1.1. Affect enzyme activity
The activity of the enzyme needs to be in a certain pH range to play the role, when it is beyond this range, it may inhibit or destroy the structure of the enzyme, thereby reducing the activity of the enzyme. When the pH value inhibits the activity of certain enzymes in the bacteria, it will hinder the metabolism of the bacterial. It is manifested in the following aspects:
① Overacid or overalkali: If the pH value of the medium is too high or too low, it may lead to changes in the structure of the enzyme, thereby inhibiting the activity of the enzyme. Too much acid or too much alkali may also destroy the active center in the enzyme molecule, thus disrupting the catalytic function of the enzyme.
② Enzymatic reaction: In some cases, the pH value can affect the rate of enzymatic reaction. For example, when the dissociation constants of the substrate and the enzyme are different, the change in pH value may affect the binding and dissociation between the enzyme and the substrate, thus affecting the activity of the enzyme.
③ Metabolic pathways: Different microbial species have different metabolic pathways, and pH may affect the enzymes in these metabolic pathways. For example, in the fermentation process of lactic acid bacteria to produce lactic acid, pH value will directly affect the activity and catalytic efficiency of lactic dehydrogenase.
1.2. Affect the charged state of the cell membrane
The charged state of the cell membrane is mainly determined by the distribution and activity of ion exchange proteins on the membrane. At the certain pH value, ion-exchange proteins bind to specific ions to form the charged ion layer on the surface of the cell membrane. In the process of microbial fermentation, it is important to control the appropriate pH value for maintaining the stability and normal physiological function of cell membrane.
When pH changes, ion-exchange proteins on the cell membrane may undergo conformational changes that alter their ability to bind to ions. For example, when pH is elevated, some ion-exchange proteins on the cell membrane may be more inclined to bind to hydrogen ions, making the surface of the cell membrane negatively charged. Conversely, when pH is lowered, some ion-exchange proteins on the cell membrane may be more inclined to detach from hydrogen ions, resulting in the positive charge on the surface of the cell membrane.
The charged state of the cell membrane has the important effect on the growth and metabolism of microorganisms. For example, the charged state of the cell membrane can affect the absorption of nutrients and the excretion of metabolites by the cell. In addition, the charged state of the cell membrane can also affect the interaction between cells, such as in the formation of biofilm , the interaction between cells plays an important role in the stability and function of biofilm.
1.3. Affect dissociation of certain components in the medium
pH has an important effect on the dissociation of some components in fermentation medium. Under certain pH conditions, some components may exist in different forms, which directly affect their solubility and availability. In fermentation medium, some nutrients such as amino acids and nucleic acids have two dissociable forms which are acidity and alkalinity. Under different pH conditions, they may exist in different forms, which directly affects their solubility and availability.
2.1. Medium components
① Carbon source: Excessive carbon source, especially excessive glucose, will lead to the accumulation of organic acids during the fermentation process, so it results in the decrease in pH.
② Nitrogen source: Too much nitrogen source may cause pH to rise, this is because the ammonium ion in the nitrogen source will be released during metabolism,so it results in the rise of pH.
③ Phosphate: The concentration of phosphate in the medium will affect the fermentation pH. High concentration of phosphate may cause pH to drop, while low concentration of phosphate may cause pH to rise.
④ Buffer solution: In some cases, in order to maintain the stability of pH in the process of fermentation, buffer solution is added to the medium. Common buffer solutions are phosphate buffers, acetate buffers and so on.
⑤ In addition, the amount of defoamer, the type and concentrations of physiological acidic substances and physiological alkaline substances will also affect the pH in the fermentation process. Therefore, when selecting and adjusting the composition of the medium, it is necessary to consider comprehensively according to the specific microbial species and fermentation requirements.
2.2. Microbial species
Different types of microorganisms have different pH requirements, and each microorganism has its optimal growth pH value and optimal production pH value. During the process of fermentation, the metabolic activity of microorganisms will change the pH value of the medium, so the optimal pH value needs to be selected and controlled according to different microbial species. For example, the optimal pH for bacteria is usually between 6.5 and 7.5, while the optimal pH for molds and yeasts is usually between 4.0 and 6.0. In addition, the metabolic process of microorganisms is also affected by the pH value, for example, when the pH value is too high or too low, the activity of enzymes may be inhibited or destroyed, thus affecting the growth of microorganisms and product formation. Therefore, in the process of microbial fermentation, it is very important to control the pH value.
