经验放大法对肺炎链球菌规模化发酵工艺条件的研究  

Investigation of conditions on scale fermentation process for Streptococcus pneumonia by experiential method

作　　者：张新庄[1] 白贵杰[1] 张轶[1] 任珍芸[1] 张丽芝[1] 张钦[1] 沈荣[1] 

机构地区：[1]兰州生物制品研究所有限责任公司第一研究室甘肃省疫苗工程技术研究中心,甘肃兰州730046

出　　处：《微生物学免疫学进展》2016年第6期48-54,共7页Progress In Microbiology and Immunology

摘　　要：目的应用经验放大法探索肺炎链球菌规模化基本发酵工艺条件。方法复苏肺炎链球菌菌种,传代培养,最后接种于30 L发酵罐中,以细菌发酵的培养时间、收获液菌浓度A600 nm值、荚膜多糖产率为指标评价发酵工艺;以纯化多糖的功能基团含量、相对分子质量大小(色谱分配系数KD值)、生物活性为指标评价多糖质量。以4型、19F型肺炎链球菌为首选菌株,用于筛选高产菌株、优化传代方法、培养基及30 L发酵罐的通气条件,并应用经验放大法至270 L及1 200 L发酵罐,并将发酵条件推演到其他血清型肺炎链球菌。结果筛选出肺炎链球菌4型和19F高产菌株,利用市售复合培养基,经3代种子液体-液体传代法,0.03 VVM通气条件下,4型和19F菌株在3 0 L发酵罐中获得良好的培养结果;并采用经验放大法至1 200 L发酵罐,获得良好结果。采用此发酵工艺条件,其他20个血清型肺炎链球菌在1 200 L发酵罐中也获得较理想结果。结论应用经验放大法初步探索了肺炎链球菌在1 200 L发酵罐中的发酵工艺条件。Objective Development of the basic conditions of scale fermentation process for Streptococcus pneumonia was carried out by means of an experiential method. Methods The frozen stock of the working seed was reconstituted,passaged,and inoculated in 30 L fermentor. The indexes were used to evaluate the fermentation process including the cultivation time of the bacteria,optical density at A600 of harvested culture and yield of capsular polysaccharides（ CPS）. The standard in estimation of the polysaccharides quality was combined with indexes,such as percentage for component in functional group,the distribution coefficient（ KD） and the biological activity of CPS. Take serotype 4 and 19 F as first choice,the fermentation process was explored for 30 L fermentor with strain screening and passages,medium selection and ventilation strategy,then the selected conditions were used for culture with 270 L and 1 200 L fermentor by the experiential method,finally,the fermentation process was extended to culture the other serotypes. Results The high-prodution strain from serotype 4 and 19 F were obtained by screening. The satisfied fermentation results were obtained in 30 L fermentor as the strain was on 3 passages in a growth liquid, by using of commercial culture medium and aeration rate of 0. 3 vvm（air volume/culture volume/min）for Streptococcus pneumonia serotype 4 and 19 F,and the similar results can be obtained when the fermentation process was scale up to 1 200 L fermentor by the experiential method,and good results can be o btained by using of the same fermetation technology in 1 200 L fermentor for 20 other serotypes. Conclusion The priliminary fermentation process was basically developed in 1 200 L fermentor. by the experiential method for Streptococcus p neumoniaein.

关 键 词：肺炎链球菌 菌种筛选 通气条件 培养基 经验放大法 

分 类 号：TQ920.6[轻工技术与工程—发酵工程]