关键词: 鹿茸, 抗氧化性, 酶水解法, 响应曲面法, UPLC/MS/MS

Abstract: It is believed that the velvet antler head has the best health benefits in traditional Chinese medicine. But its output is small, and the price is high. The first section of velvet antler is used frequently, and its protein/peptide content is highest among all sections. The polypeptide molecular weight is about 5 kDa. In this work, in order to make full use of the velvet antler resources, the water extract of the middle and low sections collagen of velvet antler was used as an enzymatic substrate to prepare an antioxidant peptide using papain. Enzymatic hydrolysis conditions (time, enzyme addition ratio, pH and temperature) were optimized by response surface methodology (RSM). The 1,1-diphenyl-2-picrylhydrazyl (DPPH?) radical-scavenging activity rate was used as an index. The target antioxidant peptide (molecular weight less than 5 kDa) obtained by separation through an ultrafiltration membrane was further separated by reversed-phase high-performance liquid chromatography. And 10 components, S1?S10, were obtained respectively and the components were lyophilized. Component S3 was further subjected by reversed-phase ultra-high performance liquid chromatography (UPLC) on-line electrospray ionization mass spectrometry (ESI-MS) for molecular weight determination. The polypeptide with the highest antioxidant activity was fractionated by ultrafiltration membranes of molecular weight cut off of 50, 10 and 5 kDa respectively. The results showed that the optimal experimental conditions were time of 56 min, enzyme addition of 1.40wt%, pH of 5.60, temperature of 60℃, DPPH? radical-scavenging activity rate was 83.09%. The molecular weight of polypeptide was 0.2?0.6 kDa. The resulting small molecule peptide has similar health benefits as the head section, is more easily absorbed by the body, and is easier to further process and store.

Key words: Cervi cornu pantotrichum, antioxidant activity, enzymatic hydrolysis, response surface methodology, UPLC/MS/MS