surgical and Trauma Patients, presented by Claudia Spies; (2) Are Serum Levels or %CDT and GGT Related to Severity of Liver Biopsy Inflammation, Fibrosis, and Steatohepatitis in Patients with Hepatitis C? by Martin Javors; (3) Biochemical Alcohol Screening in the Treatment of Hypertension, presented by Peter Miller; and (4) The Cost-Effectiveness of a New Biomarker, CDT, in a Primary Care Sample, by Michael Fleming. Presentations were discussed by Raymond Anton.

Murphy SE; Link CA; Jensen J; Le C; Puumala SS; Hecht SS et al. A comparison of urinary biomarkers of tobacco and carcinogen exposure in smokers. Cancer Epidemiology, Biomarkers & Prevention 13(10): 1617-1623, 2004. (42 refs.)

Recently, several potential harm reduction strategies, such as reduction in the number of cigarettes smoked and the use of modified cigarette products, have been discussed as possible means by which to reduce tobacco-related disease. To assess any potential reduction in harm by either of these approaches requires an accurate assessment of tobacco toxin exposure. We have recently completed a cigarette reduction study in which smokers were required to reduce the number of cigarettes smoked by 75%. This reduction took place over a 6-week period. We report here the comparison of urinary concentrations of tobacco alkaloid and tobacco carcinogen biomarkers in a subset of these same smokers during a 7-week period prior to any reduction in cigarette consumption. Urine samples were collected at four time points and analyzed for 4(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), and its glucuronide, 1-hydroxypyrene, anatabine, free nicotine, total nicotine (free plus glucuronidated), free cotinine, total cotinine (free plus glucuronidated), and total trans-3'-hydroxycotinine (free plus glucuronidated). Anatabine is a minor alkaloid that may be useful in assessing tobacco exposure in individuals using nicotine replacement therapies. Urinary anatabine levels were well correlated (P < 0.0001) with both free and total nicotine (r = 0.753 and 0.773, respectively). Anatabine levels were also correlated with free cotinine (r = 0.465; P < 0.001), total cotinine (r = 0.514; P < 0.001), and total NALL (r = 0.633; P < 0.001). These data support the role of anatabine as a biomarker of tobacco exposure. 1-Hydroxypyrene is a biomarker of polycyclic aromatic hydrocarbon exposure, but unlike NNAL it is not tobacco specific. Whereas urinary concentrations of 1-hydroxypyrene were consistent across the four visits, the levels were not correlated with NNAL, anatabine, nicotine, or any nicotine metabolites.

Mustonen TK; Spencer SM; Hoskinson RA; Sachs DPL; Garvey AJ. The influence of gender, race, and menthol content on tobacco exposure measures. Nicotine & Tobacco Research 7(4): 581-590, 2005. (44 refs.)

Research has suggested that race, gender, and menthol cigarette use influence tobacco-smoke exposure measures and smoking-related disease risk. For example, a high proportion of Black smokers prefer menthol cigarettes and, despite smoking fewer cigarettes per day (CPD) than do Whites, tend to have higher cotinine levels. Additionally, Black males are more at risk for smoking-related lung cancer. High cotinine levels and smoking menthol cigarettes may lead to higher toxin intake, which contributes to increased disease risk. We explored the relationship between tobacco exposure variables (i.e., cotinine, CPD, carbon monoxide [CO], nicotine content, and nicotine dependence) with respect to race, gender, and menthol content in a sample of 307 smokers recruited from the greater Boston area to participate in a smoking cessation treatment trial. The pattern of correlations between tobacco exposure measures and cotinine showed a consistently positive correlation between cotinine and CO in all smokers and a correlation between cotinine and CPD in those who smoked nonmenthol cigarettes. Cotinine and CPD correlations varied by gender and race among menthol cigarette smokers. Consistently, we found a significant gender x race x menthol interaction on salivary cotinine level as well as cotinine/CPD ratio. These findings suggest that the relationship between number of cigarettes consumed and salivary cotinine is more complex than previously believed. It is not sufficient to look at race alone; researchers and clinicians need to look at race and gender concurrently, as well as type of cigarette consumed.

Neumann T; Spies C. Use of biomarkers for alcohol use disorders in clinical practice. Addiction 98(Supplement 2): 81-91, 2003. (129 refs.)

Background Biomedical markers may provide additive objective information in screening and confirmation of acute or recent consumption, intoxication, relapse, heavy drinking, hazardous/harmful use/abuse and dependence and alcohol use related organ dysfunction (alcohol use-related disorders: AUDs). Aims To review the use of biomarkers in clinical practice to detect AUDs. Findings About one-fifth of the patients seen in clinical practice have AUDs, which offer a variety of treatment options if diagnosed. The diagnosis of AUDs relies on clinical and alcohol-related history, physical examination, questionnaires and laboratory values. No clinical available laboratory test [e.g. for acute abuse: alcohol in blood or breath; for chronic alcohol abuse: -glutamyl transferase (GGT), mean corpuscular volume (MCV), carbohydrate-deficient transferrin (CDT)] is reliable enough on its own to support a diagnosis of alcohol dependence, harmful use or abuse. Sensitivities, specificities and the predictive values may vary considerably according to patient and control group characteristics (e.g. gender, age or related comorbidity). In patient groups with limited cooperation markers may be helpful when considering treatment options. Conclusions More research is needed to determine the value of markers (single or combined, with questionnaires) in the context of clinical decision-making algorithms in defined settings and with defined dichotomous outcome variables.

Nicolaou C; Chatzipanagiotou S; Tzivos D; Tzavellas EO; Boufidou F; Liappas IA. Serum cytokine concentrations in alcohol-dependent individuals without liver disease. Alcohol 32(3): 243-247, 2004. (25 refs.)

In the current study, our aim was to evaluate and investigate the influence of heavy alcohol intake on serum interleukin (IL)-6, IL-8, IL-10, IL-12, and tumor necrosis factor-alpha (TNF-) concentrations. The selection of cytokines was based on their presumptive role in the pathophysiology of alcohol dependence. On admission to the Drug-Free Substance Addiction Detoxification clinic ("ATHENA"), blood samples were obtained from study participants, and serum cytokine concentrations were measured by using a commercial sandwich enzyme-linked immunosorbent assay technique. Alcohol dependence, as diagnosed according to DSM-IV [Diagnostic and Statistical Manual of Mental Disorders (4th ed.)] criteria for alcohol dependence and estimated by using the Composite International Diagnostic Interview (CIDI), was characterized by increased serum IL-6 concentration. Interleukin-8, IL-10, IL-12, and TNF- concentrations were comparable to those found in control subjects (P > .05). These results indicate that in alcohol-dependent individuals there is a significant increase in the serum IL-6 concentration (P < .05).

