Enzymatic Hydrolysis Of Velvet Antler Polypeptide And Its Anti-inflammatory Activity

Deer antler was used as a valuable Chinese medicine since ancient times.The main function of antler is to tonify the kidney,replenish the essence,strengthen the muscles,strengthen the bones,adjust the body,and have a good therapeutic effect on deafness and tinnitus.At present,the focus research was mainly focused on protein activity,growth factor.Hormone regulation,but the anti-inflammatory effect of antler was rarely reported.In this research,the anti-inflammatory activity of velvet antler was studied.Fresh Shuangyang sika deer antler?Cervus nippon?was got to extract the total polypeptides from velvet antler by ultrasonic extraction and CO₂ supercritical extraction.Compared the two metton of extraction rate of total polypeptide from velvet antler with different extraction processes,the extraction process of total polypeptide from velvet antler was further determined.Theresalt showed that:The extraction rate of total polypeptide from velvet antler by CO₂ supercritical extraction method is higher than that by ultrasonic extraction.The conclusions were as follows:The extraction rate of total pilose antler polypeptide by ultrasonic assisted extraction method was 2.81%,and the extraction rate of total pelt mushroom polypeptide by CO₂ supercritical extraction method was 3.93%.The total peptides of velvet antler were hydrolyzed by pepsin,alkaline protease,neutral protease and complex protease respectively?alkaline protease,neutral protease,trypsin complex?.The optimum conditions of enzymatic hydrolysis of velvet antler were determined by single factor analysis and formaldehyde titration.The optimum enzymatic hydrolysis conditions were obtained as the condition that antler polypeptide was used as hydrolysis substrate,substrate concentration was 3%,temperature was 50?,time was 3h.The anti-inflammatory model of mouse embryonic fibroblast cells?NIH3T3?was constructed,and the anti-inflammatory effect of enzymatic hydrolysis of velvet antler polypeptide was observed when 25mg of each enzymatic velvet antler polypeptide was administered:The anti-inflammatory activity of velvet antler polypeptide after enzymatic hydrolysis was better than that of velvet antler polypeptide after enzymatic hydrolysis.The anti-inflammatory activity of velvet antler polypeptide after neutral protease digestion is better than that of velvet antler polypeptide after alkaline protease digestion.The anti-inflammatory activity of velvet antler polypeptide after trypsin hydrolysis is better than the anti-inflammatory activity of antler polypeptide after alkaline protease digestion.The anti-inflammatory activity of velvet antler polypeptide after alkaline protease digestion is better than that of velvet antler polypeptide after pepsin digestion.The anti-inflammatory activity of velvet antler polypeptide after pepsin digestion was better than that of unenzymatic velvet antler total protein.By SDS-PAGE gel electrophoresis of the velvet antler polypeptide obtained of enzymatic hydrolysis,the molecular weight of the antler polypeptide protein after enzymatic hydrolysis was about 4¹5kDa.At the same time,a single protein was separated by gel chromatography,and the anti-inflammatory activity of the monomeric protein was examined by constructing a NIH3T3 anti-inflammatory cell model.Monomeric eggs with anti-inflammatory activity were sequenced,and the results of the sequencing were compared by NCBI data to analyze their protein functions.Comparing 13 single proteins with anti-inflammatory activity,and then performing NIH3T3 experiments in turn,we found that the anti-inflammatory effect is not good.Finally,the anti-inflammatory effect of pilose antler polypeptide is the result of the interaction of multiple polypeptide proteins.Single protein does not effectively inhibit inflammation.The results are accurate and reliable,which lays a foundation for the utilization of velvet antler polypeptide and the development of related antler products,and is of great significance to the further development of the deer industry.The extraction rate of effective substances in antler and the utilization rate of velvet antler provide ideas and theoretical basis,and lay a certain experimental foundation.