Mass Spectrometry for Metabolomics

Description

This detailed volume provides a comprehensive overview of state-of-the-art metabolomics methods based on mass spectrometry (MS), and their application in food, nutrition, and biomedical research. The chapters assembled herein cover hot topics related to sample preparation, chromatographic and electrophoretic separation, MS-based analysis, as well as data processing and analysis. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.  Authoritative and cutting-edge, Mass Spectrometry for Metabolomics serves as a timely guide for chemists, biochemists, biologists, nutritionists, clinicians, and other experts working in the growing and exciting field of metabolomics. Preface Contents Contributors Chapter 1: Comprehensive and High-Throughput Method for Quantitative Fingerprinting of Phenolic Compounds 1 Introduction 2 Materials 2.1 Metabolite Solutions 2.2 Sample Preparation and Analysis 2.3 Instrumentation 3 Methods 3.1 Sample Preparation 3.1.1 Extraction of Strawberry Samples 3.1.2 Extraction of Olive Oil Samples 3.1.3 Extraction of Red Wine Samples 3.2 Analysis of Phenolic Compounds by RP-UHPLC-DAD-MS 3.3 Data Processing 4 Notes References Chapter 2: Determination of Volatile Metabolites in Vinegar by Solid-Phase Microextraction-Gas Chromatography-Mass Spectrometr... 1 Introduction 2 Materials 2.1 Solid-Phase Extraction 2.2 GC-MS Analysis 3 Methods 3.1 Manual Solid-Phase Microextraction 3.2 Desorption Procedure 3.3 GC-MS Analysis 3.4 Obtention of Quantitative Information 4 Notes References Chapter 3: Determination of Volatile Metabolites in Vinegar by Stir Bar Sorptive Extraction-Gas Chromatography-Mass Spectromet... 1 Introduction 2 Materials 2.1 Stir Bar Sorptive Extraction 2.2 Desorption Procedure 2.3 GC-MS Analysis 3 Methods 3.1 Stir Bar Sorptive Extraction 3.2 Desorption Procedure 3.3 GC-MS Analysis 3.4 Obtention of Quantitative Information 4 Notes References Chapter 4: Discovery of Food Intake Biomarkers Using Metabolomics 1 Introduction 2 Materials 2.1 Equipment and Instrumentation 2.2 Solvents and Chemicals 2.3 Supplies 2.4 Software 3 Methods 3.1 Intervention Cross-Over Trial Study Design and Sampling 3.2 Sample Treatment 3.3 Instrumental Analysis 3.3.1 Ultra-High-Performance Liquid Chromatography (UHPLC) Conditions 3.3.2 High-Resolution Mass Spectrometry (HRMS) Setup 3.4 Data Processing 3.5 Statistical Analysis 3.6 Biomarker Identification 4 Notes References Chapter 5: Metabolomic Characterization of the Antiproliferative Activity of Bioactive Compounds from Fruit By-Products Agains... 1 Introduction 2 Materials 2.1 Pressurized Liquid Extraction 2.2 Cell Lines and Antiproliferative Activity Assay 2.3 Metabolites Extraction from the Cytosolic Fraction of HT-29 Cells 2.4 UHPLC-QTOF-MS/MS Analysis 2.5 Software and Databases for Data Processing and Interpretation 3 Methods 3.1 Extraction of Bioactives from Fruit By-Products 3.2 Cell Lines Treatment with Tropical Fruit By-Products Extracts 3.3 Metabolites Extraction Procedure 3.4 UHPLC-QTOF-MS/MS Metabolic Analysis of Human HT29 Colon Cancer Cells 3.5 Metabolomic Fingerprinting Data Analysis 3.6 Metabolomic Data Interpretation 4 Notes References Chapter 6: Untargeted Metabolomics by Liquid Chromatography-Mass Spectrometry in Biomedical Research 1 Introduction 2 Materials 2.1 Metabolite Extraction and Protein Precipitation 2.2 Liquid Chromatography and Mass Spectrometry 2.3 Software for Data Set Curation and Statistical Analysis 2.4 Databases for the Identification of Metabolites 3 Methods 3.1 Sample Preparation 3.2 Liquid Chromatography 3.3 Mass Spectrometry 3.4 Data Set Creation 3.5 Analytical Validation 3.6 Statistical Analysis (Univariate) 3.7 Identification of Metabolites 4 Notes References Chapter 7: Untargeted Metabolomics for Disease-Specific Signatures 1 Introduction 2 Materials 2.1 Solutions and Materials for Blood Collection and Processing 2.2 Liquid Chromatography-Mass Spectrometry 3 Methods 3.1 Sample Collection and Storage 3.2 Sample Processing (Plasma/Serum) 3.3 Sample Processing (Frozen Blood) 3.4 Sample Randomization 3.5 