木瓜超氧化物歧化酶粗酶提取工艺初探

目的]筛选木瓜超氧化物歧化酶(SOD)粗酶最佳提取工艺。方法]以重庆綦江皱皮木瓜为原料,以溶解SOD,去除杂蛋白,沉淀SOD为基本工艺程序,对SOD的4种提取工艺进行比较研究。同时对4种工艺提取的SOD单位蛋白、活性、比活的检测结果进行比较。结果]磷酸缓冲液的用量对提取效果有直接影响,缓冲液与木瓜之比为3∶1,测得其比活最高,为54.55 U/mg。粗酶液加2.5倍体积的氯仿-乙醇混合液去除杂蛋白效果比较明显。粗酶液添加(NH4)2SO4到90%饱和度沉淀SOD效果较好,比活较提取液上升44.52 Umg。结论]木瓜SOD粗酶最佳提取工艺条件:温度为25～30℃,pH值为7.8,0.05 mol/L磷酸缓冲液与木瓜比例为3∶1,用2.5倍体积的氯仿-乙醇混合液去杂蛋白,添加55%(NH4)2SO4至90%饱和度沉淀SOD。

Primary Exploration on the Extraction Technology of Crude Superoxide Dismutase from Chinese Quince

Objective] The purpose of the study was to screen out the optimum extraction technology of crude superoxide dismutase(SOD) in Chinese quince.Method] With Qijiang Cheanomeles speciosa from Chongqing as material,SOD dissolution,impure protein removal and SOD sedimentation as basic technology process,4 extraction technologies of SOD was studied comparatively.And the detection results on unit protein,activity and specific activity of SOD extracted by the 4 technologies were compared.Result] The dosage of phosphate buffer solution had direct influence on the extraction effect,and the determined highest specific activity was 54.55 U/mg when the ratio of buffer solution to Chinese quince was 3∶1.The impure protein removal effect in crude enzyme solution adding 2.5 time volumes of chloroform-ethanol mixed solution was more obvious.The SOD sedimentation effect of in crude enzyme solution added(NH4)2SO4 till 90 % saturation was better and the specific activity increased 44.52 U/mg in comparison with extraction solution.Conclusion] The optimum technological condition of extracting crude SOD from Chinese quince was temperature of 25～30 ℃,pH value of 7.8,ratio of 0.05 mol/L phosphate buffer solution to Chinese quince of 3∶1,removing impure protein with 2.5 time volumes of chloroform-ethanol mixed solution and precipitating SOD by adding 55 %(NH4)2SO4 till 90 % saturation.