Combining Molecular Weight Fractionation and Metabolomics to Elucidate the Bioactivity of Vegetal Protein Hydrolysates in Tomato Plants

The comprehension of the bioactive fractions involved in the biostimulant activity of plant derived protein hydrolysates (PH) is a complex task, but it can also lead to significant improvements in the production of more effective plant biostimulants. The aim of this work is to shed light onto the bioactivity of different PH dialysis fractions (PH1 < 0.5–1 kDa; PH2 > 0.5–1 kDa; PH3 < 8–10 kDa; PH4 > 8–10 kDa) of a commercial PH-based biostimulant through a combined in vivo bioassay and metabolomics approach. A first tomato rooting bioassay investigated the auxin-like activity of PH and its fractions, each of them at three nitrogen levels (3, 30, and 300 mg L−1 of N) in comparison with a negative control (water) and a positive control (indole-3-butyric acid, IBA). Thereafter, a second experiment was carried out where metabolomics was applied to elucidate the biochemical changes imposed by the PH and its best performing fraction (both at 300 mg L−1 of N) in comparison to water and IBA. Overall, both the PH and its fractions increased the root length of tomato cuttings, compared to negative control. Moreover, the highest root length was obtained in the treatment PH1 following foliar application. Metabolomics allowed highlighting a response to PH1 that involved changes at phytohormones and secondary metabolite level. Notably, such metabolic reprogramming supported the effect on rooting of tomato cuttings, being shared with the response induced by the positive control IBA. Taken together, the outcome of in vivo assays and metabolomics indicate an auxin-like activity of the selected PH1 fraction.