Nissinen AE; Makela SM; Vuoristo JT; Liisanantti MK; Hannuksela ML; Horkko S et al. Immunological detection of in vitro formed phosphatidylethanol - An alcohol biomarker - With monoclonal antibodies. Alcoholism: Clinical and Experimental Research 32(6): 921-928, 2008. (45 refs.)

Background: Phosphatidylethanol (PEth) is a promising new marker for detecting long-term alcohol abuse with excellent sensitivity and specificity. Current methods are based on the high performance liquid chromatography-mass spectrometric method and therefore require high levels of expertise and expensive instrumentation. This study was designed to generate PEth-specific monoclonal antibodies for PEth immunoassay development. Methods: C57/BL6 mice were immunized with PEth in 3 different carriers, mouse serum albumin, mouse high-density lipoproteins, and human low-density lipoprotein (LDL). Mouse splenocytes were fused with a mouse myeloma cell line using the hybridoma technique. Mouse IgM-producing cell lines were selected by limiting dilutions. Binding characteristics of the anti-PEth antibodies were studied using luminometric immunoassays and sequence analysis of the variable region mRNA sequences of the antibodies. Produced antibodies were purified by chromatographic methods. PEth was detected with these antibodies in fluorescence immunoassay and flow cytometric analysis. Results: We generated monoclonal cell lines (2B1 and 2E9) that produce IgM antibodies binding specifically to PEth but not to structurally or chemically similar phospholipids, such as phosphatidylcholine, phosphatidic acid, and cardiolipin. We show here that these anti-PEth antibodies can be used to detect PEth in a fluorescent PEth assay and FACS analysis of human red blood cell samples spiked with PEth. Conclusions: The present study shows that PEth-specific monoclonal antibodies can be generated using traditional hybridoma technique. Immunogenicity of PEth was enhanced using human LDL as an immunization carrier. The generated monoclonal anti-PEth antibodies, 2B1 and 2E9 bind to PEth in fluid phase and in biological membranes.

Page-Sharp M; Hale TW; Hackett LP; Kristensen JH; Ilett KF. Measurement of nicotine and cotinine in human milk by high-performance liquid chromatography with ultraviolet absorbance detection. Journal of Chromatography B 796(1): 173-180, 2003. (31 refs.)

A high-performance liquid chromatographic (HPLC) assay for the determination of nicotine and cotinine in human milk was developed using an extraction by liquid-liquid partition combined with back extraction into acid, and followed by reverse-phase chromatography with UV detection of analytes. The assay was linear up to 500 mug/l for both nicotine and cotinine. Intra- and inter-day relative standard deviations (R.S.D.) were <10% (25-500 mug/l) for both nicotine and cotinine. Limits of quantitation (LOQ) were 10 and 12 mug/l for nicotine and cotinine, respectively, while the limits of detection (LOD) were 8 and 10 mug/l for nicotine and cotinine, respectively. The mean recoveries were 79-93% (range 25-500 mug/l) for nicotine and 78-89% (range 25-500 mug/l) for cotinine. The amount of fat in the milk did not affect the recovery. We found that this method was sensitive and reliable in measuring nicotine and cotinine concentrations in milk from a nursing mother who participated in a trial of the nicotine patch for smoking cessation.

Parzynski CS; Jaszyna-Gasior M; Franken FH; Moolchan ET. Measuring nicotine intake among highly-dependent adolescent smokers: Comparability of saliva and plasma cotinine concentrations. Pharmacology, Biochemistry and Behavior 89(2): 145-149, 2008. (35 refs.)

Cotinine is the most common biomarker used to assess nicotine exposure and abstinence. It can be measured in various matrices including saliva, plasma, and urine. Previous research with adults has shown high correlations between saliva and plasma cotinine concentrations. However, the research has not examined this relationship in adolescents. Additionally, variability in saliva flow and metabolism across gender, ethnicity, and age may impact the relationship between saliva and plasma cotinine concentration. Our aim was to examine the relationship between saliva and plasma cotinine concentration in a group of nicotine-dependent adolescent smokers. Additionally, we examined these correlations across gender, ethnicity and age. The sample consisted of 66 adolescent smokers (age 15.1 +/- 1.3, 63.6% girls, 66.7% European American, CPD 18.3 +/- 8.5, FTND 7.1 +/- 13). Saliva and plasma specimens were collected before the treatment phase of a nicotine replacement therapy trial and analyzed. The relationship between saliva and plasma cotinine concentration was analyzed using Pearson's correlation coefficients. We performed a secondary analysis using multiple regressions to compare correlations across race, gender and age. Results indicated a positive correlation between saliva cotinine and plasma cotinine concentration (r=0.84, p<0.001). Differences in correlations across age were significant (t=3.03, p<0.01). Differences across ethnicity approached significance (t=-1.93, p=0.058). Future research should seek to further validate saliva-to-plasma cotinine concentration ratios in adolescents as well as characterize saliva-to-plasma concentration differences and their underlying mechanisms.

Peterson K. Biomarkers for alcohol use and abuse: A summary. Alcohol Research & Health 28(1): 30-37, 2004. (30 refs.)

Clinicians can use several biochemical measurements to objectively assess patients' current or past alcohol use. However, none of these currently available biomarkers-including measures of various liver enzymes and blood volume are ideal. Several more experimental markers hold promise for measuring acute alcohol consumption and relapse. These include certain alcohol byproducts, such as acetaldehyde, ethyl glucuronide (EtG), and fatty acid ethyl esters (FAEE), as well as two measures of sialic acid, a carbohydrate that appears to be altered in alcoholics. Some progress has been made in finding markers that predict people's genetic predisposition to alcoholism, such as genetic differences in several neurotransmitters, including beta-endorphin and gamma-aminobutryic acid (GABA).

Public Domain

Rosalki SB. Carbohydrate-deficient transferrin: A marker of alcohol abuse. International Journal of Clinical Practice 58(4): 391-393, 2004. (2 refs.)

Serum carbohydrate-deficient transferrin (CDT) measurement is valuable for the identification of chronic excess alcohol consumption. CDT is increased in serum in approximately 50% of those classed as heavy drinkers and 70-80% of those defined as alcoholics. With abstinence, the serum CDT concentration reverts towards normal within approximately 2 weeks. Specificity for alcohol abuse ranges from 80 to 95% in the general population and from 70 to 80% in liver clinic patients.

