Spatial lipidomics at high chemical and spatial resolutions using mass spectrometry

Imaging mass spectrometry enables the visualization of lipid distributions directly in tissue. However, the complexity of the spatial lipidome results in the presence of many isomeric compounds that complicate spectral analysis. Each individual component in this mixture may possess a unique cellular function and spatial distribution, necessitating means to effect lipid isomer separation. Conventional chromatographic methods are often incompatible with pixel-by-pixel imaging mass spectrometry timescales. Instead, investigators have turned to tandem mass spectrometry (MS/MS) to enable isomer separation in the gas-phase. Conventional MS/MS approaches performed using low-energy collision induced dissociation (CID) of protonated or deprotonated lipid ion types result in limited structural information, necessitating alternative MS/MS approaches.

MS/MS structural information is dependent on the dissociation method and ion type. Previous reports leveraged metal-cationized lipid ion types to produce complementary fragmentation information upon CID. Herein, we build on that work by forming divalent metal-ligand-lipid ion types. We have achieved metal cationization in the gas-phase by performing charge inversion ion/ion reactions between singly charge lipid analyte ions and multiply charged metal-ligand reagent ions. These gas-phase transformations are fast, efficient, and specific, making them ideally suited for implementation into imaging mass spectrometry workflows. Our prior work has shown ion/ion reactions to enable the identification of multiple sn-positional phosphatidylcholine isomers as well as the separation of isobaric phosphatidylserines and sulfatides. Building on that success, we have used divalent metal cationization to reveal lipid isomers in biological tissues. For example, we have used magnesium tris-phenanthroline reagent dications to effect a gas-phase ion/ion reaction with fatty acid analyte monoanions. The resulting [FA-H+MgPhen2]+ metal-ligand complex produces charge remote fragmentation upon subsequent CID, enabling double bond (C=C) localization. We have also demonstrated imaging of [sulfatide-H]- beta and alpha isomers using a [Sr(Phen)3]2+ ion/ion reaction reagent. These results demonstrate a new isomeric dimension to lipid imaging via mass spectrometry.