Activate Condition Optimization of Trypsin and Its Application in Animal Cell Culture

Abstract:

Trypsin was extracted from porcine pancreas，enzyme activity was used as the measurement index. On the basis of single factor experiment the orthogonal test was used to determine the optimization of parameters of trypsinogen activation. At the same time，the trypsin was purified using ion exchange chromatography. The effects of digestive BHK-21 and Vero cells were investigated. The results showed that the optimum conditions for activation of trypsinogen were as follows：activator concentration 0.2 mol/L，activation pH 8.5，16 ℃ for 1.5 h. Under these conditions，enzyme activity of trypsin was 2 953.68 U/mg. Trypsin was purified to activity get 3 980.42 U/mg with purification fold of 2.26 times onto a DEAE-sepharose-FF column. The cell culture results showed that BHK-21 and Vero cells were digestived 10 generation. The growth and proliferate well，every generation cell morphology was normal. The digestion time and cell proliferation concentration had no obvious difference with the products of Hyclone and Gibco. In conclusion，the process is simple and easy to be industrialized with short activate time. The study can be served as a scientific basis for the further exploitation，utilization and localization of trypsin.