Spectral Data Preprocessing Normalization

spectral-data-preprocessing-normalization

Summary

Standardize and clean raw MS/MS spectral data by normalizing intensity values, filtering low-quality spectra, and converting between spectral formats (MGF, CSV, mzML, mzTab) according to repository-defined quality thresholds. This skill prepares heterogeneous spectral libraries for downstream harmonization and curation.

When to use

Apply this skill when you have raw MS/MS spectral data in MGF format from multiple sources or instruments with inconsistent metadata fields, varying intensity scales, and potential low-quality spectra that need standardization before metadata harmonization or spectral library compilation. Trigger conditions include: receipt of multi-source MGF files, presence of spectra with incomplete or non-standard metadata annotations, or need to merge spectral libraries into a unified format.

When NOT to use

• Input is already a single-source, internally consistent spectral library with validated metadata and no known duplicates or quality issues.
• Raw spectra have already undergone format conversion and intensity normalization by another preprocessing pipeline.
• Use case requires real-time or streaming spectral processing rather than batch preprocessing of entire libraries.

Inputs

• Raw MS/MS spectral data in MGF (Mascot Generic Format) files
• Associated metadata files with compound identifiers, adduct annotations, collision energies, instrument types
• Configuration JSON files (config_step1.json, config_step2.json, config_step3.json) specifying script execution order and data paths
• Matchms filter configuration YAML files (library_cleaning_1.yaml, library_cleaning_2.yaml)

Outputs

• Preprocessed and harmonized MS/MS spectral library in standardized format (mzML, mzTab, or MGF)
• CSV intermediate files with standardized metadata fields
• Cleaned MGF files with validated spectra and removed duplicates
• Numpy format pairwise cosine similarity scores for duplicate candidate spectra

How to apply

Execute the preprocessing pipeline in three sequential phases using configuration-driven scripts. Phase 1 (config_step1.json): clean MGF files, standardize metadata fields, convert to CSV format, merge files, and convert back to MGF with manual metadata standardization. Phase 2 (config_step2.json): remove duplicate spectra (identical fragment intensities), standardize SMILES representations, filter by precursor/fragment mass coherence, remove low-resolution spectra, and eliminate high fragment masses exceeding precursor m/z. Phase 3 (config_step3.json): identify and remove duplicate spectra using cosine similarity scoring, remove low-intensity noise fragments, and finalize instrument and collision energy annotations. Each phase is followed by matchms library cleaning using YAML-defined filter specifications to repair metadata annotations.

Related tools

• spectraverse-analysis (Master repository containing preprocessing pipeline scripts, configuration templates, and orchestration logic for MGF cleaning, format conversion, and metadata standardization) — https://github.com/skinniderlab/spectraverse-analysis
• matchms (Library used to perform spectral preprocessing and metadata repair with YAML-specified filter chains applied between preprocessing phases) — https://github.com/matchms/matchms
• SpectralEntropy (Library used to calculate pairwise cosine similarity scores between duplicate candidate spectra for informed removal decisions) — https://github.com/YuanyueLi/SpectralEntropy.git

Examples

python run_steps.py --config config/config_step1.json && python run_steps.py --config config/config_step2.json && python run_steps.py --config config/config_step3.json

Evaluation signals

• All input MGF spectra successfully parse and convert through intermediate CSV format without data loss or format errors.
• Preprocessed library exhibits zero duplicate spectra when assessed by cosine similarity scoring above the removal threshold.
• Metadata fields (compound identifiers, adducts, collision energies, instrument types) conform to unified schema with no missing required annotations.
• Low-quality spectrum removal (low resolution, high fragment masses exceeding precursor m/z, identical fragment intensities) reduces library size by expected percentage relative to input size.
• Output spectral library validates against schema requirements (e.g., each spectrum has precursor m/z, ≥1 fragment annotation, consistent adduct annotation).

Limitations

• Pipeline execution order is strict; each phase depends on outputs of the previous phase, making parallelization or selective re-runs difficult.
• Matchms library requires custom modifications (provided as spectraverse/matchms.zip) to correctly handle spectra with [M]+ or [M]− adducts annotated with zero charge; default matchms may produce incorrect neutral mass calculations.
• Cosine similarity-based duplicate detection requires manual tuning of similarity threshold for removal decisions; no single universal threshold is provided for all spectral types.
• No changelog or version history is documented in the repository, limiting reproducibility tracking and rollback capability.

Evidence

• [intro] code used to preprocess and harmonize MS/MS spectra and their associated metadata: "This repository contains code used to preprocess and harmonize MS/MS spectra and their associated metadata in the compilation of Spectraverse."
• [readme] sequential three-phase pipeline structure with intermediate matchms application: "Step 1: Run the initial preprocessing. Step 2: Run matchms using filters specified in matchms/library_cleaning_1.yaml. Step 3: Run the second preprocessing step. Step 4: Run matchms again using"
• [readme] MGF cleaning, metadata standardization, format conversion, and merging operations: "step1-1_mgf-clean.py: MGF cleaning and metadata standardization; step1-2_mgf2csv.py: MGF to CSV conversion; step1-3_merge-csv.py: Merge CSV files; step1-4_merge-mgf.py: Merge MGF files;"
• [readme] duplicate removal, quality filtering, and adduct standardization: "step2-2_cleaning.py: Cleaning metadata and MGF files by removing spectra with identical fragment intensities; step2-3_standardization.py: Standardize SMILES; step2-5_lowres-check.py: Removal of"
• [readme] final duplicate identification and cosine scoring methodology: "step3-3_uniq-comb.py: Identification of candidate duplicate spectra; step3-4_uniq-cos-calc.py: Calculating pairwise cosine scores for duplicate spectra and saving in numpy format;"
• [readme] matchms customization requirement for charge handling: "this version fixes the function _get_neutral_mass to ignore spectra annotated with [M]+ or [M]− adducts that had a reported charge of zero."