Metabolites define the energy content, nutritional value, and the taste of food and feed. Metabolites not only define quality but are likewise essential for plant growth and health: they give rise to biomass, yield, and define interactions of plants with beneficial partners and pathogens. Soil metabolites define its ecosystem function, impact plant growth, and presence of macro- and microorganisms. Here, a pipeline to investigate plant metabolites is established to systematically and reliably identify a large variety of plant and soil metabolites, allowing to identify differences between cultivars and environments, to characterizes biodiversity, and to detect shifts in metabolite profiles when employing novel breeding technologies. We develop protocols from sample preparation to metabolomic analysis, and we generate a database comprising the ‘chemical space’ of crops and soils.