Top-down Metabolomic Approaches for Nitrogen-Containing Metabolites

Streamlining
the processes that reveal heteroatom-containing metabolites
and their biosynthetic genes is essential in integrated metabolomics
studies. These metabolites are especially targeted for their potential
pharmaceutical activities. By using a Fourier-transform ion cyclotron
resonance–mass spectrometry (FTICR–MS) instrument, we
provide top-down targeted metabolomic analyses using ultrahigh-resolution
liquid chromatography–mass spectrometry (LC–MS), high-resolution
matrix-assisted laser desorption/ionization (MALDI), and high-resolution
imaging mass spectrometry (IMS) with 15N labeling of nitrogen-containing
metabolites. In this study, we efficiently extract known and unknown
chemicals and spatial information from the medicinal plant Catharanthus roseus, which sources several cancer
drugs. The ultrahigh-resolution LC–MS analysis showed that
the molecular formula of 65 N-metabolites were identified using the
petals, peduncles, leaves, petioles, stems, and roots of the non-
and 15N-labeled Catharanthus plants. The high resolution MALDI analysis showed the molecular
formula of 64 N-metabolites using the petals, leaves, and stems of
the non- and 15N-labeled Catharanthus. The chemical assignments using molecular formulas stored in databases
identified known and unknown metabolites. The comparative analyses
using the assigned metabolites revealed that most of the organ-specific
ions are derived from unknown N-metabolites. The high-resolution IMS
analysis characterized the spatial accumulation patterns of 32 N-metabolites
using the buds, leaves, stems, and roots in Catharanthus. The comparative analysis using the non- and 15N-labeled
IMS data showed the same spatial accumulation patterns of a non- and 15N-labeled metabolite in the organs, showing that top-down
analysis can be performed even in IMS analysis.