In vivo comprehensive multiphase NMR

Traditionally, due to different hardware requirements, nuclear magnetic resonance (NMR) has developed as two separate fields: one dealing with solids, and one with solutions. Comprehensive multiphase (CMP) NMR combines all electronics and hardware (magic angle spinning [MAS], gradients, high power Radio Frequency (RF) handling, lock, susceptibility matching) into a universal probe that permits a comprehensive study of all phases (i.e., liquid, gel-like, semisolid, and solid), in intact samples. When applied in vivo, it provides unique insight into the wide array of bonds in a living system from the most mobile liquids (blood, fluids) through gels (muscle, tissues) to the most rigid (exoskeleton, shell). In this tutorial, the practical aspects of in vivo CMP NMR are discussed including: handling the organisms, rotor preparation, sample spinning, water suppression, editing experiments, and finishes with a brief look at the potential of other heteronuclei (²H, ¹⁵N, ¹⁹F, ³¹P) for in vivo research. The tutorial is aimed as a general resource for researchers interested in developing and applying MAS-based approaches to living organisms. Although the focus here is CMP NMR, many of the approaches can be adapted (or directly applied) using conventional high-resolution magic angle spinning, and in some cases, even standard solid-state NMR probes.     

Identification of metabolic markers in patients with type 2 Diabetes by Ultrafast gas chromatography coupled to electronic nose. A pilot study

Metabolomics is a potential tool for the discovery of new biomarkers in the early diagnosis of diseases. An ultra-fast gas chromatography system equipped to an electronic nose detector (FGC eNose) was used to identify the metabolomic profile of Volatile Organic Compounds (VOCs) in type 2 diabetes (T2D) urine from Mexican population. A cross-sectional, comparative, and clinical study with translational approach was performed. We recruited twenty T2D patients and twenty-one healthy subjects. Urine samples were taken and analyzed by FGC eNose. Eighty-eight compounds were identified through Kovats's indexes. A natural variation of 30% between the metabolites, expressed by study groups, was observed in Principal Component 1 and 2 with a significant difference (p < 0.001). The model, performed through a Canonical Analysis of Principal coordinated (CAP), allowed a correct classification of 84.6% between healthy and T2D patients, with a 15.4% error. The metabolites 2-propenal, 2-propanol, butane- 2,3-dione and 2-methylpropanal, were increased in patients with T2D, and they were strongly correlated with discrimination between clinically healthy people and T2D patients. This study identified metabolites in urine through FGC eNose that can be used as biomarkers in the identification of T2D patients. However, more studies are needed for its implementation in clinical practice.     

Comprehensive Profiling by Non-targeted Stable Isotope Tracing Capillary Electrophoresis-Mass Spectrometry: A New Tool Complementing Metabolomic Analyses of Polar Metabolites

Mass spectrometry (MS) driven metabolomics is a frequently used tool in various areas of life sciences; however, the analysis of polar metabolites is less commonly included. In general, metabolomic analyses lead to the detection of the total amount of all covered metabolites. This is currently a major limitation with respect to metabolites showing high turnover rates, but no changes in their concentration. Such metabolites and pathways could be crucial metabolic nodes (e.g., potential drug targets in cancer metabolism). A stable-isotope tracing capillary electrophoresis–mass spectrometry (CE-MS) metabolomic approach was developed to cover both polar metabolites and isotopologues in a non-targeted way. An in-house developed software enables high throughput processing of complex multidimensional data. The practicability is demonstrated analyzing [U-¹³C]-glucose exposed prostate cancer and non-cancer cells. This CE-MS-driven analytical strategy complements polar metabolite profiles through isotopologue labeling patterns, thereby improving not only the metabolomic coverage, but also the understanding of metabolism.     

Antidepressant-like effect of triterpenoids extracts from Poria cocos on the CUMS rats by 16S rRNA gene sequencing and LC–MS metabolomics

Abstract

Poria cocos is an edible medicinal mushroom with the effects of inducing diuresis, excreting dampness, invigorating the spleen and tranquilizing the mind. The triterpenoids of Poria cocos as the main active ingredients have shown health benefits of the central nervous system of the human body. Accordingly, this study aimed at further understanding the antidepressant-like effect and a potential mechanism of the triterpenoids extracts from Poria cocos (TPC). As a result, the TPC significantly ameliorated depression-like behaviors on chronic unpredictable mild stress (CUMS) rats, and restored the levels of brain-derived neurotrophic factor and nerve growth factor in the hippocampus of rats. Gut microiota analysis revealed that TPC could increase the biodiversity and markedly regulate the relative abundance of [Prevotella], Allobaculum and Ochrobactrum of CUMS rats. The cecal contents metabolomics pointed to thirteen biomarkers associated with TPC antidepressant effect, which involved in primary bile acid biosynthesis, taurine and hypotaurine metabolism, arginine and proline metabolism. Correlation analysis further showed that there was a strong correlation relationship between the gut microbiota and the cecal contents metabolites, especially compounds involved in energy metabolism, inflammation and immunity. In conclusion, TPC showed a potential antidepressant effect, which was possibly mediated the gut microbiota and cecal contents metabolism.     

