Research on Key Parameters and Control Methods for Scaling up Biological Fermentation Processes

August 24, 2023
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Biological fermentation refers to the process of using organisms (usually microorganisms or cells) to convert raw materials into human products through specific metabolic pathways under suitable conditions. These useful metabolites, protein expression products, and other products are widely used in the pharmaceutical industry, food industry, energy industry, chemical industry, agriculture, and other fields, and are inseparable from people's daily lives. In biological fermentation engineering, how to effectively amplify the laboratory research conditions and directly apply them to production has always been a problem that troubles the application of biological fermentation. Due to the many influencing factors of biological reactions, the reaction inside the fermentation tank is a closed reaction. How to effectively control parameters so that the amplification process does not have an impact on the biological fermentation process has always been a focus of research in the biological fermentation industry. During the amplification process of biological fermentation reaction, the flow state inside the tank changes significantly with the increase of volume. Changes in the flow field can also lead to changes in a series of parameters such as temperature and dissolved oxygen, resulting in changes in the entire reaction system. The article briefly organizes and introduces the influencing factors and corresponding control parameters of biological fermentation process amplification, providing reference for parameter selection in actual biological fermentation amplification process.

Factors affecting the amplification of biological fermentation process
1.1 Mass Transfer and Mixing
Mass transfer process is the process of material transfer, and mass transfer activities in biological fermentation process occur simultaneously with biological reactions. The main mass transfer processes are divided into gas-liquid absorption and liquid mass transfer. The transfer of substances in the liquid phase is mainly due to the eddy diffusion driven by the stirring paddles of the biological fermentation tank. In commonly used biological fermentation culture, mass transfer process is very important. Good mass transfer can ensure the necessary oxygen, nutrients, and metabolites for microbial and cell culture and development. The volumetric dissolved oxygen coefficient is the most important factor affecting mass transfer, but due to the complex flow field inside the biological fermentation tank, there are many influencing factors, making it difficult to analyze the volumetric dissolved oxygen coefficient.
Another key parameter that directly affects the amplification of the biological fermentation tank process is the mixing process. Common biological fermentation reaction mixing includes liquid-liquid mixing, solid-liquid mixing, gas-liquid mixing, and gas-liquid solid three-phase mixing. Due to the increase in the volume of the fermentation tank and the increase in fermentation products and raw materials, the mixing inside the tank is uneven. For example, the mixing of substances at the top of the fermentation tank is relatively difficult compared to the bottom. Scientifically increasing the mixing of various substances in the fermentation tank can improve the efficiency of biological fermentation.
1.2 Cutting
The traditional view is that increasing the stirring rate of the biological fermentation tank can enhance mass transfer and mixing during the fermentation process. However, with in-depth research, it has been found that many biological fermentation failures are caused by excessive shear force on the target material of the biological fermentation, leading to microbial and cell damage. For example, in a microbial fermentation system, excessive shear force can cause harm to the growth of the bacterial body; Low shear force is not conducive to bubble breakage and affects the efficiency of air propagation. How to scientifically increase the mixing of various substances in the fermentation tank and control shear stress within an acceptable range for microorganisms and cells is an important factor in the fermentation amplification process.
1.3 Heat transfer
Temperature is also an important factor in the biological fermentation process. The temperature control of biological fermentation tanks is mainly achieved through the jacket layer. However, as the volume inside the large biological fermentation tank increases, the surface area per unit of heating decreases. Therefore, the efficiency of heat transfer will directly affect the production efficiency of biological fermentation target substances.
1.4 Other factors
There are other factors in the amplification process of biological fermentation tanks that can affect the fermentation process, such as the parameters of air replenishment, feeding speed, and sample inlet setting, which can all affect the fermentation process. Due to process limitations, production type biological fermentation tanks cannot detect the concentration of various substrates, products, and metabolites in real-time like the biological fermentation process in the laboratory. Therefore, it is crucial to scientifically design the speed and quantity of feeding and intake. At the same time, it is necessary to comprehensively consider the apparent gas velocity for material replenishment and air replenishment to avoid the phenomenon of "liquid flooding".

