Influence on the growth of four different plant species and inhibition of pathogenic fungi by humic acids from soil and lignite in relation to their chemical structure

Background: Humic acids (HA) represent the major fraction of humic substances, which are supramolecular mixtures formed through humification of organic matter in soils, sediments, and natural waters. Rich in carboxylic and phenolic groups, HA influence soil fertility, nutrient availability, plant growth, and stress tolerance. Their biostimulant effects depend on origin, chemical structure, and concentration, and, although still poorly understood, their fungistatic activity against phytopathogenic fungi is increasingly recognized. This study investigated the relationship between HA chemical features and biological activity by comparing two natural HA isolated from lignite (LHA) and soil (SHA) with a synthetic HA (HALP). The assays focused on germination and early growth of lettuce, tomato, lentil, and durum wheat, as well as inhibition of six pathogenic fungi. Results: Structural characterization highlighted distinct profiles. SHA contained abundant aliphatic C, unbound fatty acids, and sugars, whereas LHA and HALP were richer in aromatic and phenolic groups. Biostimulation trials revealed species- and dose- dependent responses. Lettuce and wheat showed consistent enhancement of germination and growth, especially with LHA and SHA at up to 100 ppm, while tomato and lentil were more sensitive, exhibiting reduced growth, in particular with HALP at higher concentrations. Redundancy analysis indicated that low-dose stimulation correlated with COOH-rich aliphatic fractions, while higher concentrations emphasized phenolic and aromatic domains, which promoted biomass in tolerant species but induced phytotoxicity in sensitive ones. Fungistatic assays showed that SHA exerted the strongest inhibition, particularly against Fusarium spp. and Botrytis cinerea, outperforming LHA and HALP. Correlation analyses identified carboxyl and aliphatic carbons as positive drivers of antifungal activity, whereas phenolic and aromatic fractions correlated negatively. Conclusions: HA biological efficacy is strongly linked to chemical structure and applied dose. SHA, enriched in aliphatic and carboxyl moieties and containing unbound fatty acids, displayed the best dual functionality, combining plant growth stimulation with effective inhibition of major fungal pathogens. These results emphasize the importance of dose optimization and structural tailoring of HA for use as sustainable biostimulants and natural antifungal agents in crop production.