title: Profiling of lipid species by normal-phase liquid chromatography, nanoelectrospray ionization, and ion trap-orbitrap mass spectrometry.
abstract: Detailed analysis of lipid species can be challenging due to their structural diversity and wide concentration range in cells, tissues, and biofluids. To address these analytical challenges, we devised a reproducible, sensitive, and integrated lipidomics workflow based on normal-phase liquid chromatography-Fourier transform mass spectrometry (LC-FTMS) and LC-ITMS(2) (ion trap tandem mass spectrometry) for profiling and structural analysis of lipid species. The workflow uses a normal-phase LC system for efficient separation of apolar and polar lipid species combined with sensitive and specific analysis powered by a chip-based nanoelectrospray ion source and a hybrid ion trap-orbitrap mass spectrometer. The workflow was executed using a primary LC-FTMS survey routine for identification and profiling of lipid species based on high-mass accuracy and retention time followed by a targeted LC-ITMS(2) routine for characterizing the fatty acid moieties of identified lipid species. We benchmarked the performance of the workflow by characterizing the chromatographic properties of the LC-MS system for general lipid analysis. In addition, we demonstrate the efficacy of the workflow by reporting a study of low-abundant triacylglycerol and ceramide species in mouse brain cerebellum and 3T3-L1 adipocytes, respectively. The workflow described here is generic and can be extended for detailed lipid analysis of sample matrices having a wide range of lipid compositions.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense, Denmark.
authors.forename: Elena
authors.initials: E
authors.lastname: Sokol
authors.forename: Reinaldo
authors.initials: R
authors.lastname: Almeida
authors.forename: Hans Kristian
authors.initials: HK
authors.lastname: Hannibal-Bach
authors.forename: Dorota
authors.initials: D
authors.lastname: Kotowska
authors.forename: Johannes
authors.initials: J
authors.lastname: Vogt
authors.forename: Jan
authors.initials: J
authors.lastname: Baumgart
authors.forename: Karsten
authors.initials: K
authors.lastname: Kristiansen
authors.forename: Robert
authors.initials: R
authors.lastname: Nitsch
authors.forename: Jens
authors.initials: J
authors.lastname: Knudsen
authors.forename: Christer S
authors.initials: CS
authors.lastname: Ejsing
mesh_terms: D041721:3T3-L1 Cells / Q000737:chemistry*; D000818:Animals; D002518:Ceramides / Q000145:classification / Q000302:isolation & purification*; D002531:Cerebellum / Q000737:chemistry*; D002853:Chromatography, Liquid; D057927:Hydrophobic and Hydrophilic Interactions; D051379:Mice; D008810:Mice, Inbred C57BL; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry; D014280:Triglycerides / Q000145:classification / Q000302:isolation & purification*
chemical_list: D002518:Ceramides; D014280:Triglycerides
keywords: Lipidomics; Nanoelectrospray ionization; Normal-phase LC; Orbitrap mass spectrometry