猪凝血酶的分离纯化和部分性质研究

STUDIES ON PURIFICATION AND PROPERTIES OF PORCINE THROMBIN

以猪血为材料，采用等电点沉淀和７２４树脂离子交换柱层析分离提纯，得凝血酶原，经ＣａＣｌ２激活后，通过ＤＥＡＥ－纤维素离子交换柱层析，获得比活为１２００ＩＵ／ｍｇ的凝血酶制剂，其收率为３２．１％。研究发现，ＮａＣｌ浓度对猪凝血酶活力有较大的影响，加１ｍｏｌ／ＬＮａＣｌ于凝血酶溶液中，室温（２５℃）放置３ｈ后，活力丧失５０％。猪凝血酶的最适温度和最适ｐＨ分别是３７℃和ｐＨ７．０。猪凝血酶在ｐＨ７．０、４０℃下放置２７ｈ，活力丧失８４％。通过ＳｅｐｈａｄｅｘＧ－１００凝胶层析，测出猪凝血酶的分子量为３０ｋｄ。

The porcine prothrombin was isolated by using isoelectric fractionation and 724 resin ion exchange column chromatography. The thrombin was obtained by DEAE cellulose ion exchange column chromatography form the prothrombin which was activated by CaCl 2. The purified thrombin had specific activity 1200 IU/mg and the recovery was 32.1%. The concentration of NaCl significantly affected the activity of porcine thrombin. In 0.6 mol/L NaCl at 25℃ for 3 h, it lost 50% activity. The optimum pH and temperature of porcine thrombin were pH 7 and 37℃ respectively. In pH7, 40℃, for 27 h, the enzyme lost 84% activity. The molecular weight of thrombin was 30 kd determined by Sephadex G 100 gel filtration.