Sarkar M; Kapur S; Frost-Pineda K; Feng SX; Wang JZ; Liang QW et al. Evaluation of biomarkers of exposure to selected cigarette smoke constituents in adult smokers switched to carbon-filtered cigarettes in short-term and long-term clinical studies. Nicotine & Tobacco Research 10(12): 1761-1772, 2008. (35 refs.)

Cigarette smoke is a complex aerosol that includes a gas vapor phase and a particulate phase. Inclusion of activated carbon in the cigarette filter can reduce some of the gas-phase smoke constituents implicated as toxicologically relevant. The present study evaluated exposure to selected gas-phase constituents when adult smokers switched to prototype cigarettes with a highly activated carbon filter. Smokers (N=160) in two separate studies were randomized to continue to smoke conventional cigarettes (either a 6-mg or 11-mg FTC tar product), to smoke test cigarettes containing carbon filters (comparable tar levels), or to stop smoking. After completing 8 days in controlled smoking conditions (short-term studies), smokers had the option to continue in 24-week long-term ambulatory studies with unrestricted smoking. Urinary excretion of mercapturic acid metabolites of 1,3-butadiene, acrolein, and benzene; nicotine and five of its metabolites, total NNAL, and 1-hydroxypyrene were measured at baseline in the conventional cigarette group, in all groups in the short-term studies, and every 4 weeks in the long-term studies. In the short-term studies, statistically significant reductions (70%, p.001) in gas-phase biomarker levels were observed in the test cigarette group for both tar level products compared with the conventional cigarette group. These reductions were similar to those observed in the stop-smoking groups. The reductions continued consistently (p.001) throughout the long-term studies. Switching to test cigarettes minimally affected the particulate-phase biomarkers. Statistically significant and consistent reductions in selected gas vapor phase biomarkers were observed when smokers switched to activated carbon filter cigarettes.

Schwan R; Albuisson E; Malet L; Loiseaux MN; Reynaud M; Schellenberg F et al. The use of biological laboratory markers in the diagnosis of alcohol misuse: an evidence-based approach. Drug and Alcohol Dependence 74(3): 273-279, 2004. (41 refs.)

Background: A large number of patients seen in clinical practice have an underlying alcohol problem. There is a pressing need for accurate methods to diagnose alcohol over-consumption objectively. Our aim was to determine how best to use biological markers to objectify alcohol problems in patients with clinical suspicion of alcohol misuse. Methods: A 6-month longitudinal multicenter trial was conducted, using four study groups (alcohol abusers, alcohol-dependents, healthy controls and consulting controls). CDT, GGT and MCV were measured. Statistical analyses used a computer learning system that created classification systems displayed in decision trees. Results: In 379 subjects the marker that best discriminated those with alcohol problems from controls was CDT. GGT then helped to differentiate between alcohol abuse and alcohol dependence in cases of high CDT. MCV, age and gender provided no extra information. Discussion: We recommend CDT as a first-line biological marker to confirm or disprove suspected alcohol misuse. High CDT plus GGT above normal points to alcohol dependence, while high CDT plus GGT below normal is evidence of alcohol abuse.

Segal M; Avital A; Rusakov A; Sandbank S; Weizman A. Serum creatine kinase activity differentiates alcohol syndromes of dependence, withdrawal and delirium tremens. European Neuropsychopharmacology 19(2): 92-96, 2009. (43 refs.)

Previous reports described significant differences in serum creatine kinase (CK) activity in bipolar disorder and various forms of depression. The comorbidity of depression and alcohol syndromes was also widely described. We aim to examine potential differences in serum CK level in different alcohol-related syndromes. We assessed morning serum CK activity in 114 inpatients, diagnosed by the Structured Clinical Interview for DSM-IV: Fifty-five subjects with alcohol dependence, 28 with alcohol withdrawal and 31 with delirium tremens (DTs). We found low normal CK activity for the alcohol dependence, higher for alcohol withdrawal and the highest for DT's. Peripheral CK activity of four patients that were admitted during each of the three phases showed similar pattern. These findings may be related to enhanced dopamine activity in alcohol dependence and conversely, to a significant decrease in dopamine activity during withdrawal syndromes. We suggest a supplementary simple laboratory tool for the detection of alcohol-related states.

Stevens KR; Munoz LR. Cigarette smoking: Evidence to guide measurement. Research in Nursing & Health 27(4): 281-292, 2004. (80 refs.)

Smoking cessation programs measure outcomes in terms of abstinence from or reduction in smoking. These outcomes can be measured through self-report by the smoker, through measurement with a biological marker of smoking, or through a combination of both. Consideration of the relative advantages of self-report and biomarker approaches is important in the selection of measurement strategies to evaluate outcomes in smoking cessation interventions. In this article both ways of measuring smoking behavior, self-report and biomarkers of carbon monoxide, cotinine, nicotine, thiocyanate, and alkaloids of nicotine, are explored. Measurement approaches are discussed in light of research evidence and their physiologic bases.

Swift R. Direct measurement of alcohol and its metabolites. (review). Addiction 98(Supplement 2): 73-80, 2003. (34 refs.)

Background: Ethanol can be easily and accurately measured in body fluids or vapours by several chemical and enzymatic methods. However, the persistence of ethanol in the body is short lived and ethanol's metabolism and distribution within the body are extremely complex. Aim: This article discusses the complex pharmacokinetics of ethanol and how pharmacokinetics can affect measured ethanol concentrations when ethanol is sampled across different individuals or from different body fluids or vapors, such as whole blood, plasma, breath, urine, saliva, or transdermal ethanol. Findings: Several physical methods for the quantification of ethanol concentrations are described, including gas chromatography, electrochemical detection, infrared detection and enzymatic methods, along with their advantages and limitations. Also discussed, is the utility of the measurement of several immediate metabolites of ethanol, including acetaldehyde, acetate, and fatty acid ethyl esters. Conclusions: The complex distribution and metabolism of ethanol and its metabolites can obscure the relationship between the ethanol concentration that is measured in body fluids or vapours and the amount of ethanol that is actually taken into the body.

Taracha E; Habrat B; Lehner M; Wislowska A; Woronowicz BT; Bogulas M et al. Alanine aminopeptidase activity in urine: A new marker of chronic alcohol abuse? Alcoholism: Clinical and Experimental Research 28(5): 729-735, 2004. (32 refs.)

