Comprehensive untargeted lipidomic analysis using core-shell C30 particle column and high field orbitrap mass...

Area of Science:

• Analytical Chemistry
• Biochemistry
• Mass Spectrometry

Background:

• Untargeted lipidomics aims for high-throughput, comprehensive, and unambiguous lipid identification and quantification.
• Advancements in liquid chromatography (LC) stationary phases and high-resolution mass spectrometry (MS) are crucial for achieving these goals.

Purpose of the Study:

• To compare the performance of four different reversed-phase LC columns when coupled with a high-field quadrupole orbitrap mass spectrometer (Q Exactive HF) for untargeted lipidomics.
• To evaluate column efficiency, peak capacity, reproducibility, and lipid identification capabilities using complex lipid standards and biological samples (rat plasma and liver).

Main Methods:

• Four reversed-phase LC columns (C30 and C18 functionalizations on core-shell or totally porous silica particles, 1.7–2.6 μm particle size) were tested.
• A high-field quadrupole orbitrap mass spectrometer (Q Exactive HF) was used for high-resolution mass analysis.
• Complex lipid mixtures, rat plasma, and rat liver samples were analyzed using a 24-minute gradient.

Main Results:

• The Accucore C30 column demonstrated superior performance with the narrowest peaks, highest theoretical plate number, excellent peak capacity, and high retention time reproducibility (<1% SD).
• This setup enabled the identification of 430 lipid species with high confidence from rat plasma and liver samples.
• High-resolution LC and MS allowed discrimination of cis/trans isomers and isobaric species, and unambiguous assignment of sn-positional isomers for lysophospholipids.

Conclusions:

• High-efficiency LC separation, particularly with the Accucore C30 column, combined with high-resolution orbitrap mass spectrometry, significantly enhances the comprehensiveness and confidence of lipid identification in untargeted lipidomics.
• These optimized analytical approaches are highly promising for advancing lipidomics research.