肺炎球菌无动物源性冻干保护剂对肺炎球菌冷冻干燥过程中的保护作用评价  

Evaluation of protective effect of animal-free freeze-drying protective agent on Streptococcus pneumoniae during freeze-drying

作　　者：王研研 汪辉 曹欣[1] 刘艳宾 周也驰 徐永学 张钰东 刘建凯[1] WANG Yanyan;WANG Hui;CAO Xin;LIU Yanbin;ZHOU Yechi;XU Yongxue;ZHANG Yudong;LIU Jiankai(Beijing Minhai Biotechnology Co.,Ltd.,Beijing Key Laboratory of New Technology Research of Conjugate Vaccine,Beijing 102600,China)

机构地区：[1]北京民海生物科技有限公司结合疫苗新技术研究北京市重点实验室,北京102600

出　　处：《中国生物制品学杂志》2024年第12期1415-1420,1427,共7页Chinese Journal of Biologicals

基　　金：国家科技攻关计划(2020ZX09301034)。

摘　　要：目的 评价一种无动物源物料的肺炎球菌菌种冻干保护剂YTG对肺炎球菌冷冻干燥过程中的保护作用。方法 首先通过对YTG冻干保护剂共晶点与共熔点的分析确定其冻干程序,随后通过菌种的冻干存活率及加速稳定性试验初步确定YTG冻干保护剂在冻干与保存过程中对肺炎球菌的保护效果,再通过菌种检定与荚膜多糖抗原基因序列分析,判断YTG冻干保护剂对菌种检测特性与基因序列的影响。最后用YTG冻干保护剂冻干的菌种按生产程序进行发酵培养,分析培养情况及荚膜多糖抗原产量及质量,确定YTG冻干保护剂的生产适用性。结果 冻干存活率为50%~90%,加速稳定性试验中活菌数趋于1.2×10^(9)CFU/支。菌种检定、抗原基因序列与对照(羊血冻干菌种)相比无差异。用YTG冻干保护剂冻干的菌种发酵培养后,菌体生长与荚膜多糖产量略有提高,且荚膜多糖相关检定指标均符合《中国药典》三部(2020版)标准。结论 YTG冻干保护剂具有良好的适用性,可用于冻干保护肺炎球菌。Objective To evaluate the protective effect of YTG,an animal-free freeze-drying protective agent for pneumococcal strains,on Streptococcus pneumoniae(Spn)during freeze-drying.Methods First,the freeze-drying procedure was determined by analyzing the eutectic point and plait point of YTG freeze-drying protective agent,then the protective effect of YTG freeze-drying protective agent on pneumococcus during freeze-drying and preservation was preliminarily determined by freeze-drying survival rate and accelerated stability test of the strains.Furthermore,the effect on the characteristics of strain and polysaccharide antigen gene sequence was judged by strain verification and capsular polysaccharide antigen gene sequence analysis.Finally,the freeze-dried strains with YTG freeze-drying protective agent were used for fermentation culture according to the production procedure,and the culture situation,the yield and quality of capsular polysaccharide antigen were analyzed to determine the production applicability of YTG freeze-drying protective agent.Results The survival rate of freeze-drying was 50%-90%,and the number of viable bacteria tended to 1.2×10^(9)CFU/branch in the accelerated stability test.There was no difference in strain appearance and antigen gene sequence compared with the control(sheep blood freeze-dried strain).After the fermentation of the freeze-dried strain with YTG,the growth of bacteria and the yield of capsular polysaccharide increased slightly,and the related verification indexes of capsular polysaccharide were in line with the standards of Chinese Pharmacopoeia(VolumeⅢ,2020 edition).Conclusion YTG freeze-drying protective agent has good applicability and can be used for freeze-drying protection of pneumococcus.

关 键 词：肺炎球菌 无动物源性物料 YTG冻干保护剂 加速稳定性 生产适用性 

分 类 号：Q819[生物学—生物工程]