A method for purifying biological dust suppressant by two-stage ultrafiltration is provided, which utilizes the differences of physical characteristics such as solubility and molecular morphology of each substance in fermentation broth to separate and purify the target product by two-stage ultrafiltration. Firstly, the fermentation broth is centrifugally pretreated to separate insoluble substances such as microbial cells; after centrifugation, the supernatant passes through a primary ultrafiltration membrane, and the polysaccharide, lactic acid and aggregated biological dust suppressant molecules with molecular weights more than 20 kilodaltons are cut off; organic solvents such as methanol and ethyl acetate are added to the retentate for dilution, further filtering is carried out with ultrafiltration membrane with a cutoff molecular weight more than 10 kilodaltons to obtain liquid product, and finally preparing solid purified product is conducted through rotary evaporation and constant temperature drying.