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目的从中性乙醇提取胰岛素后的残渣中提取制备高活性的胰酶,并从中分离纯化出胰蛋白酶(T)、糜蛋白酶(CT)。方法在胰渣中加入活化剂活化后,用PEG-6000沉淀、丙酮脱脂干燥得到胰酶;胰酶经CM-纤维素纯化得到胰糜蛋白酶混合制剂,再经CM-sepharose FF层析分离得胰蛋白酶和糜蛋白酶。结果胰酶回收率为12.1%,胰蛋白酶、胰脂肪酶、胰淀粉酶的活性分别为5.40、39.72、76.72 U·mg-1,3种酶的活性比例达1∶7.4∶14.2。胰蛋白酶、糜蛋白酶的效价分别为2505、1003 U·mg-1;相对于胰酶,二者的活性回收分别为48.93%、61.73%。结论从胰渣中制得的胰酶活性较高(约9倍于《中国药典》2010年版中的标准);制得的T及CT的活力也达到《中国药典》2010年版中的标准。
Objective To extract and purify trypsin with high activity from the residue after the extraction of insulin by neutral ethanol, and to isolate and purify trypsin (T) and chymotrypsin (CT). Methods Pancreatin was activated by activated sludge and precipitated with PEG-6000 and degreased with acetone to give trypsin. Chymotrypsin was purified by CM-cellulose to obtain a mixed preparation of chymotrypsin, which was separated by CM-sepharose FF chromatography Protease and chymotrypsin. Results The recovery of trypsin was 12.1%. The activities of trypsin, pancreatic lipase and pancreatic amylase were 5.40, 39.72 and 76.72 U · mg-1, respectively. The activity ratio of the three enzymes reached 1: 7.4: 14.2. The titer of trypsin and chymotrypsin was 2505,1003 U · mg-1, respectively. Compared with trypsin, the activity recoveries of the two proteins were 48.93% and 61.73%, respectively. Conclusion The trypsin activity obtained from pancreatic residue is relatively high (about 9 times that of the Chinese Pharmacopoeia 2010 edition). The T and CT vitality obtained also reached the standard of Chinese Pharmacopoeia 2010 edition.