Data Acquisition 3.6 Data Preprocessing and Analysis 4 Notes References Chapter 8: Metabolomics Analysis of Blood, Urine, and Saliva Samples Based on Capillary Electrophoresis-Mass Spectrometry 1 Introduction 2 Materials 2.1 Blood Sample 2.2 Urine Sample 2.3 Saliva Sample 3 Methods 3.1 Blood Samples 3.2 Urine Samples 3.3 Saliva Samples 3.4 Cation Measurement Setting 3.5 Anion Measurement Setting 4 Notes References Chapter 9: Capillary Electrophoresis-Mass Spectrometry for the Direct Analysis of Metabolites in Highly Saline Samples Using I... 1 Introduction 2 Materials 2.1 Solutions and Samples for Analysis 2.2 Rat Brain Microdialysis Samples 2.3 Analytical Equipment 3 Methods 3.1 Preparation of Microdialysates/Perfusion Fluid 3.2 CE-MS Analysis 3.2.1 Capillary Conditioning and System Performance Check 3.2.2 CE-MS Analysis of Biological Samples with Online Sample Preconcentration 4 Notes References Chapter 10: Chemical Derivatization to Enhance Capillary Electrophoresis Mass Spectrometric Analysis of Acidic Metabolites in ... 1 Introduction 2 Materials 2.1 Solutions and Samples for Analysis 2.2 Derivatization Reagent 2.3 Mammalian (HepG2) Cells 2.4 Analytical Equipment 3 Methods 3.1 Mammalian (HepG2) Cell Lysate Sample Preparation 3.2 Derivatization of Metabolite Standards and Mammalian (HepG2) Cell Extracts 3.3 Sample Analysis of Derivatized Metabolite Standards and Mammalian (HepG2) Cell Extracts Using CE-MS 4 Notes References Chapter 11: Untargeted Metabolomics Based on Liquid Chromatography-Mass Spectrometry for the Analysis of Plasma and Erythrocyt... 1 Introduction 2 Materials 2.1 Sample Collection and Preparation 2.2 Metabolomics Analysis 3 Methods 3.1 Sample Collection 3.2 Extraction of Plasma Samples 3.3 Extraction of Erythrocyte Samples 3.4 Metabolomics Analysis by UHPLC-MS 4 Notes References Chapter 12: Characterization of the Plasmatic and Erythroid Multielemental Biodistribution in Childhood Obesity Using a High-T... 1 Introduction 2 Materials 2.1 Sample Collection and Preparation 2.2 Metallomics Analysis 3 Methods 3.1 Sample Collection 3.2 Size Fractionation of Metal Species by Protein Precipitation Under Non-denaturing Conditions 3.3 Determination of the Total Metal Content 3.4 Multielemental Analysis by ICP-MS 3.5 Data Processing and Quality Control 4 Notes References Chapter 13: Analysis of Brain Metabolites Using Two Complementary Ultrahigh-Performance Liquid Chromatography-Mass Spectrometr... 1 Introduction 2 Materials 2.1 Chemicals 2.2 Ultrahigh-Performance Liquid Chromatography Columns and Vials 2.3 Ultrahigh-Performance Liquid Chromatography and Mass Spectrometry Instruments 3 Methods 3.1 Preparation of Tissue Samples 3.1.1 Dissection and Extraction 3.1.2 Post-extraction 3.1.3 Preparation of Pooled Quality Control (QC) Samples 3.2 Preparation of Extraction Blanks 3.3 Mobile Phase Preparation 3.3.1 Preparation of Mobile Phases A and B for Reversed-Phase C18+ UHPLC Positive and Negative Ion Modes 3.3.2 Preparation of Mobile Phases A and B for HILIC UHPLC Positive and Negative Ion Modes 3.4 UHPLC and MS Methods 3.4.1 Reversed-Phase C18+ UHPLC Gradient 3.4.2 HILIC UHPLC Gradient 3.4.3 HESI-II Ion Source Settings 3.5 Sequence Setup 4 Notes References Chapter 14: Analysis of Cholesterol from the Liver Using Gas Chromatography-Mass Spectrometry 1 Introduction 2 Materials 2.1 Tissue Preparation 2.2 Lipid Extraction 2.2.1 Bligh and Dyer Lipid Extraction 2.2.2 Methyl Tertiary Butyl Ether (MTBE) Lipid Extraction 2.3 GC-MS 3 Methods 3.1 Tissue Preparation and Lipid Extraction 3.1.1 Bligh and Dyer Lipid Extraction Method 3.1.2 Methyl Tertiary Butyl Ether (MTBE) Lipid Extraction Method 3.2 Gas Chromatography-Mass Spectrometry Analysis 3.2.1 Preparation of Instrument Settings 3.2.2 Sample Preparation 4 Notes References Chapter 15: Metabolomics Analysis of Aqueous Humor Based on Liquid Chromatography-Mass Spectrometry 1 Introduction 2 Materials 2.1 Samples and Solvents 2.2 Equipment 3 Methods 3.1 AH Sample Collection 3.2 AH Sample Preparation 3.3 Quality Assurance 3.4 LC-RP-MS Method 3.5 LC-HILIC-MS Method 4 Notes References Chapter 