花生清蛋白的分离纯化及抗氧化、DNA酶活性研究

为了解花生清蛋白的生物学活性,本试验采用硫酸铵沉淀法初步分离花生种子中的花生清蛋白,并用DEAE-52柱纯化得到花生清蛋白,分析其体外抗氧化性及DNA酶活性。结果表明,65%硫酸铵分离花生清蛋白效果最好;纯化后的花生清蛋白由3个亚基组成,其相对分子质量分别为14.5、15.5、17.2 kDa。花生清蛋白体外抗氧化活性较高,对DPPH自由基和羟基自由基的清除率与花生清蛋白浓度呈正相关,清蛋白浓度从0.02 mg·mL^-1升至0.10 mg·mL^-1时,DPPH自由基清除率从26.3%增加到45.0%,羟基自由基清除率从10.8%增加到93.5%。DNA酶活性也与花生清蛋白浓度呈正相关,清蛋白浓度从0.1 mg·mL^-1提高到0.6 mg·mL^-1,清蛋白和质粒混合液的电泳条带逐渐变暗;当花生清蛋白浓度为0.8 mg·mL^-1时,电泳条带消失,质粒被完全降解;Ca²⁺、Mg²⁺、K⁺、Na⁺4种金属离子都有增强清蛋白DNA酶活性的作用,增强能力由大到小依次是Mg²⁺>Ca²⁺>Na⁺>K⁺。本研究结果为花生清蛋白功能性质研究及开发利用提供了一定的理论基础。

Study on Isolation,Purification and Antioxidant and DNase Activities of Arachis hypogaea Albumin

In order to understand the biological activity of peanut albumin, the storage protein in peanut seeds was preliminarily separated by ammonium sulfate precipitation method, and the peanut albumin was purified by DEAE-52 column. In addition, the antioxidant and DNase activities of peanut albumin in vitro were analyzed. The results showed that 65% ammonium sulfate had the best separation effect. The purified peanut albumin was composed of three subunits with relative molecular masses of 14.5, 15.5, and 17.2 kDa, respectively. Peanut albumin had higher antioxidant activity in vitro, and the scavenging rate of DPPH and hydroxyl radicals was positively correlated with the concentration of peanut albumin. As the albumin concentration increased from 0.02 mg·mL^-1 to 0.10 mg·mL^-1, the DPPH free radical scavenging rate increased from 26.3% to 45.0%, and the hydroxyl radical scavenging rate increased from 10.8% to 93.5%. The DNase activity was also positively correlated with the concentration of peanut albumin. As the albumin concentration increased from 0.1 mg·mL^-1 to 0.6 mg·mL^-1, the electrophoretic bands of albumin and plasmid mixture gradually darkened. Peanut albumin at the concentration of 0.8 mg·mL^-1 caused the electrophoresis band of mixture to disappear, and the plasmid was completely degraded. All of the four metal ions of Ca²⁺, Mg²⁺, K⁺ and Na⁺ enhanced the activity of albumin DNase, and the descending order of enhanced ability was Mg²⁺>Ca²⁺>Na⁺>K⁺. This study provides a theoretical basis for the function research and application of peanut albumin.