CN114949202A - Probiotic and protein composition and application thereof in resisting helicobacter pylori - Google Patents
Probiotic and protein composition and application thereof in resisting helicobacter pylori Download PDFInfo
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- CN114949202A CN114949202A CN202210561760.4A CN202210561760A CN114949202A CN 114949202 A CN114949202 A CN 114949202A CN 202210561760 A CN202210561760 A CN 202210561760A CN 114949202 A CN114949202 A CN 114949202A
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- helicobacter pylori
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Abstract
The invention provides a composition of probiotics and protein and application of the composition in resisting helicobacter pylori, belonging to the technical field of microbial preparations. The composition comprises animal bifidobacterium Bb-12, lactobacillus acidophilus NCFM, yolk globulin powder and resveratrol, has good capability of inhibiting helicobacter pylori, can obviously eliminate the helicobacter pylori in a human body, and reduces the occurrence of gastric cancer.
Description
Technical Field
The invention belongs to the technical field of microbial preparations, and particularly relates to a composition of probiotics and protein and application of the composition in resisting helicobacter pylori.
Background
The helicobacter pylori belongs to gram-negative bacteria, has a length of about 2.5-4.0 μm and a width of about 0.5-1.0 μm, and has spiral, S-shaped or gull-shaped thallus, acid resistance and capophily. Helicobacter pylori was first discovered and isolated from the stomach tissue of patients with chronic gastritis in 1983 by Warren and Marshall. Existing studies have shown that the colonization of the gastric mucosa by helicobacter pylori stimulates a strong local and systemic inflammatory and immune response which, together with the toxic substances produced by the bacteria, causes peptic ulcers in 10% to 20% of infected individuals and gastric cancers in 1% to 3% of infected individuals (subebaum S, Michetti P).
Helicobacter pylori is infectious and generally transmitted through the digestive tract, and mainly comprises a feces-oral route and an oral-oral route, wherein the feces-oral route means that feces containing bacteria of an infected person directly or indirectly pollute food or tableware, the person eating the food is infected, and the oral-oral route means that saliva containing bacteria of the infected person pollutes the food or tableware, and the food or the tableware is infected after being eaten. In addition, there is a special approach, i.e. iatrogenic infection, which is mainly caused by infection of helicobacter pylori by other people during examination because the infected people are not completely sterilized after digestive endoscopy.
Nowadays, the common therapy for helicobacter pylori is "triple therapy", which is further developed into "quadruple therapy". The combination of two antibiotics, combined with either a proton pump inhibitor or bismuth citrate, is currently the predominant formulation for "triple" therapy in medicine (continuous administration for one and a half weeks to two weeks). The 'four-combination therapy' is to add bismuth citrate on the basis of two antibacterial drugs and proton pump for continuous administration for 10-14 days. In vitro experiments, many antibiotics have some inhibitory effect on helicobacter pylori, but only a few antibiotics can be used for in vivo treatment. The incomplete and repeated relapse of the conventional therapy of the helicobacter pylori, combined with limited antibiotic selection, causes the helicobacter pylori to be easy to generate mutation and drug resistance, and can also cause the appearance of super bacteria under the condition of poor treatment. This greatly impairs the effectiveness and persistence of the relevant therapy in inhibiting H.pylori, delaying the treatment of the relevant disease (Gerrits et al 2006). In fact, in clinical treatment of diseases, excessive use of antibiotics does not benefit human bodies, and can cause a series of potential hazards to human bodies, and the abuse of antibiotics can influence normal bacteria in human bodiesPopulation distribution and quantity, which may inhibit probiotic growth, even more likely cause a range of endogenous diseases caused by a disturbed microbiota, such as c. Compared with antibiotics, probiotics have obvious advantages in helicobacter pylori treatment, for example, the probiotics have the functions of strengthening the biological barrier of the gastric mucosa, releasing cytotoxin and producing short-chain fatty acid to resist helicobacter pylori infection, competitively inhibiting the colonization of the helicobacter pylori, balancing the levels of various cytokines and the like. Patent CN201910101339.3 discloses a probiotic composition for resisting helicobacter pylori, the composition comprises lactobacillus plantarum (CCTCC NO: M2018268) powder and lactobacillus plantarum metabolite powder as active ingredients, lactobacillus plantarum zymocyte liquid is obtained by performing expanded strain culture, primary seed culture, seed tank culture and fermentation culture on lactobacillus plantarum, helicobacter pylori can be effectively inhibited until helicobacter pylori in a human body is eliminated, gastric cancer is reduced, and no harm is caused to the human body. The patent CN202010574273.2 provides a composite probiotic composition for inhibiting helicobacter pylori and application thereof, wherein the composite probiotic composition comprises the viable count of lactobacillus reuteri of 1-2 multiplied by 10 10 CFU/g, viable count of Lactobacillus johnsonii 3-4 × 10 10 CFU/g, and other components in parts by weight: 80-160 parts of fructo-oligosaccharide, 400 parts of resistant dextrin, 400 parts of galacto-oligosaccharide, 8-12 parts of licorice extract, 4-6 parts of strawberry extract and the balance of maltodextrin, wherein the total amount of the composition is 2000 parts, the composition has strong capability of inhibiting helicobacter pylori, and each added component is beneficial to the field planting of probiotics and improving the activity of the probiotics, so that the stomach environment can be obviously improved, the helicobacter pylori can be effectively eradicated, and the composition has great application potential in the aspect of treating diseases caused by the helicobacter pylori.
