Urine tested positive for ethyl glucuronide and ethyl sulphate after the consumption of "non-alcoholic" beer

In abstinence maintenance programs, for reissuing the driving licence and in workplace monitoring programs abstinence from ethanol and its proof are demanded. Various monitoring programs that mainly use ethyl glucuronide (EtG) as alcohol consumption marker have been established. To abstain from ethanol, but not from the taste of alcoholic beverages, in particular non-alcoholic beer has become more and more popular. In Germany, these "alcohol-free" beverages may still have an ethanol content of up to 0.5vol.% without the duty of declaration. Due to severe negative consequences resulting from positive EtG tests, a drinking experiment with 2.5L of non-alcoholic beer per person was performed to address the question of measurable concentrations of the direct metabolites EtG and EtS (ethyl sulphate) in urine and blood. Both alcohol consumption markers - determined by LC-MS/MS - were found in high concentrations: maximum concentrations in urine found in three volunteers were EtG 0.30-0.87mg/L and EtS 0.04-0.07mg/L, i.e., above the often applied cut-off value for the proof of abstinence of 0.1mg EtG/L. In the urine samples of one further volunteer, EtG and EtS concentrations cumulated over-night and reached up to 14.1mg/L EtG and 16.1mg/L EtS in the next morning's urine. Ethanol concentrations in blood and urine samples were negative (determined by HS-GC-FID and by an ADH-based method).

Genetic variation in the PNPLA3 gene is associated with alcoholic liver injury in caucasians

A recent genome-wide study revealed an association between variation in the PNPLA3 gene and liver fat content. In addition, the PNPLA3 single-nucleotide polymorphism rs738409 (M148I) was reported to be associated with advanced alcoholic liver disease in alcohol-dependent individuals of Mestizo descent. We therefore evaluated the impact of rs738409 on the manifestation of alcoholic liver disease in two independent German cohorts. Genotype and allele frequencies of rs738409 (M148I) were determined in 1,043 alcoholic patients with or without alcoholic liver injury and in 376 at-risk drinkers from a population-based cohort. Relative to alcoholic patients without liver damage (n = 439), rs738409 genotype GG was strongly overrepresented in patients with alcoholic liver cirrhosis (n = 210; OR 2.79; P(genotype) = 1.2 × 10(-5) ; P(allelic) = 1.6 × 10(-6) ) and in alcoholic patients without cirrhosis but with elevated alanine aminotransferase levels (n = 219; OR 2.33; P(genotype) = 0.0085; P(allelic) = 0.0042). The latter, biochemically defined association was confirmed in an independent population-based cohort of at-risk drinkers with a median alcohol intake of 300 g/week (OR 4.75; P(genotype) = 0.040; P(allelic) = 0.022), and for aspartate aminotransferase (AST) levels. Frequencies of allele PNPLA3 rs738409(G) in individuals with steatosis and normal alanine aminotransferase (ALT) and AST levels were lower than in alcoholics without steatosis and normal ALT/AST (P(combined) = 0.03). The population attributable risk of cirrhosis in alcoholic carriers of allele PNPLA3 rs738409(G) was estimated at 26.6%. CONCLUSION: Genotype PNPLA3 rs738409(GG) is associated with alcoholic liver cirrhosis and elevated aminotransferase levels in alcoholic Caucasians.

Metabolomic tissue signature in human non-alcoholic fatty liver disease identifies protective candidate metabolites

Background Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disorder in industrialized countries, yet its pathophysiology is incompletely understood. Small-molecule metabolite screens may offer new insights into disease mechanisms and reveal new treatment targets. Methods Discovery (N = 33) and replication (N = 66) of liver biopsies spanning the range from normal liver histology to non-alcoholic steatohepatitis (NASH) were ascertained ensuring rapid freezing under 30 s in patients. 252 metabolites were assessed using GC/MS. Replicated metabolites were evaluated in a murine high-fat diet model of NAFLD. Results In a two-stage metabolic screening, hydroquinone (HQ, pcombined = 3.0 × 10−4) and nicotinic acid (NA, pcombined = 3.9 × 10−9) were inversely correlated with histological NAFLD severity. A murine high-fat diet model of NAFLD demonstrated a protective effect of these two substances against NAFLD: Supplementation with 1% HQ reduced only liver steatosis, whereas 0.6% NA reduced both liver fat content and serum transaminase levels and induced a complex regulatory network of genes linked to NALFD pathogenesis in a global expression pathway analysis. Human nutritional intake of NA equivalent was also consistent with a protective effect of NA against NASH progression. Conclusion This first small-molecular screen of human liver tissue identified two replicated protective metabolites. Either the use of NA or targeting its regulatory pathways might be explored to treat or prevent human NAFLD.

TM6SF2 rs58542926 influences hepatic fibrosis progression in patients with non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) is an increasingly common condition, strongly associated with the metabolic syndrome, that can lead to progressive hepatic fibrosis, cirrhosis and hepatic failure. Subtle inter-patient genetic variation and environmental factors combine to determine variation in disease progression. A common non-synonymous polymorphism in TM6SF2 (rs58542926 c.449 C>T, p.Glu167Lys) was recently associated with increased hepatic triglyceride content, but whether this variant promotes clinically relevant hepatic fibrosis is unknown. Here we confirm that TM6SF2 minor allele carriage is associated with NAFLD and is causally related to a previously reported chromosome 19 GWAS signal that was ascribed to the gene NCAN. Furthermore, using two histologically characterized cohorts encompassing steatosis, steatohepatitis, fibrosis and cirrhosis (combined n=1,074), we demonstrate a new association, independent of potential confounding factors (age, BMI, type 2 diabetes mellitus and PNPLA3 rs738409 genotype), with advanced hepatic fibrosis/cirrhosis. These findings establish new and important clinical relevance to TM6SF2 in NAFLD.