CN111518741B - Method for improving oral adhesion of lactic acid bacteria - Google Patents
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Abstract
The invention discloses a method for improving the oral adhesion capacity of lactic acid bacteria, which relates to the technical field of biology and comprises the following steps: (1) culturing lactobacillus seed liquid; (2) Inoculating the cultured lactobacillus seed liquid into a liquid culture medium for culture in an inoculation amount of 2-5% (v/v); (3) After the culture is finished, reducing the pH value in the liquid culture medium, and adding sodium lactate for standing culture; (4) After the static culture is finished, the required thalli with high oral cavity adhesive capacity is obtained by centrifugation. According to the invention, the low pH and sodium lactate static culture treatment is carried out after the lactobacillus culture is finished, so that the adhesion and colonization capacity of the lactobacillus in the oral cavity is improved; the method disclosed by the invention is simple to operate, good in experimental repeatability and easy to produce and amplify.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a method for improving the oral adhesion capacity of lactic acid bacteria.
Background
The human oral environment is a complex micro-ecosystem consisting of a plurality of microorganisms, and is the second biggest micro-ecosystem which is researched only next to the intestinal tract at present. The research shows that the microorganisms commonly found in the oral cavity comprise bacteria, fungi, mycoplasma, viruses and the like, and the microorganisms are widely distributed on the buccal mucosa, the back of the tongue, the coronary plaque, the gingival sulcus and the like. The balance of oral micro-ecology is influenced by the symbiosis, antagonism and the like, and the balance of the oral micro-ecology is closely related to the health and diseases of the oral cavity. For example, the generation of oral caries has a clear relationship with the imbalance of oral micro-ecology, and the relationship between streptococcus mutans and dental caries is the most widely studied.
The traditional methods for preventing and treating dental caries mainly comprise a mechanical method, a pharmaceutical method, a diet control method, a re-mining method and the like, but the methods have certain defects, such as the problems that the drug resistance of bacteria is increased easily due to the excessive use of drugs, the diet control is difficult to realize and the like, and therefore, the search for a new prevention and treatment method becomes a hot point for research. Lactic acid bacteria are the earliest microorganisms applied to human beings, and can improve the nutritive value of food, improve the flavor of the food, regulate the balance of body flora, help digestion, improve immunity and the like. In terms of Oral caries control, as found in a study on 138 healthy children aged 2-3 years, probiotic supplementation is effective in reducing the incidence of caries in early childhood (Hedayati-Hajikan et al BMC Oral Health 2015, 15. Another report examined the effect of fluoride in combination with lactobacillus rhamnosus LB21 on caries treatment and found that fluoride in combination with lactobacillus rhamnosus LB21 was effective in shortening the treatment cycle for caries (c.tack sen-Blicks et al care Research,2009, 43.
The lactic acid bacteria must be present in a certain amount to exert their probiotic effect, and successful adhesion to a specific site is the first step in the quantitative proliferation of the lactic acid bacteria. The adhesion ability of lactic acid bacteria is related to various factors, such as: adhesins, cell surface proteins, capsular polysaccharides, and the like. Generally, products containing lactic acid bacteria mainly enter oral cavities through chewing, buccal dissolution and other modes, compared with intestinal digestion, the stay time in the oral cavities is often shorter, if the adhesion capability of the lactic acid bacteria is not strong, the lactic acid bacteria can quickly enter digestive tracts along with swallowing and other actions, so that the lactic acid bacteria cannot be fixedly planted in the oral cavities, cannot be proliferated and can play a beneficial role. On the other hand, the environment of the oral cavity is more complex than that of the intestinal tract, which also increases the difficulty of adhesion of lactic acid bacteria in the oral cavity. At present, in the aspect of oral cavity probiotics, strains with high adhesion performance are mainly bred as potential research objects (Jonathan C.L.Chua et al.Critical reviews in food science and nutrition.2019, DOI: 10.1080/10408398.2019.1669528), and reports are rarely made on a method for improving the adhesion of lactic acid bacteria in the oral cavity, so that the application of the lactic acid bacteria in the aspect of oral cavity dental caries is greatly limited. In contrast, studies on improving the adhesion of lactic acid bacteria in the intestinal tract are more frequent, and the adhesion of lactic acid bacteria to intestinal epithelial cells is effectively improved by changing the components of the culture medium (chinese patent publication No. CN 103275920B). However, it should be noted that the difference between the oral environment and the intestinal environment is large, and the objects, sites and environments for permanent planting are all significantly different, so it is very important to develop a method for improving the adhesion of lactic acid bacteria in the oral cavity.
