Keil T, Dittrich B, Lattermann C, Habicher T, Büchs J
1AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074 Aachen, Germany.
2DWI - Leibniz Institute for Interactive Materials, RWTH Aachen University, Forckenbeckstraße 50, 52074 Aachen, Germany.
J Biol Eng. 2019 Feb 22;13:18. doi: 10.1186/s13036-019-0147-6. eCollection 2019.
Fed-batch conditions are advantageous for industrial cultivations as they avoid unfavorable phenomena appearing in batch cultivations. Those are for example the formation of overflow metabolites, catabolite repression, oxygen limitation or inhibition due to elevated osmotic concentrations. For both, the early bioprocess development and the optimization of existing bioprocesses, small-scale reaction vessels are applied to ensure high throughput, low costs and prompt results. However, most conventional small-scale procedures work in batch operation mode, which stands in contrast to fed-batch conditions in large-scale bioprocesses. Extensive expenditure for installations and operation accompany almost all cultivation systems in the market allowing fed-batch conditions in small-scale. An alternative, more cost efficient enzymatic glucose release system is strongly influenced by environmental conditions. To overcome these issues, this study investigates a polymer-based fed-batch system for controlled substrate release in microtiter plates.
Immobilizing a solid silicone matrix with embedded glucose crystals at the bottom of each well of a microtiter plate is a suitable technique for implementing fed-batch conditions in microtiter plates. The results showed that the glucose release rate depends on the osmotic concentration, the pH and the temperature of the medium. Moreover, the applied nitrogen source proved to influence the glucose release rate. A new developed mathematical tool predicts the glucose release for various media conditions. The two model organisms and were cultivated in the fed-batch microtiter plate to investigate the general applicability for microbial systems. Online monitoring of the oxygen transfer rate and offline analysis of substrate, product, biomass and pH confirmed that fed-batch conditions are comparable to large-scale cultivations. Furthermore, due to fed-batch conditions in microtiter plates, product formation could be enhanced by the factor 245 compared to batch cultivations.
The polymer-based fed-batch microtiter plate represents a sophisticated and cost efficient system to mimic typical industrial fed-batch conditions in small-scale. Thus, a more reliable strain screening and early process development can be performed. A systematical scale-down with low expenditure of work, time and money is possible.
补料分批培养条件对工业培养有利，因为它们可避免分批培养中出现的不利现象。例如，这些现象包括溢流代谢产物的形成、分解代谢物阻遏、氧气限制或因渗透压升高导致的抑制作用。对于早期生物工艺开发和现有生物工艺的优化，均采用小规模反应容器以确保高通量、低成本和快速获得结果。然而，大多数传统的小规模程序采用分批操作模式，这与大规模生物工艺中的补料分批培养条件形成对比。市场上几乎所有允许小规模补料分批培养条件的培养系统，其安装和运行成本都很高。一种替代的、更具成本效益的酶促葡萄糖释放系统受环境条件的影响很大。为克服这些问题，本研究探讨了一种基于聚合物的补料分批系统，用于在微孔板中控制底物释放。
在微孔板每个孔的底部固定嵌入葡萄糖晶体的固体硅氧烷基质，是在微孔板中实现补料分批培养条件的合适技术。结果表明，葡萄糖释放速率取决于培养基的渗透压、pH值和温度。此外，所应用的氮源被证明会影响葡萄糖释放速率。一种新开发的数学工具可预测各种培养基条件下的葡萄糖释放量。在补料分批微孔板中培养两种模式生物，以研究其对微生物系统的普遍适用性。对氧气传递速率的在线监测以及对底物、产物、生物量和pH值的离线分析证实，补料分批培养条件与大规模培养相当。此外，由于微孔板中的补料分批培养条件，与分批培养相比，产物形成可提高245倍。
基于聚合物的补料分批微孔板是一种复杂且具成本效益的系统，可在小规模下模拟典型的工业补料分批培养条件。因此，可以进行更可靠的菌株筛选和早期工艺开发。实现低工作量、时间和金钱投入的系统规模缩小是可行的。
