The purposes of speed control in the microbial fermentation process are multifaceted, which can mainly be summarized as follows:
1.1 Optimizing growth conditions:
microbial growth and metabolism are affected by a variety of environmental factors, including temperature, pH, dissolved oxygen, substrate concentration, etc. By controlling the speed, these environmental factors can be adjusted and optimized to provide the optimal growth conditions for microorganisms, thereby promoting their rapid growth and efficient metabolism.
1.2 Increasing product yield:
In the fermentation process, microorganisms will synthesize target products (such as antibiotics, enzymes, organic acids, etc.). Speed control can ensure that the microbial metabolic activity in the optimal state, thereby increasing the rate of product synthesis and production. If the fermentation rate is too fast, it may lead to rapid substrate depletion, produce byproducts that inhibit microbial growth, or make microorganisms enter into decline period earlier, which results in reducing product yield.
1.3 Maintaining fermentation stability:
The stability of the fermentation process is essential to ensure the quality and yield of the product. By controlling the speed, various parameters in the fermentation process can be maintained within a stable range to avoid adverse effects of violent fluctuations on microbial growth and metabolism. This helps to reduce the risk of fermentation failure and improve production efficiency and economic efficiency.
1.4 Saving energy and resources:
In the fermentation process, a lot of energy and resources (such as electricity, steam, substrate, etc.) need to be consumed. Through reasonable speed control, energy and resource utilization efficiency can be optimized, and waste and cost can be reduced. For example, in terms of dissolved oxygen control, the precise adjustment of ventilation and stirring speed can ensure that the concentration of dissolved oxygen in the fermentation liquid is maintained within the appropriate range, so as to avoid energy waste and substrate oxidation loss caused by excessive ventilation.
1.5 To meet the needs of different fermentation stages:
the fermentation process usually includes different stages such as adaptation period, logarithmic growth period, stable period and decline period. Each stage has different requirements for environmental conditions and nutrients. By controlling the speed, fermentation conditions can be adjusted according to the needs of different fermentation stages, ensuring that microorganisms can obtain the best growth and metabolic environment at each stage.
2.1 Shear force
Due to the brittle skin of some microorganisms, high rotational speed may lead to the rupture of microorganisms:Cell rupture: High shear forces may directly lead to the rupture of the microbial cell wall, releasing material inside the cell. This not only affects the survival rate of the microorganisms, but may also adversely affect the subsequent fermentation or treatment process.Metabolite leakage: After the cell ruptures, metabolites within the cell leak into the environment. These metabolites may be biologically active or toxic, which will result in potential threats to the environment and human health.Fermentation efficiency declines: If a large number of microbial cells rupture during the fermentation process, it will lead to the reduction in the concentration of cells in the fermentation liquid, which will affect the fermentation efficiency and product yield.
2.2 Oxygen Poisoning
For aerobic microorganisms, the mechanism of oxygen poisoning mainly involves the excessive generation and damage of oxygen free radicals and the imbalance of antioxidant defense system. Under high oxygen concentrations or stress, aerobic microorganisms may be at risk of oxidative stress, leading to disruption of cell structure and function and death of cell. Therefore, when cultivating and applying aerobic microorganisms, it is necessary to control the appropriate oxygen concentration and pressure conditions to ensure their normal growth and metabolic activities. Therefore, it is necessary to control the rate of fermentation by limiting the ventilation ratio.
In the hyperventilation state, if the oxygen concentration in the environment of microorganisms is too high, it can cause damage to them and even lead to death. This damage mechanism may be related to the formation and attack of oxygen free radicals.
Controlling ventilation amount: During the fermentation process, ventilation amount should be reasonably controlled according to the needs and tolerance of microorganisms to avoid too high or too low oxygen concentration.
Monitoring oxygen concentration: Regularly monitor the oxygen concentration in the fermentation solution to ensure that it is within the appropriate range.
Optimization of fermentation conditions: In addition to ventilation amount, other fermentation conditions such as temperature, pH, substrate concentration, etc. should be paid attention to to create the most suitable environment for microbial growth and product synthesis.
Strengthening equipment maintenance: ensure that the fermentation equipment is in good condition to avoid abnormal ventilation caused by equipment failure.
2.3 Substrate inhibition
High concentration of substrate will inhibit microbial metabolism, mainly because of the following aspects:
a. The increase of substrate concentration will cause the osmotic pressure of the solution to rise, thus affecting the growth and reproduction of microorganisms. The high concentration of substrate will inhibit the growth and reproduction of microorganisms, because too high osmotic pressure will destroy the cell membrane, resulting in cell water loss or excessive water absorption, which will have an impact on the normal function of the cell.
b. High concentrations of substrates may lead to saturation of metabolic pathways. When the substrate concentration reaches the certain level, the corresponding enzyme may reach the saturated state, resulting in the further increase in the substrate concentration, and the reaction speed no longer increases.
c. High concentrations of substrates may also induce metabolic overflow. When the substrate concentration is too high, the microorganism may initiate unnecessary metabolic pathways, resulting in energy waste and by-product generation, which also affects the learning of the target product. In the study of microbial metabolism, controlling substrate concentration is of great significance to regulate metabolic pathways and improve the yield and efficiency of target products.
