The widespread application of nanoparticles (NPs) in agriculture has not only enhanced the efficiency of agrochemical use but also introduced environmental pollution, potentially impacting human health through absorption and accumulation in edible plants. The purpose of this study was to evaluate the toxic effects and ecological risks of Cu2O nanoparticles (nCu2O) in the life cycle of soybean, and to provide a theoretical basis for the safe application of NPs in agriculture. Soybeans were grown in natural soil modified with nCu2O, bulk cuprous oxide (bCu2O) and copper sulfate (CuSO4) at concentrations of 0, 50, 200, and 800 mg/kg. Samples and grains from treated soybeans were collected at the flowering, podding, and seed-filling stages for analysis. The results indicated that treatments with nCu2O, bCu2O, and Cu2+ reduced the chlorophyll content in soybean leaves, thereby affecting photosynthesis. Significant reductions were observed in the net photosynthetic rate (Pn), the transpiration rate (Tr), stomatal conductance (Gs), the quantum yield of photosystem II (Y(II)), photochemical quenching (qP), and the electron transport rate (ETR) at high concentrations. However, the toxicity of nCu2O to photosynthesis recovers as the plant grows. Almost all treatments increased the levels of antioxidant enzymes (SOD, POD, CAT) and reduced oxidative stress. In the nCu2O and bCu2O treatments, grain protein content was significantly reduced, while fat and water content increased. Phosphorus (P) content decreased, whereas sulfur (S), potassium (K), magnesium (Mg) and calcium (Ca) contents increased. The accumulation of copper in plants followed the order nCu2O > bCu2O > Cu2+, with the bCu2O treatment being slightly more toxic than the nCu2O treatment, and both being more toxic than the Cu2+ treatment. The above data indicated that nCu2O had a dose-dependent effect, which significantly inhibited soybean growth and changed grain quality at high concentrations.