Basic eluent for rapid and comprehensive analysis of fatty acid isomers using reversed-phase high performance liquid...

Comprehensive separation of fatty acids, including various isomers, is very important for chemical analyses associated with detailed physiological function investigations. We found that a basic eluent is effective for realizing clear separation of double-bond regioisomers by reversed-phase high-performance liquid chromatography. We carried out a comprehensive analysis of fatty acids, including double-bond positional isomers, trans-fatty acids, and iso-fatty acids using reversed-phase high-performance liquid chromatography coupled with Fourier transform mass spectrometry (LC/FTMS). Clear separation of these fatty acids was achieved using a C18 column and a basic eluent. A three-pump gradient system using acetone provided rapid elution within 22 min. In a single run, 184 fatty acid molecular species contained in a dietary fish oil supplement were detected by the FTMS full scan. A previously unreported peak was also detected, which was assigned as tetratriacontadecaenoic acid by comparison with the MS² profiles of fatty acids with known chemical structures. This result demonstrates that the developed method is useful for clarifying the composition of fatty acid molecular species in target samples, providing a promising approach to discover unreported fatty acids.