Background: The purpose of this study was to evaluate the activity of urinary alanine aminopeptidase (AAP), the enzyme released from the brush border membranes of renal proximal tubules, as a new biological marker of chronic alcohol abuse.Methods: The AAP activity was assayed and compared between a group of 76 alcoholics undergoing detoxification and a group of 37 alcoholics abstaining from alcohol for at least 6 weeks. In all patients, the enzyme activity was measured both in untreated urine (uAAP) and after removal of endogenous AAP inhibitors by molecular filtration on Sephadex (eAAP).Results: There was a correlation between the uAAP and eAAP activities in both groups of patients (r = 0.61 and r = 0.81 in abstinent alcoholics and in alcoholics undergoing detoxification, respectively), and both the uAAP activity and the eAAP activity were significantly and markedly higher in alcoholics being detoxified than in their abstinent counterparts. As revealed by receiver operating characteristic analysis, the discriminative power of the eAAP activity assay was higher than that of the uAAP activity assay. The area under the corresponding receiver operating characteristic curves was 0.84 +/- 0.04 and 0.78 +/- 0.05 (mean SE), respectively.Conclusions: The results of this study demonstrate that the assays of urinary AAP activity, which relate to the nephrotoxic effects of alcohol abuse, could be a valuable complement to the other presently used markers of chronic alcohol abuse that are generally based on ethanol hepatotoxicity. Compared with the uAAP activity test, the eAAP activity test is of clear diagnostic advantage.

Thaqi A; Franke K; Merkel G; Wichmann HE; Heinrich J. Biomarkers of exposure to passive smoking of school children: Frequency and determinants. Indoor Air 15(5): 302-310, 2005. (46 refs.)

This study aims to assess the extent of children' exposure to ETS and quantify potential determinants. A total of 2767 children aged 5-14 years participated in an environmental survey in East Germany in 1998-1999 (participation rate 75.9%). A subgroup of 979 children between the ages of 11 and 14 years with complete data on nicotine and cotinine in urine were selected for this analysis. This study population consisted of 73 self-reported smokers (7.5%), 793 non-smokers (81%) and 113 children with missing data on smoking status (11.5%). Nicotine and cotinine concentrations in spontaneous urine sample were determined by high-performance liquid chromatography methods with ultraviolet-detection and corrected for creatinine. Approximately 40% of self-reported non-smokers were exposed to environmental tobacco smoke (ETS) at home. Non-smoking children exposed to parental tobacco smoke at home compared with not exposed showed in average higher nicotine and cotinine concentration (geometric mean 4.7 mu g/l vs. 1.4 mu g/l and 8.1 mu g/l vs. 2.7 mu g/l) and the adjusted odds ratio (OR) for detectable biomarkers ranged between 17 and 22. There were increased rates of detectable biomarkers in urine with increasing numbers of smoked cigarettes in the household (adjusted OR increased from 8 to 54). Maternal smoking showed a stronger effect than paternal smoking. Furthermore, low parental education, cold season, height of dwelling (<= 2.40), urine collected on Monday were statistically significant associated with high nicotine and cotinine excretion levels. Children exposed to parental smoke showed much higher biomarker levels than the non-smoking spouse of an adult smoker. Therefore, children need specifically protection from ETS at home.

Tomita M; Mizuno S; Yokota K. Increased levels of serum uric acid among ex-smokers. Journal of Epidemiology 18(3): 132-134, 2008. (10 refs.)

Background: It remains unclear whether serum uric acid level increases after the cessation of smoking. Methods: In 2000, we conducted a cross-sectional study on the effects of smoking cessation on serum uric acid levels by analyzing the results of annual health check-ups in the Japanese male working population (n = 16,642). Results: The serum uric acid level (6.18 mg/dL) was the highest in ex-smokers, followed by that in never-smokers (6.10 mg/dL) and that in current smokers (5.98 mg/dL). Ex-smokers weighed 0.6 kg more than the never-smokers and 1.5 kg more than the current smokers. The frequency of alcohol intake was closely correlated to the smoking habits. The serum uric acid levels declined in all groups, after adjustments for age, body mass index, and alcohol intake, though the levels in ex-smokers were 0.2 mg/dL higher than those in current smokers. Conclusion: The results suggested that alcohol intake contributed considerably to the serum uric acid levels and that smoking itself may have suppressed these levels via metabolic effects or the action of superoxides.

Uhl M; Sachs H. Cannabinoids in hair: Strategy to prove marijuana/hashish consumption. Forensic Science International 145(2-3): 143-147, 2004. (18 refs.)

Delta(9)-Tetrahydrocannabinol (THC) and 11-nor-Delta(9)-tetrahydrocannabinol-9-carboxylic acid (THCA) are equally used to indicate consumption of cannabis (hashish and marijuana). Publications of the early 90's demonstrate the possibilities of determining THC, cannabinol (CBN), and cannabidiol (CBD). All these substances are present in cannabis smoke and can be incorporated into the hair only by contamination. Generally, washing procedures should prevent false positive results, but finally it cannot be excluded that traces of THC may be found in hair after mere passive cannabis smoke exposure. Three authentic cases illustrate the problems originating in the exclusive determination of THC/CBN. The first example is the case of a couple living together in an apartment. Both persons' hair samples had been taken and gave positive results for THC and CBN. The male subject admitted smoking cannabis several times per day, but the female mate denied any consumption. Examination of the hair for THCA showed a high level (>6.6 pg/mg) in the sample of the male person and negative results (LOQ 0.1 pg/mg) in the sample of his mate. The second case hair is of a self admitted cannabis user's hair and was tested first by an immunoassay and GC/MS with a negative result. Nevertheless, the THCA concentration quantified in his sample was 2.7 pg/mg hair. The third hair sample is of a 2-year-old child that was tested positive for cannabis by using an immunochemical test. No THC and CBN were detectable by GUMS, however, trace amounts of THCA using GC/MS/MS. A comparative study of hair samples (screening for cannabinoids using ELISA test, THC determination by GUMS, THCA by GC/MS/MS) showed that only 26 segments of 66 were positive for both THC and THCA. Thirteen were negative for THC and positive for THCA, and six were positive for THC but negative for THCA. The cases were selected by an ELISA test or re-examined when the blood/urine results or the statement of the accused did not match with a THC outcome. The most appropriate strategy to prove cannabis consumption is immunochemical initial test followed by a GC/MS/MS confirmation of THCA.

Vaswani M; Desai NG. Alcohol dehydrogenase: An indicator of hepatic damage or marker in alcohol dependence. Addictive Disorders and Their Treatment 2(2): 53-58, 2003. (27 refs.)

