The invention discloses a method for separating bacitracin from bacitracin fermentation liquid and purifying the bacitracin and a method for preparing zinc salt bacitracin zinc of the bacitracin. The method includes the steps that the fermentation liquid is acidized and filtered, and the filtered liquid is adsorbed and desorbed through macroporous cation exchange resin; after the desorbed liquid is desalted, decolorized, extracted and precipitated, the high-activity bacitracin is prepared. The extracted liquid is subjected to back extraction and enters an aqueous phase to be reacted with zinc salt, and the bacitracin zinc is obtained. The whole technology is simple and easy to operate, and the activity of the product is retained to the maximum degree; the technology is efficient, low in energy consumption and suitable for industrial production, the yield of the bacitracin is 60% or above, and the prepared bacitracin and the prepared zinc salt both meet the United States pharmacopeia standard.