Disclosure of Invention
The invention aims to provide a method for improving the oral adhesion of lactic acid bacteria, and solves the problem that the existing lactic acid bacteria have poor oral adhesion.
In order to solve the technical problems, the technical scheme of the invention is as follows:
a method for improving the oral adhesion of lactic acid bacteria comprises the following steps:
(1) Culturing lactobacillus seed liquid;
(2) Inoculating the cultured lactobacillus seed liquid into a liquid culture medium for culture in an inoculation amount of 2-5% (v/v);
(3) After the culture is finished, reducing the pH value in the liquid culture medium, and adding sodium lactate for standing culture;
(4) After the static culture is finished, the required thalli with high oral cavity adhesive capacity is obtained by centrifugation.
Preferably, the step (1) of culturing the lactic acid bacteria seed solution specifically comprises: streaking lactobacillus on MRS solid culture medium, performing inverted culture at 36-38 deg.C for 24-28 hr to obtain single colony, selecting single colony, inoculating into MRS liquid culture medium, and performing static culture at 36-38 deg.C for 12-14 hr to obtain lactobacillus seed solution.
Preferably, the step (2) specifically comprises: inoculating lactobacillus seed solution into MRS liquid culture medium at 2-5% (v/v) for culturing at 36-38 deg.C under 200rpm and pH controlled at 5-6 for 20-24 hr.
Preferably, in the step (3), the pH of the liquid culture medium is reduced by HCl, the addition amount of sodium lactate is 3-7% of the mass fraction of the liquid culture medium, and the standing culture time is 2-3h.
Preferably, the centrifugation speed in the step (4) is 4000-6000rpm, and the centrifugation time is 15-30min.
Preferably, the lactic acid bacteria is at least one of lactobacillus plantarum, lactobacillus salivarius, bifidobacterium longum, lactobacillus rhamnosus, lactobacillus acidophilus, bifidobacterium lactis, bifidobacterium bifidum, lactobacillus paracasei, lactobacillus casei, lactobacillus reuteri, lactobacillus crispatus or pediococcus acidilactici.
Preferably, the MRS solid culture medium comprises the following raw materials in percentage by mass:
0.5 to 1.5 percent of peptone, 0.2 to 0.8 percent of beef powder, 0.2 to 0.6 percent of yeast powder, 1 to 3 percent of glucose, 800.05 to 0.15 percent of tween, 0.1 to 0.3 percent of dipotassium phosphate, 0.2 to 0.8 percent of sodium acetate, 0.1 to 0.3 percent of triammonium citrate, 0.01 to 0.03 percent of magnesium sulfate, 0.002 to 0.008 percent of manganese sulfate, 1.0 to 2.0 percent of agar powder and the balance of water.
Preferably, the MRS liquid culture medium comprises the following raw materials in percentage by mass:
0.5 to 1.5 percent of peptone, 0.2 to 0.8 percent of beef powder, 0.2 to 0.6 percent of yeast powder, 1 to 3 percent of glucose, 800.05 to 0.15 percent of tween, 0.1 to 0.3 percent of dipotassium phosphate, 0.2 to 0.8 percent of sodium acetate, 0.1 to 0.3 percent of triammonium citrate, 0.01 to 0.03 percent of magnesium sulfate, 0.002 to 0.008 percent of manganese sulfate and the balance of water.
