IntroductionBoiling ethanol extraction is a frequently used method for metabolomics studies of biological samples. However, the stability of several central carbon metabolites, including nucleotide triphosphates, and the influence of the cellular matrix on their degradation have not been addressed.ObjectivesTo study how a complex cellular matrix extracted from yeast (Saccharomyces cerevisiae) may affect the degradation profiles of nucleotide triphosphates extracted under boiling ethanol conditions.MethodsWe present a double-labelling LC–MS approach with a 13C-labeled yeast cellular extract as complex surrogate matrix, and 13C15N-labeled nucleotides as internal standards, to study the effect of the yeast matrix on the degradation of nucleotide triphosphates.ResultsWhile nucleotide triphosphates were degraded to the corresponding diphosphates in pure solutions, degradation was prevented in the presence of the yeast matrix under typical boiling ethanol extraction conditions.ConclusionsExtraction of biological samples under boiling ethanol extraction conditions that rapidly inactivate enzyme activity are suitable for labile central energy metabolites such as nucleotide triphosphates due to the stabilizing effect of the yeast matrix. The basis of this phenomenon requires further study.Graphical abstract Electronic supplementary materialThe online version of this article (doi:10.1007/s11306-016-1140-4) contains supplementary material, which is available to authorized users.

Recent advances in analytical chemistry have set the stage for metabolite profiling to help understand complex molecular processes in physiology. Despite ongoing efforts, there are concerns regarding metabolomics workflows, since it has been shown that internal (enzyme activity, blood contamination, and the dynamic nature of metabolite concentrations) as well as external factors (storage, handling, and analysis method) may affect the metabolome profile. Many metabolites are intrinsically instable, particularly some of those associated with central carbon metabolism. While enzymatic conversions have been studied in great detail, nonenzymatic, chemical conversions received comparatively little attention. This review aims to give an in-depth overview of nonenzymatic energy metabolite degradation/interconversion chemistry focusing on a selected range of metabolites. Special attention will be given to qualitative (degradation pathways) as well as quantitative aspects, that may affect the acquisition of accurate data in the context of metabolomics studies. Problems related to the use of isotopically labeled internal standards hindering the quantitative analysis of common metabolites will be presented with an experimental example. Finally, general conclusions and perspectives are given.

Lipidomics is a rapidly developing field in modern biomedical research. While LC-MS systems are able to detect most of the known lipid classes in a biological matrix, there is no single technique able to extract all of them simultaneously. In comparison with two-phase extractions, one-phase extraction systems are of particular interest, since they decrease the complexity of the experimental procedure. By using an untargeted lipidomics approach, we explored the differences/similarities between the most commonly used two-phase extraction systems (Folch, Bligh and Dyer, and MTBE) and one of the more recently introduced one-phase extraction systems for lipid analysis based on the MMC solvent mixture (MeOH/MTBE/CHCl3). The four extraction methods were evaluated and thoroughly compared against a pooled extract that qualitatively and quantitatively represents the average of the combined extractions. Our results show that the lipid profile obtained with the MMC system displayed the highest similarity to the pooled extract, indicating that it was most representative of the lipidome in the original sample. Furthermore, it showed better extraction efficiencies for moderate and highly apolar lipid species in comparison with the Folch, Bligh and Dyer, and MTBE extraction systems. Finally, the technical simplicity of the MMC procedure makes this solvent system highly suitable for automated, untargeted lipidomics analysis.Electronic supplementary materialThe online version of this article (10.1007/s00216-018-1200-x) contains supplementary material, which is available to authorized users.

In response to heart failure (HF), the heart reacts by repressing adult genes and expressing fetal genes, thereby returning to a more fetal-like gene profile. To identify genes involved in this process, we carried out transcriptional analysis on murine hearts at different stages of development and on hearts from adult mice with HF. Our screen identified , encoding for 5-oxoprolinase, a member of the γ-glutamyl cycle that functions by scavenging 5-oxoproline. OPLAH depletion occurred as a result of cardiac injury, leading to elevated 5-oxoproline and oxidative stress, whereas OPLAH overexpression improved cardiac function after ischemic injury. In HF patients, we observed elevated plasma 5-oxoproline, which was associated with a worse clinical outcome. Understanding and modulating fetal-like genes in the failing heart may lead to potential diagnostic, prognostic, and therapeutic options in HF.

Oxidative stress induced by 5-oxoproline results in a murine phenotype reminiscent of the clinical manifestation of HFpEF without the need for surgical or pharmacological interference. Better understanding of the role of oxidative stress in HFpEF may potentially lead to novel therapeutic options.