OBJECTIVE： To optimize microwave-assisted aqueous two-phase system of total flavonoids from Glycyrrhiza uralensis. METHODS： The ammonium sulfate and ethanol were used as aqueous two-phase extraction system， the method of microwave was applied to extract total flavonoids from G. uralensis. The effects of 5 factors on extraction rate of total flavonoids were investigated by single factor test and orthogonal test， such as the mass concentration of ammonium sulfate， ethanol volume fraction， solid-liquid ratio， microwave power and extraction time. Validation test was conducted. RESULTS： The optimal extraction condition was as follows as the mass concentration of ammonium sulfate of 0.40 g/ml， ethanol volume fraction of 70％， solid-liquid ratio of 1 ∶ 25， extraction time of 3 min and microwave power of 300 W； in validation test， the extraction rate of total flavonoids from G. uralensis was 2.038％ （RSD＝0.69％， n＝3）， and the extraction rate of two-phase system was 94.46％ （RSD＝1.30％， n＝3）. CONCLUSIONS： The technology is simple in operation and stable in results， and also has gentle reaction conditions and high extraction rate， which is suitable for industrial production.