2.3. Cultivation conditions
Temperature, humidity, and oxygen concentration have relatively small effects on pH, but they may also have certain effect on pH during fermentation.
① Temperature: Temperature affects pH mainly by affecting enzyme activity and microbial growth rate. In most cases, raising the temperature will promote the growth and metabolism of microorganisms, resulting in the decline in pH. However,when the temperature is too high, it can also lead to the inactivation of enzymes or the death of microorganisms, causing the pH to rise.
② Humidity: Humidity has an important impact on the fermentation process of some microorganisms. For example, for the fermentation process that requires the substrate with low water activity, increasing humidity can improve the solubility of the substrate and microbial absorption of the substrate, thus affecting pH.
③ Oxygen concentration: Oxygen concentration has an important impact on the fermentation process of aerobic microorganisms. Under aerobic conditions, microorganisms will perform aerobic respiration and produce more metabolites, which affects pH. In addition, the change in oxygen concentration may also lead to change in metabolic pathways , so it will affect pH.
2.4. Growth stage of bacteria
The influence of bacteria on fermentation pH at different growth stages is complex and it needs to be monitored and controlled in practical operation. Different bacteria and different culture conditions will affect the change of pH, so it is necessary to make corresponding adjustments according to the specific situation.
① Early stage of bacterial growth: In the early stage of bacteria access to the medium, the rate of bacterial growth is rapid, cells absorb a large number of nutrients, produce a large number of acidic or neutral metabolites, such as lactic acid, acetic acid, etc., resulting in the decline in pH.
② Bacterial logarithmic growth period: With the bacterial growth into the logarithmic growth period, bacterial metabolic activities are vigorous, producing a large number of acidic metabolites, such as acetic acid, butyric acid, etc., while also producing some alkaline metabolites, such as ammonia, urea, etc. Therefore, the pH value may fluctuate at this stage.
③ Late stage of bacterial growth: In the late stage of bacterial growth, bacterial metabolic activity is weakened, the production of metabolic products is reduced, and the pH value may rebound.
④ Stage of product formation: In some fermentation processes, product formation may occur at the late stage of bacterial growth. The pH at this stage may be affected by product formation, for example, in the process of acetone butanol fermentation, the pH declines as acetone butanol accumulates.
The selection and adjustment & control of optimal pH is one of the key steps in microbial fermentation. The optimal pH means that under the condition of this pH value, the growth, metabolism and product formation of microorganisms are the best. However, during the process of fermentation , the pH value may be constantly changing due to changes in the metabolic activity of microorganisms and culture conditions. Therefore, in order to maintain the optimal pH value, the adjustment & control of pH is required.
The following is the optimal pH selection and adjustment & control method for microbial fermentation:
① Select the most suitable pH: According to the growth characteristics of microorganisms and fermentation requirements, it is necessary to select the most suitable pH value. The rate of microbial growth and product formation at different pH values can be measured experimentally, and the best pH value can be selected.
② pH adjustment & control: During the fermentation process, the pH value is adjusted by adding acid or alkali . The commonly used method of adjustment & control is to directly supplement acid or alkali substances, such as H2SO4, NaOH, etc. However, when the pH value deviation is not large, the use of strong acid substances and alkali substances can easily destroy the buffer system, and will cause hydrolysis of the culture solution components, so this method has been less used.
③ Ventilation volume adjustment: pH value can be controlled by adjusting the ventilation volume. This method is mainly used in the individual case of adding more defoamer, and increasing the air flow volume can accelerate the oxidation of fatty acids to form the decline of pH which was caused by the accumulation of fatty acids.
④ Supplement physiological acidity or alkaline salt matrix: pH value is adjusted through metabolism. The addition of physiological acidic substance and alkaline substance not only adjusts the pH value of the fermentation liquid, but also replenishes nutrients and reduces inhibition. The method of replenishment depends on the actual production situation, which can be direct addition, flow addition, multiple flow addition and so on.
⑤ pH value is adjusted in the manner of supplement material: For example, when the pH value rises to exceed the most appropriate value, it means that the bacteria is in the state of hunger, which can be adjusted by adding sugar, and excessive sugar will cause the pH value to decline.
The selection and adjustment & control of the optimal pH for microbial fermentation is one of the important steps in the fermentation process. By selecting the optimal pH value and adopting the appropriate pH adjustment & control method, the growth, metabolism and product formation rate of microorganisms can be optimized, and the fermentation efficiency can be improved.