metabolomics-metabolite-annotation

name	metabolomics-metabolite-annotation
description	Metabolite identification from m/z and retention time. Covers database matching, MS/MS spectral matching, and confidence level assignment. Use when assigning compound identities to detected features in untargeted metabolomics.
tool_type	mixed
primary_tool	HMDB

Version Compatibility

Reference examples tested with: pandas 2.2+, xcms 4.0+

Before using code patterns, verify installed versions match. If versions differ:

• Python: pip show <package> then help(module.function) to check signatures
• R: packageVersion('<pkg>') then ?function_name to verify parameters
• CLI: <tool> --version then <tool> --help to confirm flags

If code throws ImportError, AttributeError, or TypeError, introspect the installed package and adapt the example to match the actual API rather than retrying.

Metabolite Annotation

Database Matching by m/z

Goal: Generate putative metabolite identifications by matching observed m/z values against HMDB.

Approach: Convert m/z to neutral mass by subtracting adduct mass, then query HMDB within a specified ppm tolerance.

"Annotate my metabolomics features with compound identities" -> Match detected features against metabolite databases by exact mass, MS/MS spectra, and retention time to assign compound identities with confidence levels.

library(MetaboAnalystR)

# Load feature table
features <- read.csv('feature_table.csv')

# Search HMDB by exact mass
search_hmdb <- function(mz, adduct = '[M+H]+', ppm = 10) {
    # Calculate neutral mass from m/z
    adduct_masses <- list(
        '[M+H]+' = 1.007276,
        '[M+Na]+' = 22.989218,
        '[M-H]-' = -1.007276,
        '[M+Cl]-' = 34.969402
    )

    neutral_mass <- mz - adduct_masses[[adduct]]

    # Query HMDB (or local database)
    # Returns putative matches
    matches <- QueryHMDB(neutral_mass, ppm)
    return(matches



annotations  lapplyfeaturesmz m search_hmdbm  

MS/MS Spectral Matching

from matchms import calculate_scores
from matchms.importing import load_from_mgf
from matchms.similarity import CosineGreedy

# Load query spectra
queries = list(load_from_mgf('sample_msms.mgf'))

# Load reference library (e.g., GNPS, MassBank)
references = list(load_from_mgf('reference_library.mgf'))

# Calculate similarity scores
similarity = CosineGreedy(tolerance=0.01)
scores = calculate_scores(references, queries, similarity)

# Get best matches
for query_idx, query in enumerate(queries):
    best_match_idx = scores.scores[:, query_idx].argmax()
    best_score = scores.scores[best_match_idx, query_idx]

    if best_score > 0.7:
        ref = references[best_match_idx]
        print(f'{query.get("precursor_mz")}: {ref.get("compound_name")} (score={best_score:.2f})')

SIRIUS + CSI:FingerID

# Molecular formula and structure prediction
sirius \
    --input sample.ms \
    --output sirius_results \
    --database hmdb \
    formula \
    fingerid

# Output structure:
# sirius_results/
#   compound_1/
#     formula_candidates.tsv
#     fingerid_candidates.tsv

MetFrag In Silico Fragmentation

library(metfRag)

# Configure MetFrag search
settings <- list(
    DatabaseSearchRelativeMassDeviation = 10,
    FragmentPeakMatchAbsoluteMassDeviation = 0.01,
    FragmentPeakMatchRelativeMassDeviation = 10,
    MetFragDatabaseType = 'HMDB',
    NeutralPrecursorMass = 147.0532
)

# Run fragmentation prediction
results <- run.metfrag(settings, spectrum_file = 'query_spectrum.txt')

RT Prediction for Validation

from deepchem.models import GraphConvModel
import pandas as pd

# Use predicted RT to validate annotations
# Compare observed RT with predicted RT from chemical structure

def validate_annotation(observed_rt, smiles, rt_model):
    '''Check if observed RT matches prediction'''
    predicted_rt = rt_model.predict(smiles)
    rt_error = abs(observed_rt - predicted_rt)

    if rt_error < 30:  # seconds
        return 'confident'
    elif rt_error < 60:
        return 'probable'
    else:
        return 'unlikely'

Confidence Levels (MSI)

# Metabolomics Standards Initiative levels
assign_confidence <- function(annotation) {
    if (!is.null(annotation$authentic_standard)) {
        return(1)  # Identified by authentic standard
    } else if (!is.null(annotation$msms_match) && annotation$msms_score > 0.8) {
        return(2)  # MS/MS match to database
    } else if (!is.null(annotation$formula_match)) {
        return(3)  # Formula confirmed
       annotationmass_match 
          
      
          
    



featuresconfidence_level  sapplyannotations assign_confidence

CAMERA Adduct Annotation

library(CAMERA)

# Identify adduct and isotope patterns
xsa <- xsAnnotate(xcms_set)
xsa <- groupFWHM(xsa, perfwhm = 0.6)
xsa <- findIsotopes(xsa, mzabs = 0.01, ppm = 10)
xsa <- findAdducts(xsa, polarity = 'positive',
                   rules = c('[M+H]+', '[M+Na]+', '[M+K]+', '[M+NH4]+'))

# Get annotated features
annotated <- getPeaklist(xsa)
annotated$adduct  # Adduct assignment
annotated$isotopes  # Isotope group
annotated$pcgroup  # Correlation group

Batch Annotation Pipeline

library(tidyverse)

annotate_features <- function(feature_table, ppm = 10, polarity = 'positive') {
    results <- feature_table %>%
        rowwise() %>%
        mutate(
            # Calculate possible neutral masses
            mass_h = ifelse(polarity == 'positive', mz - 1.007276, mz + 1.007276),

            # Query databases
            hmdb_match = list(query_hmdb(mass_h, ppm)),
            kegg_match = list(query_kegg(mass_h, ppm)),

            # Best match
            best_match = get_best_match(hmdb_match, kegg_match
            compound_name  best_matchname
            compound_id  best_matchid
            mass_error_ppm  mz  best_matchmz  mz  
        

    results



query_hmdb  mass ppm 
    
    

Export Annotated Results

# Create annotation report
annotation_report <- features %>%
    select(feature_id, mz, rt, compound_name, compound_id,
           formula, confidence_level, mass_error_ppm, adduct) %>%
    arrange(confidence_level, desc(intensity))

write.csv(annotation_report, 'annotated_features.csv', row.names = FALSE)

# Summary
cat('Annotation summary:\n')
cat('  Level 1 (confirmed):', sum(annotation_report$confidence_level == 1), '\n')
cat('  Level 2 (MS/MS match):', sum(annotation_report$confidence_level == 2), '\n')
cat annotation_reportconfidence_level   
cat annotation_reportconfidence_level   
cat annotation_reportconfidence_level   

Related Skills

• xcms-preprocessing - Generate feature table
• pathway-mapping - Map annotated metabolites to pathways
• proteomics/spectral-libraries - Similar spectral matching concepts