<p>Light conditions inside a greenhouse vary greatly in different seasons, and there is no optimal fertilization strategy for cultivating high-quality pepper seedlings under different light conditions. In this study, the effects of different combinations of daily light integral (DLI, 7.2, 14.4, and 21.6 mol m⁻² d⁻¹) and nitrogen concentration (7.5, 15, and 22.5 mmol L⁻¹) on the morphological characteristics, chlorophyll content, antioxidant enzyme activity, and chlorophyll fluorescence of pepper seedlings were evaluated. The results show that the shoot fresh weight, seedling quality index, chlorophyll a content, and root volume of pepper seedlings exposed to DLI at 14.4 mol m⁻² d⁻¹ combined with nitrogen at a concentration of 15 mmol L⁻¹ were the highest and were increased by 15.4%, 18.8%, 18.9%, and 28.6% compared with those under the combination of DLI at 14.4 mol m⁻² d⁻¹ and nitrogen at 7.5 mmol L⁻¹. Under a DLI of 21.6 mol m⁻² d⁻¹ combined with a nitrogen concentration of 7.5 mmol L⁻¹, the leaf area, soluble protein content, and photochemical quenching of pepper seedlings were increased by 16.4%, 15.5%, and 35.7% compared with those of 21.6 mol m⁻² d⁻¹ DLI combined with 15 mmol L⁻¹ nitrogen concentration, respectively. In conclusion, the seedling quality can be altered by adjusting the nitrogen concentration under different DLI conditions. A combination of DLI at 14.4 mol m⁻² d⁻¹ and nitrogen concentration at 15 mmol L⁻¹ can improve the seedling quality of pepper seedlings. Our study has provided valuable insights related to the annual production of high-quality pepper seedlings via regulating daily light integral and nitrogen supply.</p>