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首页> 外文学位 >Fermentation control and modeling with application in production of thermo-stable alpha-amylase using recombinant Bacillus subtilis.
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Fermentation control and modeling with application in production of thermo-stable alpha-amylase using recombinant Bacillus subtilis.

机译：发酵控制和建模，并应用重组枯草芽孢杆菌生产热稳定的α-淀粉酶。

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Bacillus subtilis is an attractive host for the production of heterologous proteins. Fed-batch fermentation can attain a high cell density by avoiding substrate inhibition and accumulation of inhibitory metabolites such as acetate. In a fed-batch fermentation, the concentration of glucose or the specific growth rate is commonly used as an index in controlling the addition of the substrate (i.e., glucose) in order to maintain a stable, low concentration of the substrate during the entire process. A computer controlled system is required to monitor or control the limiting index when the specific fed-batch strategy is applied to the high cell density culture.; In this work, a computer controlled system using LabVIEW software was established. It achieved flexible and automatic monitoring and control of process parameters.; A dual exponential fed-batch culture strategy was applied to minimize the formation of acetate and to control the glucose concentration and specific growth rate at the predetermined values. Due to low solubilities of tyrosine and tryptophan in Feed Stream 1 containing concentrated glucose and other nutrients, tyrosine and tryptophan were removed from Feed Stream 1 and were dissolved in 14.4% ammonium water to form Feed Stream 2. By dual feeding both Stream 1 and Stream 2 at different exponential feed rates, the cells grew exponentially and a high cell density of 24.2 g/l and a final alpha-amylase activity of 71.4 U/ml were achieved. The overall biomass yield was 0.39 g cell/g glucose. In comparison, for the batch culture with the initial glucose concentration of 8 g/l, the final cell density was 2.3 g/l, and alpha-amylase concentration was 1.5 U/ml. The corresponding biomass yield was 0.28 g cell/g glucose.; A mathematical model was developed to investigate the inherent relationships between growth, substrate consumption, differentiation and product formation. The model includes three distinguishable cell states and the transition from the vegetative phase to sporangium and finally to mature spore. An age-based population balance model was applied to describe the maturity of sporangium toward the formation of spores. The model was able to predict the transient behavior of B. subtilis in both batch and fed-batch cultures satisfactorily.
机译：枯草芽孢杆菌是产生异源蛋白质的有吸引力的宿主。分批补料发酵可通过避免底物抑制和抑制性代谢产物（例如乙酸盐）的积累来获得高细胞密度。在分批补料发酵中，葡萄糖的浓度或特定的生长速率通常用作控制底物（即葡萄糖）添加的指标，以便在整个过程中维持稳定的低底物浓度。当特定的分批补料策略应用于高细胞密度培养时，需要一个计算机控制的系统来监测或控制极限指数。在这项工作中，建立了使用LabVIEW软件的计算机控制系统。它实现了对过程参数的灵活，自动的监视和控制。采用双指数分批补料培养策略以最小化乙酸盐的形成并将葡萄糖浓度和比生长速率控制在预定值。由于酪氨酸和色氨酸在含有浓葡萄糖和其他营养物质的原料流1中的溶解度低，因此酪氨酸和色氨酸从原料流1中除去，并溶解在14.4％的铵水中形成原料流2。如图2所示，在不同的指数进料速率下，细胞呈指数增长，并达到24.2 g / l的高细胞密度和71.4 U / ml的最终α-淀粉酶活性。生物质总产量为0.39g细胞/ g葡萄糖。相比之下，对于初始葡萄糖浓度为8 g / l的分批培养，最终细胞密度为2.3 g / l，α-淀粉酶浓度为1.5 U / ml。相应的生物量产量为0.28g细胞/ g葡萄糖。建立了一个数学模型来研究生长，底物消耗，分化和产物形成之间的内在联系。该模型包括三个可区分的细胞状态，以及从营养期到孢子囊，最后到成熟孢子的过渡。应用基于年龄的种群平衡模型来描述孢子囊对孢子形成的成熟度。该模型能够令人满意地预测枯草芽孢杆菌在分批和补料分批培养中的瞬时行为。

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