Expanded Coverage of Phytocompounds by Mass Spectrometry Imaging Using On-Tissue Chemical Derivatization by 4-APEBA

Probing the entirety of any species metabolome is ananalyticalgrand challenge, especially on a cellular scale. Matrix-assisted laserdesorption/ionization mass spectrometry imaging (MALDI-MSI) is a commonspatial metabolomics assay, but this technique has limited molecularcoverage for several reasons. To expand the application space of spatialmetabolomics, we developed an on-tissue chemical derivatization (OTCD)workflow using 4-APEBA for the confident identification of severaldozen elusive phytocompounds. Overall, this new OTCD method enabledthe annotation of roughly 280 metabolites, with only a 10% overlapin metabolic coverage when compared to analog negative ion mode MALDI-MSIon serial sections. We demonstrate that 4-APEBA outperforms otherderivatization agents by providing: (1) broad specificity toward carbonyls,(2) low background, and (3) introduction of bromine isotopes. Notably,the latter two attributes also facilitate more confidence in our bioinformaticsfor data processing. The workflow detailed here trailblazes a pathtoward spatial hormonomics within plant samples, enhancing the detectionof carboxylates, aldehydes, and plausibly other carbonyls. As such,several phytohormones, which have various roles within stress responsesand cellular communication, can now be spatially profiled, as demonstratedin poplar root and soybean root nodule.