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2 Key control parameters of biological fermentation process methods
2.1 Mixing
The commonly used mixing mode of stirred biological fermentation tanks is the rotation of the stirring paddle to drive the mixing of the entire fermentation liquid. The control of fermentation tank stirring parameters is mainly achieved through speed control. Speed control should not only consider increasing the speed and improving mixing efficiency, but also control the speed within a reasonable range. Excessive speed can lead to increased heat generation, increased shear force on cells, and fermentation failure. In addition, research has found that the flow pattern of the fermentation volume system, the selection of stirring paddles, and the diameter can all affect the efficiency of biological fermentation. In the process of large-scale biological fermentation, in addition to rotational speed, the selection of stirring paddle type and spatial position is also very important. It is necessary to choose the appropriate stirring paddle type based on the fluid properties of the culture material. Currently, a combination of axial flow and radial flow paddles is commonly used, which combines micro liquid flow and macro flow fields to improve the degree of material mixing in the entire biological fermentation system. The mixing paddle generally adopts a bottom runoff type and an upper axial flow type, which can effectively ensure that the nutrients added to the top are quickly distributed to the bottom of the tank under the action of axial flow slurry, and the air introduced to the bottom of the tank can also be dispersed in a timely manner, ensuring the overall circulation and flow of the entire tank, providing a suitable environment for the entire microbial fermentation.
2.2 Temperature
The biological fermentation tank is generally at 26~37 ℃ depending on the type of bacteria being cultivated, while the cultivation of special bacteria may be at 65 ℃. During the amplification process of the fermentation process, the temperature field inside the entire fermentation tank will undergo significant changes. In the small-scale and pilot stages, due to the small size of the tank, the temperature field is relatively uniform. In the production type fermentation tank, the temperature probes of the fermentation tank are usually distributed in the lower part of the tank, with a length of 100mm, and the soaking part in the fermentation liquid is 50-60mm. The heating and cooling methods of biological fermentation tanks are usually carried out through a jacket water layer, so the heat transfer efficiency of the fermentation tank directly affects the temperature distribution inside the entire tank. The temperature of the jacket water layer and the temperature display value of the fermentation tank temperature probe cannot truly reflect the temperature of the liquid in the fermentation tank. Scientifically arranging temperature probes and scientifically setting fermentation temperature based on the heat transfer coefficient of the fermentation system can effectively ensure the reaction temperature.
2.3 Other chemical parameters
The control of chemical parameters in biological fermentation, such as pH value and dissolved oxygen, can have an impact on the fermentation results. Taking dissolved oxygen parameters as an example, in aerobic biological processes, oxygen is an important nutrient for microbial growth. However, due to its bottom solubility, oxygen becomes a key substrate for biochemical processes. Therefore, maintaining an adequate supply of oxygen from the gas phase to the liquid phase is crucial. In theory, increasing ventilation and increasing the culture medium column can effectively prolong the time of bubbles in the culture medium and improve gas-liquid exchange efficiency. However, these parameters are also limited by the cost of the fermentation tank, and the degree and size of bubble dispersion can also affect the efficiency of oxygen propagation. Therefore, it is necessary to comprehensively consider various factors and provide the most suitable process design amplification method.

3. Scaling up method for biological fermentation process
3.1 Empirical amplification method
The scaling up process of traditional biological fermentation processes is mostly based on traditional empirical methods. In the biological fermentation process, a series of parameters such as the speed, arrangement, air ventilation rate, feed flow rate, and other chemical parameters of the stirring paddles can affect the fermentation yield. Users will choose similar fermentation processes based on previous or other fermentation process settings, and select corresponding fermentation process amplification parameters; Alternatively, based on traditional experience, the fluid dynamics in the fermentation tank can be predicted, and the various parameters inside the tank can be geometrically enlarged while maintaining their relative positions to expand the fermentation volume. Empirical amplification methods mainly focus on key parameters within the fermentation system, such as volumetric mass transfer coefficient, unit volume power consumption, mixing time, etc. This method is usually only suitable for simple amplification and cannot effectively predict the fluid dynamics and kinematic characteristics in fermentation tanks.
3.2 Amplification Method Based on Computational Fluid Dynamics
Computational Fluid Dynamics (CFD) refers to the simulation and calculation of fluid motion laws in a computer based on microscopic equations such as mass transfer, momentum transfer, and energy transfer in fluid mechanics. Compared with experiential biological fermentation process schemes, using CFD simulation technology has the characteristics of low cost and size independence, and is widely used in the field of fluid engineering. CFD simulation mainly simulates the flow field, stirring power, and gas holdup inside the biological fermentation tank. At the same time, a dissolved oxygen mass transfer model is coupled in the gas-liquid two-phase flow model of the biological fermentation tank, which can simulate the dissolved oxygen mass transfer process and biochemical reaction process during the same fermentation process.
With the development of simulation computing technology, CFD is increasingly applied in the simulation of biological fermentation amplification processes. However, due to the complexity of gas-liquid two-phase flow, further research on parameters such as gas holdup and bubbles in the prediction process is still needed.

4 Conclusion
The process of biological fermentation is a complex and multifactorial process. Although traditional stirred tank bioreactors are relatively simple in structure, the actual process of the fluid inside the fermentation tank is very complex in the actual reaction process. Especially in the amplification process of biological fermentation, multiple factors need to be comprehensively considered for methodological amplification. The traditional experiential amplification process can only perform simple amplification and cannot truly simulate the real data of various systems in the fermentation tank. In addition to ensuring that the growth environment of fermentation products is consistent with the laboratory, attention should also be paid to energy conservation. Based on computational fluid dynamics, more scientific analysis and simulation methods can more accurately predict and simulate the biological fermentation amplification process, providing effective basis and reference for the selection of biological fermentation amplification processes.


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