Effect of egg ingestion on trimethylamine-N-oxide production in humans: a randomized, controlled, dose-response study

Background: It is important to understand whether eating eggs, which are a major source of dietary choline, results in increased exposure to trimethylamine-N-oxide (TMAO), which is purported to be a risk factor for developing heart disease

Quantitative Metabolomics by 1H-NMR and LC-MS/MS Confirms Altered Metabolic Pathways in Diabetes

Insulin is as a major postprandial hormone with profound effects on carbohydrate, fat, and protein metabolism. In the absence of exogenous insulin, patients with type 1 diabetes exhibit a variety of metabolic abnormalities including hyperglycemia, glycosurea, accelerated ketogenesis, and muscle wasting due to increased proteolysis. We analyzed plasma from type 1 diabetic (T1D) humans during insulin treatment (I+) and acute insulin deprivation (I-) and non-diabetic participants (ND) by 1H nuclear magnetic resonance spectroscopy and liquid chromatography-tandem mass spectrometry. The aim was to determine if this combination of analytical methods could provide information on metabolic pathways known to be altered by insulin deficiency. Multivariate statistics differentiated proton spectra from I- and I+ based on several derived plasma metabolites that were elevated during insulin deprivation (lactate, acetate, allantoin, ketones). Mass spectrometry revealed significant perturbations in levels of plasma amino acids and amino acid metabolites during insulin deprivation. Further analysis of metabolite levels measured by the two analytical techniques indicates several known metabolic pathways that are perturbed in T1D (I-) (protein synthesis and breakdown, gluconeogenesis, ketogenesis, amino acid oxidation, mitochondrial bioenergetics, and oxidative stress). This work demonstrates the promise of combining multiple analytical methods with advanced statistical methods in quantitative metabolomics research, which we have applied to the clinical situation of acute insulin deprivation in T1D to reflect the numerous metabolic pathways known to be affected by insulin deficiency

Recommendations for the Generation, Quantification, Storage, and Handling of Peptides Used for Mass Spectrometry-Based Assays

BACKGROUND: For many years, basic and clinical researchers have taken advantage of the analytical sensitivity and specificity afforded by mass spectrometry in the measurement of proteins. Clinical laboratories are now beginning to deploy these work flows as well. For assays that use proteolysis to generate peptides for protein quantification and characterization, synthetic stable isotope-labeled internal standard peptides are of central importance. No general recommendations are currently available surrounding the use of peptides in protein mass spectrometric assays. CONTENT: The Clinical Proteomic Tumor Analysis Consortium of the National Cancer Institute has collaborated with clinical laboratorians, peptide manufacturers, metrologists, representatives of the pharmaceutical industry, and other professionals to develop a consensus set of recommendations for peptide procurement, characterization, storage, and handling, as well as approaches to the interpretation of the data generated by mass spectrometric protein assays. Additionally, the importance of carefully characterized reference materials-in particular, peptide standards for the improved concordance of amino acid analysis methods across the industry-is highlighted. The alignment of practices around the use of peptides and the transparency of sample preparation protocols should allow for the harmonization of peptide and protein quantification in research and clinical care

Biomarker discovery using NMR and MS-based metabolomics: Applications to diabetes

The emerging field of “metabolomics,” in which a large number of small molecule metabolites from body fluids or tissues are detected quantitatively in a single step, promises immense potential for early diagnosis, therapy monitoring and for understanding the pathogenesis of many diseases. Metabolomics methods are mostly focused on the information rich analytical techniques of nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS). Analysis of the data from these high-resolution methods using advanced chemometric approaches provides a powerful platform for translational and clinical research as well as diagnostic applications. This work mainly focuses on developing sensitive and specific biomarker discovery using NMR and MS-based metabolomics approach. This work is divided into two parts: methodology development and application to diabetes. In the first part, statistical analytical methods were developed to enhance the detection accuracy and detection scope of metabolite biomarkers: a statistical method was developed to evaluate the appropriateness of current data processing and analysis methods used for NMR-based metabolomics to minimize the false discovery rate of potential biomarkers; and a new analytical approach with combined headspace SPME-GC/MS analysis with 1H NMR for metabolic profiling of human body fluids. This approach provides a markedly improved ability for characterizing subtle metabolite-based pathophysiological differences which are exemplified in the reported gender study. The second part of this thesis focuses on establishing new metabolite biomarkers for understanding diabetes from a system biology standpoint. Among the various types of diabetes including type 1, type 2 and gestational diabetes, we started our investigation with characterization of insulin deficiency in type 1 diabetes. We studied the insulin deficiency-induced metabolic disturbances in both an animal model and clinical subjects. For the clinical study, the insulin-treatment effect on the type 1 diabetic patients was assessed. The results suggest that type 1 diabetes could induce significant disturbances to many metabolic pathways such as glucose metabolism, TCA cycle carboxylic acids, proteins/amino acids metabolism and fatty acid metabolism. Therapeutic interventions such as insulin can effectively, although still not perfectly, adjust the disturbed metabolism back to normal. We have also explored diabetes in Zucker fatty rats, a commonly used type 2 diabetes animal model. Specifically, we investigated the effect of green tea and its formulation in ameliorating the characteristic type 2 diabetes insulin insensitivity. Two new hypotheses were generated that green tea can noticeably enhance the body’s metabolic control and that green tea formulation with ascorbic acid has a potential synergistic effect in alleviating acidosis. As shown by this work, metabolite profiling provides important information to help understand systems biology. Advanced methods in NMR, MS, and statistics are useful in identifying and quantifying selected components in complex biological samples. Implications for the future include improved chemical-based early disease detection, therapy feedback, disease monitoring, and personalized therapies

Identification and quantification of metabolites in 1H NMR spectra by Bayesian model selection

Motivation: Nuclear magnetic resonance (NMR) spectroscopy is widely used for high-throughput characterization of metabolites in complex biological mixtures. However, accurate interpretation of the spectra in terms of identities and abundances of metabolites can be challenging, in particular in crowded regions with heavy peak overlap. Although a number of computational approaches for this task have recently been proposed, they are not entirely satisfactory in either accuracy or extent of automation

Alteration of bile acid metabolism in the rat induced by chronic ethanol consumption

Our understanding of the bile acid metabolism is limited by the fact that previous analyses have primarily focused on a selected few circulating bile acids; the bile acid profiles of the liver and gastrointestinal tract pools are rarely investigated. Here, we determined how chronic ethanol consumption altered the bile acids in multiple body compartments (liver, gastrointestinal tract, and serum) of rats. Rats were fed a modified Lieber-DeCarli liquid diet with 38% of calories as ethanol (the amount equivalent of 4–5 drinks in humans). While conjugated bile acids predominated in the liver (98.3%), duodenum (97.8%), and ileum (89.7%), unconjugated bile acids comprised the largest proportion of measured bile acids in serum (81.2%), the cecum (97.7%), and the rectum (97.5%). In particular, taurine-conjugated bile acids were significantly decreased in the liver and gastrointestinal tract of ethanol-treated rats, while unconjugated and glycine-conjugated species increased. Ethanol consumption caused increased expression of genes involved in bile acid biosynthesis, efflux transport, and reduced expression of genes regulating bile acid influx transport in the liver. These results provide an improved understanding of the systemic modulations of bile acid metabolism in mammals through the gut-liver axis.—Xie, G., Zhong, W., Li, H., Li, Q., Qiu, Y., Zheng, X., Chen, H., Zhao, X., Zhang, S., Zhou, Z., Zeisel, S. H., Jia, W. Alteration of bile acid metabolism in the rat induced by chronic ethanol consumption