Combined effect of humic substances and nitrogen fertilizers on gene expression levels and enhanced phytoremediation of cadmium contaminated soil by Solanum nigrum L

Purpose Cadmium contamination in agricultural soils poses significant risks to food safety and ecosystem health. This study investigated the synergistic effects of humic substances (fulvic acid [FA] or humic acid [HA]) and nitrogen (N) fertilisers on enhancing cadmium (Cd) phytoextraction by Solanum nigrum L. ( S. nigrum ) in moderately contaminated soils. Methods A pot experiment tested ten treatments combining FA/HA with ammonium sulphate ((NH 4 ) 2 SO 4 ), urea (CH 4 N 2 O), ammonium chloride (NH 4 Cl), or ammonium bicarbonate (NH 4 HCO 3 ), measuring plant biomass, Cd uptake, physiological responses, the expression levels of three antioxidant enzyme genes and soil parameters. Results Co-application of FA with (NH 4 ) 2 SO 4 yielded the highest shoot biomass and Cd extraction relative to the control. Biomass augmentation drove these improvements, as Cd concentrations in shoots remained statistically unchanged across treatments. Physiological analysis revealed reduced oxidative stress (H 2 O 2 and MDA levels), elevated antioxidant enzyme activities (SOD and POD) in shoots under combined treatments, meanwhile, the relative expression levels of SnCAT , SnPOD and SnSOD were clearly changed. Soil pH and bioavailable Cd concentrations remained stable, minimising risks of Cd leaching or mobilisation. The results demonstrate that FA or HA combined with ammonium-based fertilisers significantly accelerates Cd phytoextraction by circumventing the “dilution through growth” effect observed in other species, maintaining high Cd uptake efficiency. Conclusions The FA + (NH 4 ) 2 SO 4 treatment emerged as the optimal strategy, offering a practical solution for remediating moderately Cd-contaminated soils within a decade. These findings provide a scalable framework for integrating S. nigrum -based phytoremediation into agricultural systems, particularly where soil buffering capacity limits amendment-induced pH fluctuations.