Effect of Green Synthesized FeONP Priming on Alfalfa Seed Germination Under Drought Stress.

Drought stress is one of the key environmental factors restricting the germination of alfalfa seeds ( L.). Nanopriming is an innovative seed-priming technology able to meet economic, agronomic, and environmental needs in agriculture. However, the use of conventional nanomaterials is hampered by high costs, environmental risks, and biotoxicity. In this study, we synthesized iron oxide nanoparticles (FeONPs) using seasonal leaf extracts (collected from August to November) obtained via an enzymatic ultrasonic-assisted method. The synthesized FeONPs were characterized using SEM, EDS, DLS, FTIR, UV-Vis, and XRD. To investigate the effects of FeONP priming on alfalfa seed germination under drought stress, germination and pot experiments were conducted with five FeONP priming concentrations (unprimed, 0, 20, 40, and 60 mg/L) and three PEG-6000 concentrations (0%, 10%, and 15%) to simulate normal, moderate, and severe drought conditions. The results showed that leaf extracts collected in November exhibited the highest flavonoid content (12.8 mg/g), successfully yielding bioactive-capped spherical FeONPs with a particle size of 369.5 ± 100.6 nm. Germination experiments revealed that under severe drought stress (15% PEG-6000), the 40 mg/L FeONP treatment most effectively enhanced seed vigor, increasing the germination rate, vigor index, and α-amylase activity by 22.1%, 189.4%, and 35.5% ( < 0.05), respectively, compared to controls. Under moderate drought stress (10% PEG-6000), the 20 mg/L FeONP treatment optimally improved germination traits, increasing the germination rate by 25.5% and seedling elongation by 115.6%. The pot experiments demonstrated morphological adaptations in alfalfa seedlings: under moderate drought stress, the 40 mg/L FeONPs significantly increased lateral root numbers, while under severe drought stress, the 60 mg/L FeONPs increased the root surface area by 20.5% and preserved the roots' structural integrity compared to controls. These findings highlight that FeONPs synthesized via Ginkgo leaf extracts and enzymatic ultrasonic methods exhibit promising agricultural potential. The optimal FeONP priming concentrations enhanced seed vigor, germination traits, and drought resistance by modulating root morphology, with concentration-specific efficacy under varying drought intensities.