Background Phosphorus (P), iron (Fe) and zinc (Zn) are essential elements for plant growth and development, but their availability in soil is often limited. Intercropping contributes to increased P, Fe and Zn uptake and thereby increases yield and improves grain nutritional quality and ultimately human health. A better understanding of how intercropping leads to increased plant P, Fe and Zn availability will help to improve P-fertilizer-use efficiency and agronomic Fe and Zn biofortification.

Ammonium (NH₄⁺) is one of the most important nutrients required by plants. However, a high concentration of NH₄⁺ as the sole nitrogen source suppresses plant growth. Although nitrate (NO₃⁻) can alleviate NH₄⁺ toxicity, the mechanisms underlying this ability have not been fully elucidated. In this study, wheat plants were cultivated in hydroponic solution with 7.5 mM NO₃⁻ (control), 7.5 mM NH₄⁺ (sole ammonium, SA) or 7.5 mM NH₄⁺ plus 1.0 mM NO₃⁻ (ammonium and nitrate, AN). The results showed that compared with the control, the SA treatment significantly decreased root growth, protein content and the concentrations of most intermediates and the activity of enzymes from the tricarboxylic acid (TCA) cycle. Moreover, increased the activity of plasma membrane H⁺-ATPase and the rate of H⁺ efflux along roots, caused solution acidification, and increased the activity of mitochondrial respiratory chain complexes I–IV and the contents of protein-bound carbonyls and malondialdehyde in roots. SA treatment induced ultrastructure disruption and reduced the viability of root cells. Compared with the SA treatment, the AN treatment increased root growth, protein content, the concentrations of most intermediates and the activity of enzymes from the TCA cycle. Furthermore, AN treatment decreased the rate of H⁺ efflux, retarded medium acidification, decreased protein carbonylation and lipid peroxidation in roots and relieved ultrastructure disruption and increased the viability of root cells. Taken together, these results indicate that NO₃^–-dependent alleviation of NH₄⁺ toxicity in wheat seedlings is closely associated with physiological processes that mediate TCA cycle, relieve rhizospheric acidification and decrease the production of ROS and oxidative damage.