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