A lipidomic workflow capable of resolving <i>sn</i>- and CC location isomers of phosphatidylcholines

Zhao, Xue; Zhang, Wenpeng; Zhang, Donghui; Li, Xinwei; Cao, Wenbo; Chen, Qinhua; Ouyang, Zheng; Xia, Yu

doi:10.1039/c9sc03521d

Cited by 67 publications

(82 citation statements)

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“…In addition to the C=C locations, sn -positions of PCs in single cells could also be identified. It has been reported that the sn -position of PCs could be identified by negative mode MS/MS analysis with the adduct of bicarbonate anion (HCO 3 − ) 25 . This method was adapted in this study for the single-cell lipid analysis by changing the assisted solvent to 50 mM ammonium bicarbonate solvent in methanol:acetonitrile:water = 4.5:4.5:1 (v/v).…”

Section: Resultsmentioning

confidence: 99%

“…The chemical structures of lipids are very complex and a significant amount of efforts have been put in the discovery of interesting chemistry (e.g., photochemical reactions and epoxidation) for elucidating lipid structures using MS 21 . Paternò-Büchi (PB) reaction combined with tandem MS is one of the most effective protocols for the detailed characterization and relative quantitation of lipids at carbon-carbon double bond (C=C) location and sn-position levels [22][23][24][25][26] . Epoxidation is also common in deep lipidomic analysis with C=C location specificity, owing to its universal accessibility [27][28][29][30][31] .…”

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confidence: 99%

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Single-cell lipidomics with high structural specificity by mass spectrometry

et al. 2021

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Single-cell analysis is critical to revealing cell-to-cell heterogeneity that would otherwise be lost in ensemble analysis. Detailed lipidome characterization for single cells is still far from mature, especially when considering the highly complex structural diversity of lipids and the limited sample amounts available from a single cell. We report the development of a general strategy enabling single-cell lipidomic analysis with high structural specificity. Cell fixation is applied to retain lipids in the cell during batch treatments prior to single-cell analysis. In addition to tandem mass spectrometry analysis revealing the class and fatty acyl-chain for lipids, batch photochemical derivatization and single-cell droplet treatment are performed to identify the C=C locations and sn-positions of lipids, respectively. Electro-migration combined with droplet-assisted electrospray ionization enables single-cell mass spectrometry analysis with easy operation but high efficiency in sample usage. Four subtypes of human breast cancer cells are correctly classified through quantitative analysis of lipid C=C location or sn-position isomers in ~160 cells. Most importantly, the single-cell deep lipidomics strategy successfully discriminates gefitinib-resistant cells from a population of wild-type human lung cancer cells (HCC827), highlighting its unique capability to promote precision medicine.

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Section: Resultsmentioning

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Single-cell lipidomics with high structural specificity by mass spectrometry

et al. 2021

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“…PB reaction converts the C=C to an oxetane which can be preferentially fragmented by low-energy collisioninduced dissociation (CID). Other approaches include ion mobility spectrometry (IMS) 36,37 and alternative gas-phase ion activation [38][39][40][41][42][43][44] . These lipid characterizing methods were also compatible with mass spectrometry imaging (MSI) [45][46][47][48][49][50] .…”

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Large-scale lipid analysis with C=C location and sn-position isomer resolving power

Cao¹,

Cheng²,

Yang³

et al. 2020

Nat Commun

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Lipids play a pivotal role in biological processes and lipid analysis by mass spectrometry (MS) has significantly advanced lipidomic studies. While the structure specificity of lipid analysis proves to be critical for studying the biological functions of lipids, current mainstream methods for large-scale lipid analysis can only identify the lipid classes and fatty acyl chains, leaving the C=C location and sn-position unidentified. In this study, combining photochemistry and tandem MS we develop a simple but effective workflow to enable large-scale and near-complete lipid structure characterization with a powerful capability of identifying C=C location(s) and sn-position(s) simultaneously. Quantitation of lipid structure isomers at multiple levels of specificity is achieved and different subtypes of human breast cancer cells are successfully discriminated. Remarkably, human lung cancer tissues can only be distinguished from adjacent normal tissues using quantitative results of both lipid C=C location and sn-position isomers.

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“…For instance, studies suggested positive associations between high ratios of dietary ω‐6:ω‐3 PUFAs and production of inflammatory mediators, while increased consumption of ω‐3 PUFAs appeared to suppress inflammation 6,7 . Moreover, changes in C18 (∆9):(∆11) were observed in breast cancer 8–10 . Mass spectrometry (MS) is a primary tool for lipid structure characterization.…”

Section: Introductionmentioning

confidence: 99%

In situ detection of fatty acid C=C positional isomers by coupling on‐tissue mCPBA epoxidation with infrared matrix‐assisted laser desorption electrospray ionization mass spectrometry

Garrard

Said

et al. 2021

Rapid Comm Mass Spectrometry

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Rationale Unsaturated fatty acids (UFAs) play vital roles in regulating cellular functions. In‐depth structural characterization of UFAs such as localizing carbon–carbon double bonds is fundamentally important but poses considerable challenges in mass spectrometry (MS) given that the most widely accessible ion activation method, low‐energy collision‐induced dissociation (CID), primarily generates uninformative fragments (e.g., neutral loss of CO2) that are not suggestive of the double‐bond positions. Methods m‐Chloroperoxybenzoic acid (mCPBA) was uniformly deposited onto the sample slides using a TM Sprayer, converting the carbon–carbon double bonds into epoxides under ambient conditions. The epoxidation product was ionized in situ by infrared matrix‐assisted laser desorption electrospray ionization mass spectrometry (IR‐MALDESI‐MS), and subsequently cleaved via CID, generating a diagnostic ion pair associated with the double‐bond position. The reaction efficiency, sensitivity and relative quantification capability of the method were validated with five UFA standards dried on glass slides, and then this strategy was demonstrated on thin tissue sections of rat liver and human bladder. Results The mCPBA reaction yielded conversion rates in the range of 44–60% in 10 min with high specificity and sensitivity. Further tandem mass spectrometry (MS/MS) of the mono‐epoxidized products generated informative fragment ions specific to the double‐bond positions, and relative quantification of positional isomers in binary mixtures was performed across a wide mole fraction from 0 to 1. An innovative spiral scan pattern was utilized during data acquisition, elucidating the major isomeric compositions of multiple UFAs from a tissue section in a single run. Conclusions The on‐tissue mCPBA epoxidation was implemented into an ambient MS imaging workflow to offer a rapid and simple way for in situ identification and relative quantification of double‐bond positional isomers without the requirement for instrument modification. The method can be readily implemented on many other MS platforms to reveal the role of double‐bond positional isomers in lipid biology and to discover potential biomarkers.

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A lipidomic workflow capable of resolving sn- and CC location isomers of phosphatidylcholines

Abstract: Large-scale profiling of phosphatidylcholines at the isomer level is achieved by incorporating gas-phase radical-directed fragmentation into an LC-MS/MS workflow.

Cited by 67 publications

References 48 publications

Single-cell lipidomics with high structural specificity by mass spectrometry

Single-cell lipidomics with high structural specificity by mass spectrometry

Large-scale lipid analysis with C=C location and sn-position isomer resolving power

In situ detection of fatty acid C=C positional isomers by coupling on‐tissue mCPBA epoxidation with infrared matrix‐assisted laser desorption electrospray ionization mass spectrometry

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