响应面法优化北豆根粗多糖的除蛋白工艺及其神经保护活性研究

为研究北豆根粗多糖的最佳除蛋白方法和工艺,探究北豆根粗多糖的神经保护活性。实验比较了酶法、Sevage法、酶-Sevage法、三氯乙酸(TCA)-正丁醇法、酶-TCA-正丁醇法除蛋白效果,筛选出了北豆根粗多糖的最佳除蛋白方法,并用响应面法对该方法进行优化。经响应面分析,以综合评分为指标,考查TCA与正丁醇体积比、北豆根粗多糖溶液与TCA-正丁醇溶液体积比和振摇时间对北豆根粗多糖TCA-正丁醇法除蛋白工艺的影响。用H_2O_2诱导PC12细胞损伤,探究了北豆根粗多糖的神经保护作用。结果表明,北豆根粗多糖的最佳除蛋白方法为TCA-正丁醇法。经响应面优化,确定最优除蛋白工艺为TCA:正丁醇=1∶10.4,北豆根粗多糖溶液:TCA-正丁醇溶液=1∶1.77,振摇时间为36.5 min。采用该工艺对北豆根多糖进行除蛋白,其蛋白清除率为73.1%,综合评分为92.1。神经保护研究结果表明,北豆根粗多糖对H_2O_2诱导的PC12细胞损伤具有明显的保护作用。本研究表明TCA-正丁醇法可以有效的除去北豆根粗多糖中的蛋白质且北豆根粗多糖具有神经保护活性。

Optimization of deproteinization process by response surface method and the neuroprotective activity study of Rhizoma Menispermi crude polysaccharide

To optimize the deproteinization process and to determine the neuroprotective activity of Rhizoma Menispermi crude polysaccharide,five methods were compared to screen out the best protein removing method for the Rhizoma Menispermi crude polysaccharide,including enzymatic method,Sevage method,enzyme-Sevage method,TCA-n-butanol method and enzyme-TCA-n-butanol method.The effects of the volume ratio of TCA to n-butanol 、the ratio of the polysaccharide solution to the TCA-n-butanol solution and the shake time on the deproteinization of Rhizome Menispermi crude polysaccharide in TCA-n-butanol method were investigated by single-factor experiment combined with central combination design method of Box-Behnken.PC12 cell injury was induced by H 2O 2 to study the neuroprotective activity of Rhizoma Menispermi crude polysaccharides.The results showed that the best method for deproteinization process of Rhizoma Menispermi crude polysaccharide was TCA-n-butanol method.The optimal conditions were as follows ∶the ratio of TCA to n-butanol was 1 ∶10.4,the ratio of solution of Rhizoma Menispermi crude polysaccharide to TCA-n-butanol solution was 1 ∶1.77,and the shaking time was 36.5 min.Under this condition the remove rate of protein was 73.1 %,the comprehensive score was 92.1.Neuroprotective studies have showed that Rhizoma Menispermi crude polysaccharide has obvious protective effect on H 2O 2-induced PC12 cell injury.This study showed that the TCA-n-butanol method can effectively remove the protein in the Rhizoma Menispermi crude polysaccharides,and the Rhizoma Menispermi crude polysaccharides had neuroprotective activity.