By adopting the technical scheme, the invention improves the adhesion and colonization capacity of the lactic acid bacteria in the oral cavity by carrying out low pH and sodium lactate static culture after the lactic acid bacteria culture is finished; the method disclosed by the invention is simple to operate, good in experimental repeatability and easy to produce and amplify.
Drawings
FIG. 1 is a graph showing the results of Lactobacillus plantarum adherence KB cell index for examples 1-3 and comparative examples 1, 2 and 8;
FIG. 2 is a graph showing the results of cell indexes of Lactobacillus plantarum adhesion KB cell for examples 1, 4 and 5 and comparative examples 3, 4 and 8;
FIG. 3 is a graph showing the results of Lactobacillus plantarum adherence KB cell index for examples 1, 6 and 7 and comparative examples 5, 6 and 8;
FIG. 4 is a graph showing the results of Lactobacillus plantarum adherence KB cell index for example 1 and comparative examples 7 and 8.
Detailed Description
The following description of the embodiments of the present invention will be made with reference to the accompanying drawings. It should be noted that the description of the embodiments is provided to help understanding of the present invention, but the present invention is not limited thereto. In addition, the technical features involved in the respective embodiments of the present invention described below may be combined with each other as long as they do not conflict with each other.
Example 1
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which takes lactobacillus plantarum as an investigation object, and comprises the following specific operation processes:
(1) Marking a line on a MRS solid culture medium for lactobacillus plantarum preserved in 15% glycerol, carrying out inverted culture at 37 ℃ for 26h until a single colony grows out, selecting the single colony, inoculating the single colony into the MRS liquid culture medium, and carrying out standing culture at 37 ℃ for 14h to obtain lactobacillus plantarum seed liquid;
(2) Inoculating the lactobacillus plantarum seed solution into an MRS liquid culture medium in an inoculation amount of 3%, culturing at 37 ℃ and 200rpm for 23h under the condition that the pH is controlled at 5.5 until the lactobacillus plantarum strain grows completely;
(3) After the lactobacillus plantarum strain is cultured, adding HCl to reduce the pH value of a liquid culture medium to 3.5, adding sodium lactate with the mass fraction of 5% of the liquid culture medium, and standing and culturing for 2.5h;
(4) After the static culture is finished, centrifuging at 5000rpm for 20min to collect the thalli, and obtaining the required thalli with high oral adhesion capacity.
Example 2
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which specifically comprises the following steps:
the remaining steps were the same as in example 1, except that HCl was added in step (3) to lower the pH of the liquid medium to 3.
Example 3
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which specifically comprises the following steps:
the remaining steps were the same as in example 1, except that HCl was added in step (3) to lower the pH of the liquid medium to 4.
Example 4
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which specifically comprises the following steps:
the procedure was as in example 1 except that sodium lactate was added in an amount of 3% by mass based on the total amount of the liquid medium in step (3).
Example 5
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which specifically comprises the following steps:
the rest steps are the same as the method in the example 1, but sodium lactate with the mass fraction of 7 percent of the liquid medium is added in the step (3).
Example 6
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which specifically comprises the following steps:
the rest of the procedure was the same as in example 1 except that the standing culture time in step (3) was 2 hours.
Example 7
The embodiment provides a method for improving the oral adhesion performance of lactic acid bacteria, which specifically comprises the following steps:
the rest of the procedure was the same as in example 1 except that the standing culture time in step (3) was 3 hours.
Example 8
This example provides a method for improving the oral adhesion properties of lactic acid bacteria, which comprises culturing Lactobacillus salivarius, bifidobacterium longum, lactobacillus rhamnosus, lactobacillus acidophilus, bifidobacterium lactis, bifidobacterium bifidum, lactobacillus paracasei, lactobacillus casei, lactobacillus reuteri, lactobacillus crispatus and Pediococcus acidilactici alone in the same manner as in example 1.
Comparative example 1
The remaining steps of this comparative example were the same as in example 1, except that HCl was added in step (3) to lower the pH of the liquid medium to 2.5.
