蝉蜕蛋白的提取工艺优化及体外抗氧化活性研究

Optimization of Extraction Technology of Protein from Cryptotympana pustulata and in vitro Antioxidant Activity Study

目的:优化蝉蜕蛋白的提取工艺,并考察其体外抗氧化活性,为蝉蜕蛋白的深入研究提供参考。方法:以蛋白提取量为响应值,在单因素试验基础上,采用Box-Behnken响应面法对超声波辅助提取蝉蜕蛋白的液料比、超声时间、提取次数等条件进行优化,并进行验证试验。以维生素C(VC)为阳性对照,以对1,1-二苯基-2-三硝基苯肼(DPPH)和2,2′-连氮基-双-(3-乙基苯并二氢噻唑啉-6-磺酸)(ABTS)自由基的清除率为指标,评价蝉蜕蛋白的体外抗氧化活性。结果:蝉蜕蛋白的最佳提取条件为液料比28∶1(mL/g),超声时间65 min,提取2次。验证试验中,蝉蜕蛋白的平均提取量为65.45 mg/g(RSD=1.68%,n=3),与预测值的相对误差为5.48%。蝉蜕蛋白对ABTS和DPPH自由基均有一定的清除效果,当蝉蜕蛋白质量浓度为0.2 mg/ L时对ABTS自由基的清除率即达97.0%,其作用与VC相当;蝉蜕蛋白对DPPH自由基的清除能力稍弱,半数清除浓度为0.96 mg/mL,其作用不及VC。结论:采用Box-Behnken响应面法优化蝉蜕蛋白的提取条件准确、可靠;蝉蜕蛋白具有一定的体外抗氧化活性。

OBJECTIVE:To optimize extraction technology of protein from Cryptotympana pustulata and investigate its in vitro antioxidant activity,so as to provide reference for further research of protein from C.pustulata.METHODS:Using extraction amount of protein as response value,based on single factor test,Box-Behnken response surface methodology was used to optimize the ratio of liquid to material,ultraonic time and extraction times.Validation test was conducted.Using Vitamin C(VC)as positive control,in vitro antioxidant activity of protein from C.pustulata was evaluated by using scavenging rate of 1,1-diphenyl-2-picrylhydrazyl(DPPH)and 2,2′-azino-bis(3-ethyl benzothiazoline-6-sulfonic acid)(ABTS)free radical as index.RESULTS:The optimal extraction condition of protein from C.pustulata was as follows as the ratio of liquid to material 28∶1(mL/g),ultrasonic time of 65 min,extracting for twice.In validation test,average extraction amount of protein from C.pustulata was 65.45 mg/g(RSD=1.68%,n=3),relative error of which to predicted value was 5.48%.The protein from C.pustulata showed strong scavenging effect on ABTS and DPPH free radicals.When the concentration of protein from C.slough was 0.2 mg/mL,and the scavenging rate of it to ABTS free radicals was 97%,the effect of which was similar to VC.The protein from C.pustulata showed weak scavenging ability to DPPH free radical,IC50 was 0.96 mg/mL,the effect of which was not as good as VC.CONCLUSIONS:The extraction technology of protein from C.pustulata optimized by Box-Behnken response surface methodology shows high accuracy and good reliability.The protein from C.pustulata shows certain antioxidant activity in vitro.