Abstract: A series of experiments were carried out to explore the effects of combined immersion freeze-drying protectants (trehalose CaCl₂₎ and ultra-low temperature freeze-thaw pretreatment on the active substances and quality of blueberries. Directly dried blueberries were taken as the control group. Three groups were selected as the ordinary freeze-thaw (-20℃) , ultra-low temperature rapid freeze-thaw (-80 ℃), and ultrasound-assisted freeze-drying protectants immersion freeze-thaw (-20 ℃) , and (-80 ℃). A systematic comparison was implemented to determine various indicators of blueberry, including the active substances (PPO and POD enzyme activity), nutrients (VC, anthocyanins, total phenols, and flavonoids), and texture characteristics (hardness and chewiness). The experimental results show that the higher quality of blueberries was protected in the combination of a single freeze-drying protectant immersion or ultra-low temperature treatment, compared with the single one. Especially, the hardness was effectively maintained for blueberries that soaked and frozen at -80℃, while the retention of nutrients was significantly improved with less duration in vacuum freeze-drying. Three-factor three-level Box Behnken design was adopted to clarify the effects of soaking time, freeze-thaw frequency and freezing time on the comprehensive indicators of blueberries. It was found that the soaking time was 3.7 h, and the single freezing time was 4.2 d when the number of freeze-thaw cycles was 2. The highest comprehensive indicators of blueberries were achieved in the retention of nutrients. Feature and sum normalizations were performed on the experimental data. All data was between (0, 1) and the sum was 1. The lipid membrane interaction of calcium pectinate and trehalose was formed by the interaction between Ca²⁺ and the cell wall. There was a great reduction in the structural damage caused by freeze-thaw. At the same time, the finer ice crystals were formed in the ultra-low temperature environments. The quality of freeze-dried blueberries was significantly improved when combined with the freeze-dried protective agents. The freeze-drying protectant impregnation pretreatment and ultra-low temperature freeze-thaw were combined to improve the quality of vacuum freeze-drying blueberries. A systematic investigation was implemented to clarify the effects of immersion time, freeze-thaw frequency, and freezing time on the quality of dried products. The optimal parameters were then determined for the vacuum freeze-drying pretreatment. In summary, the combination of immersion freeze-drying protectant (trehalose CaCl₂) and ultra-low temperature freeze-thaw pretreatment can be expected to improve the active substances and quality of blueberries. The pretreatment conditions and parameters were optimized to successfully improve the nutrient retention and quality of blueberries. The findings can provide useful and practical references for the processing of blueberry products. The application scope can be further expanded for vacuum freeze-drying in the field of food processing.