Due to paucity of Indian data on ADH, this study examined its usefulness as a biological marker for alcohol dependence, and as an indicator of alcoholic liver dysfunction. The study did not investigate the presence or non-presence of alcoholic or non-alcoholic liver disease. Seventy alcoholic patients and 50 normal healthy controls were studied. A single point estimation of serum levels of alcohol dehydrogenase, aspartate amino transferase, alanine amino transferase, and gamma glutamyl transferase was performed. On the basis of severity of liver dysfunction, the study group was divided in two subgroups. Alcohol dehydrogenase levels were higher in patients of alcohol dependence compared with controls but did not reach significance levels. Using gamma glutamyl transferase and aspartate amino transferase as a reference standard, levels of alcohol dehydrogenase were significantly higher in alcoholic patients with severe liver dysfunction compared with those without severe liver dysfunction. Stepwise discriminant analysis revealed that aspartate amino transferase and gamma glutamyl transferase best identified alcoholic patients from normal controls with good diagnostic accuracy. Serum levels of alcohol dehydrogenase showed good sensitivity but poor specificity and low kappa value. The results provides evidence for use of serum ADH level as a useful indicator of alcoholic liver dysfunction, but does not support the hypothesis for it being a useful marker for alcohol dependence.

Whincup PH; Gilg JA; Emberson JR; Jarvis MJ; Feyerabend C et al. Passive smoking and risk of coronary heart disease and stroke: Prospective study with cotinine measurement. British Medical Journal 329(7459): 200-205, 2004. (30 refs.)

Objective: To examine the associations between a biomarker of overall passive exposure to tobacco smoke (serum cotinine concentration) and risk of coronary heart disease and stroke. Design Prospective population based study in general practice (the British regional heart study). Participants 4729 men in 18 towns Who provided baseline blood samples (for cotinine assay) and a detailed smoking history in 1978-80. Main outcome measure Major coronary heart disease and stroke events (fatal and non-fatal) during 20 years of follow up. Results: 2105 men Who said they did not smoke and who had cotinine concentrations <14.1 ng/ml were divided into four equal sized groups on die basis of cotinine concentrations. Relative hazards (95% confidence intervals) for coronary heart disease in the second (0.8-1.4 ng/ml), third (1.5-2.7 ng/ml), and fourth (2.8-14.0 ng/ml) quarters of cotinine concentration compared with the first (&LE;0.7 ng/ml) were 1.45 (1.01 to 2.08),1.49 (1.03 to 2.14), and 1.57 (1.08 to 2.28), respectively, after adjustment for established risk factors for coronary heart disease. Hazard ratios (for cotinine 0.8-14.0 v &LE;0.7 ng/ml) were particularly increased during the first (3.73, 1.32 to 10.58) and second five year follow up periods (1.95, 1.09 to 3.48) compared with later periods. There was no consistent association between cotinine concentration and risk of stroke. Conclusion: Studies based on reports of smoking in a partner alone seem to underestimate the risks of exposure to passive smoking. Further prospective studies relating biomarkers of passive smoking to risk of coronary heart disease are needed.

Wilhelm J; Bayerlein K; Hillemacher T; Reulbach U; Frieling H; Kromolan B et al. Short-term cognition deficits during early alcohol withdrawal are associated with elevated plasma homocysteine levels in patients with alcoholism. Journal of Neural Transmission 113(3): 357-363, 2006. (36 refs.)

Higher plasma homocysteine levels have been found in actively drinking alcoholics as well as in early abstinent patients. Furthermore, elevated homocysteine levels are associated with cognitive decline in dementia and in healthy elderly people. The aim of this prospective study was to investigate a possible association between homocysteine serum levels and clinically well known cognitive deficits during alcohol withdrawal. We examined 89 patients (67 men, 22 women) during early withdrawal treatment. Cognitive function was assessed using the c.f.-test. Patients with cognitive deficits showed significantly higher homocysteine serum levels (Mann-Whitney-U, p = 0.004) than patients without cognitive deficits, while the difference in blood alcohol concentration was not significant. Using logistic regression analysis, cognitive deficits were best predicted by high homocysteine serum levels (Wald chi(2) = 4.071, OR = 1.043, 95% CI 1.001-1.086, p < 0.05), which was confirmed by Receiver Operating Curves (AUC = 0.68, 95% CI = 0.57-0.79, p = 0.004). The present results show first evidence of an association between elevated plasma homocysteine levels in alcoholics and cognition deficits in patients undergoing alcohol withdrawal.

Wurst FM; Alling C; Araclottir S; Pragst F; Allen JP; Weinmann W. Emerging biomarkers: New directions and clinical applications. Alcoholism: Clinical and Experimental Research 29(3): 465-473, 2005. (43 refs.)

This article summarizes content proceedings of a symposium held at the 2004 Research Society on Alcoholism Scientific Annual Meeting in Vancouver, Canada. The chairs were Friedrich M. Wurst and Raye Litten. The presentations were (1) Introduction, by Raye Litten; (2) Direct Ethanol Metabolites-On the Threshold From Science to Routine Use, by Friedrich M. Wurst; (3) Sialic Acid Index of Plasma Apolipoprotein J (SIJ) as a Viable Marker for Chronic Alcohol Consumption, by Philippe Marmillot; (4) The Emergence of Ethyl Glucuronide (EtG) Testing as a Tool in Monitoring Healthcare Professionals, by Gregory E. Skipper; (5) Application of Biomarkers for Alcohol Use Disorders in Clinical Practice, by Tim Neumann; (6) Utility of Biomarkers in Assessing the Efficacy of Medications for Treating Alcoholism, by Marty Javors; and (7) Discussion, by Raye, Litten.

Wurst FM; Bechtel G; Forster S; Wolfersdorf M; Huber P; Scholer A et al. Leptin levels of alcohol abstainers and detoxification patients are not different. Alcohol and Alcoholism 38(4): 364-368, 2003. (38 refs.)

