The invention discloses a method for separating and preparing potato protein powder from potato juice, which comprises pretreating fresh potato juice by defoaming and crude separation, performing membrane separation to obtain trapped liquid, subjecting the trapped liquid to diafiltration desalting, sterilization and drying to obtain potato protein powder, reusing permeate liquid from membrane separation for washing precipitate obtained by pretreatment, mixing liquid obtained by washing the precipitate with fresh potato juice, and delivering to a membrane separation system. The invention well retains activity of natural potato protein by relatively mild membrane separation technology, improves the membrane separation efficiency by defoaming, improves the protein recovery rate by recovering residual protein in the solid obtained by pretreatment with the permeate liquid, and increases the protein purity by performing diafiltration on the trapped liquid. The application of intensive technology can obtain potato protein powder with certain physiological activity, which not only provides new profit growth point for industry, but also effectively reduces the pollutant concentration of industrial wastewater.