食品中化学性有害物的质谱软电离裂解规律及筛查技术研究进展

食品中化学性有害物是导致食品安全问题的重要原因，质谱是对食品中化学性有害物进行定性定量分析的有效方法。该文按照化合物结构类别，综述了食品中重要化学性有害物的质谱软电离裂解机理，包括农药、兽药、真菌毒素，以及其他化学污染物。对于每类化合物，重点综述化合物质谱裂解产生的特征碎片、中性丢失等，以及这些质谱软电离裂解机理在食品中化学性有害物筛查及发现中的应用。研究化合物的质谱裂解机理可以帮助研究者对化合物进行结构解析和结构确证，为食品中同类结构新型化学性有害物的发掘提供理论依据。     

蛋白质组学和代谢组学在微生物代谢工程中的应用

构建微生物细胞工厂是化学品、生物能源以及药物分子可持续生产的可行性策略。然而，微生物的代谢复杂、调控严谨，制约着目标产物高效合成。蛋白质组学和代谢组学可以从系统生物学角度分析酶和代谢物组分，从而理解复杂的生物系统，为微生物代谢工程改造提供重要线索。该文介绍了蛋白质组学和代谢组学在微生物代谢工程中的应用，包括基因组尺度代谢模型构建、菌株生物合成优化、指导菌株耐受性改造、限速步骤预测、植物次级代谢途径挖掘，从而为微生物合成天然产物提供新的基因或途径。在此基础上，该文还展望了生物大数据未来的发展方向。     

Targeted metabolomic analysis of 33 amino acids and biogenic amines in human urine by ion‐pairing HPLC‐MS/MS: Biomarkers for tacrolimus nephrotoxicity after renal transplantation

Calcineurin inhibitor nephrotoxicity, especially for the widely used tacrolimus, has become a major concern in post‐transplant immunosuppression. Multiparametric amino acid metabolomics is useful for biomarker identification of tacrolimus nephrotoxicity, for which specific quantitative methods are highlighted as a premise. This article presents a targeted metabolomic assay to quantify 33 amino acids and biogenic amines in human urine by high‐performance liquid chromatography coupled with tandem mass spectrometry. Chromatographic separation was carried out on an Agilent Zorbax SB‐C₁₈ column (3.0 × 150 mm, 5 μm) with addition of an ion‐pairing agent in the mobile phase, and MS/MS detection was achieved in both the positive and negative multiple reaction monitoring modes. Good correlation coefficients (r² > 0.98) were obtained for most analytes. Intra‐ and inter‐day precision, stability, carryover and incurred sample reanalysis met with the acceptance criteria of the guidance of the US Food and Drug Administration. Analysis on urine from healthy volunteers and renal transplantation patients with tacrolimus nephrotoxicity confirmed symmetric dimethylarginine and serine as biomarkers for kidney injury, with AUC values of 0.95 and 0.81 in receiver operating characteristic analysis, respectively. Additionally, symmetric dimethylarginine exhibited a tight correlation with serum creatinine, and was therefore indicative of renal function. The targeted metabolomic assay was time and cost prohibitive for amino acid analysis in human urine, facilitating the biomarker identification of tacrolimus nephrotoxicity.     

The impact of blood on liver metabolite profiling – a combined metabolomic and proteomic approach

Metabolomics has entered the well‐established omic sciences as it is an indispensable information resource to achieve a global picture of biological systems. The aim of the present study was to estimate the influence of blood removal from mice liver as part of sample preparation for metabolomic and proteomic studies. For this purpose, perfused mice liver tissue (i.e. with blood removed) and unperfused mice liver tissue (i.e. containing blood) were compared by two‐dimensional gas chromatography time of flight mass spectrometry (GC × GC‐TOFMS) for the metabolomic part, and by liquid chromatography tandem mass spectrometry (LC‐MS/MS) for the proteomic part. Our data showed significant differences between the unperfused and perfused liver tissue samples. Furthermore, we also observed an overlap of blood and tissue metabolite profiles in our data, suggesting that the perfusion of liver tissue prior to analysis is beneficial for an accurate metabolic profile of this organ. Copyright © 2013 John Wiley & Sons, Ltd.     

Phytochemical Differentiation of Saffron (Crocus sativus L.) by High Resolution Mass Spectrometry Metabolomic Studies

The metabolite profiling of saffron (Crocus sativus L.) from several countries was measured by using ultra-performance liquid chromatography combined with high resolution mass spectrometry (UPLC-HR MS). Multivariate statistical analysis was employed to distinguish among the several samples of C. sativus L. from Greece, Italy, Morocco, Iran, India, Afghanistan and Kashmir. The results of this study showed that the phytochemical content in the samples of C. sativus L. were obviously diverse in the different countries of origin. The metabolomics approach was deemed to be the most suitable in order to evaluate the enormous array of putative metabolites among the saffron samples studied, and was able to provide a comparative phytochemical screening of these samples. Several markers have been identified that aided the differentiation of a group from its counterparts. This can be important for the selection of the appropriate saffron sample, in view of its health-promoting effect which occurs through the modulation of various biological and physiological processes.     

The use of untargeted and widely targeted metabolomics to distinguish between asphyxia and sudden cardiac death as the cause of death in rats: A preliminary study

It is important to determine the cause of death in the case of asphyxia. However, it is difficult to conclude death by asphyxia, especially when the deceased has underlying heart disease, because there are often no specific and representative corpse signs for both asphyxia and sudden cardiac death (SCD). The aim of the present work was to investigate the potential of metabolomics to discriminate asphyxia from SCD as the cause of death. A total of thirty male Sprague–Dawley rats were used to construct models of asphyxia, SCD (interfering cause of death), and cervical dislocation (control). Untargeted and widely targeted metabolomics approaches were used to obtain rat pulmonary metabolic profiles in this study. First, the metabolic alterations resulting from asphyxia were explored. There were significant changes found in carbohydrate metabolism, the endocrine system, and the sensory system. Second, we screened potential biomarkers and built classification models to determine the cause of death. Moreover, some biomarkers remained differentiated at 24 h and 48 h postmortem, so the cause of death could still be determined after death. This study showed the application potential of metabolomics to investigate the metabolic changes occurring in the process of death, as well as to determine the cause of death on the basis of metabolic differences even after death.