The invention relates to a method for preparing aglycone from soybean isoflavone glucoside. The method comprises the following steps: galli stomachichum corium powder is mixed with 0.2 milligram per liter phosphoric acid buffer solution or acetic acid buffer solution with a pH value between 3.0 and 7.0 according to the mass ratio of 1 to 3-10; the mixture is leached for 3 to 36 hours at a temperature of between 4 and 40 DEG C and then waste slag is centrifuged; supernatant is added with ammonia sulfate or ethanol, and precipitated galli stomachichum corium zymoprotein is dissolved into the 0.2 milligram per liter phosphoric acid buffer solution or the acetic acid buffer solution with a pH value between 3.0 and 7.0, and then galli stomachichum corium enzyme solution is obtained; enzyme, the soybean isoflavone glucoside and the buffer solution are mixed and reacted for 3 to 30 hours at a temperature of between 10 and 50 DEG C and with a pH value between 3.0 and 7.0; and the soybean isoflavone aglycone is extracted. The method overcomes the defects of large destructiveness, large pollution and poor purposiveness of the acid and alkali hydrolysis method on the aglycone and simultaneously overcomes the defect that the enzyme activity of the microbial enzyme hydrolysis method is generally low. The method has easy, simple and convenient technology, complete zymohydrolysis, high conversion and high edible safety, and is suitable for industrial production.