Abstract: Coenzyme Q10 is also known as ubiquinone, molecular formula is C59H90O4, molecular weight is 863, belongs to the fat-soluble type of quinone compounds, under the normal temperature environment will show orange-yellow crystalline objects, melting point is 49 degrees Celsius above zero, no taste, its physiological function is directly determined by the quinone base of the redox characteristics, and isoprene-like side chain features there is a direct link between the reduced state CoQ10 and isoprene monomer belongs to the trans type structure, and the oxidized state CoQ10 and monomer belong to the cis type structure, high activity, strong pharmacological effects. Its physiological functions are directly determined by the redox characteristics of the quinone group and also by the characteristics of the isoprene-like side chain, and the structure of the reduced CoQ10 and the isoprene monomer belongs to the trans type, while that of the oxidized CoQ10 and the monomer belongs to the cis type, which is highly active and has strong pharmacological effects. Currently, microbial fermentation has been used for the production of CoQ10 for its value, and this paper investigates the microbial fermentation method to provide assistance for the subsequent production.
At present, microbial fermentation is used in the production process of Coenzyme Q10, mainly in the microbial organisms to extract the effective material components, in the production process need to be reasonable selection of strains of bacteria, the strains of bacteria for modification and processing, to achieve the final production purpose.
1 Progress of research on rational selection of strains
Although microbial organisms in the natural environment generally contain high levels of CoQ10, the fermentation products are often a mixture of homologues, which can be costly to purify. Therefore, it is important to select suitable strains of bacteria during the fermentation process, e.g. Bacteroides phototrophicus, which contains a high content of CoQ10 and belongs to the phylum Bacteroidetes in the prokaryotic world, where the bacteria of the family Bacteroidaceae are the most ideal for the efficient and high quality production of CoQ10.
2 Progress in genetic modification of strains
2.1 Breeding using techniques of metabolic regulation:
Rongguang [1] experts in the study that the process of CoQ10 microbial synthesis can be dealt with in the form of biosynthesis using aromatic rings, isoprenyl side chains, etc. According to the synthesis pathway, the characteristics of bacterial metabolic regulation, etc., the selection and breeding of fermentation strains of CoQ10 need to focus on the following points: (1) nutritional defects in the type of mutant strains selection and breeding measures.
It has been investigated that carotenoids and CoQ10 are anabolically related to each other by using polyisoprene as a precursor, and that a reduction in the amount of carotenoids produced can accelerate the rate of biosynthesis of CoQ10. Therefore, mutant strains with carotenoid-deficient types can be selected to increase the amount of CoQ10 in the production process. (ii) Selection and breeding of mutant strains resistant to metabolic antagonists. In general, the elimination of inhibitors in the CoQ10 synthesis process can lead to a continuous increase in CoQ10 content, for example, screening for precursors, inhibitors or structural analogues resistant to CoQ10 synthesis, such as daunorubicin mutant strains, which can increase the efficiency of fermentation production by about 15%.
2.2 Creation of genetically engineered strains:
During the actual production, we can use molecular biology technology to find the key enzyme genes of CoQ10 production strains, and use advanced sufficient DNA technology to incorporate such genes into the production strains, so as to copy the key enzyme genes, increase the quantity and form a highly efficient expression pattern, so as to ensure the synthesis performance of CoQ10.
In general, the rate-limiting step in the biosynthesis of CoQ10 in microbial cells is the condensation of hydroxybenzoic acid with polyisoprene pyrophosphate by p-hydroxybenzoic acid-polyisoprene pyrophosphate transferase, and it was learned that this enzyme is capable of forming a wide range of substrate mobility, and according to this principle, it is possible to generate high-quality CoQ10 components by cloning the ubia gene in E. coli and introducing it into PSB in the form of enhanced gene expression. Based on this principle, the ubia gene in E. coli was cloned and introduced into PSB to produce high-quality CoQ10 components through enhanced gene expression.
