A metabolomics investigation on experimental interventions of acute alcohol consumption

Abstract

This thesis, titled: "A metabolomics investigation on experimental interventions of acute alcohol consumption", deals with a current topic of global interest, namely, alcohol use and abuse. Alcohol abuse is associated with many serious, and even detrimental, health, social and economic consequences, and is one of the world’s leading risk factors for disability, morbidity and mortality. For these reasons it is a topic of growing concern in developing, as well as developed, countries. Alcohol is metabolized mainly in the liver by two nicotinamide adenine dinucleotide (NAD)-dependent enzymes — alcohol dehydrogenase and, subsequently, aldehyde dehydrogenase. In both of these reactions oxidized NAD (NAD+) is reduced to NADH, which increases the NAHD:NAD+ ratio in hepatocytes. This ratio controls the activity of several key metabolic enzymes and the direction of many reversible metabolic reactions, and its disruption is known to result in perturbations of various metabolic pathways. Various studies examining the effects of, and diseases related to, chronic alcohol abuse have been performed in the last few decades. However, to date, no comprehensive metabolomics study on the effects of acute alcohol consumption has been done. Thus, with the guidance of experts in the fields of metabolism, metabolomics and biostatistics, the first extensive, multidisciplinary metabolomics cross-over intervention study into the effects of acute alcohol consumption on the urinary metabolite profiles of healthy, young males was designed, and is presented in this thesis. The study consisted of analysing urine samples, collected from experimental participants over a defined period of time following four interventions, on two different analytical platforms — proton nuclear magnetic resonance (1H-NMR) spectroscopy as an untargeted approach, and gas chromatography—mass spectrometry (GC—MS) as a semi-targeted approach. The results from these investigations demonstrated the power of applying metabolomics to this area of research and provided the opportunity to obtain a holistic view of the urinary metabolic profile resulting from acute alcohol consumption. From both of these approaches a list of metabolites perturbed by acute alcohol consumption could be compiled with the use of statistical analyses. Various metabolic pathways were seen to be disrupted, most of them due to the known alcohol-induced increased NADH:NAD+ ratio. Additionally, two urinary metabolites — sorbitol, from the 1H-NMR analysis, and 2-hydroxyisobutyric acid, from the GC—MS investigation — not previously known to be associated with the consequences of acute alcohol consumption were identified in the metabolic profiles of the experimental participants following acute alcohol consumption. These novel findings could possibly be used as a basis for determining biomarkers of acute alcohol consumption, which could have various health, economic and legal benefits. This thesis, the eventual product of a skilfully designed and diligently carried out scientific study, is compiled and presented in article format as per the requirements of North-West University. The scientific contributions made during this study to the existing alcohol-related scientific knowledge resulted in three publications. Two (1 and 3) have already been published, and one (2) has been accepted for publication. 1. Irwin, C., Van Reenen, M., Mason, S., Mienie, L.J., Westerhuis, J.A. & Reinecke, C.J. 2016. Contribution towards a metabolite profile of the detoxification of benzoic acid through glycine conjugation: an intervention study. PLOS ONE, 11(12):e0167309. doi:10.1371/journal.pone. 0167309. 2. Irwin, C., Van Reenen, M., Mason, S., Mienie, L.J., Wevers, R.A., Westerhuis, J.A. & Reinecke, C.J. The 1H-NMR-based metabolite profile of acute alcohol consumption: a metabolomics intervention study. 3. Irwin, C., Mienie, L.J., Wevers, R.A., Mason, S., Westerhuis, J.A., Van Reenen, M. & Reinecke, C.J. 2018. GC—MS-based urinary organic acid profiling reveals multiple dysregulated metabolic pathways following experimental acute alcohol consumption. Scientific Reports, 8:5775. doi:10.1038/s41598-018-24128-1.