Multifunctional Reactive MALDI Matrix Enabling High-Lateral Resolution Dual Polarity MSI and Lipid C=C Position-Resolved MS2I

Local lipid variations in tissues are readily revealed with mass spectrometry imaging (MSI) methods and resulting lipid distributions serve as bioanalytical signatures to reveal cell- or tissue-specific lipids. Comprehensive MSI lipid mapping requires measurements in both ion polarities. Additionally, structural lipid characterization is necessary to link lipid structure to lipid function. Whereas some structural elements of lipids are readily derived from high-resolution mass spectrometry (MS) and tandem-MS (MSn), the localization of C=C double bonds (DBs) requires specialized fragmentation and/or functionalization methods. In this work, we identify a multifunctional matrix-assisted laser desorption/ionization (MALDI) matrix for spatially-resolved lipidomics investigations that reacts with lipids in Paternò-Büchi (PB) reactions during laser irradiation facilitating DB position assignment and allows dual polarity high-resolution MALDI-MSI and MALDI MS2I studies. By screening twelve compounds for improved ionization efficiency in positive/negative ion mode and PB functionalization yield compared to the previously introduced reactive MALDI matrix benzophenone, benzoylpyridine (BzPy) is identified as the best candidate. The multifunctional character of the new matrix enables DB localization of authentic standards belonging to twelve lipid classes and helps to assign 506/365 lipid features in positive/negative ion mode from mouse cerebellum tissue. The analytical capabilities of BzBy as a multifunctional MALDI-MSI matrix are demonstrated by imaging endogenous and PB-functionalized lipids in mouse kidney sections with 7 µm lateral resolution in both ion modes. Tracking diagnostic lipid DB position fragment ions in mouse pancreas tissue with down to 10 µm pixel size allows to identify islets of Langerhans associated lipid isomer upregulation or depletion.