Comparative example 2
The remaining steps of this comparative example were the same as in example 1, except that HCl was added in step (3) to lower the pH of the liquid medium to 4.5.
Comparative example 3
In this comparative example, the procedure was carried out in the same manner as in example 1 except that in step (3), sodium lactate was added in an amount such that the amount of the liquid culture medium was 2%.
Comparative example 4
In this comparative example, the procedure was carried out in the same manner as in example 1 except that sodium lactate was added in an amount of 8% by mass in the liquid medium in step (3).
Comparative example 5
The remaining steps of this comparative example were the same as those in example 1 except that the standing culture time in step (3) was 1 hour.
Comparative example 6
The remaining steps of this comparative example were the same as those in example 1, except that the standing culture time was set to 4 hours.
Comparative example 7
The comparative example refers to an authorization publication No. CN103275920B, and the patent name is a method for improving the ability of lactobacillus to adhere to intestinal epithelial cells, and the specific steps are as follows:
(1) Inoculating lactobacillus plantarum preserved in 15% glycerol into an MRS liquid culture medium, performing static culture at 37 ℃ for 6 hours, dipping a small amount of culture solution by an inoculating needle, performing streak-line reverse culture at 37 ℃ on the MRS solid culture medium for 26 hours until a single colony grows out, selecting the single colony, inoculating the single colony into the MRS liquid culture medium, and performing static culture at 37 ℃ for 10 hours to obtain a seed culture solution;
(2) Inoculating the seed culture solution into MRS liquid culture medium containing 8% NaCl in an inoculation amount of 3%, and performing static culture at 37 ℃ for 10h;
(3) The cells were collected by centrifugation at 5000rpm for 5 min.
Comparative example 8
This comparison provides a method, specifically as follows:
(1) Marking a line of lactobacillus plantarum preserved in 15% glycerol on an MRS solid culture medium, carrying out inverted culture at 37 ℃ for 26h until a single colony grows out, selecting the single colony, inoculating the single colony into the MRS liquid culture medium, and carrying out standing culture at 37 ℃ for 14h to obtain a seed culture solution;
(2) Inoculating the seed culture solution into an MRS liquid culture medium by 3 percent of inoculation amount, culturing at 37 ℃ and 200rpmp for 23h under the pH control of 5.5 until the growth of the strain is finished;
(3) The cells were collected by centrifugation at 5000rpm for 20 min.
The process differences for examples 1-8 and comparative examples 1-8 are shown in the following table:
adhesion test of Lactobacillus plantarum
Washing the collected Lactobacillus plantarum with 0.5% physiological saline for 3 times, re-suspending the cells with physiological saline, and adjusting the cell concentration to 5 × 10 7 CFU/mL. Oral KB cells were cultured in 1640 cell culture medium containing 10% heat-inactivated newborn calf serum, and the cell concentration was 5% CO at 37 ℃ in the medium 2 Adjusting the cell concentration to 5 × 10 after incubation in a constant temperature incubator 5 CFU/mL, inoculating KB cells into 6-well plates covered with glass slides overnight for culture until complete adherence, removing culture medium by aspiration, rinsing with fresh culture medium for 2 times, and adding 5 × 10 7 CFU/mL of the cells at 37 ℃ and 5% CO 2 After incubation at constant temperature for 2 hours, the cells were washed 3 times with PBS solution to remove non-adhered cells. First of allGram staining is carried out after alcohol is fixed for 15min, 50 cells are randomly selected under an oil lens, the number of bacteria adhered to the cells is calculated, and the average number of adhered bacteria on each cell is the adhesion index.
