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目的:比较无动物源性成分培养基和动物源性培养基培养肺炎球菌的差异;摸索不使用有机溶剂的肺炎球菌多糖纯化新工艺。方法:用无动物源性的原料替代肺炎发酵培养基中动物源性的成分,考察菌体密度和多糖收量的变化;采用柱层析纯化工艺收获肺炎球菌多糖,测定各阶段核酸、蛋白、多糖含量。结果:与疫苗生产所采用动物源性发酵培养基相比,无动物源性发酵培养基收获的1,7F,19A型菌体密度分别提高了1倍左右;发酵液中多糖含量分别提高了48%,50%和200%;多糖收量提高了30%～90%。纯化新工艺收获的17F,18C,19F精糖,核酸和蛋白的含量均远远低于企业的注册标准,HPLC检测多糖纯度分别为99.24%,98.92%和99.02%。结论:无动物源性成分发酵培养基适合于肺炎球菌生长,不仅增加了多糖产量,也提高疫苗的安全性;纯化新工艺能够收获合格的肺炎精制多糖。
OBJECTIVE: To compare the differences of pneumococci cultured in animal-derived medium and animal-derived medium, and to explore the new technology of purifying pneumococcal polysaccharide without using organic solvents. Methods: Animal-derived material was replaced by animal-free raw material to investigate the changes of cell density and polysaccharide yield. The pneumococcal polysaccharide was harvested by column chromatography and purified. The nucleic acid, protein, Polysaccharide content. Results: Compared with the animal-derived fermentation medium used in the vaccine production, the density of 1,7F and 19A-type cells harvested without animal-derived fermentation medium increased by about 1 time respectively; the content of polysaccharides in the fermentation broth increased by 48% %, 50% and 200%; polysaccharide yield increased by 30% to 90%. Purification of new technology harvested 17F, 18C, 19F refined sugar, nucleic acid and protein content were far below the registration standards of enterprises, HPLC purity polysaccharides were 99.24%, 98.92% and 99.02%. Conclusion: The culture medium without animal origin is suitable for the growth of pneumococcus, which not only increases the yield of polysaccharides but also improves the safety of the vaccine. The purification of new technology can harvest qualified pneumonia refined polysaccharides.