CN114010626A - Application of citric acid in preparation of product for directionally promoting proliferation of beneficial bacteria in intestinal tract - Google Patents
Application of citric acid in preparation of product for directionally promoting proliferation of beneficial bacteria in intestinal tract Download PDFInfo
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Abstract
The invention discloses application of citric acid in preparation of a product for directionally promoting proliferation of beneficial bacteria in intestinal tracts, and relates to the technical field of biology. The inventor creatively discovers that citric acid can directionally proliferate various beneficial intestinal bacteria, can prepare products for promoting the proliferation of beneficial intestinal bacteria (multiple or single genus), and can prepare medicaments (such as bacteria liquid or capsules for fecal transplantation (FMT)) by utilizing the citric acid based on that many diseases are caused by the reduction of the beneficial intestinal bacteria content, so as to improve or treat the diseases caused by the reduction of the beneficial intestinal bacteria content.
Description
Technical Field
The invention relates to the technical field of biology, in particular to application of citric acid in preparation of a product for directionally promoting the proliferation of beneficial bacteria in intestinal tracts.
Background
Citric acid is an important organic acid, and natural citric acid is widely distributed in nature, and is contained in fruits such as plants of lemon, orange, pineapple and the like, or bones, muscles and blood of animals. The artificially synthesized citric acid is prepared by fermenting sugar-containing materials such as granulated sugar, molasses, starch, and fructus Vitis Viniferae. The citric acid has wide application, has multiple applications in various industries such as food, chemical industry, cosmetics, medicine and the like, simultaneously has rich medicinal value, and has the effects of eliminating and preventing skin pigment from being dark, eliminating phlegm, whitening skin, moistening skin, reducing cholesterol and the like. In addition, citric acid also has the effect of inhibiting various pathogenic bacteria.
The intestinal microorganisms can be functionally divided into three major groups, namely symbiotic bacteria, beneficial bacteria and pathogenic microorganisms, and the three groups keep a dynamic balance in the intestinal tract of a human body. The group of microorganisms has various functions including substance metabolism, biological barrier, immune regulation, host defense and the like, and the intestinal microorganisms not only help the human body to absorb nutrition from food, but also can synthesize amino acid, organic acid, vitamin, antibiotic and the like for us to utilize, and can metabolize the generated toxin to reduce the toxicity to the human body.
Different dietary habits and life styles have great influence on the types of intestinal microorganisms of human bodies, and various diseases of human bodies are closely related to the intestinal microorganisms. Therefore, the intestinal microorganisms and the human body have a mutual and beneficial symbiotic relationship, and play an important role in maintaining the health of the human body. Roseburia has been reported in the literature to be associated with Crohn, gout, cirrhosis, depression and type II diabetes.
FMT is an emerging medical technology and it is one of the most important rings to provide patients with a healthy flora that has an improvement in their disease. The flora condition of the patient can be detected by a second-generation sequencing technology, and higher requirements are provided for accurate FMT. Therefore, the donor intestinal bacteria beneficial to the patient through the directed proliferation are the technologies which are urgently needed by the future FMT, but how to realize the directed proliferation of the beneficial bacteria is a key link.
In view of this, the invention is particularly proposed.
Disclosure of Invention
The invention aims to provide application of citric acid in preparation of a product for directionally promoting the proliferation of beneficial bacteria in intestinal tracts.
Another object of the present invention is to provide a product for promoting the proliferation of beneficial intestinal bacteria, which aims to significantly promote the proliferation of beneficial intestinal bacteria.
The third purpose of the invention is to provide the application of citric acid in preparing the medicine for treating diseases caused by the reduction of the content of beneficial bacteria in the intestinal tract.
The invention is realized by the following steps:
in a first aspect, the present invention provides the use of citric acid in the manufacture of a product for promoting the proliferation of beneficial gut bacteria.
In alternative embodiments, the beneficial gut bacteria include Roseburia, Clostridium, antiaestropes, Turicibacter, Faecalibacterium, and Blautia.