By taking the adhesion performance of lactobacillus plantarum of a conventionally cultured control group as a control, as shown in fig. 1, the influence of different pH values on the adhesion index of lactobacillus plantarum to oral KB cells is compared, and the adhesion index of lactobacillus plantarum to oral KB cells is found to be greatly improved after the method provided by the invention is adopted, which indicates that the adhesion capacity of lactobacillus plantarum to oral KB cells can be improved by the method provided by the invention. In addition, the results of comparative examples 1 to 3 and comparative examples 1 and 2 show that the results of examples 1 to 3 are superior to those of comparative examples 1 and 2, and it can be seen that the effect of promoting adhesion of the strain to the oral KB cells is not significant when the pH is out of the range disclosed in the present invention. The reason is probably that the lactobacillus has certain low-acidity tolerance, but when the environmental acidity is too low, the growth capacity of the strain is greatly attenuated, so that the activity of the strain is reduced, and when the environmental pH value is too high, the environmental pH value is more suitable for the growth of the strain, and the adhesion performance of the strain cannot be improved. In addition, as a result of comparing examples 1 to 3, the effect of example 1 to improve the adhesive property of lactobacillus plantarum is most significant.
Compared with the effect of different sodium lactate concentrations on the adhesion index of lactobacillus plantarum to oral cavity KB cells, the adhesion index of lactobacillus plantarum to oral cavity KB cells is greatly improved by using the method disclosed by the invention as shown in figure 2 by taking the adhesion performance of the conventionally cultured lactobacillus plantarum in a control group as a control, which indicates that the adhesion ability of lactobacillus plantarum to oral cavity KB cells can be improved by the method disclosed by the invention. In addition, comparing the results of examples 1, 4, 5 and comparative examples 3 and 4, it was found that the results of examples 1, 4, 5 were superior to those of comparative examples 3 and 4, and it can be seen that the effect of promoting adhesion of the strain to oral cavity KB cells was not significant when the sodium lactate concentration was outside the range disclosed in the present invention. It is presumed that the improvement in adhesion performance of the strain cannot be induced when the sodium lactate concentration is too low, while the reduction in strain viability and the reduction in adhesion performance of the strain may be caused when the sodium lactate concentration is too high by inhibiting the growth of the strain. In addition, as a result of comparing examples 1, 4, and 5, the effect of example 1 to improve the adhesion property of lactobacillus plantarum was most significant.
The adhesion performance of lactobacillus plantarum in a conventionally cultured control group is used as a control, as shown in fig. 3, and compared with the influence of different culture times on the adhesion index of lactobacillus plantarum to oral cavity KB cells, the adhesion index of lactobacillus plantarum to oral cavity KB cells is greatly improved after the method disclosed by the invention is adopted, which indicates that the method disclosed by the invention can improve the adhesion capacity of lactobacillus plantarum to oral cavity KB cells. In addition, the results of comparative examples 1, 6 and 7 and comparative examples 5 and 6 show that the results of examples 1, 6 and 7 are superior to those of comparative examples 5 and 6, and it can be seen that the effect of promoting adhesion of the strain to the oral KB cells is not significant when the range of the static culture time is not within the range disclosed in the present invention. It is presumed that the improvement of the adhesion property of the strain cannot be induced when the standing culture time is too short, and the growth of the strain is inhibited when the standing culture time is too long, which may decrease the viability of the strain and, on the contrary, the adhesion property of the strain. In addition, as a result of comparing examples 1, 6, and 7, the effect of example 1 to improve the adhesion property of lactobacillus plantarum was most significant.
In addition, as shown in fig. 4, by comparing the experimental results of the method of the present invention with the previously reported method for promoting the adhesion of lactic acid bacteria to intestinal epithelial cells, it is found that the method of the present invention can effectively improve the adhesion performance of the strain to oral KB cells, whereas the previously reported method for promoting the adhesion of lactic acid bacteria to intestinal epithelial cells cannot significantly improve the adhesion performance of lactobacillus plantarum to oral KB cells. The reason is that the difference between the oral cavity environment and the intestinal environment is large, and objects, sites and the like planted by the oral cavity environment and the intestinal environment are obviously different, which shows that the method is more favorable for improving the adhesion performance of the lactobacillus to the oral cavity.