16: Analyses and Localization of Serotonin and L-DOPA in Ocular Tissues by Imaging Mass Spectrometry 1 Introduction 2 Materials 2.1 Solvents 2.2 MALDI/MS Instrument or Accessories 2.3 Software 2.4 Other Materials 2.5 Matrix Solutions for Calibration 2.6 Calibration Mix Solutions 2.7 Matrix Solution for Tissue 3 Methods 3.1 Tissue Preparation Using Fresh Frozen Tissue Samples 3.2 Matrix Application 3.3 MALDI Source Preparation 3.4 Calibration of Mass Spectrometer 3.5 Data Acquisition from Sample 3.6 Data Analysis 4 Notes References Chapter 17: Isobaric Incorporation of C13-Histidine for the Assessment of Remyelination 1 Introduction 2 Materials 2.1 Experimental Autoimmune Encephalomyelitis (EAE) Models 2.2 Optic Nerve Injections 2.3 Immunoprecipitation 2.4 Sample Preparation for Mass Spectrometry 2.5 Mass Spectrometry Analysis 3 Methods 3.1 Generation of Experimental Autoimmune Encephalomyelitis (EAE) Models 3.2 Optic Nerve Injections 3.3 Immunoprecipitation and Enrichment of MBP 3.4 Sample Preparation for Mass Spectrometry 3.5 Mass Spectrometry and Proteomics Analysis 4 Notes References Chapter 18: Isolation and Lipidomic Screening of Human Milk Extracellular Vesicles 1 Introduction 2 Materials 3 Methods 3.1 HM Sample Collection and Storage (See Note 3) 3.2 Isolation of EVs from HM 3.3 Extraction of the HM-EV Lipid Fraction 3.4 LC-MS Lipidomics Method (See Note 15) 3.5 Data Processing and Analysis 4 Notes References Chapter 19: Mass Spectrometry-Based Untargeted Metabolomics and Lipidomics Platforms to Analyze Cell Culture Extracts 1 Introduction 2 Materials 2.1 Solutions and Mixtures for Sample Preparation and Instrumental Analysis 2.2 Solutions and Mixtures for Cell Culturing 2.3 LC-MS System 2.4 Additional Materials and Equipment for Sample Preparation 2.5 Additional Materials and Equipment for Cell Culturing 3 Methods 3.1 Incubation and Exposure 3.2 Sample Preparation 3.2.1 Extraction 3.2.2 Reconstitution of Dried Extracts 3.3 LC-MS 4 Notes References Chapter 20: Data Processing and Analysis in Mass Spectrometry-Based Metabolomics Abbreviations 1 Introduction 2 Experimental Devices for Metabolomics 2.1 NMR 2.2 CE-MS 2.3 GC-MS 2.4 LC-MS 2.5 Ion Mobility 3 Metabolomics Workflow 3.1 The Influence of Experimental Design and Data Acquisition on the Processing and Analysis of Metabolomics Data 3.1.1 Standard Operating Procedures (SOPs) 3.1.2 Confounding Factors 3.1.3 Quality Control in the Analytical Stage 3.1.4 Randomization 3.2 Data Processing 3.2.1 Peak Picking 3.2.2 Peak Alignment 3.2.3 Quality Assurance and Quality Control in Metabolomics Analysis 3.2.4 Missing Value Imputation 3.2.5 Data Transformation, Scaling, and Normalization 3.3 Data Analysis 3.3.1 Data Visualization 3.3.2 Univariate Analysis 3.3.3 Multivariate Analysis Methods 3.3.4 Peak Annotation 3.4 Functional Analysis 3.4.1 Pathway Enrichment 3.4.2 Network Analysis 4 Computational Tools 4.1 R-Based Tools 4.2 XCMS Online 4.3 MetaboAnalyst 4.4 Workflow4metabolomics 4.5 MZmine 2 4.6 MS-Dial 4.7 IP4M 4.8 iMAP 5 Metabolomic Databases 5.1 HMDB 5.2 KEGG References Chapter 21: Data Processing and Analysis in Liquid Chromatography-Mass Spectrometry-Based Targeted Metabolomics 1 Introduction 2 Methods 2.1 Designing Sample Sequence 2.2 Raw Data Processing 2.3 Statistical Analysis of Quantified Data 3 Notes References Chapter 22: Instrument-Agnostizing Methodology for Liquid Chromatography-Mass Spectrometry Systems 1 Introduction 2 Materials 2.1 External Standard Solutions (ES) 2.2 Internal Standard Solutions (IS) 2.3 Mass Calibration 2.4 Separation Systems 2.5 Mass Spectrometry Systems 2.5.1 MS System #1 2.5.2 MS System #2 3 Methods 3.1 Sample Treatment 3.2 LC Conditions 3.3 MS Parameters 3.3.1 MS System #1 3.3.2 MS System #2 3.4 Step 1: Setting up Standard Retention Scores (SRS) 3.5 Step 2: Liquid Chromatography-Mass Spectrometry Agnostizing 3.5.1 Chromatographic Analyses of Samples 3.5.2 Normalization of Signal Intensities 3.5.3 Normalization of Retention Scores: Transfer to SRS Domain 3.6 Instrument-Agnostizing of Samples Chromatograms: General Overview 4 Notes References Index