The invention provides an etimicin sulfate preparation method. The method comprises the following steps: taking gentamycin C1a base, dissolving the gentamycin C1a base in a certain proportion of a solvent, adding a certain proportion of a complexing agent to complex, adding a certain proportion of an amino group protection agent, keeping for a certain time, adding a certain proportion of a precipitating agent, stirring above materials, filtering the obtained mixture, adding a certain proportion of an acetylation reagent to the above obtained filtrate, reacting for a certain time, adding a certain proportion of a reducing agent, reacting for a certain time, removing an amino protection group, adding a certain proportion of water, carring out vacuum concentration under certain conditions to remove the solvent, adjusting the pH value with ammonia water, adsorbing with macro-porous resin, carrying out gradient separating purification with diluted ethanol, collecting an etimicin solution with a certain purity, carrying out vacuum concentration under certain conditions, adding sulfuric acid to adjust the pH value, adding a certain proportion of active carbon, decolorizing, filtering, and drying to obtain etimicin sulfate according with relevant standards.