Ambient fine particles (PM_2.5) have been shown to have adverse health effects by inducing oxidative stress. Here, dithiothreitol (DTT)-based oxidative potential (OP) was used to assess the capacity of oxidative stress caused by PM_2.5. In this study, PM_2.5 samples were collected in the Nanjing area in 2016, and physicochemical properties and DTT activity were investigated. The annual mean PM_2.5 mass concentration was 73 μg m⁻³ and greatly varied among seasons (spring > winter > summer > autumn). Three fluorescent substances were identified by the excitation-emission matrix (EEM) spectrum. The annual mean mass-normalized DTT activity (DTT_m; 0.02 nmol min⁻¹ μg⁻¹) was similar to that documented for cities of some developed countries. The annual mean volume-normalized DTT activity (DTT_v) showed a relatively high value of 1.16 nmol min⁻¹ m⁻³, and the seasonal mean DTT_v was highest in winter, followed by spring, autumn, and summer, whose pattern is different from PM_2.5 mass concentration. Correlation and multiple linear regression analysis suggested that transition metals may have a greater effect on OP in autumn and winter, humic-like substances and UV absorbing aromatic substances may have a strong effect on OP in spring and summer. Generally, this study enhances our understanding of seasonal variation in health effects associated with PM_2.5.