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Int J Mol Sci. 2015 Apr 14;16(4):8351-63. doi: 10.3390/ijms16048351.

Determination of oxidized phosphatidylcholines by hydrophilic interaction liquid chromatography coupled to Fourier transform mass spectrometry.

Author information

1
Core Facility for Mass Spectrometry, Medical University of Graz, Stiftingtalstrasse 24, Graz 8010, Austria. lankmayr@tugraz.at.
2
Institute of Analytical Chemistry and Food Chemistry, Graz University of Technology, Stremayrgasse 9/II, Graz 8010, Austria. lankmayr@tugraz.at.
3
Core Facility for Mass Spectrometry, Medical University of Graz, Stiftingtalstrasse 24, Graz 8010, Austria. martina.brunner@medunigraz.at.
4
Core Facility for Mass Spectrometry, Medical University of Graz, Stiftingtalstrasse 24, Graz 8010, Austria. martin.troetzmueller@medunigraz.at.
5
Core Facility for Mass Spectrometry, Medical University of Graz, Stiftingtalstrasse 24, Graz 8010, Austria. alexander.triebl@medunigraz.at.
6
Bioinformatics Group, Institute for Knowledge Discovery, Graz University of Technology, Petersgasse 14, Graz 8010, Austria. juergen.hartler@tugraz.at.
7
Omics Center Graz, Stiftingtalstrasse 24, Graz 8010, Austria. juergen.hartler@tugraz.at.
8
Core Facility for Mass Spectrometry, Medical University of Graz, Stiftingtalstrasse 24, Graz 8010, Austria. harald.koefeler@medunigraz.at.
9
Omics Center Graz, Stiftingtalstrasse 24, Graz 8010, Austria. harald.koefeler@medunigraz.at.

Abstract

A novel liquid chromatography-mass spectrometry (LC-MS) approach for analysis of oxidized phosphatidylcholines by an Orbitrap Fourier Transform mass spectrometer in positive electrospray ionization (ESI) coupled to hydrophilic interaction liquid chromatography (HILIC) was developed. This method depends on three selectivity criteria for separation and identification: retention time, exact mass at a resolution of 100,000 and collision induced dissociation (CID) fragment spectra in a linear ion trap. The process of chromatography development showed the best separation properties with a silica-based Kinetex column. This type of chromatography was able to separate all major lipid classes expected in mammalian samples, yielding increased sensitivity of oxidized phosphatidylcholines over reversed phase chromatography. Identification of molecular species was achieved by exact mass on intact molecular ions and CID tandem mass spectra containing characteristic fragments. Due to a lack of commercially available standards, method development was performed with copper induced oxidation products of palmitoyl-arachidonoyl-phosphatidylcholine, which resulted in a plethora of lipid species oxidized at the arachidonoyl moiety. Validation of the method was done with copper oxidized human low-density lipoprotein (LDL) prepared by ultracentrifugation. In these LDL samples we could identify 46 oxidized molecular phosphatidylcholine species out of 99 possible candidates.

PMID:
25874761
PMCID:
PMC4425085
DOI:
10.3390/ijms16048351
[Indexed for MEDLINE]
Free PMC Article
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