This second edition volume brings together some of the best experts in the field of modern metabolomics to discuss the latest various techniques used to study specific metabolite classes, and metabolomics in bacterial and mammalian systems. The chapters in this book cover topics such as handling big data for metabolite identification and quantification, as well as building pathways for comparison with other omes, Isotopic Ratio Outlier Analysis (IROA) for quantitative analysis; cholesterol and derivatives in ocular tissues using LC-MS/MS methods; microbial metabolites analysis by mass spectrometry; the metabolomic study of tissues in different diseases; and NMR analysis in livestock metabolomics. It also includes several chapters on the emerging area of spatial metabolomics. Written in the highly successful Methods in Molecular Biology series format, the chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding pitfalls. Cutting-edge and authoritative, Metabolomics: Methods and Protocols, Second Edition is a valuable resource for any researcher looking to expand their knowledge about this important and advancing field.