Calcium regulation of nitrogen fixation and metabolism in soybean (Glycine max L.)

Calcium (Ca) is pivotal for symbiotic nitrogen fixation (SNF) and the overall growth and development of legumes. However, the mechanisms by which varying Ca supply levels coordinate the homeostasis between nodular N fixation and nitrogen (N) metabolism remain poorly understood. This study conducted a two-year sand culture experiment to investigate the effects of low (Ca0.1), optimal (Ca1), and high (Ca10) calcium regimes on soybean (Glycine max L.). Optimal Ca supply (1 mmol·L−1, Ca1) significantly promoted plant growth and increased grain yield by 54.5–91.0% in 2024 and 55.3–74.7% in 2025 compared to other regimes. It maximized nitrogenase activity (SNA and ARA)—with ARA being 1.63–2.51 fold higher than in stressed plants in 2025—and significantly upregulated N-metabolism enzymes (GS, GOGAT, GLS, and L-ASNase) in both leaves and nodules. Furthermore, amino acid profiling substantiated that optimal Ca improved N utilization efficiency by preventing the atypical buildup of free amino acids (FAAs) observed under Ca-stress conditions. In summary, an optimal Ca supply maximizes soybean productivity by synergistically enhancing biological nitrogen fixation capacity and N-use efficiency. This coordination maintains carbon–nitrogen (C/N) homeostasis and improves key yield components, providing a theoretical framework for refined Ca fertilization management in soybean production.