Aims: Leptin is a cytokine-type peptide hormone, recently implicated as a putative state marker of alcohol use and in craving. Our goal was to evaluate the potential of leptin as a state and trait marker and to rule out the role of current alcohol intoxication on leptin levels. Methods: Eighteen alcohol withdrawal patients (16 males, 2 females) whose blood contained 202 mg/dl (median) of ethanol at hospitalization, who had a median age of 43.5 years and had consumed 1075 g of ethanol (median) in the last 7 days were included in the study. Leptin was determined in samples at day 1 (when still intoxicated) and day 7 of withdrawal. Expected leptin levels were calculated with a formula. For comparison, 27 blood samples of 18 abstinent persons, matched for gender, age and body mass index were used. Furthermore, mean cell volume, gamma-glutamyl transferase (GGT), blood glucose, cholesterol, triglycerides and body composition (bioimpedance device) were determined. For statistical analysis, SPSS 11 was used. Results: Expected leptin levels were 1.71 ng/ml (median), leptin measured at day 1 was 2.65 ng/ml (median) and 2.85 ng/ml on day 7 for the alcohol withdrawal patients and 2.2 ng/ml (median) for the abstainers. These concentrations were not significantly different. Significant correlations were found between leptin day 1 and expected leptin levels, percentage fat body mass, cigarettes smoked per day, GGT and blood alcohol concentration. Conclusions: Our preliminary data do not support the hypothesis of leptin as a state or trait marker and suggest only a minor influence of acute intoxication on leptin levels in alcohol detoxification patients.

Wurst FM; Dresen S; Allen JP; Wiesbeck G; Graf M; Weinmann W. Ethyl sulphate: A direct ethanol metabolite reflecting recent alcohol consumption. Addiction 101(2): 204-211, 2006. (37 refs.)

Background: Ethyl sulphate (EtS), a direct ethanol metabolite, appears to offer potential as a biomarker for recent alcohol consumption. Although its window of assessment is similar to that of ethyl glucuronide (EtG), there are differences between the two markers in their pathways for formation and degradation. Aims: (a) To assess the excretion of EtS compared to EtG and ethanol in drinking experiments with healthy volunteers, and (b) to elucidate the possibility of using the two metabolites for monitoring abstinence in substance use disorder patients during rehabilitation treatment. Design, setting, participants (a) Nine drinking experiments were performed by six healthy volunteers (two females, four males), with a mean age of 34.1 years (20-62), average oral intake of 0.2 g/kg ethanol (0.1-0.61), and having 74 spot urine samples. (b) Thirty-six substance abuse patients (mean age 41.9 years, 20-59; 22 males, 14 females) in a rehabilitation programme after withdrawal, producing 98 urine samples. Ethyl glucuronide and ethyl sulphate were measured using liquid chromatography tandem mass spectrometry (LC-MS/MS) using d(5)-EtG and d(5)-EtS, respectively, as an internal standard. Findings: (a) Volunteers: EtG and EtS were detectable for up to 36 hours and reached the limits of determination in urine at 20.6 hours and 21.2 hours (median), respectively, after ethanol intake. EtG-100 (standardized to a creatinine of 100 mg/dl) reached its maximum level at 2.8 hours and EtS-100 at 2.1 hours (median) after the beginning of the experiment. Of the ethanol ingested, 0.022% was excreted as EtS in one volunteer. Eight samples were positive for EtS only and six for EtG only. Spearman's rank correlation coefficients of 0.84 (P < 0.0001) between EtG and EtS and 0.87 (P < 0.0001) between EtG-100 and EtS-100 were found. (b) Patients: of the 98 urine samples evaluated, 27 were positive for EtS and of these only 20 were also positive for EtG. Spearman's rank correlation coefficients of 0.84 (P < 0.0001) between EtG and EtS and 0.82 (P < 0.0001) between EtG-100 and EtS-100 were found. Conclusions: The data from patients and volunteers suggest that the direct ethanol metabolite ethyl sulphate has the potential to serve as a biomarker of recent ethanol intake. Because EtG and EtS are formed via different pathways they might be used conjointly, thereby increasing sensitivity.

Wurst FM; Dursteler-MacFarland KM; Auwaerter V; Ergovic S; Thon N; Yegles M et al. Assessment of alcohol use among methadone maintenance patients by direct ethanol metabolites and self-reports. Alcoholism: Clinical and Experimental Research 32(9): 1552-1557, 2008. (54 refs.)

Background: Heavy alcohol consumption may accelerate the progression of hepatitis C (HCV)-related liver disease and/or limit efforts at antiviral treatment. As most of the patients in methadone maintenance treatment (MMT) suffer from hepatitis C infection, this study was conducted to identify the alcohol intake among these patients at a Swiss Psychiatric University Clinic by self-reports and direct ethanol metabolites as biomarkers of ethanol consumption. Patients and Methods: A convenience sample of 40 MMT patients (15 women, 25 men; median age 39 years) of the total 124 patients was asked and consented to participate in this study. This sample was not different in age, gender distribution, and rate of hepatitis C infection from the total sample. The Alcohol Use Disorders Identification Test (AUDIT) and self-reported ethanol intake during the previous 7 days were assessed. In addition, ethyl glucuronide (EtG) in urine, and fatty acid ethyl esters (FAEEs) and EtG in hair were determined using LC-MS/MS and gas chromatograph/mass spectrometer. The limit of quantitation for UEtG, HEtG, and FAEEs were 0.1 mg/l, 2.3 pg/mg, and 0.1 ng/mg, respectively. Results: Fourteen participants reported abstinence from alcohol for the previous 7 days. AUDIT scores were >= 8 in 15 male and > 5 in 5 female participants. Direct ethanol metabolites were as follows (median, min, max, standard deviation): UEtG (19 positives; 9.91, 1.38 to 251, 62.39 mg/l); the values of HEtG were 17.65, 0 to 513, 105.62 pg/mg [in 2 cases no material, 8 abstinent (up to 7 pg/mg), 15 social drinkers (up to 50 g per day), and 15 excessive users (> 50/60 g/d)]. For the 13 cases, where enough material for additional determination of HFAEEs was available, the values were 0.32, 0 to 1.32, 0.44 ng/mg. Among the 30 HEtG-positive participants, 20 had not reported the corresponding ethanol intake using question 1 (frequency) and 2 (quantity) of the AUDIT. Of the 14 participants reporting no alcohol intake during the previous 7 days, 4 were UEtG-positive. HEtG and AUDIT correlated significantly (r = 0.622, p < 0.0001), but this was not the case for UEtG and self-reported ethanol intake during the previous 7 days. Conclusion: (1) HEtG identified 20 cases of daily ethanol intake of more than 20 g, that would have been missed by the sole use of question 1 (frequency) and 2 (quantity) of the AUDIT. (2) Using the total score of the AUDIT, HEtG confirmed 10 more cases positive for alcohol intake. (3) Episodic heavy drinking is with 22.5% more frequent than in general population, and (4) of the 14 participants who reported no alcohol intake during the previous 7 days, 4 were UEtG positive. Improved detection of alcohol consumption, which is hazardous or harmful in the context of HCV and opiate dependence, would allow for earlier intervention in this population which is at particular risk of liver disease and fatal respiratory-depressed overdose. The combined use of self-reports and direct ethanol metabolites seems promising.

