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Untargeted Lipidomics: An Unbiased High Coverage Lipid Profiling Workflow

Untargeted Lipidomics

March 17, 2021

Mass spectrometry (MS) based lipidomics, the study of lipids present & quantitated within a given biological system, comprises many different approaches for lipid profiling. However, it becomes a daunting task to select an ideal approach that can be robust and provide a high lipid coverage. With the recent advances in the field of mass spectrometry, untargeted liquid chromatography (LC)-MS approach has emerged as one of the front liners for discovery research. Untargeted lipidomics provides an unbiased broader coverage of all the lipids present in a sample. In general, this approach can be performed either by direct infusion or LC-MS. Due to its high sensitivity, the LC-MS approach is often preferred for identification and quantification of lipids.

The LC-MS approach provides the ability to separate isobaric and isomeric species that is not possible with the shotgun approach. Additionally, this approach allows the implementation of different types of LC columns (normal/reverse phase, HILIC, HPLC/UHPLC) that improves the selectivity of lipid identification. The application of high resolving power on both HPLC and MS are critical for this approach. This workflow involves acquiring both MS and MS/MS spectra for lipid extracts after separation using LC coupled with MS. The MS/MS spectra are compared across the diverse lipid database to identify the lipids present based on the fragments from the spectra. Upon identification, these lipids can be quantitated relatively based on their extracted ion chromatograms. This approach is heavily dependent on bioinformatic tools for faster and accurate lipid identification and quantitation. SimLipid® software comes into play at this juncture with following abilities:

  • LC-peak picking & detection
  • Peak alignment
  • Robust database for high-throughput search
  • Quantitation using internal standards
  • Report generation with lipidIDs

A schematic workflow for this untargeted approach is shown below.

Untargeted Macrolipidomic workflow for plant-based oils

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