The enhancement of specific antibody production rate in glucose- and glutamine-controlled fed-batch culture

The concentration effects of certain amino acids (Asp, Ile, Leu, Lys, Met, Val, Phe and Gln which were highly consumed during cultivation), and glucose on cell growth and antibody productivity were investigated using dish culture. From these experiments, it was found that only glutamine enrichment enhanced the specific antibody production rate. The other amino acids described above did not affect either the specific growth rate or specific antibody production rate. Thus we investigated the quantitative effects of glutamine concentration in the range of 0.4∼33.3 mmol·1⁻¹ on kinetic parameters in fed-batch culture which kept both glucose and glutamine concentration constant. As a result the specific growth rate decreased with increase in glutamine concentration in the range larger than 20 mmol·1⁻¹. The specific antibody production rate had a maximum value at about 25 mmol·1⁻¹ glutamine concentration.     

Physiological and technological aspects of large-scale heterologous-protein production with yeasts

Commercial production of heterologous proteins by yeasts has gained considerable interest. Expression systems have been developed forSaccharomyces cerevisiae and a number of other yeasts. Generally, much attention is paid to the molecular aspects of heterologous-gene expression. The success of this approach is indicated by the high expression levels that have been obtained in shake-flask cultures. For large-scale production however, possibilities and restrictions related to host-strain physiology and fermentation technology also have to be considered. In this review, these physiological and technological aspects have been evaluated with the aid of numerical simulations. Factors that affect the choice of a carbon substrate for large-scale production involve price, purity and solubility. Since oxygen demand and heat production (which are closely linked) limit the attainable growth rate in large-scale processes, the biomass yield on oxygen is also a key parameter. Large-scale processes impose restrictions on the expression system. Many promoter systems that work well in small-scale systems cannot be implemented in industrial environments. Furthermore, large-scale fed-batch fermentations involve a substantial number of generations. Therefore, even low expression-cassette instability has a profound effect on the overall productivity of the system. Multicopy-integration systems may provide highly stable expression systems for industrial processes. Large-scale fed-batch processes are typically performed at a low growth rate. Therefore, effects of a low growth rate on the physiology and product formation rates of yeasts are of key importance. Due to the low growth rates in the industrial process, a substantial part of the substrate carbon is expended to meet maintenance-energy requirements. Factors that reduce maintenance-energy requirements will therefore have a positive effect on product yield. The relationship between specific growth rate and specific product formation rate (kg product·[kg biomass]^–1·h^–1) is the main factor influencing production levels in large-scale production processes. Expression systems characterized by a high specific rate of product formation at low specific growth rates are highly favourable for large-scale heterologous-protein production.     

Xanthomonas campestris as a host for the production of recombinantPseudomonas aeruginosa lipase

Recombinant plasmid pBP13, which expresses the alkaline lipase fromPseudomonas aeruginosa IGB83 under thetac promoter was transferred toXanthomonas campestris pvcampestris IBT148. Different fermentation conditions were tested for lipase productivity by strain IBT148 carrying plasmid pBP13, and a fermentation process was established in an instrumented bioreactor, where lipase production was increased more than 12-fold with respect to the initial culture conditions in shake flasks. Xanthan gum stabilized the activity of the alkaline lipase.     

Growth kinetics of the photosynthetic bacterium Chlorobium thiosulfatophilum in a fed-batch reactor

