Nickel Supplementation for Enhanced Soybean Growth: A Micrometric-to-Nanometric Investigation of Biological Nitrogen Fixation and Metabolism

High Resolution Image Download MS PowerPoint Slide Nickel (Ni) plays a central role in plant nitrogen (N) metabolism, yet the long-term physiological impact of nanostructured Ni fertilizers on soybean ( Glycine max L.) remains insufficiently characterized. This study assessed the effects of Ni seed treatment and foliar application, using macro-, micro-, and nanosized Ni sources, on soybean cv. “IPRO Flecha” N metabolism and biological nitrogen fixation (BNF) throughout the crop life cycle. Ni was applied as seed coating (45 mg Ni kg –1 seed) and/or foliar spray (20 g Ni ha –1 at V4) using NiSO 4 ·6H 2 O (macro), micrometric Ni(OH) 2 (∼24 μm), or nanometric Ni(OH) 2 (∼5 nm). Key physiological, enzymatic, and isotopic parameters were measured at V6, R2, and R7 stages, including nitrogenase, urease, and nitrate reductase activities, BNF contribution (δ 15 N), and N metabolites (N-NH 3, N-NO 3 –, N-ureides). At R2, nano-Ni seed + foliar increased leaf dry weight by 28% compared with seed-only and 49% compared with the control, whereas at R5 the same treatment reduced leaf dry weight by 62% relative to nano-Ni seed-only, indicating a strong stage-dependent response. At maturity (R7), Ni application increased seed yield across sources, with mean yield rising from 12 g pot –1 in the control to 19–22 g pot –1, with the highest yield under nano-Ni (22 g pot –1 ). Ni supply increased urease and nitrate reductase activities relative to the control; however, nano-Ni reduced nitrate reductase at V6 (≈41–43% lower than macro- and micro-Ni), and nano-Ni seed-only reduced nitrogenase activity at R2 by ∼33% compared with the control. BNF in shoots ranged from 60.5–87.4% at R2 and was generally higher under Ni treatments at R7 (≈85–92%), except for nano-Ni seed + foliar (67.3%). Overall, Ni improved soybean N metabolism and yield, but nano-Ni effects were highly dependent on the growth stage and application method, highlighting the need for mechanistic studies on nanoparticle behavior over time to support consistent agronomic outcomes.