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

Probing the entirety of any species metabolome is an
analytical
grand challenge, especially on a cellular scale. Matrix-assisted laser
desorption/ionization mass spectrometry imaging (MALDI-MSI) is a common
spatial metabolomics assay, but this technique has limited molecular
coverage for several reasons. To expand the application space of spatial
metabolomics, we developed an on-tissue chemical derivatization (OTCD)
workflow using 4-APEBA for the confident identification of several
dozen elusive phytocompounds. Overall, this new OTCD method enabled
the annotation of roughly 280 metabolites, with only a 10% overlap
in metabolic coverage when compared to analog negative ion mode MALDI-MSI
on serial sections. We demonstrate that 4-APEBA outperforms other
derivatization 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 bioinformatics
for data processing. The workflow detailed here trailblazes a path
toward spatial hormonomics within plant samples, enhancing the detection
of carboxylates, aldehydes, and plausibly other carbonyls. As such,
several phytohormones, which have various roles within stress responses
and cellular communication, can now be spatially profiled, as demonstrated
in poplar root and soybean root nodule.