Identification and analyses of ether-containing ethanolamine phospholipid molecular species in lipid extracts of mouse cerebelluma.

aMouse cerebellar lipids were extracted by using a modified Bligh and Dyer procedure [21]. Analyses of PtdEtn molecular species were performed in the negative-ion mode using an LTQ-Orbitrap mass spectrometer with an electrospray ion source by using a shotgun lipidomics approach. The determined monoisotopic masses (column 1) of each PtdEtn molecular species were externally calibrated relative to the selected internal standard at m/z 686.476. The molecular formulas listed in column 2 were derived from accurate mass analyses of monoisotopic mass and were grouped into each isobaric mass. The prefix “a”, “d”, and “p” stand for alkyl-acyl PtdEtn, diacyl PtdEtn, and plasmalogen PtdEtn, respectively. The content of each individual molecular species listed in column 3 was determined through ratiometric comparison of the ion peak intensity with that of the internal standard after 13C de-isotoping. The data represent X±SD of at least four separate animals. The notation m∶n represents the fatty acyl (or ether aliphatic) chain containing m carbons and n double bonds which do not include the vinyl ether double bond in plasmalogens. The numbers in the parentheses represent the relative composition of each individual molecular species of an isobaric ion. The symbols of “” indicate that the data represent the best estimation from the analyses.bIdentification of individual pPtdEtn molecular species was performed based on the accurate mass analyses and acidic vapor treatment as described in the text. Identification of individual aPtdEtn molecular species was performed based on the accurate mass analyses, the paired rule, and the information of the identified pPtdEtn counterparts as discussed in the text. Identification of individual dPtdEtn molecular species was conducted based on accurate mass analyses. The abundance of each of the paired dPtdEtn molecular species cannot be accurately determined at the current stage of lipidomic technology.