The activity of trehalase (α, α-trehalose 1-D-glucohydrolase, ED 3. 2. 1. 28) in Sarcoma 180 ascites tumor cells and ascitic fluid which harvested at 7 days after transplantation has been studied. The activity of trehalase was measured by the method of Lapp and Mason. The results obtained were as follows; 1. The activity of trehalase could not be found in the Sarcoma 180 ascites tumor cells. 2. The activity of trehalase in ascitic fluid was 938.54 units/㎖. 3. In the ascitic fluid, the trehalase activity is relatively low compared with those in insect, microorganism, Ascaris suum and mouse kidney. 4. Trehalase from ascitic fluid has been purified about 58 fold by ammonium sulfate precipitation, DEAE-cellulose column chromatography and Sephadex G-200 gel filtration. 5. The optimal pH for enzymatic activity was 5.4, and optimal temperature was 60℃. 6. The enzyme was specific for trehalose as substrate. 7. The product of the reaction was identified as glucose by paper chromatographic method.