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Four-Dimensional Lipidomic Analysis Using Comprehensive Online UHPLC × UHPSFC/Tandem Mass Spectrometry
Zuzana Lásko1, Tomáš Hájek1, Robert Jirásko1
1Department of Analytical Chemistry, University of Pardubice, Faculty of Chemical Technology, Studentská 573, Pardubice 53210, Czech Republic.
This study introduces a novel multidimensional chromatography method for lipidomic analysis, significantly increasing peak capacity for complex biological samples. The advanced technique enables detailed lipidome characterization and identification of numerous lipid species.
Area of Science:
- Analytical Chemistry
- Biochemistry
- Chromatography
Background:
- Complex biological samples require advanced separation techniques for comprehensive analysis.
- Multidimensional chromatography enhances resolution and peak capacity, crucial for lipidomics.
- Existing methods may lack the resolution needed for detailed lipidome profiling.
Purpose of the Study:
- To develop a novel comprehensive online multidimensional chromatography method for lipidomic analysis.
- To combine reversed-phase ultrahigh-performance liquid chromatography (RP-UHPLC) and ultrahigh-performance supercritical fluid chromatography (UHPSFC) for enhanced separation.
- To significantly increase peak capacity for detailed characterization of lipid species in biological samples.
Main Methods:
- A two-dimensional chromatography system combining RP-UHPLC (150 mm C18 column) and UHPSFC (10 mm silica column) with sub-2 μm particles.
- Coupling the system with a high-resolution quadrupole-time-of-flight (QTOF) mass analyzer using electrospray ionization (ESI).
- Utilizing both full-scan and tandem mass spectrometry (MS/MS) with positive and negative ion polarities, including MSE and fast data-dependent acquisition.
Main Results:
- Achieved a peak capacity 10 times higher than RP-UHPLC and 18 times higher than UHPSFC alone.
- Enabled ultrafast analysis in the second dimension (UHPSFC) with a sampling time of 0.55 min.
- Successfully characterized the lipidomic profile of human plasma, identifying 298 lipid species from 16 subclasses with high confidence.
Conclusions:
- The developed multidimensional chromatography method offers superior peak capacity and resolution for lipidomic analysis.
- This approach allows for detailed characterization and confident identification of a wide range of lipid species.
- The method is effective for analyzing complex biological matrices like human plasma.
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