Comparative test of adhesion properties of different lactic acid bacteria
The method of example 1 was used to perform stationary culture on lactobacillus salivarius, bifidobacterium longum, lactobacillus rhamnosus, lactobacillus acidophilus, bifidobacterium lactis, bifidobacterium bifidum, lactobacillus paracasei, lactobacillus casei, lactobacillus reuteri, lactobacillus crispatus and pediococcus acidilactici, and the results are shown in the following table comparing the effect of the method on the improvement of the adhesion performance of different lactic acid bacteria after conventional culture:
| bacterial strains | Adhesion index improvement factor | P value |
| Lactobacillus plantarum (control) | 2.95±0.32 | — |
| Lactobacillus salivarius | 3.09±0.42 | 0.667 |
| Bifidobacterium longum | 2.45±0.21 | 0.087 |
| Lactobacillus rhamnosus | 2.87±0.19 | 0.455 |
| Lactobacillus acidophilus | 3.16±0.45 | 0.068 |
| Bifidobacterium lactis | 2.53±0.31 | 0.181 |
| Bifidobacterium bifidum | 2.63±0.27 | 0.155 |
| Lactobacillus paracasei | 2.33±0.27 | 0.447 |
| Lactobacillus casei | 3.30±0.32 | 0.173 |
| Lactobacillus reuteri | 2.13±0.21 | 0.368 |
| Lactobacillus crispatus | 2.81±0.24 | 0.395 |
| Pediococcus acidilactici | 2.15±0.30 | 0.929 |
From the above results, it can be seen that the adhesion of different lactic acid bacteria to oral KB cells can be effectively improved by the method of the present invention. In addition, statistical analysis shows that the improvement effect of the method on different strains is not significantly different from the improvement effect on lactobacillus plantarum (P > 0.05), and the method is applicable to the strains.
The embodiments of the present invention have been described in detail with reference to the accompanying drawings, but the present invention is not limited to the described embodiments. It will be apparent to those skilled in the art that various changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, and the scope of protection is still within the scope of the invention.
Claims (7)
1. A method for improving the oral adhesion capability of lactic acid bacteria, which is characterized by comprising the following steps: the method comprises the following steps:
(1) Culturing lactobacillus seed liquid;
(2) Inoculating the cultured lactobacillus seed liquid into a liquid culture medium for culture in an inoculation amount of 2-5% (v/v);
(3) After the culture is finished, reducing the pH value in the liquid culture medium, and adding sodium lactate for standing culture;
(4) After the static culture is finished, centrifuging to obtain the required thalli with high oral cavity adhesion capacity;
the lactobacillus is at least one of lactobacillus plantarum, lactobacillus salivarius, bifidobacterium longum, lactobacillus rhamnosus, lactobacillus acidophilus, bifidobacterium lactis, bifidobacterium bifidum, lactobacillus paracasei, lactobacillus casei, lactobacillus reuteri, lactobacillus crispatus or pediococcus acidilactici;
when the lactic acid bacteria are lactobacillus plantarum, in the step (3), the pH of the liquid culture medium is reduced to 3, 3.5 or 4, the addition amount of sodium lactate is 3-7% of the mass fraction of the liquid culture medium, and the standing culture time is 2-3 h;
when the lactic acid bacteria are other bacteria, in the step (3), the pH of the liquid culture medium is reduced to 3.5, the addition amount of the sodium lactate is 5% of the mass fraction of the liquid culture medium, and the standing culture time is 2.5 h.
2. The method for improving the oral adhesion of lactic acid bacteria according to claim 1, wherein: the step (1) of culturing the lactobacillus seed solution specifically comprises the following steps: streaking lactobacillus on MRS solid culture medium, performing inverted culture at 36-38 deg.C for 24-28 hr to obtain single colony, selecting single colony, inoculating into MRS liquid culture medium, and performing static culture at 36-38 deg.C for 12-14 hr to obtain lactobacillus seed solution.