Therefore, there is a need to provide a probiotic product with a better anti-helicobacter pylori effect.
Disclosure of Invention
In view of the above problems, the present invention provides a composition of probiotics and protein and its application in resisting helicobacter pylori. The composition comprises animal bifidobacterium Bb-12, lactobacillus acidophilus NCFM, yolk globulin powder and resveratrol, has good capability of inhibiting helicobacter pylori, can obviously eliminate the helicobacter pylori in a human body, and reduces the occurrence of gastric cancer.
In order to achieve the above object, the technical solution of the present invention is as follows:
in one aspect, the present invention provides a composition comprising: bifidobacterium animalis Bb-12, Lactobacillus acidophilus NCFM and egg yolk globulin powder.
Specifically, the composition also comprises resveratrol.
Specifically, the viable count of the animal bifidobacterium Bb-12 is 10 6 -10 10 CFU/mL。
Specifically, the viable count of the lactobacillus acidophilus NCFM is 10 5 -10 9 CFU/mL。
Specifically, the concentration of the egg yolk globulin powder is 0.1-10mg/L, preferably 1 mg/L.
Specifically, the concentration of the resveratrol is 5-20mg/L, preferably 10-15 mg/L.
In another aspect, the invention provides the application of the composition in preparing products for resisting helicobacter pylori.
Further specifically, the product is a health product, food or medicine.
Further specifically, the types of the health care product include, but are not limited to: medicinal liquor, capsules, tablets, medicinal granules, tea products, fruit juice, fruit vinegar, oral liquid, soft capsules, granules, fermented milk products, fermented grain products, fermented bean products, honey paste, distillate, powder, fresh juice, meal replacement powder and the like.
The health product also comprises health product additives.
The nutraceutical additives include, but are not limited to: essence, spice, colorant, sweetener, sour agent, freshener, emulsifier, thickener, antiseptic, antioxidant, nutrition enhancer, etc.
More specifically, the types of food products include, but are not limited to: cookies, dairy products, meal replacements, meat products, sauces, baked goods, yoghurts, ice creams, fermented cereal-based products, fruit juices, rice wine, candies, syrups, canned foods, marinades, condiments, soy products, chocolates, fillings, tea products, puffed foods, and the like.
The food also comprises food additives.
The food additive comprises, but is not limited to, preservatives, acidity regulators, anticaking agents, antifoaming agents, antioxidants, bleaching agents, leavening agents, base materials in gum-based candies, coloring agents, color retention agents, emulsifiers, enzyme preparations, flavoring agents, flour treatment agents, coating agents, moisture retention agents, nutrition enhancers, preservatives, stabilizers and coagulants, sweeteners, thickeners, natural flavors for food, synthetic flavors for food, and the like.
More specifically, the dosage form of the drug includes but is not limited to: a gastrointestinal administration form or a parenteral administration form.
The dosage form of the gastrointestinal administration includes but is not limited to powder, tablets, granules, capsules, solutions, emulsions, suspensions and oils.
The parenteral dosage forms include, but are not limited to: injection, respiratory tract, skin, mucosa, and cavity tract.
The injection administration forms include but are not limited to: intravenous injection, intramuscular injection, subcutaneous injection, intradermal injection, intracavity injection.
The administration form of the respiratory tract comprises but is not limited to spray, aerosol and powder spray.
The skin administration dosage forms include but are not limited to external solution, lotion, liniment, ointment, plaster, paste and patch.
