Abstract: Paeonia qiui is one of the important ancestors of cultivated species of tree peony, serving as an important genetic resource in peony cross breeding, due to its purple-red leaves. The pollen is widely known to directly influence reproductive success and genetic structure of the main plant population. This study aims to investigate the storage characteristics of Paeonia qiui pollen, further to elucidate the physiological mechanism of pollen storage at different temperatures, in order to determine the accurate and efficient storage method for the viability of Paeonia qiui pollen. In this study, the pollen morphology of Paeonia qiui was characterized using scanning electron microscopes. Four factors (sucrose, boric acid, Ca (NO3)2, GA3) at three different levels were selected in the orthogonal test to investigate the characteristics of Paeonia qiui in vitro germination. The experiment was conducted to explore the effect of storage temperatures and duration on the pollen germination rate, malondialdehyde (MDA), and the activity of superoxide dismutase (SOD), peroxidase(POD), catalase (CAT). The results show that the rates of abnormal and shrunken pollen were as high as 19.5%, indicating the low viability of Paeonia qiui pollen. The important factors affecting the germination rate of Paeonia qiui pollen were listed in order, sucrose, boric acid, Ca (NO3) 2, GA3. The optimum concentrations of media were achieved in vitro pollen germination, when containing 110 g/L sucrose, 45 mg/L boric acid, 55 mg/L GA3, 30 mg/L Ca(NO3)2. The optimum temperatures were 2℃ and -20℃ for the pollen storage of 1-2 months and 4-6 months, respectively. The optimum condition was -80℃ for the intermediate storage within one year, whereas the promising temperature was -196℃ for the long-term preservation of Paeonia qiui pollen. The germination rate was 75.5% after 365 days of storage, indicating that the optimum condition can effectively prolong pollen longevity. The germination rate of pollen was positively correlated with three protective enzymes, while negatively correlated with malondialdehyde content. The influence of three protective enzymes on pollen germination rate was ranked in order: superoxide dismutase (SOD) , catalase (CAT), peroxidase (POD). The sensitivity of three protective enzymes was different at different storage temperatures. Specifically, SOD served as a sensitive protective enzyme at 2, -20℃ and -80℃, whereas POD served as sensitive protective enzymes at room temperature. The protective enzyme activity decreased, while the malondialdehyde content increased, leading to the active oxygen can not be effectively removed. This process was one of the main reasons for the decrease in pollen viability after storage at room temperature, 2, -20 and -80℃. The malondialdehyde content and the activity of three protective enzymes in pollen remained basically stable. The three protective enzymes can effectively maintain the internal balance of oxidative metabolism, and thereby reduce the levels of oxidative damage. This can be the internal cause to keep high germination rate after cryopreservation. This finding can provide a promising experimental and theoretical basis for cross-temporal pollination hybridization and germplasm resource conservation.