2.4 Product inhibition
Excessive concentration of the product will significantly inhibit the growth and metabolism of microorganisms, slow down the fermentation rate, and even lead to fermentation failure. High concentration of products may directly poison cells, affect enzyme activity and metabolic pathways, increase osmotic pressure of fermentation fluid, and destroy cell balance. Therefore, it is very important to control the fermentation rate. By adjusting the culture conditions, adding inhibitors,etc we can maintain the appropriate rate of product formation and prevent excessive accumulation. At the same time, the product separation rate is improved to realize the coupling process of fermentation and separation, which can effectively reduce the product concentration, reduce the inhibition effect, ensure the stable fermentation process, and improve the product yield and quality. The main solutions are coupling fermentation or perfusion culture.
Coupling fermentation technology, that is, the technique of separating while fermenting, is an advanced technology that closely combines the fermentation process with the product separation process. By using appropriate separation technology in the fermentation process, such as membrane separation, extraction, etc., the product is separated from the fermentation liquid in real time, so as to effectively reduce the product concentration, relieve the inhibition of microorganisms, and improve the fermentation efficiency and product yield. This process not only optimizes the fermentation process, but also improves the purity and recovery rate of the product, which is an important development direction of modern biological fermentation industry.
2.5 Deformed Synthesis
a. Decreased stability of mRNA: Under conditions of rapid growth, mRNA stability within cells may be affected, resulting in an accelerated rate of mRNA degradation associated with protein synthesis. This will reduce the amount of mRNA available for translation, which in turn affects protein synthesis.
b. Impaired ribosome function: Although the ribosome itself is not directly reduced due to excessive growth rate, the increased metabolic pressure and unequal resource distribution that may result from rapid growth may indirectly affect ribosome function. For example, ribosomes may not be able to obtain enough energy or amino acids to support efficient protein synthesis.
c. Abnormalities of protein folding and modification : Due to the lack of Golgi apparatus and endoplasmic reticulum, the rapidly growing cells may not be able to provide an adequate folding and modification environment for nascent proteins. This may result in the protein not folding properly into a biologically active conformation, or not being able to obtain the necessary modifications such as glycosylation and phosphorylation, thus affecting its function and stability.
Due to the above reasons, for prokaryotic microorganisms, controlling the growth rate of microorganisms by adjusting medium composition, feeding strategy, temperature and other ways is conducive to maintaining the stability of intracellular metabolic balance and gene expression regulation, thus improving the efficiency and quality of protein synthesis.
2.6 Energy distribution and material distribution
In the microbial process, controlling fermentation rate is the key to ensure the balance of all stages. By fine-tuning the fermentation rate, the allocation of time, biomass, nutrients and energy can be optimized. Through the segmented process design, the growth cycle of microorganisms is comprehensively considered, and the appropriate growth conditions are maintained to avoid resource waste or product inhibition caused by too fast or too slow. Rate-controlled fermentation is an important strategy in modern biological fermentation industry to maximize the utilization of resources and improve product quality and yield.
The logic of speed control of microbial fermentation largely revolves around controlling the rate of enzyme-catalyzed reactions in microorganisms. Enzymes are proteins that catalyze chemical reactions in living organisms, and they are able to accelerate the process of the reaction without changing the total energy change of the reaction. In the process of microbial fermentation, the rate of enzyme-catalyzed reaction directly affects the production rate of metabolites and fermentation efficiency. Fast and stable enzyme-catalyzed reaction rates help to increase fermentation efficiency, reduce fermentation time, and potentially increase product purity and yield.
3.1 Temperature control
The enzyme activity is significantly affected by temperature. In general, each enzyme has its optimal temperature range, in which the enzyme activity is the highest and the reaction rate is the fastest. Therefore, by controlling the fermentation temperature, the rate of enzyme-catalyzed reaction can be regulated.For example, for some microorganisms requiring high temperature fermentation, the suitable high temperature environment can be provided to accelerate the enzyme-catalyzed reaction; For microorganisms that require low temperature fermentation, the low temperature is needed to maintain enzyme activity.
3.2 pH value regulation
Enzyme activity is also affected by pH. Each enzyme has its optimal pH range, if it is deviated from this range,the enzyme activity will decrease or even be inactivated. Therefore, the rate of enzyme-catalyzed reaction can be affected by adjusting the pH value of fermentation solution. The pH value of the fermentation solution can be tested regularly, and the acid-base regulator can be added as needed to maintain the optimal fermentation state.
