Trehalose (TRE) was used to improve the gastrointestinal tolerance of Lactobacillus plantarum embedded with whey protein concentrate/pullulan (WPC/PUL) hydrogel and the embedded L. plantarum was applied to juice. The study indicated that 5% TRE significantly increased the viable counts of L. plantarum embedded in WPC/PUL hydrogel from (8.83 ± 0.03) to (9.14 ± 0.04) (lg (CFU/g)) in simulated gastric juice (SGJ) and from (9.13 ± 0.04) to (9.38 ± 0.04) (lg (CFU/g)) in simulated intestinal juice, respectively. The addition of TRE improved the glass transition temperature of WPC/PUL hydrogel and decreased the hardness and its solubility in SGJ, which may be responsible for the improved protection of WPC/PUL hydrogels on L. plantarum. In addition, TRE increased the viable counts of L. plantarum in WPC/PUL probiotic microcapsule juice at low pH and high temperature during storage.