Preparation of marine plasmalogen and selective identification of molecular species by LC-MS/MS

In this study, we investigated the laboratory-scale preparation and characteristics of ethanolamine plasmalogen from marine invertebrates. The preparation method consists of fractionation by acetone and ether treatment, and separation using column chromatography with silica gel and different eluents. Plasmalogen fractions (Pls fraction) were obtained from the viscera of the ascidian Halocynthia roretzi, and the prominent fatty acids were present as 20:5 (33.0%) and 22:6 (29.6%) n-3 polyunsaturated fatty acids (PUFA). The plasmalogen purity was 40%, and the alkenyl chains consisted of 18:0 (86.1%), 16:0 (5.9%) and 18:1 (4.9%). Precursor ion scanning in negative and positive ion modes using liquid chromatography-tandem mass spectrometry (LC-MS/MS) enabled the profiling of phosphatidylethanolamine (PE) molecular species in ascidian viscera. Following LC-MS/MS with multiple reaction monitoring (MRM), the prominent plasmalogen species were found to be 18:0p/20:5 (30.4%) and 18:0p/22:6 (24.6%) (p at sn-1 position indicates alkenyl linkage). In conclusion, this preparative procedure using ascidian viscera as a source achieved 40% pure plasmalogen that was rich in n-3 PUFA. In addition, an LC-MS/MS assay enabled rapid analysis of plasmalogen species with selectivity and sensitivity. The present results will contribute to the understanding of dietary plasmalogen absorption and metabolism.