摘要
为获得侵袭性抗肿瘤靶向工程菌株EcNA高菌体浓度的培养基配方,在单因素试验的基础上,通过Plackett Burman试验筛选出对菌体浓度影响最显著的因素为酵母抽提物、K_(2)HPO_(4)用量,对显著影响因素进行中心组合试验响应面分析。结果表明最佳培养基成分为:可溶性淀粉2 g/L、酵母抽提物50.3 g/L、K_(2)HPO_(4)14.26 g/L、KH_(2)PO_(4)3 g/L、MgSO_(4)0.3 g/L、微量元素母液2 mL/L、接种量3%。发酵后溶液中菌体OD_(600 nm)达到7.602,菌体生物量达到2.96×10^(9)CFU/mL,比优化前提高了78.3%。以冻干保护剂在冷冻干燥过程中对工程菌株的保护效果为研究对象,通过正交试验得到制成菌粉的最佳保护剂配方为脱脂乳14.25 g/100 mL、蔗糖3 g/100 mL、抗坏血酸钠3 g/100 mL,优化后菌体冻干粉存活率可达86.32%,利于制成延长贮存期的口服菌粉胶囊。
The present study aimed to optimize the medium formulation for high cell density culture of an invasive tumortargeting engineered strain,Ec NA.Using one-factor-at-a-time method and a Plackett-Burman design,yeast extract and dipotassium phosphate were found to be the most important factors affecting cell concentration.The optimal medium,as determined using central composite design and response surface methodology,was composed of soluble starch 2 g/L,yeast extract 50.3 g/L,K_(2)HPO_(4)14.26 g/L,KH_(2)PO_(4)3 g/L,MgSO_(4)0.3 g/L,and trace element stock solution 2 mL/L,and the inoculum amount was 3%.The optical density at 600 nm(OD_(600 nm))of the bacterial culture obtained using the optimized medium reached 7.602 and the biomass was 2.96×10^(9)CFU/mL,which was 78.3%higher than that before optimization.Moreover,using orthogonal array design,a lyoprotectant formulation consisting of skim milk 14.25 g/100 mL,sucrose 3 g/100 mL,VC-Na 3 g/100 mL was found to be optimal for the freeze-drying of the strain.The survival rate of the engineered bacterium was up to 86.32%during freeze-drying using the lyoprotectant,being beneficial for of prolonged storage life of oral capsules.
作者
王灿
莫湘涛
张峰
吴柳娟
李躺
夏立秋
WANG Can;MO Xiangtao;ZHANG Feng;WU Liujuan;LI Tang;XIA Liqiu(Hunan Provincial Key Laboratory of Microbial Molecular Biology,State Key Laboratory of Developmental Biology of Freshwater Fish,College of Life Science,Hunan Normal University,Changsha 410081,China)
出处
《食品科学》
EI
CAS
CSCD
北大核心
2018年第2期105-111,共7页
Food Science
基金
国家重点基础研究发展计划(973计划)项目(2012CB722301)
湖南省生物发育工程及新产品研发协同创新中心项目(20134486)
关键词
抗肿瘤靶向工程菌
高密度发酵
响应面分析法
冻干保护剂
菌粉制剂
tumor-targeting engineered strain
high cell density fermentation
response surface analysis
cryoprotectant
bacterial powder