3.3 Substrate concentration and nutrient conditions
Substrate concentration is one of the important factors that affect the reaction rate of enzyme catalysis. High substrate concentration may lead to substrate inhibition effect and decrease the reaction rate; However, too low substrate concentration may limit the reaction speed. Therefore, the dosage and acceleration of the substrate need to be controlled to maintain the suitable substrate concentration range.In addition, microorganisms also need certain nutrients during the fermentation process to maintain their growth and metabolic activities. Providing adequate nutrients helps to increase the rate and efficiency of enzyme-catalyzed reactions.
3.4 Dissolved oxygen and ventilation amount
For aerobic microorganisms, dissolved oxygen is one of the important factors affecting their metabolic activities. The level of dissolved oxygen in fermentation liquid can be controlled by regulating ventilation amount and stirring speed, which can affect the rate of enzyme-catalyzed reaction. Generally speaking, it is necessary to maintain the high level of dissolved oxygen in the process of aerobic fermentation to promote the growth and metabolic activities of microorganisms. In the process of anaerobic fermentation, it is necessary to keep the low level of dissolved oxygen to avoid the inhibition of oxygen on anaerobic microorganisms.
3.5 Cumulative control of fermentation time
Fermentation time is also one of the important factors that affect the rate of enzyme-catalyzed reaction and the amount of product produced. By controlling the length of fermentation time, the metabolic activities of microorganisms and the amount of products produced can be regulated. In practice, it is necessary to determine the appropriate fermentation time range according to the specific fermentation process and product requirements.
4.1 High oxygen consumption strains
High oxygen consumption strains are those microorganism species that require large amounts of oxygen during their growth and metabolism. These strains are extremely efficient at consuming oxygen and are commonly found in a variety of genetically engineered bacteria in industrial applications that require high-oxygen environments, such as biodegradation and wastewater treatment. For example, Bacillus subtilis, as one typical strain which is high oxygen consumption, can rapidly consume a large amount of oxygen in wastewater treatment, effectively decompose organic pollutants, and significantly reduce the chemical oxygen demand of water. Its high oxygen consumption capacity makes it an important microbial resource in the field of environmental protection.
4.2 Shear sensitive strains
Microorganisms that are sensitive to shear forces mainly include but are not limited to the following categories:
a.Plant cells, which are relatively sensitive to shear force due to their rigid and fragile cell walls and the presence of large vacuoles inside the cells. Appropriate shear force can promote cell growth and enhance metabolism, but too much shear force will cause mechanical damage to cells, reduce cell activity or destroy cell membrane structure, thus affecting cell growth and metabolism. Specific examples: safflower cells (Carthamus tinctorius L.), their tolerance to shear force threshold is low, when the shear force exceeds the certain range, it will affect the growth and metabolism of cells.
b.Mammalian cells, mammalian cells do not have the cell walls, and their membranes are relatively fragile and susceptible to external mechanical forces. In addition, mammalian cells typically have the large size, which makes them more susceptible to shear forces.
c.Aspergillus Niger in citric acid fermentation: As the main producer of citric acid fermentation, Aspergillus citrate is a shear-sensitive microorganism. The yield of citric acid was significantly affected by its morphological characteristics. In the process of citric acid fermentation, excessive shear force may affect the growth and metabolism of Aspergillus Niger, thus affecting the yield of citric acid.
d.Cells at different growth stages: Plant cells respond differently to shearing at different growth stages. For example, Taxus chinensis cells in the logarithmic phase are sensitive to shear forces.
The effect of shear force on microorganisms is a complex process, which is not only related to the species and growth stage of microorganisms, but also related to many factors such as the intensity, duration and environmental conditions of shear force. In practical applications, the shear force needs to be controlled and optimized according to the specific situation to ensure that the growth and metabolism of microorganisms are in the best condition.
4.3 Prokaryotic microorganisms
Prokaryotic microorganisms do not have formed nuclei and lack organelles such as endoplasmic reticulum and Golgi apparatus, which limits their ability and speed of post-translational protein modification and secretion. Therefore, the process of expression must be controlled for implementation.
4.4 Segmented controlled fermentation process
Feeding process, temperature sensitive strain, induction process, mixed strains fermentation, oxygen switching fermentation, substrate switching fermentation, etc., need to be segmented process design, and the segmented process must be controlled in some periods.
5.1 Full program control speed
For example, the fermentation process of some wines pursues low-temperature and long-term fermentation, because the fermentation rate is slow, the microorganisms have enough time to decompose and use the ingredients in the raw materials, thus producing more flavor and aroma substances. These substances can give fermented products the unique flavor and aroma, making the product more attractive and competitive. For example, in the process of wine making, slow fermentation at low temperature is conducive to the formation of alcohol-sweet substances and ester substances, making the wine more mellow and rich aroma.
5.2 Speed control of growth period
During the production of large tanks, full-program speed control may be required for growth-coupled microbial fermentation to lengthen the growth period of the growth curve, so that microorganisms spend more time in the growth period, thereby increasing the expression time.
5.3 Speed control of seed stage
Too fast growth will lead to aging, and sometimes it is necessary to adjust the physical and chemical parameters to fit the inoculation time point,etc.