A quali/quantitative LC–ESI–MS and LC–ESI–MS/MS approach for steviol glycosides in S. rebaudiana plants, cultured under stressed conditions (chilling stress), underlined that the different signalling molecules affect the production of secondary metabolites (steviol glycosides). The different metabolisms were evaluated by using both metabolomic targeted and untargeted approaches. In particular Stevia plants characterised by higher contents of rebaudioside A (1) and stevioside (6), were observed, giving encouraging results from plants treated with salicylic acid.