Metabolomics Spectral Processing, Data Analysis, and Multiomics Integration Using MetaboAnalyst 6.0.

MetaboAnalyst is a comprehensive, user-friendly web-based platform for metabolomics data analysis and interpretation. It offers a unified analytical workflow with an intuitive interface designed to streamline data analysis for both targeted and untargeted metabolomics, while supporting integration with other omics datasets. MetaboAnalyst 6.0 represents a significant advancement toward this vision by supporting LC-MS raw spectral processing and MS2 spectral annotation. This chapter builds on our 2019 protocols, which presented 12 step-by-step procedures for statistical and functional analysis using MetaboAnalyst 4.0, with emphasis on targeted metabolomics. Here, we extend that foundation with eight new or substantially revised protocols for MetaboAnalyst 6.0, centered mainly on LC-MS untargeted metabolomics, advanced statistics, and functional analysis. The main topics covered include: Basic Protocol 1: LC-MS raw spectral processing with or without MS2 spectra Basic Protocol 2: MS2 spectral annotation Basic Protocol 3: Functional analysis of untargeted LC-MS metabolomics data Basic Protocol 4: Advanced statistical analysis for studies with multiple factors Basic Protocol 5: Dose-response analysis Basic Protocol 6: Pathway and joint-pathway analysis Basic Protocol 7: Causal analysis Basic Protocol 8: Using MetaboAnalystR for batch processing.