A few studies in the past have shown that plant diversity in terms of species richness and functional composition can modify plant defense chemistry. However, it is not yet clear to what extent genetic differentiation of plant chemotypes or phenotypic plasticity in response to diversity-induced variation in growth conditions or a combination of both is responsible for this pattern. We collected seed families of ribwort plantain (Plantago lanceolata) from six-year old experimental grasslands of varying plant diversity (Jena Experiment). The offspring of these seed families was grown under standardized conditions with two levels of light and nutrients. The iridoid glycosides, catalpol and aucubin, and verbascoside, a caffeoyl phenylethanoid glycoside, were measured in roots and shoots. Although offspring of different seed families differed in the tissue concentrations of defensive metabolites, plant diversity in the mothers' environment did not explain the variation in the measured defensive metabolites of P. lanceolata offspring. However secondary metabolite levels in roots and shoots were strongly affected by light and nutrient availability. Highest concentrations of iridoid glycosides and verbascoside were found under high light conditions, and nutrient availability had positive effects on iridoid glycoside concentrations in plants grown under high light conditions. However, verbascoside concentrations decreased under high levels of nutrients irrespective of light. The data from our greenhouse study show that phenotypic plasticity in response to environmental variation rather than genetic differentiation in response to plant community diversity is responsible for variation in secondary metabolite concentrations of P. lanceolata in the six-year old communities of the grassland biodiversity experiment. Due to its large phenotypic plasticity P. lanceolata has the potential for a fast and efficient adjustment to varying environmental conditions in plant communities of different species richness and functional composition.