Systematic mass-spectrometry-guided metabolic fingerprinting elucidates diversity of specialized metabolites across the Brassicaceae

Abstract Background Plants produce diverse bouquets of specialized metabolites (SMs), yet only a fraction of the vast phytochemical space hasbeen explored to date.Comparative analysis of SM profiles can reveal hotspots of biochemical novelty, while systematic profilingacross taxonomic levels does presently not cover large plant families. Materials and methods To study core and accessory SM profiles in the Brassicaceae plant family, we fingerprinted 14 species by Liquid-ChromatographyMass-Spectrometry (LC-MS/MS). We develop standardized experimental and computational workflows integrating in silicoannotation tools to study consensus compound class and substructure distributions of SMs. Furthermore, we investigate thecongruence of chemotaxonomy and species phylogeny across an extended panel of 17 species. Results Unique metabolite profiles were outstanding in Camelina sativa, Capsella rubella, and Barbarea vulgaris , with the largest uniqueterpenoid profile annotated in C. sativa, accounting for 33.5% and 55.6% in positive and negative ionization mode, respectively.Substructure motifs were found to overlap with compound class predictions, highlighted for triterpenoids in Camelinodae. Furthermore, dual-tissue chemotaxonomic clustering resembled relationships of Brassica nigra , B. oleracea , B. carinata , and B. juncea , and corresponding B and C subgenomes across tissues. Conclusion We anticipate that our systematic approach can serve as a blueprint for investigating biochemical diversity in other plant lineagesand can boost the characterization of plant natural product pathways.