In an alternative embodiment, the gut beneficial bacteria is selected from at least one of Roseburia, Clostridium, antiaestropes, turibacter and Faecalibacterium.
In an alternative embodiment, the gut beneficial bacteria is selected from at least one of Roseburia, Clostridium, Faecalibacterium and Turicibacter.
In an alternative embodiment, the gut beneficial bacteria is selected from at least one of Roseburia, Clostridium and Faecalibacterium.
In alternative embodiments, the gut beneficial bacteria is selected from at least one of Roseburia and Clostridium.
In an alternative embodiment, the gut beneficial bacteria is Clostridium.
In a second aspect, the present invention provides a product for promoting the proliferation of certain specific beneficial bacteria in the gut, comprising citric acid.
In a third aspect, the invention provides the use of citric acid in the manufacture of a medicament for the treatment of a condition resulting from a reduced level of certain beneficial bacteria in the gut;
preferably, the medicament comprises a bacterial solution or capsule for fecal bacteria transplantation.
In alternative embodiments, the disease resulting from a reduced level of certain beneficial gut bacteria comprises at least one of crohn's disease, gout, cirrhosis, depression, and type ii diabetes.
The invention has the following beneficial effects: the inventor creatively discovers that citric acid can play a role in the proliferation of various beneficial bacteria in the intestinal tract, can prepare products for promoting the proliferation of the beneficial bacteria in the intestinal tract, and can prepare medicaments for treating diseases caused by the reduction of the content of certain beneficial bacteria in the intestinal tract by utilizing the citric acid based on that many diseases are caused by the reduction of the content of the beneficial bacteria in the intestinal tract.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained according to the drawings without inventive efforts.
FIG. 1 is a schematic representation of the abundance changes of a bacterial population before and after citric acid intervention in a method of applying citric acid to promote the proliferation of beneficial bacteria.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.
The features and properties of the present invention are described in further detail below with reference to examples.
The inventor finds that the citric acid can promote the proliferation of various beneficial intestinal bacteria and can be applied to the preparation of products for promoting the proliferation of the beneficial intestinal bacteria through long-term research.
Citric acid promotes the proliferation of several beneficial gut bacteria including Roseburia, Clostridium, antianestipes, turkibacter, Faecalibacterium and Blautia. Can play a role in proliferating the flora formed by the above bacteria and has good market application prospect.
In some embodiments, the gut beneficial bacteria is selected from at least one of Roseburia, Clostridium, antiaestropes, Turicibacter, and Faecalibacterium. The inventor finds that citric acid has a remarkable proliferation promoting effect on the above 5 bacteria, and the effect is better than that of Blautia.
In a preferred embodiment, the beneficial gut bacteria is selected from at least one of Roseburia, Clostridium, Faecalibacterium and Turicibacter. The inventor finds that citric acid has a remarkable proliferation promoting effect on the above 4 bacteria, and the effect is better than that of Anaerothrips.
In a more preferred embodiment, the beneficial gut bacteria is selected from at least one of Roseburia, Clostridium and Faecalibacterium, and citric acid has a significant proliferation-promoting effect on the above 3 bacteria, the overall effect being better than that of helicobacter. Although citric acid promotes the highest degree of the Turicibacter, the whole content is very low, and the market application prospect is not ideal.
In a further preferred embodiment, the beneficial intestinal bacteria are at least one selected from Roseburia and Clostridium, citric acid has a remarkable proliferation promoting effect on the above 2 bacteria, and is better than Faecalibacterium, the two beneficial bacteria are better in content after proliferation, and the proliferation rate can reach more than 85 times.
In the most preferred embodiment, the intestinal beneficial bacteria are Clostridium, the multiplication rate of citric acid to Clostridium can reach more than 350 times, and the content of citric acid is high, so that the application prospect is good.
The embodiment of the invention provides a product for promoting the proliferation of beneficial intestinal bacteria, which comprises citric acid, and the product can be a single citric acid agent or a composition formed by citric acid and other components for promoting the proliferation of the beneficial intestinal bacteria.
