Acetonitrile covalent adduct chemical ionization tandem mass spectrometry of non-methylene-interrupted pentaene fatty acid methyl esters.

Acetonitrile covalent adduct chemical ionization tandem mass spectrometry (CACIMS/MS) has shown to be an efficient method for the identification of double-bond position in homoallylic, conjugated and several polyene non-methylene-interrupted (NMI) fatty acid methyl esters. However, it has not been thoroughly evaluated for NMI highly unsaturated fatty acids (HUFA) with more than four double bonds. Docosahexaenoic acid (DHA)-rich single cell oil (DHASCO(®); Martek Biosciences, Corp.) was partially hydrogenated (partially hydrogenated DHASCO; PHDO) producing ten novel 22:5 and 22:6 HUFA isomers. In single-stage MS, the ratio of [M+54](+)/[M+54-32](+) for the 22:5 and 22:6 isomers indicated the presence of homoallylic or partially conjugated double-bond systems. The CACIMS/MS spectra revealed six 22:5 isomers with diagnostic ions corresponding to the homoallylic 22:5n-6 and 22:5n-3 isomers, and four distinct NMI 22:5 isomers. Diagnostic ions for four 22:6 isomers were identical to the native DHA illustrating that CACIMS/MS is sensitive to double-bond position but not geometry. Three gas chromatography (GC) peaks for partially conjugated 22:6 isomers were also detected and clearly distinguishable from homoallylic 22:6 isomers, but their CACIMS/MS spectra did not yield prominent ions indicative of double-bond position, possibly due to co-elution. Overall, CACIMS/MS was effective for determining double-bond position in NMI 22:5 isomers. Further investigations are warranted to evaluate and determine fragmentation patterns for partially conjugated and NMI 22:6 HUFA.
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