Wiesner Philipp, Leidl Katharina, Boettcher Alfred, Schmitz Gerd, Liebisch Gerhard
Institute of Clinical Chemistry and Laboratory Medicine, University of Regensburg, Germany.
Institute of Clinical Chemistry and Laboratory Medicine, University of Regensburg, Germany.
J Lipid Res. 2009 Mar;50(3):574-585. doi: 10.1194/jlr.D800028-JLR200. Epub 2008 Oct 1.
Glycerophospholipid and sphingolipid species and their bioactive metabolites are important regulators of lipoprotein and cell function. The aim of the study was to develop a method for lipid species profiling of separated lipoprotein classes. Human serum lipoproteins VLDL, LDL, and HDL of 21 healthy fasting blood donors were separated by fast performance liquid chromatography (FPLC) from 50 microl serum. Subsequently, phosphatidylcholine (PC), lysophosphatidylcholine, sphingomyelin (SM), ceramide (CER), phosphatidylethanolamine (PE), PE-based plasmalogen (PE-pl), cholesterol, and cholesteryl ester (CE) content of the separated lipoproteins was quantified by electrospray ionization tandem mass spectrometry (ESI-MS/MS). Analysis of FPLC fractions with PAGE demonstrated that albumin partially coelutes with HDL fractions. However, analysis of an HDL deficient serum (Tangier disease) showed that only lysophosphatidylcholine, but none of the other lipids analyzed, exhibited a significant coelution with the albumin containing fractions. Approximately 60% of lipoprotein CER were found in LDL fractions and 60% of PC, PE, and plasmalogens in HDL fractions. VLDL, LDL, and HDL displayed characteristic lipid class and species pattern. The developed method provides a detailed lipid class and species composition of lipoprotein fractions and may serve as a valuable tool to identify alterations of lipoprotein lipid species profiles in disease with a reasonable experimental effort.
甘油磷脂和鞘脂种类及其生物活性代谢产物是脂蛋白和细胞功能的重要调节因子。本研究的目的是开发一种用于分离脂蛋白类别的脂质种类分析方法。通过快速高效液相色谱（FPLC）从50微升血清中分离出21名健康空腹献血者的人血清脂蛋白极低密度脂蛋白（VLDL）、低密度脂蛋白（LDL）和高密度脂蛋白（HDL）。随后，通过电喷雾电离串联质谱（ESI-MS/MS）对分离出的脂蛋白中的磷脂酰胆碱（PC）、溶血磷脂酰胆碱、鞘磷脂（SM）、神经酰胺（CER）、磷脂酰乙醇胺（PE）、基于PE的缩醛磷脂（PE-pl）、胆固醇和胆固醇酯（CE）含量进行定量。用聚丙烯酰胺凝胶电泳（PAGE）分析FPLC级分表明，白蛋白部分与HDL级分共洗脱。然而，对一种HDL缺陷血清（丹吉尔病）的分析表明，只有溶血磷脂酰胆碱，而不是所分析的其他脂质，与含白蛋白的级分有显著的共洗脱。约60%的脂蛋白CER存在于LDL级分中，60%的PC、PE和缩醛磷脂存在于HDL级分中。VLDL、LDL和HDL呈现出特征性的脂质类别和种类模式。所开发的方法提供了脂蛋白级分详细的脂质类别和种类组成，并且可以作为一种有价值的工具，通过合理的实验工作量来识别疾病中脂蛋白脂质种类谱的改变。