Specifically, the product can be food, health product or medicine, and the form is not limited.
In some embodiments, the drug may be a bacterial solution or capsule for fecal transplantation, and targeting the donor population of beneficial gut bacteria with citric acid provides more options for accurate FMT.
The embodiment of the invention also provides application of citric acid in preparing a medicine for treating diseases caused by reduction of beneficial bacteria content in intestinal tract, and according to the record of the existing literature, Roseburia is related to Crohn, gout, liver cirrhosis, depression and type II diabetes.
Therefore, citric acid can be used for preparing and forming medicaments for treating diseases such as Crohn, gout, liver cirrhosis, depression, type II diabetes and the like.
Based on the records of the existing documents, the characteristic that citric acid promotes the proliferation of beneficial intestinal bacteria such as Roseburia, Clostridium, Anaerostipes and Turcibacter can be utilized, and the citric acid can be used for preparing and forming a medicament for treating diseases such as Crohn, gout, liver cirrhosis, depression and type II diabetes.
Test example 1
In order to ensure the accuracy of detection, the in-vivo simulation experiment is carried out in vitro by utilizing the intestinal flora of a human body, the human excrement is taken as an intestinal flora sample, the effect of citric acid intervention on the intestinal flora is observed, and the specific operation process is as follows:
(1) preparation of basic culture medium
60g of modified GAM broth powder was weighed and poured into a 1000mL beaker. The measuring cylinder measures 800mL of ultrapure water, pours the ultrapure water into a beaker, and puts the ultrapure water on a magnetic heating stirrer to stir until the ultrapure water is completely dissolved. The above completely dissolved solution was drained to a 1000mL volumetric flask, and the beaker was rinsed with a small amount of ultrapure water and poured into the volumetric flask, and repeated 3 times. The liquid in the volumetric flask was made to a constant volume with ultra pure water, and after closing the cap, the flask was inverted up and down 10 times to ensure thorough mixing of the solution.
Pouring the solution with constant volume into a glass bottle, putting 500mL of culture medium in each 1000mL bottle, unscrewing the bottle cap of the glass bottle with the culture medium, putting the glass bottle into a pulse autoclave, and using a liquid program for 15 minutes at 121 ℃. After sterilization, the bottle cap is screwed down and cooled to room temperature.
(2) Preparation of primary enteric bacteria
A 10g fecal sample was weighed out and an appropriate amount of phosphate buffered saline (pH 7.4) was added to the centrifuge tube and mixed well on a shaker. And then evenly subpackaging the completely mixed samples into new 50mL centrifuge tubes, and adding a proper amount of phosphate buffer solution into each tube.
And (3) filtering the diluted sample by a filter screen of 20 meshes, 50 meshes, 100 meshes and 200 meshes in sequence, collecting filtrate into a centrifugal tube, putting the centrifugal tube into a centrifugal machine, balancing, centrifuging at 6000G and 4 ℃ for 15min, and removing supernatant. After weighing the precipitate, the final concentration is 5%, and the volume is determined by phosphate buffer solution to obtain the intestinal microorganism sample.
(3) Citric acid-GAM cultured intestinal bacteria
Measuring the GAM culture medium, subpackaging into glass tubes with 2mL each, taking citric acid solution (citric acid is commercially available, purity is 99%, and dissolved with phosphate buffer solution to mass percent concentration of 5%), adding into the glass tubes with the GAM culture medium with 2mL each, and using as a test group. Meanwhile, a blank control group is also arranged, each group has 3 repetitions, and the grouping is as follows:
test groups: 2mL of GAM medium +2mL of citric acid solution +1mL of phosphate buffered saline;
control group: 2mL GAM medium +3mL phosphate buffered saline;
the glass tube cap is screwed down and transferred to an anaerobic box transfer box, and disinfection treatment with 84 disinfectant is needed before placement. After the anaerobic tank is placed, the bottle cap is unscrewed to replace oxygen, and the replacement is carried out for 12 hours. The intestinal microorganism samples obtained after the above treatment were inoculated into a test group (citric acid-GAM medium) and a blank control group (GAM medium-phosphate buffered saline) 1mL each tube, respectively. After culturing for 72 hours in an anaerobic chamber, the growth of the flora was observed.
