Skip to content

Identifying Lipid Chromatographic Peaks

Jump to bottom
Paul Hutchins edited this page Feb 14, 2019 · 5 revisions

Identifying Lipid Chromatographic Peaks (Peak Finder)

Overview

Peak Finder requires .csv peak tables generated from either Compound Discoverer or mzMine 2 and .csv MS/MS result files generated by LipiDex and identifies lipid chromatographic peaks. The resulting peak table is extensively filtered to generate high-confidence lipid identifications with minimal requires hand-annotation. Results are exported as .csv files to the same directory as the peak table.

Instructions

  • Open Peak Finder

  • Select the Peak Table Type

  • Upload Peak Tables

    • For each field, select “Add” and locate the applicable peak tables.

    • For Compound Discover, the required peak tables are an aligned and an unaligned table. For MzMine 2, a peak table for both positive and negative polarity are required.

Loading Peak Tables for Compound Discoverer uploading_sheet.gif

Loading Peak Tables for MZmine 2 mzmine_peaktables.gif

  • Select MS/MS Result Files

    • Select the “Load Files” button

    • In the “Results Uploader” window, select the appropriate data acquisition type.

    • Select “Add Files” to locate the Spectrum Searcher .csv result files

    • If “Separate Polarity Analysis” is selected, edit the “File ID” field in the table. The positive and negative polarity experiments to be merged should share the same File ID number.

    • Select “Upload”

loading_results.gif

  • Update MS/MS Parameters as Needed

    • The “Min. Lipid Spectral Purity” parameter specifies the minimum spectral purity needed to annotate a lipid at the molecular composition level (PC 16:1_18:1) rather than at the sum composition level (PC 34:2).

    • The “Min. MS2 Search Dot Product” parameter specifies the minimum spectral similarity score needed to use an MS/MS identification.

    • The “Min. MS2 Search Rev. Dot Product” parameter specifies the minimum reverse spectral similarity score needed to use an MS/MS identification.

    • The “FWHM Window Multiplier” parameter specifies the maximum allowed retention difference between the apex of the chromatographic peak and the MS/MS spectra in terms of the FWHM of the chromatographic peak.

    • The “Max. Mass Difference” parameter specifies the maximum relative mass difference (ppm) allowed to associate a lipid identification with a chromatographic peak.

  • Update Result Filtering Parameters as Needed

    • Select possible adducts for feature merging

    • Select “Configure”

    • Add all adducts observed in the LC-MS/MS experiment using the same formatting described in the “Creating and Editing a Lipid Library” section of this document.

    • The “Max. RT M.A.D. Factor” parameter specifies the maximum allowed retention time difference between a lipid identification and the all other identified lipids of the same class in terms of multiples of the median absolute retention time deviation of the lipid class.

    • The “Feature Found in n Files” parameter specifies the minimum number of times the specific feature was identified to be included in the final peak table.

adding_adduct_filter.gif

  • Identify Lipid Chromatographic Peaks

    • Select the “Identify Chromatographic Peaks” button

    • Upon completion all result files are written to the same directory as the peak tables

    • The filtered peak table is written to Final_Results.csv

    • The peak table which includes all peaks which have been filtered is written to Unfiltered_Results.csv

    • Sample-specific metrics are written to Sample_Information.csv

Tutorials

Clone this wiki locally