At the same time, PSB is not a mature recipient strain for genetic engineering, so it needs to be transformed into E. coli as a recipient strain to create a perfect metabolic pathway. Moreover, there are some differences in the length of side chains in various microorganisms, which have different control effects on gene formation, and there are also some differences in the length of side chains of CoQ components, and there is a direct link between the length of side chains and the control of genes, e.g., the ispB of E. coli and the coql of yeast belong to this principle, and E. coli is easy to operate in high-density cultivation, and it has a very mature exogenous gene expression system. E. coli is easy to operate in high-density culture and has a very mature exogenous gene expression system, however, the main component of the synthesized CoQ is CoQ8, so the side chain length gene of CoQ10 cloned from PSB needs to be inactivated in E. coli cells in order to realize the high-quality production of CoQ10 effectively [2].
2.3 Focus on optimizing conditions for fermentation:
The production of CoQ10 requires not only the application of metabolic control techniques, the rational selection of mutant strains and the creation of adequate strain patterns, but also the optimization of fermentation conditions to ensure improved yield and quality.
(1) Enhance the optimization of culture medium. The optimization of fermentation is carried out by using various sources of carbon, nitrogen, growth factors, etc., to ensure the optimization of the composition of the selected medium. For example, metal ions can be used as the culture medium for CoQ10 fermentation production, and the addition of 12.2 mmol/L of magnesium sulfate metal ions and 1.8 mmol/L of iron sulfate metal ions to the culture medium can effectively increase the yield of CoQ10. The addition of 12.2 mmol/L MgSO4 and 1.8 mmol/L FeSO4 can effectively increase the yield of CoQ10, and the precursors can effectively increase the yield by using the metal ions as the medium, which can also control the flow of the synthesized metabolites of the bacterium to a certain extent [3] .
Optimization of culture conditions. Optimization of culture conditions directly affects the effect of fermentation production. For example, the use of stirring and aeration to optimize the culture conditions may promote or inhibit CoQ10 fermentation due to the differences in the strains, so the use of stirring and aeration should be selected according to the characteristics of the strains and their specific conditions during the optimization of culture conditions. Another example is that the optimization of culture conditions should be realized by using light, especially for Rhodobacter sphaeroides, which not only has the characteristics of photosynthesis of specialized anaerobic bacteria, but also has the characteristics of aerobic respiration and fermentation. Therefore, good anaerobic conditions of light should be created in the process of cultivation according to the actual situation, so as to increase the quality and effect of the production and avoid the lowering of the yield due to the effect of the aerobic environment in the darkness. In addition, it is also necessary to strictly control the cultivation time, mainly because in the process of microbial fermentation production of CoQ10, the yield will be increased when the time is increased, so it is necessary to rationally optimize the conditions in accordance with the characteristics and conditions of production.
2.4 Improvement of extraction mode: At present, in the process of microbial fermentation production of CoQ10, the extraction of CoQ10 from microbial cells mainly involves saponification and non-saponification forms, of which the non-saponification form is less than the saponification form of the extract, but will not have a destructive effect on CoQ10. The saponified form is easy to use, but very costly and can easily cause damage to CoQ10.
Huang Yuqing [4] experts in the study put forward to practice the application of saponification extraction technology recommendations, omit the traditional saponification extraction work in the pyrogallic gallic acid add link, ethanol solvent add link, the use of acid crushing cell rapid saponification processing, in reducing production costs at the same time can also achieve industrialization, batch production purposes, with a certain degree of popularization advantages and value [5]. In addition, in recent years, in the process of rapid development of science and technology, but also developed organic solvent stirring broken type, grinding broken type, ultrasonic broken type of extraction technology, achieved good results, especially ultrasonic crushing extraction technology, in the process of application according to the production characteristics of saponification technology and ultrasonic crushing technology integrated with each other, or through the enzyme enzyme digestion form, can enhance the crushing extraction effect, has certain advantages and value. In particular, ultrasonic crushing and extraction technology can be integrated with saponification technology and ultrasonic crushing technology according to the production characteristics, or through the auxiliary form of dissolving enzyme digestion, which can enhance the effect of crushing and extraction, and it has a certain value of popularization.
concluding remarks
In summary, in the process of microbial fermentation production of coenzyme Q10, strains and bacteria should be scientifically selected according to the specific production requirements and characteristics, the selection and cultivation of strains should be done well, the medium should be optimized, the specific cultivation environment and light conditions should be perfected, and advanced extraction technology measures should be scientifically selected to improve the quality and yield of coenzyme Q10 production. In the process of future development, we should strengthen the research and innovation of technology to ensure the production of coenzyme Q10.
References:
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