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Two protein-hydrolyzate-based fertilizers (PHFs), one from alfalfa (AH) and one from meat flour (MFH), were studied chemically and biologically. AH and MFH revealed a different degree of hydrolysis and a different amino acid composition. The biostimulant activity was investigated using two specific and sensitive bioassays of auxins and gibberellins. Extracts of AH and MFH elicited a gibberellin-like activity and a weak auxin-like one. To improve our understanding of the biostimulant activity, AH and MFH were supplied to maize plants and their effect on growth and nitrate metabolism was studied. Both PHFs increased root and leaf growth and induced morphological changes in root architecture. Besides, the treatments increased nitrate reductase (NR) and glutamine synthetase (GS) activities, suggesting a positive role of the two hydrolyzates in the induction of nitrate conversion into organic nitrogen. Moreover, treatments enhanced GS1 and GS2 isoforms in maize leaves. The latter isoform, amounting to 5- to 7-fold the level of the former, appears to be a superior form in the assimilation of ammonia. The high NR and GS activities together with the high induction of GS isoforms indicate a stimulatory effect of the two PHFs on the assimilation of nitrate. In addition, a role of amino acids and small peptides of the two PHFs is suggested in the regulation of the hormone-like activity and nitrogen pathway

In recent years, the use of biostimulants in sustainable agriculture has been growing. Biostimulants can be obtained from different organic materials and include humic substances (HS), complex organic materials, beneficial chemical elements, peptides and amino acids, inorganic salts, seaweed extracts, chitin and chitosan derivatives, antitranspirants, amino acids and other N-containing substances. The application of biostimulants to plants leads to higher content of nutrients in their tissue and positive metabolic changes. For these reasons, the development of new biostimulants has become a focus of scientific interest. Among their different functions, biostimulants influence plant growth and nitrogen metabolism, especially because of their content in hormones and other signalling molecules. A significant increase in root hair length and density is often observed in plants treated with biostimulants, suggesting that these substances induce a "nutrient acquisition response" that favors nutrient uptake in plants via an increase in the absorptive surface area. Furthermore, biostimulants positively influence the activity and gene expression of enzymes functioning in the primary and secondary plant metabolism. This article reviews the current literature on two main classes of biostimulants: humic substances and protein-based biostimulants. The characteristic of these biostimulants and their effects on plants are thoroughly described.

In this study, a detailed characterization of earthworm low molecular size humic substances (LMS) was performed and these substances were used to study their effect on the nitrate influx in roots, tissue nitrate content, and expression of maize genes putatively involved in nitrate uptake in maize (Zea mays L.). The results show that the humic fraction with low molecular size used in this study is endowed with the characteristic structural network described for most humic substances so far isolated and confirm the presence of IAA in this fraction. The results also show that the LMS fraction of humic substances stimulates the uptake of nitrate by roots and the accumulation of the anion at the leaf level. Moreover, the analysis of the expression of genes encoding two putative maize nitrate transporters (ZmNrt2.1 and ZmNrt1.1) and of two maize H(+)-ATPase isoforms (Mha1 and Mha2) show that these substances may exert direct effects on gene transcription in roots, as shown for the Mha2 gene, and long-distance effects in shoots, as observed for the ZmNrt2.1 gene.

Two biostimulants, one derived from alfalfa plants (AH) and the other obtained from red grape (RG), were chemically characterized using enzyme linked immuno-sorbent assays, Fourier transform infrared (FT-IR) and Raman spectroscopies. Two doses (50 and 100 mL L−1 for RG, and 25 and 50 mL L−1 for AH) of biostimulants were applied to Capsicum chinensis L. plants cultivated in pots inside a tunnel. The experimental design consisted of the factorial combination of treatment (no biostimulant, plus AH, plus RG) at three doses (zero, low, and high) and two time-course applications (at the second and fourth week after transplantation) and the effects were recorded at flowering and maturity. Both biostimulants contained different amounts of indoleacetic acid and isopentenyladenosine; the AH spectra exhibited amino acid functional groups in the peptidic structure, while the RG spectra showed the presence of polyphenols, such as resveratrol. These results revealed that at flowering, RG and AH increased the weights of fresh leaves and fruits and the number of green fruits, whereas at maturity, the biostimulants most affected the fresh weight and number of red fruits. At flowering, the leaves of the treated plants contained high amounts of epicatechin, ascorbic acid, quercetin, and dihydrocapsaicin. At maturity, the leaves of the treated plants exhibited elevated amounts of fructose, glucose, chlorogenic, and ferulic acids. Moreover, green fruits exhibited a high content of chlorogenic acid, p-hydroxybenzoic acid, p-coumaric acid and antioxidant activity, while both AH- and RG-treated red fruits were highly endowed in capsaicin. The 1H high-resolution magic-angle spinning (HRMAS)-nuclear magnetic resonance (NMR) spectra of red fruits revealed that both products induced a high amount of NADP+, whereas RG also increased glucose, fumarate, ascorbate, thymidine and high molecular weight species. Our results suggested that AH and RG promoted plant growth and the production of secondary metabolites, such as phenols.