Objective: A peptide with antioxidant activity was screened from whey protein hydrolysate and used to protect Probio-M8 during the freeze-drying process. The whey proteins were hydrolyzed by alkaline protease and separated by ultrafiltration and three different molecular weight peptide samples were obtained， k1 (>8 ku)， k2 (3-8 ku)， and k3 (<3 ku). The peptides were then purified and identified by strong cation exchange chromatography， reversed-phase high-performance liquid chromatography (RP-HPLC)， and liquid chromatography-mass spectrometry (LC-MS). 1，1-Diphenyl-2-trinitrophenylhydrazine (DPPH) scavenging， resistance to superoxide anion radicals， and absorption of oxidative radicals were used as assays to assess antioxidant activity and were screened. Results: Samples 3-2 and 4-6 showed high antioxidant activity. Five and six antioxidant peptide sequences were identified for samples 3-2 and 4-6， respectively. Sample 3-2 was able to protect B. lactis Probio-M8 in vacuum freeze-drying， increasing the survival rate of the strain to (88.31±0.02)% and also reducing intracellular reactive oxygen species levels and protecting cellular enzyme activity. Conclusion: Sample 3-2 was effective in reducing cell damage caused by the vacuum freeze-drying process of B. lactis Probio-M8.