固定化胰蛋白酶制备条件的优化及其在纯化抑肽酶中的应用

目的优化固定化胰蛋白酶的制备条件,并探讨固定化胰蛋白酶在纯化抑肽酶中的应用。方法以介孔分子筛SBA-15作为载体,戊二醛作为交联剂,对胰蛋白酶进行固定化,并对固定化条件进行优化。将优化条件制备的固定化胰蛋白酶作为亲和吸附剂,对抑肽酶进行分离纯化。结果固定化胰蛋白酶制备的最适条件为:戊二醛浓度0.8%,加酶量3.0mg,反应温度35℃,反应时间3h。以此条件制备的固定化胰蛋白酶活力可达15.6U/mg。以此酶作为亲和吸附剂纯化抑肽酶,纯化倍数可达420倍以上,活性回收率超过80%。结论优化了固定化胰蛋白酶的制备条件,以此酶作为亲和吸附剂用于抑肽酶的分离纯化,效果较好。

Optimization of Condition for Immobilization of Trypsin and Application of Immobilized Trypsin in Purification of Aprotinin

Objective To optimize the condition for immobilization of trypsin and apply the immobilized trypsin to the purification of aprotinin. Methods Immobilize trypsin using SBA-15 as carrier and glutaraldehyde as crosslinking agent and optimize the condition for immobilization. Isolate and purify aprotinin using the trypsin immobilized under the optimized condition as affinity adsorbent. Results The optimal glutraldehyde concentration, sample loading quantity, temperature and time for immobilization of trypsin were 0. 8%, 3. 0 mg, 35℃ and 3 h respectively. The activity of trypsin immobilized under the optimized condition reached 15. 6 U / mg. The purification fold of aprotinin with the immobilized trypsin was more than 420, and the recovery of activity was more than 80%. Conclusion The condition for immobilization of trypsin was optimized, and the purification effect of aprotinin with the immobilized trypsin was satisfactory.