title: Rapid and Reliable Identification of Phospholipids for Untargeted Metabolomics with LC-ESI-QTOF-MS/MS.
abstract: Lipids are important components of biological systems, and their role can be currently investigated by the application of untargeted, holistic approaches such as metabolomics and lipidomics. Acquired data are analyzed to find significant signals responsible for the differentiation between the investigated conditions. Subsequently, identification has to be performed to bring biological meaning to the obtained results. Lipid identification seems to be relatively easy due to the known characteristic fragments; however, the large number of structural isomers and the formation of different adducts makes it challenging and at risk of misidentification. The inspection of data, acquired for plasma samples by a standard metabolic fingerprinting method, revealed multisignal formations for phosphatidylcholines, phosphatidylethanolamines, and sphingomyelins by the formation of ions such as [M + H](+), [M + Na](+), and [M + K](+) in positive ionization mode and [M - H](-), [M + HCOO](-), and [M + Cl](-) in negative mode. Moreover, sodium formate cluster formation was found for [M + H·HCOONa](+) and [H-H·HCOONa](-). The MS/MS spectrum obtained for each of the multi-ions revealed significant differences in the fragmentation, which were confirmed by the analysis of the samples in two independent research centers. After the inspection of an acquired spectra, a list of characteristic and diagnostic fragments was proposed that allowed for easy, quick, and robust lipid identification that provides information about the headgroup, formed adduct, and fatty acyl composition. This ensures successful identification, which is of great importance for the contextualization of data and results validation.
authors.affiliation: †CEMBIO, Centre for Metabolomics and Bioanalysis, San Pablo CEU University, 28668 Boadilla del Monte, Spain.
authors.forename: Joanna
authors.initials: J
authors.lastname: Godzien
authors.affiliation: ‡Clinical Research Centre, Medical University of Bialystok, 15-089 Białystok, Poland.
authors.forename: Michal
authors.initials: M
authors.lastname: Ciborowski
authors.affiliation: †CEMBIO, Centre for Metabolomics and Bioanalysis, San Pablo CEU University, 28668 Boadilla del Monte, Spain.
authors.forename: María Paz
authors.initials: MP
authors.lastname: Martínez-Alcázar
authors.affiliation: ‡Clinical Research Centre, Medical University of Bialystok, 15-089 Białystok, Poland.
authors.forename: Paulina
authors.initials: P
authors.lastname: Samczuk
authors.affiliation: ‡Clinical Research Centre, Medical University of Bialystok, 15-089 Białystok, Poland.
authors.forename: Adam
authors.initials: A
authors.lastname: Kretowski
authors.affiliation: †CEMBIO, Centre for Metabolomics and Bioanalysis, San Pablo CEU University, 28668 Boadilla del Monte, Spain.
authors.forename: Coral
authors.initials: C
authors.lastname: Barbas
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D007477:Ions / Q000737:chemistry / Q000378:metabolism; D055432:Metabolomics / Q000379:methods*; D015394:Molecular Structure; D010713:Phosphatidylcholines / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D010743:Phospholipids / Q000097:blood / Q000737:chemistry / Q000378:metabolism*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013109:Sphingomyelins / Q000097:blood / Q000737:chemistry / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013997:Time Factors
chemical_list: D007477:Ions; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010743:Phospholipids; D013109:Sphingomyelins
keywords: CID; adduct formation; fingerprinting; fragmentation; ionization; phosphatidylcholine; phosphatidylethanolamine; phospholipids; sphingomyelin; structural elucidation