Wurst FM; Haber PS; Wiesbeck G; Watson B; Wallace C; Whitfield JB et al. Assessment of alcohol consumption among hepatitis C-positive people receiving opioid maintenance treatment using direct ethanol metabolites and self-report: A pilot study. Addiction Biology 13(3/4): 416-422, 2008. (42 refs.)

This study was conducted to identify the alcohol consumption among hepatitis C-positive people receiving opioid maintenance therapy using self-report and biomarkers. A total of 49 people (28 male, 21 female) were hepatitis C virus (HCV) positive and were included. The alcohol use disorder identification test (AUDIT) and self-reported ethanol intake in the last 28 days were assessed. In addition to gamma-glutamyl-transferase (GGT) and mean corpuscular volume (MCV), ethyl glucuronide (EtG) and ethyl sulphate (EtS) were determined in serum and urine (UEtG, UEtS, SEtG) using liquid chromatography/tandem mass-spectroscopy (LC/MS-MS) with deuterated internal standards. Abstinence from alcohol was reported for the last 28 days by 13 participants and for the last 7 days by 22. AUDIT was > 8 in 27 cases. The maximum values were 34.8 mg/l for UEtG, 5.3 mg/l for UEtS and 0.15 for SEtG. Among the 19 UEtG positives, 8 had not reported any ethanol intake in the 7 days prior to the study. Six participants reported intake of up to 320 g of ethanol in the last 7 days, but were negative for SEtG, UEtG and UEtS. Self-reported ethanol intake in the last 28 days correlated with AUDIT score (r = 0.733, P < 0.001), with the direct ethanol metabolites and MCV. In this population, abstinence and episodic heavy drinking are more common than in the general population. Episodic heavy drinking is a significant cause of acute risk in this population. Results from biomarker testing could indicate cases of under- as well as over-reporting of alcohol consumption. Further research on the diagnostic accuracy of direct ethanol metabolites, including the use of phosphatidylethanol (PEth), in this setting is needed.

Wurst FM; Skipper GE; Weinmann W. Ethyl glucuronidethe direct ethanol metabolite on the threshold from science to routine use. Addiction 98(Supplement 2): 51-62, 2003. (58 refs.)

Aims: Current biological state markers remain suboptimal with regard to sensitivity and specificity for monitoring recent alcohol consumption in various settings. Furthermore, these biomarkers can be influenced by age, gender and a variety of substances and non-alcohol-associated diseases and do not cover fully the time axis for alcohol intake. Ethyl glucuronide (EtG) is a non-volatile, water-soluble, stable, direct metabolite of ethanol that can be detected in various body fluids, tissues and hair. Shortly after the consumption even of small amounts of ethanol, EtG becomes positive. It can detect ethanol intake up to 80 hours after the complete elimination of alcohol from the body, covering a unique and important time spectrum for recent alcohol use. EtG seems to meet the need for a sensitive and specific marker to elucidate alcohol use not detected by standard testing. Design, setting, participants, methods and findings: The literature was re-viewed with a focus on possible diagnostic and therapeutic applications, currently available methods and future perspectives. To date, more than 4000 samples of body fluids, tissues and hair from approximately 1500 individuals have been assessed. Conclusions: The data suggest that EtG is a useful tool in numerous settings, including alcohol and drug treatment (to detect lapse/relapse and for motivational feedback), in safety sensitive work settings where use is dangerous or in other settings where alcohol use may be risky (e.g. such as driving, work-place, pregnancy or monitoring physicians or other professionals who are in recovery and working) or for resolving forensic questions. If the question of recent alcohol consumption has to be answered in a binary way (yes/no), such as for determining lapses, the use of EtG in urine is among the preferred tests. The use of this marker alone and complementary with other biological state markers and self-reports is expected to lead to significant improvement in treatment outcome, therapy efficacy and cost reduction.

Wurst FM; Tabakoff B; Alling C; Aradottir S; Wiesbeck GA; Muller-Spahn F et al. World Health Organization/International Society for Biomedical Research on Alcoholism study on state and trait markers of alcohol use and dependence: Back to the future. Alcoholism: Clinical and Experimental Research 29(7): 1268-1275, 2005. (50 refs.)

This article summarizes content proceedings of a symposium held at the 2004 International Society for Biomedical Research on Alcoholism Congress in Mannheim, Germany. The chairs were Boris Tabakoff and Friedrich M. Wurst. The presentations were (1) Genetic associations with alcoholism and affective disorders, by Paula Hoffman; (2) Proteomic analysis of blood constituents in alcoholism, by Boris Tabakoff; (3) Contrasts between the responses of GGT and CDT to high alcohol intake, and a test of their combined use, by John Whitfield; (4) Direct ethanol metabolites such as ethyl glucuronide, fatty acid ethyl esters, phosphatidylethanol and ethyl sulfate: a new line of sensitive and specific biomarkers, by Friedrich Martin Wurst; and (5) Genetic studies of alcoholism subtypes in a Han Taiwanese population, by Ru-Band Lu.

Wurst FM; Wiesbeck GA; Metzger JW; Weinmann W; Graf M; WHO ISBRA Study Biological State. On sensitivity, specificity, and the influence of various parameters on ethyl glucuronide levels in urine: Results from the WHO/ISBRA study. Alcoholism: Clinical and Experimental Research 28(8): 1220-1228, 2004. (50 refs.)

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.

Yegles M; Labarthe A; Auwarter V; Hartwig S; Vater H; Wennig R et al. Comparison of ethyl glucuronide and fatty acid ethyl ester concentrations in hair of alcoholics, social drinkers and teetotallers. Forensic Science International 145(2-3): 167-173, 2004. (14 refs.)

