Integrating Ion Mobility Spectrometry and Mass Spectrometry Imaging for Characterizing the Distribution of Biologically Active Disaccharide Isomers

The ability to differentiate disaccharide isomers within plant tissues, in a spatially resolved fashion, is required to render a more precise understanding of their biological function within these systems. In this study, we report drift tube ion mobility spectrometry (DTIMS) and trapped ion mobility spectrometry (TIMS) profiling of two important plant disaccharides, sucrose and trehalose, along with their 6-phosphate derivatives. We then map these disaccharides in poplar root and soybean root nodules using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) with premass analysis TIMS. This approach bridged an analytical gap for the straightforward visualization and discrimination of disaccharides within plant tissues at a spatial resolutions approaching a single plant cell size. We found direct infusion DTIMS and TIMS MS worked congruently to obtain the collision cross section (CCS) values of disaccharides, where we measured ΔCCS < 1.5% between DTCCSN2 and TIMSCCSN2 across all ions compared. This approach provided a baseline comparison for subsequent MSI measurements. We observed the disaccharide ion at m/z 381.08 ([M + K]+) displayed a single CCS value in poplar roots, whereas two different CCS values were measurable in soybean root nodules, which exemplified coexisting disaccharide isomers in this plant root tissue. We found the high-resolution ion mobility fingerprinting and ion imaging for the disaccharide isomer 6-phosphate derivatives in plant tissues were more challenging compared to those for the nonphosphorylated disaccharide species. This may be in part due to their lower abundance and MALDI-induced in-source fragmentation. Empowered by the integration of ion mobility spectrometry and MSI, this study provides a new avenue for easier characterization and direct visualization of disaccharide isomers within plant tissue.