Abstract:

BACKGROUND: The recombinant antihypertensive peptide, Val-Lys-Pro-Val-Pro-Arg (VLPVPR), was prepared using genetic engineering technology and enzymolysis method. The enzymolysis products are polypeptide compound, which is prone to be infested and deteriorated by microorganism.
OBJECTIVE: To analyze enzymolysis technology and degradation causes of the recombinant antihypertensive peptide, VLPVPR, during trypsin-catalyzed hydrolysis process.
METHODS: The content of antihypertensive peptide, VLPVPR, was detected by reversed phase high-performance liquid chromatography. The engineering germ products were sterilized by 0.22 μm membrane filter, and the content information of VLPVPR in the enzymolysis process was observed. Compared with non-filtered sterilization enzymatic process, the degradation of endoenzymes released from the engineered strain cells and bacterial contamination on VLPVPR was approached. The enzymolysis technology conditions, such as temperature, pH, final concentration of trypsin, and duration, were optimized by orthogonal experiment.
RESULTS AND CONCLUSION: The content of VLPVPR standard remained unchanged after 4 hours’ action with trypsin, which showed no significant difference (P > 0.05), demonstrating that VLPVPR could not be degraded by trypsin. The VLPVPR of the cell disruption which had been sterilized was degraded rapidly after reaching the maximum value, and the VLPVPR of the sample without sterilized was degraded faster, which showed significant difference (P < 0.05). Results showed the endoenzymes released from the engineered strain cells and bacterial contamination is the main reasons for VLPVPR degradation during preparation process. The best hydrolyzing condition of trypsin was optimized by orthogonal experiment: 30 ℃, pH 9.5, final concentration of trypsin at 180 U/mL, and reaction time for 1 hour.