Abstract: We explored the mechanism by which exogenous spermidine (Spd) mitigates copper oxide nanoparticle (CuO-NPs) stress in rice seedlings with a series of seed germination and hydroponic experiments. We assessed the effects of different Spd concentrations (0, 500, and 1000 μmol·L^-1) on germination traits, seedling growth performance, antioxidant defense system activity, osmoregulatory substance accumulation, copper ion sequestration, and the integrity of subcellular structures under varying levels of CuO-NPs stress (0, 50, 100, and 300 mg·L^-1). The results showed that CuO-NPs stress markedly suppressed germination potential, germination rate, and seedling biomass accumulation; caused excessive accumulation of reactive oxygen species (ROS); aggravated membrane lipid peroxidation with a notable increase in malondialdehyde (MDA) levels; and impaired the functional homeostasis of the antioxidant enzyme system. In contrast, exogenous Spd treatment effectively alleviated the adverse impacts of CuO-NPs stress. Under 300 mg·L^-1 CuO-NPs stress, the addition of 1000 μmol·L^-1 of Spd resulted in notable improvements. Seed germination rate, germination index, and vigor index were increased by 7.06%, 3.56%, and 27.94%, respectively. The activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were enhanced by 40.12%, 16.44%, and 20.08%, respectively. The contents of superoxide anion (O₂^-_·), hydrogen peroxide (H₂O₂), and MDA were reduced by 46.43%, 25.46%, and 32.85%, respectively. Furthermore, Spd treatment enhanced copper fixation in shoot cell walls and alleviated copper toxicity. Fourier transform infrared spectroscopy (FT-IR) analysis revealed that exogenous Spd strengthened the physical barrier function of cell wall by modulating the structural configuration and relative proportion of polysaccharides, proteins, and lipid functional groups. This regulatory effect, in turn, alleviated the destructive damage to subcellular structures induced by CuO-NPs. Collectively, those results confirm that exogenous Spd can effectively enhance the tolerance of rice seedlings to CuO-NPs stress through multiple synergistic pathways, including boosting antioxidant capacity, regulating osmotic balance, and reinforcing cell wall barrier function. These findings provide a theoretical basis for deciphering the physiological mechanisms by which polyamines alleviate the toxicity of nano-copper oxide, and thus lay a scientific foundation for the development of agronomic management strategies to mitigate the adverse effects of heavy metal nanoparticle stress in crops.

Key words: nano-copper oxide, spermidine, rice, antioxidant system, Fourier transform infrared spectroscopy (FT-IR), cell wall functional group