3. The method of improving the oral adhesion of lactic acid bacteria according to claim 1, wherein: the step (2) specifically comprises the following steps: inoculating lactobacillus seed solution into MRS liquid culture medium at 2-5% (v/v) for culturing at 36-38 deg.C under 200rpm and pH controlled at 5-6 for 20-24 hr.
4. The method of improving the oral adhesion of lactic acid bacteria according to claim 1, wherein: the pH of the liquid medium is lowered by HCl in step (3).
5. The method for improving the oral adhesion of lactic acid bacteria according to claim 1, wherein: in the step (4), the centrifugation speed is 4000-6000rpm, and the centrifugation time is 15-30min.
6. The method for improving the oral adhesion of lactic acid bacteria according to claim 2, wherein: the MRS solid culture medium comprises the following raw materials in percentage by mass:
0.5 to 1.5 percent of peptone, 0.2 to 0.8 percent of beef powder, 0.2 to 0.6 percent of yeast powder, 1 to 3 percent of glucose, 0.05 to 0.15 percent of tween 80, 0.1 to 0.3 percent of dipotassium phosphate, 0.2 to 0.8 percent of sodium acetate, 0.1 to 0.3 percent of triammonium citrate, 0.01 to 0.03 percent of magnesium sulfate, 0.002 to 0.008 percent of manganese sulfate, 1.0 to 2.0 percent of agar powder and the balance of water.
7. The method for improving the oral adhesion of lactic acid bacteria according to claim 2 or 3, wherein: the MRS liquid culture medium comprises the following raw materials in percentage by mass:
0.5 to 1.5 percent of peptone, 0.2 to 0.8 percent of beef powder, 0.2 to 0.6 percent of yeast powder, 1 to 3 percent of glucose, 0.05 to 0.15 percent of tween 80, 0.1 to 0.3 percent of dipotassium phosphate, 0.2 to 0.8 percent of sodium acetate, 0.1 to 0.3 percent of triammonium citrate, 0.01 to 0.03 percent of magnesium sulfate, 0.002 to 0.008 percent of manganese sulfate and the balance of water.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101198689A (en) * | 2005-04-15 | 2008-06-11 | 北卡罗来纳州大学 | Methods and compositions to modulate adhesion and stress tolerance in bacteria |
| CN103275920A (en) * | 2013-05-16 | 2013-09-04 | 江南大学 | Method for improving capacity of lactobacillus adhered to epithelial cells of intestinal tract |
| JP2015073483A (en) * | 2013-10-09 | 2015-04-20 | 独立行政法人農業・食品産業技術総合研究機構 | Lactic acid bacteria which has adhesiveness to alimentary canal mucosa |
-
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101198689A (en) * | 2005-04-15 | 2008-06-11 | 北卡罗来纳州大学 | Methods and compositions to modulate adhesion and stress tolerance in bacteria |
| CN103275920A (en) * | 2013-05-16 | 2013-09-04 | 江南大学 | Method for improving capacity of lactobacillus adhered to epithelial cells of intestinal tract |
| JP2015073483A (en) * | 2013-10-09 | 2015-04-20 | 独立行政法人農業・食品産業技術総合研究機構 | Lactic acid bacteria which has adhesiveness to alimentary canal mucosa |
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Address after: 215000 No. 1033, long Qiao Road, Wujiang economic and Technological Development Zone, Suzhou, Jiangsu Applicant after: Weikang probiotics (Suzhou) Co.,Ltd. Address before: 215000 No. 1033, long Qiao Road, Wujiang economic and Technological Development Zone, Suzhou, Jiangsu Applicant before: JIANGSU WECARE BIOTECHNOLOGY Co.,Ltd. |
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Denomination of invention: A method for improving the oral adhesion ability of lactic acid bacteria Effective date of registration: 20230705 Granted publication date: 20221108 Pledgee: China Construction Bank Corporation Suzhou Yangtze River Delta integration Demonstration Zone Branch Pledgor: Weikang probiotics (Suzhou) Co.,Ltd. Registration number: Y2023980047337 |