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Recent Advances on Mass Spectrometry Analysis of Nitrated Phospholipids
- Melo, Tânia, Domingues, Pedro, Ferreira, Rita, Milic, Ivana, Fedorova, Maria, Santos, Sérgio M., Segundo, Marcela A., Domingues, M. Rosário
- Analytical chemistry 2016 v.88 no.5 pp. 2622-2629
- animal models, electrospray ionization mass spectrometry, esterification, fatty acids, hydrophilic interaction chromatography, hydrophobicity, insulin-dependent diabetes mellitus, ionization, ions, mitochondria, phosphatidylcholines, phosphatidylethanolamines, phosphatidylserines
- In recent years, there has been an increasing interest in nitro fatty acids (NO₂-FA) as signaling molecules formed under nitroxidative stress. NO₂-FA were detected in vivo in a free form, although it is assumed that they may also be esterified to phospholipids (PL). Nevertheless, insufficient discussion about the nature, origin, or role of nitro phospholipids (NO₂-PL) was reported up to now. The aim of this study was to develop a mass spectrometry (MS) based approach which allows identifying nitroalkenes derivatives of three major PL classes found in living systems: phosphatidylcholines (PCs), phosphatidylethanolamine (PEs), and phosphatidylserines (PSs). NO₂-PLs were generated by NO₂BF₄ in hydrophobic environment, mimicking biological systems. The NO₂-PLs were then detected by electrospray ionization (ESI-MS) and ESI-MS coupled to hydrophilic interaction liquid chromatography (HILIC). Identified NO₂-PLs were further analyzed by tandem MS in positive (as [M + H]⁺ ions for all PL classes) and negative-ion mode (as [M – H]⁻ ions for PEs and PSs and [M + OAc]⁻ ions for PCs). Typical MS/MS fragmentation pattern of all NO₂-PL included a neutral loss of HNO₂, product ions arising from the combined loss of polar headgroup and HNO₂, [NO₂-FA + H]⁺ and [NO₂-FA – H]⁻ product ions, and cleavages on the fatty acid backbone near the nitro group, allowing its localization within the FA akyl chain. Developed MS method was used to identify NO₂-PL in cardiac mitochondria from a well-characterized animal model of type 1 diabetes mellitus. We identified nine NO₂-PCs and one NO₂-PE species. The physiological relevance of these findings is still unknown.