Examining the collision‐induced decomposition spectra of ammoniated triglycerides as a function of fatty acid chain length and degree of unsaturation. II. The PXP/YPY series

Abstract

A series of positionally pure triglycerides (TAGs) of the form PXP and YPY, where P is the palmitate moiety and X and Y are large arrays of different fatty acid moieties, is synthesized and analyzed by reversed-phase high-performance liquid chromatography/tandem mass spectrometry. The intensities of the collision-induced decomposition (CID) products of ammoniated TAGs were examined as a function of chain length, degree of unsaturation, double-bond position, and cis/trans configuration of X and Y. The major CID products, the diglyceride (DAG) fragment ions and the MH(+) ions, are plotted as functions of chain length for the saturated and monounsaturated series of X and Y. Different trends for each of these series are observed. Trends in the intensities of these fragment ions are also characterized as a function of degree of unsaturation in the TAGs. In general, the fractional intensities of MH(+) increase with increasing degree of unsaturation in the TAGs. MH(+) is absent in the CID spectra of triglycerides containing three saturated fatty acid moieties, suggesting that the presence of double bonds fosters the formation of MH(+). Double bonds positioned close to the carbonyl carbon along the fatty acid chain promote the formation of the DAG fragment ion corresponding to the loss of the fatty acid. The collection of PXP/YPY data produced in this work is used to test the mechanisms of the formation and decomposition of ammoniated TAGs that were previously presented. The YPY data are used to predict the intensities of the fragment ions in the CID spectra of YPX-type TAGs. The limitations of the mathematical approach used in these predictions are discussed in context of a broader plan to develop a software platform for comprehensive analysis of complex TAG mixtures.

Detection of the abundance of diacylglycerol and triacylglycerol molecular species in cells using neutral loss mass spectrometry

“…The next stage would be to uniquely identify those molecular species containing the fatty acyl group by MS 3 as demonstrated here (Table 4) perhaps before and after stimulation. As previously reported, changes in fatty acyl position can significantly alter the ion yield of the loss of the fatty acyl group (17,22); thus, it would be expected that remodeling of a molecular species to an isomeric molecular species containing the same fatty acyl groups may even be detected if the fatty acyl group that undergoes the neutral loss moves from sn-1 (3) to sn-2. If remodeling of a TAG or DAG species takes place with substitution of different fatty acyl groups, there will be an increase or decrease in molecular weight with corresponding loss of neutral loss abundance or increase in neutral loss abundance.…”

“…In the RAW cell neutral loss experiment for 273 u, an ion at m/z 857.8 was observed (Figure 4). From previous work of Evans [17,22], it was clear that absolute abundances of each neutral loss of RCOOH + NH 3 from a synthetic TAG (and DAG) depended significantly on acyl position on the glycerol backbone, number of fatty acyl carbon atoms, and number of double bonds in the fatty acyl group. Thus it would not be valid to assume the abundance of each ion appearing in the neutral loss scan unambiguously indicated the quantitative measure of each component relative to the abundance of others.…”

“…The second assumption is that the unique molecular species observed in the neutral loss, form the fragment ions in a reproducible fashion. There is evidence that this does occur in that the position of the acyl group largely determines abundance of the neutral fragment (17,22); therefore, if position of the fatty acyl group does not change, then abundance of the fragment ion will not change. Specifically, the relative yield of neutral loss 273 from the species 16:0/18:1/18:1 from m/z 876.8 ( Figure 3B) would be constant relative to the yield of neutral loss 273 from the internal standard D 5 -16:0/18:0/16:0 observed at m/z 857.8.…”

“…One confounding factor is that ionization efficiencies between different molecular species and the extent of acyl loss (RCOOH+NH 3 ) could vary for each fatty acyl group [17,22]. Analyses of other classes of lipids using mass spectrometry typically engage a method to quantify the molecular abundance of the lipid species of interest, which involves simultaneous analysis of one or more internal standards [23,24].…”

Sequential Collision- and Ozone-Induced Dissociation Enables Assignment of Relative Acyl Chain Position in Triacylglycerols

“…12,15 Further complications arise where the FA identity influences the product ion abundance. [17][18][19] Rather than relying on ion abundance ratios in CID spectra, an explicit marker ion for assigning positional isomerism in TGs is desirable. This approach was first demonstrated with high energy (> 1 keV) CID of [TG + M] + ions on a multi-sector instrument and has since been replicated on tandem time-offlight platforms.…”