Capillary electrophoresis-mass spectrometry based metabolomics approaches for volume-restricted applications

Metabolomics is a powerful tool that can provide a comprehensive insight into the complexity of human biology and the pathophysiology of diseases. By analyzing the metabolome, which refers to the complete set of small (endogenous) molecules (with a mass NWO723.016.003Pharmacolog

Mass spectrometry based metabolomics of volume-restricted in-vivo brain samples: actual status and the way forward

Brain metabolomics is gaining interest because of the aging of the population, resulting in more central nervous system disorders such as Alzheimer's and Parkinson's disease. Most often these diseases are studied in vivo, such as for example by analysing cerebrospinal fluid or brain extracellular fluid. These sample types are often considered in pre-clinical studies using animal models. However, the scarce availability of both matrices results in some challenges related to sampling, sample preparation and normalization. Much effort has been made towards the development of alternative, less invasive sampling techniques for collecting small sample volumes (pL till mid mL range) over the past years. Despite recent advances, the analysis of low volumes is still a tremendous challenge. Therefore, proper pre-concentration and sample pretreatment strategies are necessary together with sensitive analysis and detection techniques suitable for low-volume samples. In this review, an overview is given of the stateof-the-art mass spectrometry-based analytical workflows for probing (endogenous) metabolites in volume-restricted in-vivo brain samples. In this context, special attention is devoted to challenges related to sampling, sample preparation and preconcentration strategies. Finally, some general conclusions and perspectives are provided. (C) 2021 The Author(s). Published by Elsevier B.V.Analytical BioScience

Profiling of polar ionogenic metabolites in Polish wines by capillary electrophoresis-mass spectrometry

The composition of wine is determined by a complex interaction between environmental factors, genetic factors (i.e., grape varieties), and winemaking practices (including technology and storage). Metabolomics using NMR spectroscopy, GC-MS, and/or LC-MS has shown to be a useful approach for assessing the origin, authenticity, and quality of various wines. Nonetheless, the use of additional analytical techniques with complementary separation mechanisms may aid in the deeper understanding of wine's metabolic processes. In this study, we demonstrate that CE-MS is a very suitable approach for the efficient profiling of polar ionogenic metabolites in wines. Without using any sample preparation or derivatization, wine was analyzed using a 10-min CE-MS workflow with interday RSD values for 31 polar and charged metabolites below 3.8% and 23% for migration times and peak areas, respectively. The utility of this workflow for the global profiling of polar ionogenic metabolites in wine was evaluated by analyzing different cool-climate Polish wine samples.Analytical BioScience

Metabolomic analysis of dietary-restriction-induced attenuation of sarcopenia in prematurely aging DNA repair-deficient mice

Background: Sarcopenia is characterized by loss of skeletal muscle mass and function, and is a major risk factor for disability and independence in the elderly. Effective medication is not available. Dietary restriction (DR) has been found to attenuate aging and aging-related diseases, including sarcopenia, but the mechanism of both DR and sarcopenia are incompletely understood. Methods: In this study, mice body weight, fore and all limb grip strength, and motor learning and coordination performance were first analysed to evaluate the DR effects on muscle functioning. Liquid chromatography–mass spectrometry (LC–MS) was utilized for the metabolomics study of the DR effects on sarcopenia in progeroid DNA repair-deficient Ercc1∆/− and Xpg−/− mice, to identify potential biomarkers for attenuation of sarcopenia. Results: Muscle mass was significantly (P &lt; 0.05) decreased (13–20%) by DR; however, the muscle quality was improved with retained fore limbs and all limbs grip strength in Ercc1∆/− and Xpg−/− mice. The LC–MS results revealed that metabolites and pathways related to oxidative-stress, that is, GSSG/GSH (P &lt; 0.01); inflammation, that is, 9-HODE, 11-HETE (P &lt; 0.05), PGE2, PGD2, and TXB2 (P &lt; 0.01); and muscle growth (PGF2α) (P &lt; 0.01) and regeneration stimulation (PGE2) (P &lt; 0.05) are significantly downregulated by DR. On the other hand, anti-inflammatory indicator and several related metabolites, that is, β-hydroxybutyrate (P &lt; 0.01), 14,15-DiHETE (P &lt; 0.0001), 8,9-EET, 12,13-DiHODE, and PGF1 (P &lt; 0.05); consumption of sources of energy (i.e., muscle and liver glycogen); and energy production pathways, that is, glycolysis (glucose, glucose-6-P, fructose-6-P) (P &lt; 0.01), tricarboxylic acid cycle (succinyl-CoA, malate) (P &lt; 0.001), and gluconeogenesis-related metabolite, alanine (P &lt; 0.01), are significantly upregulated by DR. The notably (P &lt; 0.01) down-modulated muscle growth (PGF2α) and regeneration (PGE2) stimulation metabolite and the increased consumption of glycogen in muscle and liver may be related to the significantly (P &lt; 0.01) lower body weight and muscle mass by DR. The downregulated oxidative stress, pro-inflammatory mediators, and upregulated anti-inflammatory metabolites resulted in a lower energy expenditure, which contributed to enhanced muscle quality together with upregulated energy production pathways by DR. The improved muscle quality may explain why grip strength is maintained and motor coordination and learning performance are improved by DR in Ercc1∆/− and Xpg−/− mice. Conclusions: This study provides fundamental supporting information on biomarkers and pathways related to the attenuation of sarcopenia, which might facilitate its diagnosis, prevention, and clinical therapy.</p

