Fed-batch mode in shake flasks by slow-release technique
Fed-batch mode in shake flasks by slow-release technique
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DOI：
10.1002/bit.21012
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发表时间：
2006-10-20
影响因子：
3.8
通讯作者：
Buechs, J.
中科院分区：
文献类型：
--
作者：
Jeude, M.;Dittrich, B.;Buechs, J.
大多数工业生产过程都是在补料分批操作模式下进行的。相比之下，微生物生产菌株的筛选以分批模式进行，这导致与生产条件相关的生理条件完全不同。这可能会导致菌株的错误选择。开发了含有葡萄糖晶体的有机硅弹性体圆盘，以实现摇瓶中的补料分批发酵。不需要其他喂食装置。以这种方式将葡萄糖加入到受扩散控制的汉逊酵母培养物中。在摇瓶中研究了两种多形菌株：野生型菌株 (DSM 70277) 和重组菌株 pC10-FMD (P-FMD-GFP)。使用呼吸活动监测系统（RAMOS）在线监测摇瓶中培养物的氧转移率（OTR）和呼吸商（RQ）。离线测量生物量和绿色荧光蛋白 (GFP) 的形成、pH 漂移以及葡萄糖、乙醇和乙酸的代谢动态。通过缓释技术，可以减少溢出代谢，从而使生物量产量增加 85%。迄今为止，在摇瓶中可以达到23.4g/L的多形菌细胞干重。获得了0.38-0.47的生物质产量，这与配备底物进料泵的实验室规模发酵罐的产量相同。在 Syn6-MES 矿物培养基中，GFP 产量可增加 35 倍。在补料分批模式下，用 35.9 g/L 补料葡萄糖合成 88 mg/L GFP。相比之下，在批次模式下仅显示 2.5 mg/L 和 40 g/L 代谢葡萄糖。在 YNB 矿物培养基中，在补料分批模式下，421 mg/L GFP 在 41.3 g/L 补料葡萄糖下实现了超过 420 倍的改进，而在分批模式下在 40 g/L 葡萄糖补料下的改进不到 1 mg/L。 (c) 2006 年 Wiley 期刊公司
Most industrial production processes are performed in fed-batch operational mode. In contrast, the screenings for microbial production strains are run in batch mode which results in completely different physiological conditions than relevant for production conditions. This may lead to wrong selections of strains. Silicone elastomer discs containing glucose crystals were developed to realize fed-batch fermentation in shake flasks. No other device for feeding was required. Glucose was fed in this way to Hansenula polymorpha cultures controlled by diffusion. Two strains of H. polymorpha were investigated in shake flasks: the wild-type strain (DSM 70277) and a recombinant strain pC10-FMD (P-FMD-GFP). The oxygen transfer rate (OTR) and respiratory quotient (RQ) of the cultures were monitored online in shake flasks with a Respiration Activity Monitoring System (RAMOS). Formation of biomass and green fluorescent protein (GFP), pH-drift and the metabolite dynamics of glucose, ethanol and acetic acid were measured offline. With the slow-release technique overflow metabolism could be reduced leading to an increase of 85% in biomass yield. To date, 23.4g/L cell dry weight of H. polymorpha could be achieved in shake flask. Biomass yields of 0.38-0.47 were obtained which are in the same magnitude of laboratory scale fermentors equipped with a substrate feed pump. GFP yield could be increased by a factor of 35 in Syn6-MES mineral medium. In fed-batch mode 88 mg/L GFP was synthesized with 35.9 g/L fed glucose. In contrast, only 2.5 mg/L with 40 g/L metabolized glucose was revealed in batch mode. In YNB mineral medium over 420-fold improvement in fed-batch mode was achieved with 421 mg/L GFP at 41.3 g/L fed glucose in comparison to less than 1 mg/L in batch mode with 40 g/L glucose. (c) 2006 Wiley Periodicals, Inc.