Phospholipids are an important class of lipids in cell membranes and food. Several high-performance liquid chromatography (HPLC) methods have been developed to analyze phospholipids at the molecular species level. We developed a two-dimensional HPLC system with a charged aerosol detector and mass spectrometry (MS) to analyze phosphatidylethanolamine (PE), phosphatidylcholine (PC), and their plasmalogens (pls) extracted from food materials. Accordingly, the phospholipid molecular species can be analyzed in a single step despite using smaller samples. We confirmed that chromatogram peaks from soybean lecithin are mostly baseline separated, assigned, and quantified (24 molecular species for PE and 27 for PC). In addition, it was confirmed that chromatograms of lipids extracted from chicken breast meat include plasmalogen peaks. The PE fraction in lipids extracted from chicken breast meat contained 17 types of ethanolamine plasmalogens, corresponding to approximately 57% of the total by weight. The PC fraction contained only four choline plasmalogens, corresponding to approximately 11% of the total weight. The composition of the pls-PC molecular species differed from that of pls-PEs. The polyunsaturated fatty acids connected at the sn-2 positions of the pls-PEs consisted of 20.5% 20:4 fatty acid and were independent of the carbon chain at the sn-1 position. However, the 18:1 fatty acid at the sn-2 position was dependent on the carbon chain at the sn-1 position.
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