Application of a capillary electrophoresis-mass spectrometry metabolomics workflow in zebrafish larvae reveals new effects of cortisol

Animal science

Metabolomic analysis of dietary-restriction-induced attenuation of sarcopenia in prematurely aging DNA repair-deficient mice

Background: Sarcopenia is characterized by loss of skeletal muscle mass and function, and is a major risk factor for disability and independence in the elderly. Effective medication is not available. Dietary restriction (DR) has been found to attenuate aging and aging-related diseases, including sarcopenia, but the mechanism of both DR and sarcopenia are incompletely understood. Methods: In this study, mice body weight, fore and all limb grip strength, and motor learning and coordination performance were first analysed to evaluate the DR effects on muscle functioning. Liquid chromatography–mass spectrometry (LC–MS) was utilized for the metabolomics study of the DR effects on sarcopenia in progeroid DNA repair-deficient Ercc1∆/− and Xpg−/− mice, to identify potential biomarkers for attenuation of sarcopenia. Results: Muscle mass was significantly (P &lt; 0.05) decreased (13–20%) by DR; however, the muscle quality was improved with retained fore limbs and all limbs grip strength in Ercc1∆/− and Xpg−/− mice. The LC–MS results revealed that metabolites and pathways related to oxidative-stress, that is, GSSG/GSH (P &lt; 0.01); inflammation, that is, 9-HODE, 11-HETE (P &lt; 0.05), PGE2, PGD2, and TXB2 (P &lt; 0.01); and muscle growth (PGF2α) (P &lt; 0.01) and regeneration stimulation (PGE2) (P &lt; 0.05) are significantly downregulated by DR. On the other hand, anti-inflammatory indicator and several related metabolites, that is, β-hydroxybutyrate (P &lt; 0.01), 14,15-DiHETE (P &lt; 0.0001), 8,9-EET, 12,13-DiHODE, and PGF1 (P &lt; 0.05); consumption of sources of energy (i.e., muscle and liver glycogen); and energy production pathways, that is, glycolysis (glucose, glucose-6-P, fructose-6-P) (P &lt; 0.01), tricarboxylic acid cycle (succinyl-CoA, malate) (P &lt; 0.001), and gluconeogenesis-related metabolite, alanine (P &lt; 0.01), are significantly upregulated by DR. The notably (P &lt; 0.01) down-modulated muscle growth (PGF2α) and regeneration (PGE2) stimulation metabolite and the increased consumption of glycogen in muscle and liver may be related to the significantly (P &lt; 0.01) lower body weight and muscle mass by DR. The downregulated oxidative stress, pro-inflammatory mediators, and upregulated anti-inflammatory metabolites resulted in a lower energy expenditure, which contributed to enhanced muscle quality together with upregulated energy production pathways by DR. The improved muscle quality may explain why grip strength is maintained and motor coordination and learning performance are improved by DR in Ercc1∆/− and Xpg−/− mice. Conclusions: This study provides fundamental supporting information on biomarkers and pathways related to the attenuation of sarcopenia, which might facilitate its diagnosis, prevention, and clinical therapy.</p

Profiling acidic metabolites by capillary electrophoresis‐mass spectrometry in low numbers of mammalian cells using a novel chemical derivatization approach

Medicinal ChemistryAnalytical BioScience

A sample preparation method for the simultaneous profiling of signaling lipids and polar metabolites in small quantities of muscle tissues from a mouse model for sarcopenia

The metabolic profiling of a wide range of chemical classes relevant to understanding sarcopenia under conditions in which sample availability is limited, e.g., from mouse models, small muscles, or muscle biopsies, is desired. Several existing metabolomics platforms that include diverse classes of signaling lipids, energy metabolites, and amino acids and amines would be informative for suspected biochemical pathways involved in sarcopenia. The sample limitation requires an optimized sample preparation method with minimal losses during isolation and handling and maximal accuracy and reproducibility. Here, two developed sample preparation methods, BuOH-MTBE-Water (BMW) and BuOH-MTBE-More-Water (BMMW), were evaluated and compared with previously reported methods, Bligh-Dyer (BD) and BuOH-MTBE-Citrate (BMC), for their suitability for these classes. The most optimal extraction was found to be the BMMW method, with the highest extraction recovery of 63% for the signaling lipids and 81% for polar metabolites, and an acceptable matrix effect (close to 1.0) for all metabolites of interest. The BMMW method was applied on muscle tissues as small as 5 mg (dry weight) from the well-characterized, prematurely aging, DNA repair-deficient Ercc1 Delta/-mouse mutant exhibiting multiple-morbidities, including sarcopenia. We successfully detected 109 lipids and 62 polar targeted metabolites. We further investigated whether fast muscle tissue isolation is necessary for mouse sarcopenia studies. A muscle isolation procedure involving 15 min at room temperature revealed a subset of metabolites to be unstable; hence, fast sample isolation is critical, especially for more oxidative muscles. Therefore, BMMW and fast muscle tissue isolation are recommended for future sarcopenia studies. This research provides a sensitive sample preparation method for the simultaneous extraction of non-polar and polar metabolites from limited amounts of muscle tissue, supplies a stable mouse muscle tissue collection method, and methodologically supports future metabolomic mechanistic studies of sarcopenia.Analytical BioScience

Leerboek, der instrumentatie voor harmonie- en fanfare orkest ..

v. 1 De samenstelling van het blaasorkest.--v. 2 Orkestratie