Structural Analysis of Lipid Hydroperoxides Using Mass Spectrometry with Alkali Metals

Abstract

Lipid oxidation is involved in various biological phenomena (e.g., oxylipin generation and oxidative stress). Of oxidized lipid structures, the hydroperoxyl group position of lipid hydroperoxides (LOOHs) is a critical factor in determining their biological roles. Despite such interest, current methods to determine hydroperoxyl group positions possess some drawbacks such as selectivity. While we previously reported mass spectrometric methods using Na+ for the highly selective determination of hydroperoxyl group positions, nothing was known except for the fact that sodiated LOOHs (mainly linoleate) provide specific fragment ions. Thus, this study was aimed to investigate the effects of different alkali metals on the fragmentation of LOOHs, assuming its further application to analysis of other complex LOOHs. From the analysis of PC 16:0/18:2;OOH (phosphatidylcholine) and FA 18:2;OOH (fatty acid), we found that fragmentation pathways and ion intensities largely depend on the binding position and type of alkali metals (i.e., Li+, Hock fragmentation; Na+ and K+, α-cleavage (Na+ > K+); Rb+ and Cs+, no fragmentation). Furthermore, we proved that this method can be applied to determine the hydroperoxyl group position of esterified lipids (e.g., phospholipids and cholesterol esters) as well as polyunsaturated fatty acids (PUFAs) including n-3, n-6, and n-9 FA. We anticipate that the insights described in this study provide additional unique insights to conventional lipid oxidation research.

Investigation of Lipoproteins Oxidation Mechanisms by the Analysis of Lipid Hydroperoxide Isomers

“…First of all, to achieve accurate analyses, we prepared authentic standards. When these standards were analyzed, in accordance with our previous studies [5,13], sodiated PC 16:0/18:2;OOH and CE 18:2;OOH isomers generated the hydroperoxyl group position-specific product ions (Figure 2). On the other hand, this study newly showed that sodiated cis-trans isomers of PC 16:0/18:2;OOH (e.g., Figure 2A,B) and CE 18:2;OOH (e.g., Figure 2G,H) did not provide the differences in product ions.…”

“…Using this method, we analyzed PC 16:0/18:2;9OOH and PC 16:0/18:2;13OOH in human plasma and hypothesized the contribution of radical and/or enzymatic oxidation to PC 16:0/18:2;OOH formation in human plasma lipoproteins [5]. Recently, we further showed the evidence that CID of sodiated LOOH is available to various lipid classes, including cholesterol ester [13]. Therefore, in this study, to ensure the above hypothesis (i.e., LOOH present in plasma lipoproteins are mainly derived from radical and/or enzymatic oxidation), we firstly developed novel analytical methods for PC 16:0/18:2;OOH and CE 18:2;OOH isomers.…”

“…The hydroperoxyl group of the resulting FA 18:2;OOH isomers was then protected with MxP for subsequent esterification [12]. Protected FA 18:2;OOH isomers were esterified with LPC 16:0/0:0 or cholesterol [12,13]. Protected PC 16:0/18:2;OOH and CE 18:2;OOH isomers were finally deprotected and purified as in a previous study [12,13].…”

“…Protected FA 18:2;OOH isomers were esterified with LPC 16:0/0:0 or cholesterol [12,13]. Protected PC 16:0/18:2;OOH and CE 18:2;OOH isomers were finally deprotected and purified as in a previous study [12,13].…”

Analytical and Structural Tools of Lipid Hydroperoxides: Present State and Future Perspectives

“…Recently Kato et al [ 81 ] investigated the effects of different alkali metals on the fragmentation of LOOHs. From the analysis of PC 16:0/18:2;OOH (phosphatidylcholine) and FA 18:2;OOH (fatty acid), fragmentation pathways and ion intensities were found to depend largely on the binding position and type of alkali metals (i.e., Li + , Hock fragmentation; Na + and K + , α -cleavage (Na + > K + ); Rb + and Cs + , no fragmentation).…”