Biostimulation of Calendula officinalis with a soy protein hydrolysate induces flower and plant biomass and flower count by reversibly altering the floral metabolome

Protein-hydrolysates represent the most studied and widely used biostimulants in agriculture, although limited information on their use in the cultivation of medicinal plants has been published so far. In this study, Calendula officinalis plants were treated in greenhouse with a soy-protein hydrolysate (SPH) at different dosages (0, 1, 2.5, 5, and 10 g/L) for 21 days. SPH was applied through different methods, namely soil drenching (SPH-S) and foliar spraying (SPH-F), and its effects on flower count and plant yield (i.e., flower and plant biomasses), and on floral metabolome were documented. SPH-S induced both flower and total plant biomasses, while SPH-F had no significant effects. The effect on plant biomass was dose -dependent, while the highest effect on flower biomass was observed with the application of the lowest (1 and 2.5 g/L) SPH-S dosages. SPH-S at 1 g/L resulted also the best treatment to induce the average flower count per plant (n = 14.30 for SPH-S 1 g/L, and n = 8.00 for control). Conversely, no significant effects were observed for SPH-F. Both SPH-S and SPH-F induced changes to the flowers ' metabolome: hexadecanoyl- (16:0) and linoleoyl- (18:2) lysophosphatidylethanolamines, known as plant growth regulators, were induced, together with dipeptides, diglycerides, and saponins, while the amount of several flavonoids was decreased. Importantly, these effects were reversible, since floral metabolome resembled that of untreated plants after the suspension of SPH application. Carotenoids were significantly induced only by 10 g/L SPH in flowers collected 7 days after the suspension of treatment. Overall, this work is the first to show the efficacy of SPH in inducing the growth and metabolism of calendula plants, indicating that these products can be efficiently used to enhance the biomass and phytochemical content of medicinal and edible plant species. Furthermore, metabolomics data contribute to partially elucidate the molecular mechanisms underlying the effects of SPH on plants. These results may be useful for researchers and practitioners to optimize the application of SPH to medicinal plants in order to increase their productivity.