Analysis of lipid hydroperoxides and long-chain conjugated keto acids by negative ion electrospray mass spectrometry

Abstract

Lipid hydroperoxides are important products of enzymatic processes and autooxidation products of polyunsaturated fatty acids. Analysis of such compounds has proved difficult in the past, but negative ion electrospray ionization mass spectrometry was found to be suitable for direct analysis. Abundant [M - H](-) ions were observed in full scan mode for hydroper-oxyeicosatetraenoic (HPETE), hydroperoxyoctadecenoic acid isomers, and 5,12-diHPETE. Loss of water was observed for all species. Collisional activation and tandem mass spectrometry generated unique and characteristic spectra that shared some common features such as loss of small neutral molecules. More importantly, fragment ions that were indicative of the position of the hydroperoxide were observed. Collision-induced decomposition (CID) of [M - H2O](-) for the HPETE isomers was found to be virtually identical to the CID mass spectra of the [M - H](-) anions from corresponding keto-eicosatetraenoic acids, which suggests that the hydroperoxide anions decompose via a dehydration intermediate that resembles the keto acid molecular anion. Cleavage of the double bond allylic to the hydroperoxide formed structurally characteristic ions at m/z 129 from 5-HPETE, m/z 153 from 12-HPETE, and m/z 113 from 15-HPETE. Charge-driven allylic fragmentation led to formation of m/z 203 from 5-HPETE, m/z 179 from 12-HPETE, and m/z 219 from 15-HPETE. Mechanisms consistent with the decomposition of stable isotope analogues are proposed for the formation of these and other characteristic ions. These specific decompositions can be used in multiple reaction monitoring to measure picomolar concentrations of hydroperoxides by direct high performance liquid chromatography tandem mass spectrometry.

Mass spectrometric analysis of leukotriene A4 and other chemically reactive metabolites of arachidonic acid

“…These ions were produced at much lower abundance than the previous described ions. The ion at m/z 203 has been observed in the collisional activation of other eicosanoids with a single oxygen substitution at carbon atom 5 such as 5-HETE [16] and 5-oxo-ETE [17]. The appearance of m/z 189 has been previously observed in the mass spectra of other leukotrienes which contain a conjugated triene motif [18].…”

On the singular, dual, and multiple positional specificity of manganese lipoxygenase and its G316A mutant

“…MS/MS analysis of hydroperoxy fatty acids yields a strong signal attributable to loss of water, and the hydroperoxides are also partly dehydrated to the keto compounds in the heated capillary (200-325jC) of the mass spectrometer (44)(45)(46). The MS/MS/MS spectra of hydroperoxides are virtually identical to the MS/MS spectra of the corresponding keto fatty acids.…”

“…Genes homologous to Mn-LO occur in Aspergillus and in the Magnaporthaceae family. Whether any of these putative lipoxygenases are MS/MS analysis (m/z 31Yfull scan) of 13-ketoeicosatetraenoic acid (13-KETE), which was formed by dehydration of 13-HPETE in the ion trap (44,46). C: MS/MS/MS analysis (m/z 343Ym/z 324-325Yfull scan) of [5,6,8,9,11,12,14, H 8 ]13-HPETE.…”

Analysis of unsaturated compounds by Ag+ coordination ionspray mass spectrometry: Studies of the formation of the Ag+/lipid complex

“…Recently, Murphy and colleagues reported the use of negative ion electrospray mass spectrometry for the analysis of lipid hydroperoxides and long-chain keto acids [8]. This technique allows for the coupling of reverse phase HPLC and electrospray ionization mass spectrometry (ESI-MS), thus providing a powerful separation and structure elucidation tool.…”

“…This technique allows for the coupling of reverse phase HPLC and electrospray ionization mass spectrometry (ESI-MS), thus providing a powerful separation and structure elucidation tool. However, to generate negative ions, free acids must be used instead of cholesterol esters [8,9]. Our interest lies in the study of intact lipids.…”