Mass spectrometry-based lipidomics is the primary tool for the structural analysis of lipids but the effective localization of carbon–carbon double bonds (C=C) in unsaturated lipids to distinguish C=C location isomers remains challenging. Here, we develop a large-scale lipid analysis platform by coupling online C=C derivatization through the Paternò-Büchi reaction with liquid chromatography-tandem mass spectrometry. This provides rich information on lipid C=C location isomers, revealing C=C locations for more than 200 unsaturated glycerophospholipids in bovine liver among which we identify 55 groups of C=C location isomers. By analyzing tissue samples of patients with breast cancer and type 2 diabetes plasma samples, we find that the ratios of C=C isomers are much less affected by interpersonal variations than their individual abundances, suggesting that isomer ratios may be used for the discovery of lipid biomarkers.

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 andsn-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) andsn-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 andsn-position isomers.

Mass spectrometry has become a powerful tool in the field of biomedicine. The combination of ambient ionization and miniature mass spectrometry systems could most likely fulfill a significant need in medical diagnostics, providing highly specific molecular information in real time for clinical and even point-of-care analysis. In this review, we discuss the recent development of ambient ionization and miniature mass spectrometers as well as their potential in disease diagnosis and therapeutic monitoring, with an emphasis on their capability in analysis of biofluids and tissues. We also speculate the future development of the integrated, miniature MS systems and provide our perspectives on the challenges in technical development as well as possible solutions for path forward.

As a major class of mammalian lipids, phosphatidylcholines (PCs) often contain mixtures of structural isomers, resulting from different lipogenesis pathways. Profiling PCs at the isomer level, however, remains challenging in lipidomic settings, especially for characterizing the positions of fatty acyls on the glycerol backbone (sn-positions) and the locations of carbon–carbon double bonds (CCs) in unsaturated acyl chains. In this work, we have developed a workflow for profiling PCs down to sn- and CC locations at high coverage and sensitivity. This capability is enabled by radical-directed fragmentation, forming sn-1 specific fragment ions upon collision-induced dissociation (CID) of bicarbonate anion adducts of PCs ([M + HCO₃]⁻) inside a mass spectrometer. This new tandem mass spectrometry (MS/MS) method can be simply incorporated into liquid chromatography by employing ammonium bicarbonate in the mobile phase without any instrument modification needed. It is also compatible with the online Paternò–Büchì reaction and subsequent MS/MS for the assignment of CC locations in sn-1 fatty acyl chains of unsaturated PCs. The analytical performance of the workflow is manifested by identification of 82 distinct PC molecular species from the polar extract of bovine liver, including quantification of 19 pairs of sn-isomers. Finally, we demonstrate that five pairs of PC sn-isomers show significant compositional changes in tissue samples of human breast cancer relative to controls, suggesting a potential for monitoring PC sn-isomers for biomedical applications.

Direct sampling mass spectrometry enables high-performance clinical analysis, such as imaging, drug monitoring and point-of-care testing.