两步盐析联合双水相萃取提取纯化蓝藻中藻蓝蛋白

Extraction and Purification of Phycocyanin by the Combined Use of Two-Step Salt Precipitation and Aqueous Two-Phase Extraction from Blue Algae

以巢湖新鲜蓝藻为处理对象,采取冻融破壁的方式获取藻蓝蛋白的粗提液,采取两步盐析联合双水相萃取的方法提取纯化藻蓝蛋白。运用0.618法选点实验研究了两步盐析中(NH4)2SO4饱和度对提取纯化影响,构建了聚乙二醇(polyethylene glycol,PEG)-(NH4)2SO4的双水相体系,综合考虑PEG相对分子质量、PEG质量分数、(NH4)2SO4质量分数、p H值和藻蓝蛋白质量浓度对提取纯化藻蓝蛋白的影响。结果表明:室温25℃条件下,一步盐析时(NH4)2SO4的最佳饱和度为21%,二步盐析时(NH4)2SO4的最佳饱和度为30%。两步盐析后得到的藻蓝蛋白构建成PEG-(NH4)2SO4的双水相体系后,当PEG相对分子质量1 000、PEG 1 000质量分数10%、(NH4)2SO4质量分数25%时,藻蓝蛋白的纯度可达3.45。

The crude extracts of phycocyanin from fresh blue algal were obtained by freeze-thaw method.The purification of phycocyanin was performed by the combined use of two-step salting-out precipitation and aqueous two-phase extraction.The effect of different saturation degrees of（NH_4）_2SO_4 in two-step salt precipitation on purification efficiency was investigated using the 0.618 method,and the aqueous two-phase system of polyethylene glycol（PEG）-（NH_4）_2SO_4 was established after comprehensively considering the effect of PEG molecular weight,PEG mass fraction,（NH_4）_2SO_4 mass fraction,pH and phycocyanin mass concentration on purification efficiency.The results showed that the optimum（NH_4）_2SO_4 saturation was21%in the first step and 30%in the second step salt precipitation at 25℃.Using the aqueous two-phase system containing10%PEG 1 000 and 25%（NH_4）_2SO_4,the purity of purified phycocyanin was 3.45.