Sweet corn seeds are sensitive to storage due to their low starch content and poor seed vigor. Therefore, it is important to understand their physiological and biochemical behavior during storage to prolong their longevity and prevent the loss of vigor. The purpose of this study was to elucidate the physiological and biochemical changes in sweet corn seeds during storage. Artificial accelerated aging and natural aging were applied to 19 inbred lines. We found that GP, GR and VI decreased as aging proceeded. The physiological and biochemical characteristics of seeds three days after artificial aging (AA3d) and eight months after natural aging (NA8M) were evaluated. The contents of GA, ABA, and malondialdehyde and amylase activity in AA3d and NA8M seeds showed significant differences among the lines, while the total protein and total starch content showed nonsignificant or small differences. We found significant differences among the lines and a nonsignificant difference between AA3d and NA8M seeds in the activity of antioxidant enzymes. A correlation analysis showed that the germination rate was significantly negatively correlated with the ABA content in AA3d seeds, while SOD was positively correlated with GR and GI in NA8M seeds. This study provides a useful catalog of physiological and biochemical changes in sweet corn seeds, offering insights for the future genetic improvement of sweet corn’s storage tolerance.