Identification of Long-Chain Unsaturated Fatty Acids in Deep-Sea Fish Oil by Liquid Chromatography/Mass Spectrometry/Atmospheric Pressure Chemical Ionization
Abstract A method for the identification of long-chain unsaturated fatty acids in deep-sea fish oil using 1,2-benzo-3,4-dihydrocarbazole-9-ethyl-benzenesulfonate (BDEBS) as pre-column derivatization reagent by reversed-phase high-performance liquid chromatography/mass spectrometry (HPLC/MS) has been developed. To avoid the migration of double bond in the fatty acid chain, fish oil samples are saponificated and derivatized under nitrogen circumstance. The derivatized solution was extracted with hexane and dried under a stream of nitrogen; the residue was re-dissolved with acetonitrile prior to LC analysis. MS analysis of the fatty acid derivatives exhibited regular information regarding the position of the double bond in the fatty acid chain. With molecular ion mass at m/z [MH] + and specific fragment ions obtained with MS analysis, the position of the double bond in fatty acid chain was located (or calculated) by means of several established formulae. According to the analysis of HPLC/MS/APCI, 23 fatty acids were identified. The main fatty acids in deep-sea fish oil are C 12 –C 22 and 67.08% (area percent) of them are polyunsaturated fatty acids. The main components of the unsaturated fatty acids are C 16:1 : 9-hexadecenoic acid, 11.7%; C 16:4 : 4,7,10,13-hexadecatetraenoic acid, 2.91%; C 18:1 : 12-octadecenoic acid, 11.1%; C 18:4 : 6,9,12,15-octadecatetraenoic acid, 3.62%; C 20:1 : 13-eicosenoic acid, 1.21%; C 20:5 : 5,8,11,14,17-eicosapentaenoic acid, 16.71%; C 22:6 : 2,5,8,11,14,17-docosahexenoic acid, 10.53%. The established method provides a new technical tool for the location of double bond positions in the polyunsaturated fatty acids.