为了高效制备木瓜蛋白酶，利用三相分离法对番木瓜乳粉进行分离纯化。对影响分离纯化的主要因素有机溶剂种类及浓度、硫酸铵质量分数、三相成相时间及pH等进行了研究。实验结果表明，该酶的最佳分离条件为：粗酶液与叔丁醇的体积比为1∶1.5，硫酸铵质量分数为50%，成相时间为60 min，体系pH为7.0。在此条件下使用三相法分离出的木瓜蛋白酶纯化倍数为2.98倍，酶活回收率为76%。

In order to prepare papain efficiently,the three-phase partitioning technique is used to separate and purify papaya milk powder.The main factors affecting the separation and purification such as varieties of organic solvents and their concentration,concentration of ammonium sulfate,time of three-phase formation and the pH of system are investigated.The results show that the optimum separation conditions for papain are as follows:the volume ratio of crude papain solution to tert-butanol is 1.0:1.5,the mass fraction of ammonium sulfate is 50%,the phase formation time is 60 min,and the pH is 7.0.Under these conditions,the purification efficiency of papain is 2.98 times and the activity recovery is 76%.