In previous investigations hair analysis for ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEE) proved to be suitable for the detection of excessive alcohol consumption. The aim of this study was to compare EtG and FAEE concentrations in hair of alcoholics, social drinkers and teetotallers. Hair samples from 10 alcoholics in withdrawal treatment, 11 fatalities with documented excessive alcohol consumption, four moderate social drinkers who consumed up to 20 g ethanol per day, and three strict teetotallers were analysed. After external degreasing with n-heptane, extraction with a dimethyl sulfoxide/n-heptane mixture and headspace solid-phase microextraction of the extracts, four fatty acid ethyl esters (FAEEs) (ethyl myristate, ethyl palmitate, ethyl oleate and ethyl stearate) were analysed by gas chromatography-mass spectrometry (GC-MS) with deuterated internal standards. FIG was determined by GC-MS/NCI after ultrasonication of the samples with H2O, cleanup by SPE with aminopropyl columns and PFP derivatisation. The following concentrations were measured for the four groups: teetotallers EtG < 0.002 ng/mg, FAEE 0.05-0.37 ng/mg, moderate social drinkers EtG < 0.002 ng/mg, FAEE 0.26-0.50 ng/mg, alcoholic patients EtG 0.030-0.415 ng/mg, FAEE 0.65-20.50 ng/mg and the fatalities with alcohol history FIG 0.072-3.380 ng/mg, FAEE 1.30-30.60 ng/mg. The results confirm that by using a cut-off value of the sum of FAEE > 1 ng/mg and/or a positive EtG result in hair, excessive alcohol consumption can be identified using hair analysis. However, no significant correlation between the FIG and FAEE concentrations in the positive cases could be shown. Segmental analysis of some of the specimens did not reveal the same distribution for EtG compared to FAEE in hair, and no chronological accordance compared to the self-reported alcohol consumption could be observed for both parameters. These different results of both methods are discussed in terms of differences between EtG and FAEE in mechanism of formation and incorporation into hair and elimination from hair.

Yokoyama M; Yokoyama A; Yokoyama T; Hamana G; Funazu K; Kondo S; Yamashita T; Yoshimizu H; Nakamura H. Mean corpuscular volume and the aldehyde dehydrogenase-2 genotype in male Japanese workers. Alcoholism: Clinical and Experimental Research 27(9): 1395-1401, 2003. (27 refs.)

Background: Increased mean corpuscular volume (MCV) is common in alcohol abusers and alcoholics. MCV is higher in Japanese heavy drinkers with inactive aldehyde dehydrogenase-2 (ALDH2) encoded by ALDH2*1/2*2 than among those with active ALDH2 encoded by ALDH2*1/2*1. Inactive A-LDH2 dramatically increases blood acetaldehyde levels after alcohol intake. Because moderate and heavy drinkers with ALDH2*1/2*2 have very high risks for esophageal cancer, MCV might serve as an indicator of these high-risk drinkers. Methods: In this investigation of the association of red cell values with the ALDH2 genotype and possible confounding factors, the drinking, smoking, and dietary habits reported on a structured questionnaire by 163 Japanese working men were subjected to multivariate analyses. Results: Aging, lower body mass index (BMI), more alcohol consumption, and more smoking were positively associated with increased MCV. Among moderate to heavy drinkers (greater than or equal to9 units/week; 1 unit = 22 g of ethanol), both MCV and mean corpuscular hemoglobin were higher and the red cell count was lower in those with ADLH2*1/2*2 than in those with ALDH2*1/2*1. Multiple linear regression analysis after adjustment for age, BMI, and smoking revealed that a positive relationship between the amount of drinking and MCV but inverse relationships for drinking and red cell count, as well as hemoglobin and hematocrit values, were significantly stronger for men with ALDH2*1/2*2 than for those with ALDH2*1/2*1, demonstrating a gene-environment interaction. Drinking accounted for 19.9% of interindividual MCV variance among men with ALDH2*1/*2*2 but for only 1.3% of variance among those with ALDH2*1/2*1. Age, BMI, drinking, and smoking accounted for 52.1 and 34.7% of the variation among those with ALDH2*1/2*2 and ALDH2*1/2*1, respectively. Macrocytosis (MCV greater than or equal to100.0 fl) was observed in 18 subjects (11.0%), and use of macrocytosis as a biomarker of moderate to heavy drinkers with ALDH2*1/2*2 had a sensitivity of 54.5% (6 of 11) and a specificity of 92.1% (140 of 152).Conclusions: Alcohol-related red cell value changes associated with inactive ALDH2 in Japanese men suggest the importance of acetaldehyde's role in increasing MCV and the potential for using MCV as a marker for high-risk drinkers for esophageal cancer.

Young RM; Lawford BR; Nutting A; Noble EP. Advances in molecular genetics and the prevention and treatment of substance misuse: Implications of association studies of the A"1 allele of the D"2 dopamine receptor gene. Addictive Behaviors 29(7 (Special Issue)): 1275-1294, 2004. (110 refs.)

Substance misuse is influenced by multiple genetic and environmental factors. Recent research has identified a number of potential genetic markers of risk and those associated with drug reward substrates show particular promise. The current study reexamines the extant published data of the association between the D2 dopamine receptor (DRD2) gene minor Taq 1A (A1) allele and substance misuse risk. A series of meta-analyses was performed on 64 studies examining DRD2 A1+ allelic status and substance misuse. In addition, personality was examined as a possible endophenotype. Significant association was found between the A1 allele and severe substance dependence in both Caucasian and non-Caucasian groups. The data did not support a significant association between the A1 allele and personality features. While the specific mechanism underlying these associations requires further elucidation, this genetic marker shows promise as a marker of brain reinforcement processes. Possible ways of utilising the A1 allele to inform prevention and treatment initiatives are discussed.

Zierau F; Hardt F; Henriksen JH; Holm SS; Jorring S; Melsen T et al. Validation of a self-administered modified CAGE test (CAGE-C) in a somatic hospital ward: Comparison with biochemical markers. Scandinavian Journal of Clinical & Laboratory Investigation 65(7): 615-622, 2005. (25 refs.)

Objective. The time frame for the original CAGE questionnaire is lifetime and it does not quantify drinking frequency and may be less suitable in a population with very few teetotalers. The purpose of this study was to validate a variant of the CAGE questionnaire and compare it with the outcome of a thorough interview according to DSM-III and ICD-10 criteria and to the outcome of biochemical markers in inpatients in a somatic hospital setting. Material and methods. The questionnaire and biochemical markers were tested on a random sample of 130 patients admitted to a department of orthopedic surgery. The result of a diagnostic interview with a trained staff member from the local alcohol treatment unit was used as the gold standard. Data were analyzed by means of receiver operating characteristic (ROC) curves. Results. In this population 25% had an alcohol problem and the questionnaire proved to be valid, with a sensitivity and specificity of 0.94 and 0.88, respectively, while the positive predictive value (PVpos) was 0.73 and the negative predictive value (PVneg) was 0.98. Carbohydrate-deficient transferrin (CDT) had a sensitivity and a specificity of 0.47 and 0.96, and PVpos and PVneg of 0.80 and 0.85, respectively. Conclusions. This new diagnostic questionnaire is simple, easy to administer and suitable for screening purposes in populations with a high prevalence of at-risk drinkers.