Hydrogen sulfide dissolved in water can be converted to elementary sulfur or sulfate by the photosynthetic bacterium Chlorobium thiosulfatophilum. Substrate inhibition occurred at sulfide concentrations above 5.7 mM. Light inhibition was found at average light intensities of 40,000 lux in a sulfide concentration of 5 mM, where no substrate inhibition occurred. Light intensity, the most important growth parameter, was attenuated through both scattering by sulfur particles and absorption by the cells. Average cell and sulfur particle sizes were 1.1 and 9.4 mum, respectively. Cells contributed 10 times as much to the turbidity as sulfur particles of the same weight concentration. The light attenuation factor was mathematically modeled, considering both the absorption and scattering effects based on the Beer-Lambert law and the Rayleigh theory, which were introduced to the cell growth model. Optimal operational conditions relating feed rate vs. light intensity were obtained to suppress the accumulation of sulfate and sulfide and save light energy for 2- and 4-L fed-batch reactors. Light intensity should be greater for the same performance (H(2)S removal rate/unit cell concentration) in larger reactors due to the scaleup effect on light transmission. Knowledge of appropriate growth kinetics in photosynthetic fed-batch reactors was essential to increase feed rate and light intensity and therefore cell growth. A mathematical model was developed that describes the cell growth by considering the light attenuation factor due to scattering and absorption and the crowding effect of the cells. This model was in good agreement with the experimental results. (c) 1992 John Wiley & Sons, Inc.     

Simulation of L/A control in fed-batch culture of baker's yeast

L/A controllers have extended their use from continuous to fed-batch fermentation where the control is applied from the start of an initial batch phase. As opposed to proportional integral derivative (PID) controllers where even a startup procedure is recommended prior to fed-batch, the L/A controller is not upset by an early connection. It is easily retuned continuously by means of ethanol measurements and can cope with a large range of output conditions. The performance of an L/A algorithm, which uses biomass concentration as the controlled variable, is assessed through simulation. The self-contained algorithm is relatively simple with no greater intrinsic complexity than modern PID stand alone controllers.     

Fed-batch fermentation for production of nitrile hydratase byRhodococcus rhodochrous M33

To enhance the productivity and activity of nitrile hydratase inRhodococcus rhodochrous M33, a glucose-limited fed-batch culture was performed. In a fed-batch culture where the glucose was controlled at a limited level and cobalt was supplemented during the fermentation period, the cell mass and total activity of nitrile hydratase both increased 3.3-fold compared to that in the batch fermentation. The productivity of nitrile hydratase also increased 1.9-fold compared to that in the batch fermentation. The specific activity of nitrile hydratase in the whole cell preparation when using a fed-batch culture was 120 units/mg-DCW, which was similar to that in the batch culture.     

Xylitol production in repeated fed batch cultivation

The ability of Candida parapsilosis to produce xylitol was tested using successive substrate supplies, and the importance of the amount of viable cells in enhancing the conversion rate was demonstrated. The suitability of this yeast for the production of xylitol was investigated in repeated fed-batch cultivation, using pure xylose or mixtures of xylose and glucose. The use of this process increased productivity by about 40% compared with simple batch cultivation without loss of yield of product on substrate. The presence of glucose in the culture medium seemed to stimulate the specific growth rate, but had no influence over other fermentative parameters.     

pH及流加葡萄糖对酵母分批发酵生产谷胱甘肽的影响

在5 L的发酵罐中研究了pH及流加葡萄糖对酵母分批发酵生产谷胱甘肽(GSH)的影响。实验考察了不同浓度的流加葡萄糖和不同的恒pH值的分批发酵对于酵母生产GSH产量的变化。实验结果表明,当pH值控制为5.0,流加葡萄糖流速为5g.L-1.h-1,连续流加30 h,可使GSH产量最高,与之前未流加葡萄糖和控制pH相比,其产量提高了6倍。     

采用最小二乘支持向量机的青霉素发酵过程建模研究

生化过程通常是严重非线性和时变的复杂动态系统,而且重要过程参数缺少在线测量仪表,对其建立机理模型往往非常耗时和困难。采用最小二乘支持向量机(LS_SVM)并以Pensim仿真平台为例对青霉素发酵这一典型生化过程进行建模研究。给出了LS_SVM参数的调整策略和分析结果,建立了青霉素产物浓度、菌体浓度和底物浓度等重要过程变量的在线预报模型。仿真结果表明用LS_SVM建立的在线预报模型拟合误差小,推广性能好,可以作为发酵过程的进一步控制和优化的参考依据。