Trypsin, chymotrypsin and elastase were combined to hydrolyze scorpion Buthus marstesii Karsch powders to prepare scorpion peptides (SP). Orthogonal design experiment L16 (44) was used to optimize variables of enzymolysis temperature, time, pH and enzyme concentration. Degree of hydrolysis (DH) and inhibition rate (IR) of A549 cells were selected as analysis indicators. The optimum enzymolysis conditions were determined as follows: enzymolysis temperature 40°C, time 3h, pH 8.5 and enzyme concentration of each 2000U. The anti-tumor activity of SP was determined on a panel of representative cell lines (A549, MCF7 and EC109) using MTT assay. In vitro, SP significantly inhibited the proliferation of A549, which was more obvious than MCF-7 and EC109. Further experiments on cell apoptosis and cell cycle analysis revealed that SP induce apoptosis and arrest cell cycle progression in S phase.
Keywords: Orthogonal design experiment, enzymolysis conditions, anti-tumor activity, cell apoptosis, cell cycle analysis.