And centrifuging the test group sample and the control group sample at 10000rpm for 3min, discarding the supernatant, treating the precipitate with liquid nitrogen, and respectively determining the abundance of the intestinal flora in the test group and the control group, namely the percentage of different intestinal microorganisms in all detected strains, wherein the determination results of the abundance of part of the intestinal flora are shown in table 1.
TABLE 1 abundance of intestinal flora in different treatment groups
Note: the promotion rate is calculated according to the following formula: (test group mean-control group mean)/control group mean.
The method is characterized in that the abundance changes of partial intestinal flora before and after citric acid intervention are subjected to significance analysis, the analysis result is shown in figure 1, wherein P is less than 0.01, namely the abundance changes have extremely significant difference; p < 0.05, i.e. there was a significant difference in abundance change.
As is clear from table 1 and fig. 1, citric acid has different degrees of growth promoting effects on Roseburia, Clostridium, Faecalibacterium, Blautia, antiaestropes and Clostridium, but has no significant effect on Blautia and antiaestropes, and has a very desirable growth promoting effect on Roseburia, Clostridium and Clostridium, but has a higher application value in terms of content. The citric acid has the most ideal promoting effect on Clostridium in combination of content and promoting rate.
In conclusion, the invention simulates the intestinal environment in vitro, takes the intestinal flora collected from the feces of a healthy human body as a sample, and applies citric acid to directionally promote the proliferation and fermentation of beneficial bacteria (Roseburia, Clostridium and Clostridium), so that the citric acid can be used for preparing and obtaining a medicine to improve or treat diseases such as Crohn, gout, cirrhosis, depression and type II diabetes, thereby providing scientific basis for clinical application and achieving the purpose of accurately preparing the beneficial bacterium agent.
The above is only a preferred embodiment of the present invention, and is not intended to limit the present invention, and various modifications and changes will occur to those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. Application of citric acid in preparation of product for directionally promoting proliferation of beneficial bacteria in intestinal tract is provided.
2. The use of claim 1, wherein the beneficial gut bacteria comprises Roseburia, Clostridium, antiaestropes, turibacter, Faecalibacterium, and Blautia.
3. The use according to claim 1, wherein the beneficial gut bacteria is selected from at least one of Roseburia, Clostridium, antiaestropes, turibacter and Faecalibacterium.
4. Use according to claim 3, wherein the beneficial gut bacteria is selected from at least one of Roseburia, Clostridium, Faecalibacterium and Turcibacter.
5. Use according to claim 4, wherein the beneficial gut bacteria is selected from at least one of Roseburia, Clostridium and Faecalibacterium.
6. Use according to claim 5, wherein the beneficial gut bacteria is selected from at least one of Roseburia and Clostridium.
7. The use according to claim 6, wherein the beneficial gut bacteria is Clostridium.
8. A product for promoting the proliferation of beneficial gut bacteria comprising citric acid.
9. The application of citric acid in preparing medicine for treating diseases caused by the reduction of beneficial bacteria content in intestinal tract;
preferably, the medicament comprises a bacterial solution or capsule for coprophilous fungi transplantation.
10. The use of claim 9, wherein the disease resulting from reduced levels of beneficial gut bacteria comprises at least one of crohn's disease, gout, cirrhosis, depression, and type ii diabetes.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101467595A (en) * | 2007-12-17 | 2009-07-01 | 山东六和集团有限公司 | Fish feed formula added with citric acid and method for processing the same |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101467595A (en) * | 2007-12-17 | 2009-07-01 | 山东六和集团有限公司 | Fish feed formula added with citric acid and method for processing the same |
Non-Patent Citations (1)
| Title |
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| （日）对马琉璃子主编，刘迎、安云晓译, 安徽科学技术出版社 * |
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Application publication date: 20220208 |