槐花多糖的提取、纯化和抗氧化活性分析

Extraction,Purification and Antioxidant Activity of Polysaccharides from Sophora japonica

优化槐花多糖的提取工艺,并对分离纯化出的单一多糖组分SJP进行结构表征及抗氧化活性的研究。以粗多糖得率为评价指标,采用单因素结合响应面法探究水提法提取槐花多糖的最佳条件。在最佳条件下提取出的粗多糖通过脱蛋白、透析和Sephadex G-200凝胶柱层析得到纯化多糖组分SJP,采用高效液相色谱(HPLC)法测定其分子量,并对其进行化学成分的测定、刚果红实验和X-射线衍射(XRD)分析。以DPPH·、·OH、ABTS^(+)·清除率和总还原力为指标,探究SJP的抗氧化活性。结果表明,槐花多糖的最佳提取工艺条件为料液比1:20 g/mL,提取时间120 min,醇沉浓度60%,此条件下多糖的实际得率为3.94%。HPLC结果表明SJP是一种分子量为2.32×10^(6)Da的均质多糖。SJP中总糖含量、蛋白质含量和糖醛酸含量分别为93.47%±0.83%、1.62%±0.13%和7.13%±0.51%。刚果红实验表明SJP不含三螺旋结构,XRD结果表明SJP结晶度低,为无定型结构。SJP清除DPPH、OH、ABTS+自由基的IC_(50)值分别为1.09、4.31、1.39 mg/mL,在5 mg/mL浓度下,总还原力为0.57,说明其具有一定的抗氧化活性。综上所述,本研究提取、分离纯化槐花多糖的工艺切实可行,所得多糖纯度较高且具有一定的抗氧化能力。

The extraction process of polysaccharide from Sophora japonica was optimized,and the single polysaccharide component(named SJP)was purified,then the structure characterization and antioxidant activity of SJP were investigated.Using crude polysaccharide yield as evaluation index,single factor combined with response surface method were used to explore the optimal conditions of water extraction of Sophora japonica polysaccharide.The crude polysaccharide extracted under the optimal conditions was further purified by deproteinization,dialysis and Sephadex G-200 gel column chromatography.The molecular weight of SJP was determined by high performance liquid chromatography(HPLC),and its chemical composition was analyzed.Congo red experiment and X-ray diffraction(XRD)analysis was applied to characterize the SJP structure.With the free radical scavenging rate and total reducing power as indicators,the antioxidant activity of SJP was investigated.The results showed that the optimal extraction conditions were solid-liquid ratio of 1:20 g/mL,extraction time of 120 min and alcohol precipitation concentration of 60%.Under these conditions,the polysaccharide yield was verified with 3.94%.HPLC results showed that SJP was a homogeneous polysaccharide with the molecular weight of 2.32×10^(6) Da.The contents of total sugar,protein and uronic acid in SJP were 93.47%±0.83%,1.62%±0.13% and 7.13%±0.51%,respectively.Congo red experiment showed that SJP did not contain triple helix structure,and XRD results indicated that SJP had low crystallinity and amorphous structure.The IC_(50) values of SJP for scavenging DPPH,OH and ABTS^(+)radicals were 1.09,4.31 and 1.39 mg/mL,respectively,and the total reducing power was 0.57 at concentration of 5 mg/mL,indicating that SJP had certain antioxidant activity.In conclusion,the process of extracting and purifying polysaccharide from Sophora japonica in this study was feasible,and the obtained polysaccharide had high purity and exhibited good antioxidant activity.