Objective To explore the feasibility of cultivating Schizochytrium sp. using waste beer yeast hydrolysate to produce DHA algal oil.Methods Ultrasound-assisted composite enzyme method was selected to break the cell wall and hydrolyze waste beer yeast, optimizing the process based on reducing sugar and total nitrogen concentrations as response variables. The hydrolysate was then used to cultivate Schizochytrium sp. to produce DHA algal oil.Results Under the conditions of ultrasonic power at 400 W, temperature at 60 ℃, pH 5.0, papain at 0.5 g, β-glucanase:papain ratio of 4:5, and hydrolysis time of 40 min, the hydrolysate achieved maximum concentrations of reducing sugar and total nitrogen, which were 20.40 g/L and 6.72 g/L, respectively. When the hydrolysate volume fraction was 75% and the inoculum volume fraction of Schizochytrium sp. was 14%, after 3 days of cultivation, the biomass, oil yield, DHA content, and DHA yield reached their maximum values of 17.14 g/L, 2.88 g/L, 3.13%, and 0.55 g/L, respectively, which were higher than those obtained from traditional fermentation cultures (biomass 10.29 g/L, oil yield 1.84 g/L, DHA content 2.76%, DHA yield 0.29 g/L). The pathogenic bacteria and heavy metal levels in the algal oil were below the minimum limits specified in national food safety standard.Conclusion Using waste beer yeast hydrolysate obtained through ultrasound-assisted composite enzyme hydrolysis to cultivate Schizochytrium sp. for DHA production is feasible. This method achieves an organic integration of food production waste treatment and microalgae-based chemical product production.