title: Determination of nonpolar and polar lipid classes in human plasma, erythrocytes and plasma lipoprotein fractions using ultrahigh-performance liquid chromatography-mass spectrometry.
abstract: A novel normal-phase (NP) ultrahigh-performance liquid chromatography-mass spectrometry (UHPLC/MS) method is developed for a separation and quantitation of nonpolar lipid classes occurring in human plasma, erythrocytes and plasma lipoprotein fractions. The baseline class separation of cholesteryl esters (CE), cholesterol, triacylglycerols (TG), regioisomers of 1,2- and 1,3-diacylglycerols (DG) and 1-monoacylglycerols (1-MG) is achieved using an optimized hexane - 2-propanol-acetonitrile mobile phase within 18min for all nonpolar lipid classes or only 9min excluding monoacylglycerols not detected in studied samples. The determination of individual nonpolar lipid classes is performed by the response factor approach and the use of dioleoyl ethylene glycol as a single internal standard. Polar lipid classes, such as phosphatidylglycerols (PG), phosphatidylethanolamines (PE), phosphatidylcholines (PC), sphingomyelins (SM) and lysophosphatidylcholines (LPC), are separated by hydrophilic interaction liquid chromatography (HILIC) using 5mmol/L aqueous ammonium acetate-methanol-acetonitrile gradient within 13minutes. The quantitation of polar lipid classes is done by a similar approach as for nonpolar lipid classes, but a different internal standard (sphingosyl PE d17:1/12:0) is used. The complementary information on fatty acyl profiles after the transesterification of the total lipid extract is obtained by gas chromatography with flame ionization detection (GC/FID). The applicability of developed methodology for fast and comprehensive characterization of blood lipidome is illustrated on samples of human plasma, erythrocytes, high-density lipoprotein (HDL), low-density lipoprotein (LDL) and very low-density lipoprotein (VLDL) fractions.
authors.affiliation: Department of Analytical Chemistry, Faculty of Chemical Technology, University of Pardubice, Studentská 573, 53210 Pardubice, Czech Republic. Electronic address: Michal.Holcapek@upce.cz.
authors.forename: Michal
authors.initials: M
authors.lastname: Holčapek
authors.affiliation: Department of Analytical Chemistry, Faculty of Chemical Technology, University of Pardubice, Studentská 573, 53210 Pardubice, Czech Republic.
authors.forename: Eva
authors.initials: E
authors.lastname: Cífková
authors.affiliation: Department of Analytical Chemistry, Faculty of Chemical Technology, University of Pardubice, Studentská 573, 53210 Pardubice, Czech Republic.
authors.forename: Blanka
authors.initials: B
authors.lastname: Červená
authors.affiliation: Department of Analytical Chemistry, Faculty of Chemical Technology, University of Pardubice, Studentská 573, 53210 Pardubice, Czech Republic.
authors.forename: Miroslav
authors.initials: M
authors.lastname: Lísa
authors.affiliation: Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacký University, 77515 Olomouc, Czech Republic.
authors.forename: Jitka
authors.initials: J
authors.lastname: Vostálová
authors.affiliation: Department of Internal Medicine I-Cardiology, University Hospital Olomouc, IP Pavlova 6, 77520 Olomouc, Czech Republic.
authors.forename: Jan
authors.initials: J
authors.lastname: Galuszka
mesh_terms: D001774:Blood Chemical Analysis / Q000379:methods*; D002853:Chromatography, Liquid; D004912:Erythrocytes / Q000737:chemistry*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008055:Lipids / Q000032:analysis* / Q000737:chemistry; D008074:Lipoproteins / Q000097:blood*; D013058:Mass Spectrometry; D010713:Phosphatidylcholines / Q000737:chemistry
chemical_list: D008055:Lipids; D008074:Lipoproteins; D010713:Phosphatidylcholines
keywords: HILIC; LC/MS; Lipids; Normal-phase; Plasma, Erythrocytes