Consumption of leafy vegetables is highly recommended for meeting dietary requirements for antioxidants, vitamins, and minerals. However, the short shelf life of leaves often compromises the nutritional contents and causes economic loss for farmers and sellers. Chitosan, a natural polymer obtained from seafood waste, has been proposed to improve plant qualities in the postharvest condition. In this study, the quality of spinach leaves following chitosan treatments was analyzed to test the capacity of chitosan to improve postharvest management of spinach. Specifically, harvested spinach leaves were dipped in distilled water, 1% (v/v) acetic acid, and 0.1 and 0.5% (w/v) chitosan. The leaves were then examined for morphological, physiological, and molecular parameters following each treatment on days 0 and 3 after incubation at room temperature with a 12/12-h photoperiod. Our results suggested that the application of 0.1% (w/v) chitosan solution to postharvest spinach delayed the decaying process, possibly due to the suppression of bacterial growth. The elicitation mechanism by chitosan involved proline accumulation, retaining chlorophyll, increasing expression of antioxidant genes (Cu/Zn-SOD and CAT), and decreasing ROS (O2− and H2O2) content. In conclusion, our results support the suggested role of chitosan treatment in maintaining quality of postharvest spinach by regulating the elicitation processes and limiting bacterial growth.