AISPPs对木瓜蛋白酶的抗冻保护作用研究

Study on anti-freezing protection of papain by AISPPs

以苜蓿草粉为原材料,利用冰结合磷酸缓冲液提取法提取苜蓿冰结构蛋白(Alfalfa Ice Structuring Proteins,AISPs),并用碱性蛋白酶对其进行酶解.通过单因素及响应面法对碱性蛋白酶对AISPs的水解条件优化,优化结果为,酶解时间2.8 h,酶解pH5.2,酶解温度50.6℃.在最佳工艺条件下,经过3次重复试验验证,AISPs的水解度(DH)为15.89%,水解后得到苜蓿冰结构蛋白肽(Alfalfa Ice Structural Protein Peptides,AISPPs),探究AISPPs对木瓜蛋白酶活力的影响.于-30℃冻藏5 d条件下,发现加入AISPPs可提高木瓜蛋白酶活力;当冷冻时间为25 d时,对照组木瓜蛋白酶酶活性为2.8×10^5U/mg,实验组木瓜蛋白酶酶活性为4.1×10^5U/mg,相比对照组酶活性提高了46.43%.比较质量分数为0.2%、0.3%、0.4%、0.5%、0.6%的AISPPs对木瓜蛋白酶活力的影响,发现AISPPs质量分数为0.5%时,木瓜蛋白酶活力达到最高值6.3×10^5U/mg.

In this paper,alfalfa meal was used as the experimental raw material,and the icestructured protein was extracted by phosphate buffer solution to explore the effect of the structure protein on the activity of papain.The hydrolysis conditions of AISPs were optimized by single factor and response to alkaline protease.The optimization results were enzymatic hydrolysis time of 2.8 h,enzymatic hydrolysis pH of 5.2 and enzymatic hydrolysis temperature of 50.6.Under the optimum process conditions,after three repeated tests,the DH was 15.89%.Under the condition of freezing at-30℃for 5 days,it was found that the addition of ice-cold structural protein could increase the activity of papain.When the freezing time was 25 days,the activity of papain in the control group was 280000 U/mg,and the activity of papain in the experimental group was observed.At 410000 U/mg,the enzyme activity was increased by 46.43%compared to the control group.The effects of AISPPs at concentrations of 0.2%,0.3%,0.4%,0.5%,and 0.6%on the activity of papain were compared.When the concentration of AISPPs was 0.5%,the activity of papain reached the highest value of 630000 U/mg.