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目的:建立鱼腥草中抗补体活性多糖的整套制备工艺。方法:以多糖得率和经典抗补体活性为综合指标,利用正交试验确定鱼腥草活性多糖的最佳提取工艺和最佳醇沉条件;以蛋白清除率和多糖保留率为综合指标,进行三氯乙酸法除蛋白工艺优化;以色素去除率和多糖损失率为综合指标,利用正交试验优化最佳脱色工艺。结果:最佳制备工艺为于50倍水、90℃下煎煮3次、每次2 h;将提取液浓缩至相当于每毫升0.12 g生药,加入4倍体积的90%乙醇,静置24 h;离心去上清液,沉淀依次用无水乙醇、丙醇、无水乙醚洗涤,再用水复溶,于复溶液中加入三氯乙酸至终浓度为20%除蛋白;于50℃,pH 3.0,3%活性炭下吸附50 min脱色。采用该工艺制备三批鱼腥草抗补体活性多糖,多糖得率平均为4.03%(RSD 0.96%),糖质量分数平均为80.97%(RSD 1.5%),蛋白质量分数平均为2.02%(RSD 2.3%),补体抑制活性的CH50平均为0.079 g.L-1(RSD 3.6%)。结论:本实验系统建立的工艺稳定可靠,所得多糖的糖含量高、活性强,适合鱼腥草抗补体活性多糖的大量制备。
Objective: To establish the whole preparation process of anti-complement active polysaccharide in Houttuynia cordata. Methods: The polysaccharide yield and classical anti-complement activity as a comprehensive index, the orthogonal test was used to determine the optimum extraction process and the optimum alcohol precipitation conditions of Houttuynia cordata active polysaccharide; protein clearance and polysaccharide retention rate as a comprehensive index Trichloroacetic acid method of protein removal process optimization; to pigment removal rate and polysaccharide loss rate as a comprehensive index, the use of orthogonal test to optimize the best decolorization process. Results: The best preparation process was 50 times of water, boiling at 90 ℃ for 3 times for 2 h each time. The extract was concentrated to 0.12 g crude drug per ml, 4 times volume of 90% ethanol was added, h; centrifuged to the supernatant, the precipitate was washed with absolute ethanol, propanol, anhydrous ether, and then reconstituted with water, the complex solution was added trichloroacetic acid to a final concentration of 20% protein; at 50 ℃, pH 3.0, 3% activated carbon adsorption 50 min bleaching. Three batches of Houttuynia cordata anti-complement active polysaccharides were prepared by this process. The average yield of polysaccharides was 4.03% (RSD 0.96%), the average content of carbohydrates was 80.97% (RSD 1.5%) and the average protein content was 2.02% (RSD 2.3 %), And CH50 with complement inhibitory activity averaged 0.079 gL-1 (RSD 3.6%). Conclusion: The process established by this experimental system is stable and reliable. The polysaccharide obtained has high sugar content and high activity, and is suitable for mass preparation of active polysaccharide of Houttuynia cordata Thunb.