Identification of 1‐palmitoyl‐2‐linoleoyl‐phosphatidylethanolamine modifications under oxidative stress conditions by LC‐MS/MS

Abstract

Phosphatidylethanolamines are a major class of phospholipids found in cellular membranes. Identification of the alterations in these phospholipids, induced by free radicals, could provide new tools for in vivo diagnosis of oxidative stress. In this study, 1-palmitoyl-2-linoleoyl-phosphatidylethanolamine oxidation products, induced by the hydroxyl radical, were studied using LC-MS and LC-MS/MS. Data obtained allowed the identification and separation of isomeric oxidative products with modifications in the sn-2 acyl chain, attributed to long- and short-chain products. Among long-chain products keto, keto-hydroxy, hydroxy, poly-hydroxy, peroxy and hydroxy-peroxy derivatives were identified. Product ions formed by loss of two H2O molecules vs loss of HOOH, allowed the identification of, respectively, di- (or poli-) hydroxy vs peroxy derivatives. Location of functional groups was determined by the product ions formed by cleavage of C-C bonds, in the vicinity of the oxidation positions, allowing the identification of C9, C12 and C13 as the predominant substituted positions. Short-chain products identified comprised aldehydes, hydroxy-aldehydes and carboxylic derivatives, with modified sn-2 acyl lengths of C7-C9 and C11, C12. Among the short-chain products identified, C9 products showed higher relative abundance.

Cardiolipin and oxidative stress: Identification of new short chain oxidation products of cardiolipin in in vitro analysis and in nephrotoxic drug-induced disturbances in rat kidney tissue

“…They showed higher relative abundance when compared with the other C13 shortened chains oxidation products. During oxidation of other phospholipids, namely phosphatidylcholine and phosphatidylethanolamine with a linoleic acid, it was observed that the short chain products with C9 were most abundant than the ones in C13 [26,27] and these results are consistent with those findings.…”

“…However, these products formed by cleavage of fatty acyl chains were previously observed during phosphatidylcholine (PC) and phosphatidylethanolamine (PE) oxidation [26,27], being this an usual process during lipid peroxidation, which corroborates the formation of short chain products during CL oxidation. To date, only products formed by insertion of oxygen atoms [16], or formed by cleavage of the inner glycerol backbone with formation of phosphatidic acid and diacylphosphatidyl-hydroxyacetone [28] were mentioned to occur during CL oxidation.…”

“…This intermediate generates the correspondent C13 alkoxyl radical that can undergo cleavage with formation of an aldehyde that further oxidize to terminal carboxylic function. These shortened chain products are typical of lipid peroxidation [25] and their formation was observed under oxidative conditions in PC and PE phospholipids [26,27]. The structure of these CL short chain oxidation products were con- [30].…”

Structural Characterization of Oxidized Glycerophosphatidylserine: Evidence of Polar Head Oxidation

“…These ions correspond to oxidation products with shortened acyl chain at C9 and a carboxylic acid terminal. Short chain oxidation products originated from cleavage of fatty acyl chains have been described for PS [13] and have been studied in phosphatidylcholine, phosphatidylserine, and cardiolipin phospholipids [27][28][29]. The shortened oxidation products are generated after abstraction of the bis-allylic hydrogen atoms by the hydroxyl radical, which through a β-scission mechanism, break down to short-chain phospholipid products with terminal aldehydic or carboxylic acid function.…”

A simplified procedure for semi-targeted lipidomic analysis of oxidized phosphatidylcholines induced by UVA irradiation

“…Because of the tremendous excess of unoxidized lipids and the presence of isobaric peaks, analysis of individual OxPLs by mass spectrometry is only feasible after chromatographic separation of lipids, typically by reversed-phase HPLC. Previous publications described mass spectrometry-based methods for analysis of oxidized cardiolipin (15)(16)(17), phosphatidylserine ( 15,16,18 ), phosphatidylethanolamine ( 6,(19)(20)(21)(22), and phosphatidylcholine ( 23-25 and references below). The results show that different PL classes have variable propensity for oxidation.…”