Investigating the Quantitative Structure–Ionization Efficiency Relationship of Small Molecules and Lipids in the Presence of Ammonium Fluoride in MALDI-TIMS-QTOF Mass Spectrometry Imaging

Metabolites are essential small molecules that are naturally occurring in biological processes as end or intermediate products of various pathways. Matrix-assisted laser desorption/ionization-trapped ion mobility separation-mass spectrometry imaging (MALDI-TIMS-MSI) is an emerging technique that can be used to identify the spatial localization of endogenous compounds on tissue. We evaluated the potential of ammonium fluoride (NH₄F) to enhance the ionization efficiency of metabolites in negative polarity mode when used as a comatrix additive in <i>N</i>-(1-naphthyl)ethylenediamine dihydrochloride (NEDC), 9-aminoacridine (9AA), and 1,5-diaminonaphthalene (DAN) matrices. An extensive list of 234 isotopically labeled metabolites (IROA-IS) was used to establish a quantitative ionization efficiency model with respect to the metabolite chemical structures. In addition, we extended our evaluation to endogenous compounds observed in brain samples collected from male mice. Overall, our study demonstrates that NH₄F improves the sensitivity and ionization efficiency of metabolites and lipids in MALDI-TIMS-MSI. This effect was found to vary depending on the matrix, with the ionization efficiency of the studied metabolites increasing in the order NEDC < 9AA < DAN. The quantitative structure-ionization efficiency relationship model can facilitate the appropriate selection of the matrix in MALDI prior to the analysis of analytes of interest.