摘要
探索蒙古绵羊脾蛋白的最佳酶解条件,确定最具抗氧化活性的脾肽及其分子质量。将不同饱和度硫酸铵沉淀制得的脾蛋白分别经胰蛋白酶、木瓜蛋白酶及二者混合物(质量比1∶1,下称酶混合物)水解制得脾肽,经G-25柱层析分离后,电泳确定脾肽分子质量。分别在非脂质的DPPH.和.OH体系,及脂质的卵磷脂和猪油体系中对制得的脾肽进行体外抗氧化活性试验,结果表明:60℃,pH7.0,酶混合物添加量5.0%(质量分数),5.5 h为最佳酶解条件。饱和度为60%的硫酸铵沉淀制得的脾蛋白电泳条带最多,此脾蛋白经酶混合物水解后,柱层析分离得到F1、F2、F3、F44个峰区,其中F3(分子质量6 ku)的脾肽在非脂质体系中清除.OH的作用显著(P<0.01),且随剂量的增加对非脂质体系中DPPH.的清除能力增强。在脂质的卵磷脂体系中小分子质量脾肽在浓度较低时对TBARS仍有显著抑制作用(P<0.05),但随着浓度的增大,层析后不同脾肽对其抑制作用不尽相同。在猪油体系中,小分子质量脾肽均表现出一定的抗氧化活性(P<0.05),且随着浓度的增加抗氧化活性不断增强。脾肽的抗氧化性具有广谱性。
The purpose of the study was to explore the best condition under which the Mongolian Sheep spleen polypeptides were produced through enzymatic hydrolysis and to identify its antioxidant characteristics as well as its corresponding molecular weight. Spleen protein samples, precipitated by ammonium sulfate at different unsaturated levels, were hydrolyzed with trypsin, papain, and a mixture of both （in the ratio of 1 to 1 ）, respectively. The yielded polypeptides solutions were fractionated with G-25 column chromatography and analyzed with electrophoresis to identify its molecular weight. The inhibitory effect of the polypeptides on auto-oxidation was tested in vitro with 1, 1-Diphenyl-2- picrylhydrazyl （DPPH.） and · OH in non liposome system and with phosphatidylcholine in liposome system and swine lard system. The results showed that the optimal condition for the hydrolysis developed five and half hours after the 5.0% dosage of the enzyme mixture was added at a temperature of 60 ℃ and a pH value of 7.0 （mass fraction, ration of trypsin to papain in the mixture is 1 to 1）, We also found that spleen protein precipitated with 60% unsaturated ammonium sulfate displayed more stripes of fractions than those precipitated at other levels. This spleen protein was hydrolyzed in a mixture of trypsin and papain and fractionated. Four peaks of fractions, Ft, F2, F3, and F4, were detected. Among them F3 （molecular weight of 6 ku） inhibited the development of ; OH significantly （P 〈 0.01） in the non-liposome system and DPPH. scavenging capability grew stronger as the dosage increased. In the phosphatidylcholine liposome system, the low-molecular-weight spleen polypeptides in a small dosage could inhibit the thiobarbituric acid reactive substances （TBARS） effectively （P〈 0.05）. But as the dosage increased, fractionated spleen polypeptides had different effects on inhibiting TBARS in the liposome system. In the swine lard system, the small molecular weight demonstrated antioxidative activity （P
出处
《中国农业大学学报》
CAS
CSCD
北大核心
2006年第6期60-66,共7页
Journal of China Agricultural University
基金
国家自然科学基金资助项目(30470440)
关键词
脾脏蛋白
酶水解
脾脏多肽
体外抗氧化
spleen protein
enzymatic hydrolysis
spleen polypeptides
antioxidation in vitro
