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Identification of Double Bond Position Isomers in Unsaturated Lipids by m-CPBA Epoxidation and Mass Spectrometry Fragmentation.

ABSTRACT: Lipids are highly diverse biomolecules associated with several biological functions including structural constituent, energy storage, and signal transduction. It is essential to characterize lipid structural isomers and further understand their biological roles. Unsaturated lipids contain one or multiple carbon-carbon double bonds. Identifying double bond position presents a major challenge in unsaturated lipid characterization. Recently, several advancements have been made for double bond localization by mass spectrometry (MS) analysis. However, many of these studies require complex chemical reactions or advanced mass spectrometers with special fragmentation techniques, which limits the application in lipidomics study. Here, an innovative meta-chloroperoxybenzoic acid ( m-CPBA) epoxidation reaction coupling with collision-induced dissociation (CID)-MS/MS strategy provides a new tool for unsaturated lipidomics analysis. The rapid epoxidation reaction was carried out by m-CPBA with high specificity. Complete derivatization was achieved in minutes without overoxidized byproduct. Moreover, diagnostic ion pair with 16 Da mass difference indicated localization of carbon-carbon double bond in MS/MS spectra. Multiple lipid classes were evaluated with this strategy and generated abundant fragments for structural analysis. Unsaturated lipid analysis of yeast extract using this strategy took less than 30 min, demonstrating the potential for high-throughput lipidomics analysis by this approach. This study opens a door for high throughput unsaturated lipid analysis with minimal requirement for instrumentation, which could be widely applied in lipidomics analysis.

SUBMITTER: Feng Y

PROVIDER: S-EPMC6408215 | biostudies-other | 2019 Feb

REPOSITORIES: biostudies-other

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Project description:The position(s) of carbon-carbon double bonds within lipids can dramatically affect their structure and reactivity and thus has a direct bearing on biological function. Commonly employed mass spectrometric approaches to the characterization of complex lipids, however, fail to localize sites of unsaturation within the molecular structure and thus cannot distinguish naturally occurring regioisomers. In a recent communication [Thomas, M. C.; Mitchell, T. W.; Blanksby, S. J. J. Am. Chem. Soc. 2006, 128, 58-59], we have presented a new technique for the elucidation of double bond position in glycerophospholipids using ozone-induced fragmentation within the source of a conventional electrospray ionization mass spectrometer. Here we report the on-line analysis, using ozone electrospray mass spectrometry (OzESI-MS), of a broad range of common unsaturated lipids including acidic and neutral glycerophospholipids, sphingomyelins, and triacylglycerols. All lipids analyzed are found to form a pair of chemically induced fragment ions diagnostic of the position of each double bond(s) regardless of the polarity, the number of charges, or the adduct ion (e.g., [M - H](-), [M - 2H](2-), [M + H](+), [M + Na](+), [M + NH(4)](+)). The ability of OzESI-MS to distinguish lipids that differ only in the position of the double bonds is demonstrated using the glycerophosphocholine standards, GPCho(9Z-18:1/9Z-18:1) and GPCho(6Z-18:1/6Z-18:1). While these regioisomers cannot be differentiated by their conventional tandem mass spectra, the OzESI-MS spectra reveal abundant fragment ions of distinctive mass-to-charge ratio (m/z). The approach is found to be sufficiently robust to be used in conjunction with the m/z 184 precursor ion scans commonly employed for the identification of phosphocholine-containing lipids in shotgun lipidomic analyses. This tandem OzESI-MS approach was used, in conjunction with conventional tandem mass spectral analysis, for the structural characterization of an unknown sphingolipid in a crude lipid extract obtained from a human lens. The OzESI-MS data confirm the presence of two regioisomers, namely, SM(d18:0/15Z-24:1) and SM(d18:0/17Z-24:1), and suggest the possible presence of a third isomer, SM(d18:0/19Z-24:1), in lower abundance. The data presented herein demonstrate that OzESI-MS is a broadly applicable, on-line approach for structure determination and, when used in conjunction with established tandem mass spectrometric methods, can provide near complete structural characterization of a range of important lipid classes. As such, OzESI-MS may provide important new insight into the molecular diversity of naturally occurring lipids.

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Identification of Double Bond Position Isomers in Unsaturated Lipids by m-CPBA Epoxidation and Mass Spectrometry Fragmentation.

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