Metabolomics Pathway Analysis

Identify metabolites, map to metabolic pathways, find disease associations, and connect to enzymes/genes.

Domain Reasoning

Metabolite-to-pathway mapping requires correct, database-specific identifiers. HMDB IDs link to KEGG/Reactome but must be converted via BridgeDb; PubChem CIDs need explicit cross-referencing. Always verify metabolite identity first: the same common name can refer to structurally distinct isomers, and PubChem names frequently differ from CTD/KEGG names.

LOOK UP DON'T GUESS

• Pathway membership: call MetaCycgetcompound, KEGGgetcompound, or ReactomeContent_search
• Cross-database IDs: use BridgeDb_xrefs
• Enzyme-metabolite relationships: use CTDgetchemicalgeneinteractions or KEGGgetcompound
• Disease associations: query Metabolitegetdiseases or CTDgetchemical_diseases

COMPUTE, DON'T DESCRIBE

When analysis requires computation (statistics, data processing, scoring, enrichment), write and run Python code via Bash. Don't describe what you would do — execute it and report actual results. Use ToolUniverse tools to retrieve data, then Python (pandas, scipy, statsmodels, matplotlib) to analyze it.

Workflow

Phase 0: Identify & Resolve → Phase 1: Characterize → Phase 2: Pathway Map →
Phase 3: Enzyme/Gene Linkage → Phase 4: Disease Associations → Phase 5: Cross-DB Enrichment → Report

Phase 0: Metabolite Identification & Resolution

By Name

Metabolitesearch: query (REQUIRED), searchtype ("name"/"formula"). Returns PubChem matches with CID, name, formula, MW, SMILES. MetabolomicsWorkbenchsearchcompoundbyname: name (REQUIRED). Cross-reference with RefMet.

By Mass/Formula

MetabolomicsWorkbenchsearchbymz: mz (REQUIRED), adduct (e.g., "M+H"), tolerance. Uses moverz/REFMET/{mz}/{adduct}/{tolerance}. MetabolomicsWorkbenchsearchbyexactmass: exactmass (REQUIRED), tolerance. Uses moverz/REFMET/{mass}/M/{tolerance}.

By ID

Metabolitegetinfo: compoundname, hmdbid (e.g., "HMDB0000122"), or pubchemcid. Returns HMDB ID, CID, InChIKey, classification. KEGGgetcompound: compoundid (e.g., "C00031"). Returns linked pathways, enzymes, reactions.

ID Cross-Referencing

BridgeDbxrefs: identifier (REQUIRED), source (REQUIRED: "Ch"=HMDB, "Cs"=ChemSpider, "Ck"=KEGG, "Ce"=ChEBI), target (optional). BridgeDbsearch: query (REQUIRED), organism. Free-text metabolite search.

Phase 1: Metabolite Characterization

Metabolitegetinfo: classification (superclass/class/subclass), biologicalroles, cellularlocations. MetabolomicsWorkbenchgetrefmetinfo: refmetname (REQUIRED). Standardized RefMet classification. KEGGgetcompound: linked enzyme/reaction/pathway IDs.

Phase 2: Pathway Mapping

MetaCyc

• MetaCycsearchpathways: query (keyword search, e.g., "glycolysis")
• MetaCycgetpathway: pathway_id (e.g., "GLYCOLYSIS") -- reactions, enzymes, compounds
• MetaCycgetcompound: compound_id (e.g., "PYRUVATE") -- pathways it participates in
• MetaCycgetreaction: reaction_id -- substrates, products, enzymes

KEGG

• KEGGgetgenepathways: geneid (e.g., "hsa:5230") -- pathways for enzyme gene
• KEGGgetpathwaygenes: pathwayid (e.g., "hsa00010") -- all genes in pathway

Reactome

• ReactomeContent_search: query, types (e.g., "Pathway"), species
• Reactomegetpathway: id (e.g., "R-HSA-70171")
• ReactomeAnalysispathwayenrichment: identifiers (space-separated string, NOT array)
• Reactomemapuniprottopathways: uniprot_id

Phase 3: Enzyme & Gene Linkage

CTDgetchemicalgeneinteractions: inputterms (chemical name). Returns interacting genes. KEGGgetgenepathways: which pathways an enzyme gene participates in. BridgeDb_attributes: identifier, source, organism. Get attributes for identifier.

Workflow: KEGG compound -> enzyme IDs -> MetaCyc reaction -> enzyme names -> Reactome uniprot -> pathways -> MyGene for gene info.

Phase 4: Disease Associations

CTDgetchemicaldiseases: inputterms (chemical name, MeSH, CAS RN). Curated associations with direct/inferred evidence. CTDgetgenediseases: inputterms (gene name). For metabolite-processing genes from Phase 3. Metabolitegetdiseases: compoundname/hmdbid/pubchem_cid, limit (default 50). CTD-backed.

Phase 5: Cross-Database Enrichment

MetabolomicsWorkbenchgetstudy: studyid (e.g., "ST000001"). MetabolomicsWorkbenchgetcompoundbypubchemcid: pubchemcid. PubMedsearcharticles / EuropePMCsearch_articles: literature context.

For metabolite list enrichment: (1) convert names to gene/enzyme IDs via CTD, (2) run ReactomeAnalysispathwayenrichment with space-separated identifiers, (3) use KEGGgetgene_pathways per enzyme.

Common Mistakes to Avoid

Fallback Strategies

• Metabolitesearch empty -> MetabolomicsWorkbenchsearchcompoundbyname or KEGGget_compound
• MetaCyc not found -> KEGG or Reactome pathways
• CTD empty for disease -> Metabolitegetdiseases with HMDB/CID
• No KEGG compound ID -> BridgeDb_xrefs from HMDB/ChEBI
• exactmass fails -> searchbymz with M+H adduct
• Need enzyme genes -> CTDgetchemicalgeneinteractions

Evidence Grading

Limitations

• HMDB has no open API; use Metabolitegetinfo (PubChem-backed).
• MetaCyc pathways are reference (not organism-specific like KEGG).
• CTD can return very large sets for common metabolites (22K+ for acetaminophen).
• ReactomeAnalysis expects gene/protein IDs, not metabolite IDs directly.
• BridgeDb coverage depends on the metabolite being in mapping databases.