The mucosa administration dosage forms include but are not limited to eye drops, nose drops, eye ointment, gargle, sublingual tablets, sticking tablets and sticking films.
The cavity administration dosage forms include but are not limited to suppository, aerosol, effervescent tablets, drops and dripping pills.
The medicine also comprises pharmaceutically acceptable auxiliary materials.
The pharmaceutically acceptable auxiliary materials include but are not limited to solvents, emulsifiers, disintegrants, solubilizers, antioxidants, pH regulators, osmotic pressure regulators, bacteriostats, diluents, wetting agents, adhesives, film-forming agents and the like.
In still another aspect, the present invention provides an anti-helicobacter pylori product comprising the above composition.
Specifically, the product is a health product, food or medicine.
Further specifically, the types of the health care product include, but are not limited to: medicated liquor, capsule, tablet, granule, tea product, fruit juice, fruit vinegar, oral liquid, soft capsule, granule, fermented milk product, fermented cereal product, fermented bean product, honey paste, distillate, powder, fresh juice, meal replacement powder, etc.
The health product also comprises health product additives.
The nutraceutical additives include, but are not limited to: essence, spice, colorant, sweetener, sour agent, freshener, emulsifier, thickener, antiseptic, antioxidant, nutrition enhancer, etc.
More specifically, the types of food products include, but are not limited to: cookies, dairy products, meal replacements, meat products, sauces, baked goods, yoghurts, ice creams, fermented cereal-based products, fruit juices, rice wine, candies, syrups, canned foods, marinades, condiments, soy products, chocolates, fillings, tea products, puffed foods, and the like.
The food also comprises food additives.
The food additive comprises, but is not limited to, preservatives, acidity regulators, anticaking agents, antifoaming agents, antioxidants, bleaching agents, leavening agents, base materials in gum-based candies, coloring agents, color retention agents, emulsifiers, enzyme preparations, flavoring agents, flour treatment agents, coating agents, moisture retention agents, nutrition enhancers, preservatives, stabilizers and coagulants, sweeteners, thickeners, natural flavors for food, synthetic flavors for food, and the like.
More specifically, the dosage form of the drug includes but is not limited to: a gastrointestinal administration form or a parenteral administration form.
The dosage form of the gastrointestinal administration includes but is not limited to powder, tablets, granules, capsules, solutions, emulsions, suspensions and oils.
Such parenteral dosage forms include, but are not limited to: injection, respiratory tract, skin, mucosa, and cavity tract.
The injection administration forms include but are not limited to: intravenous injection, intramuscular injection, subcutaneous injection, intradermal injection, intracavity injection.
The administration form of the respiratory tract comprises but is not limited to spray, aerosol and powder spray.
The skin administration dosage forms include but are not limited to external solution, lotion, liniment, ointment, plaster, paste and patch.
The mucosa administration dosage forms include but are not limited to eye drops, nose drops, eye ointment, gargle, sublingual tablets, sticking tablets and sticking films.
The cavity administration dosage forms include but are not limited to suppository, aerosol, effervescent tablets, drops and dripping pills.
The medicine also comprises pharmaceutically acceptable auxiliary materials.
The pharmaceutically acceptable auxiliary materials include but are not limited to solvents, emulsifiers, disintegrants, solubilizers, antioxidants, pH regulators, osmotic pressure regulators, bacteriostatic agents, diluents, wetting agents, adhesives, film forming agents and the like.
The preparation method of the medicine, food and health care product can adopt the related preparation methods currently existing in the field and developed in the future. It should be understood that the specific preparation method is not intended to limit the scope of the present application. Whether using currently existing or future developed manufacturing methods, so long as the above microorganisms and resveratrol are included, are within the scope of what is claimed herein.
Compared with the prior art, the invention has the advantages that:
the composition comprises animal bifidobacterium Bb-12, lactobacillus acidophilus NCFM, yolk globulin powder and resveratrol, has better capability of inhibiting helicobacter pylori, can obviously eliminate the helicobacter pylori in a human body, and reduces the occurrence of gastric cancer.
Detailed Description
The present invention will be further illustrated in detail with reference to the following specific examples, which are not intended to limit the present invention but are merely illustrative thereof. The experimental methods used in the following examples are not specifically described, and the materials, reagents and the like used in the following examples are generally commercially available under the usual conditions without specific descriptions.
Experimental Material
1. Bifidobacterium animalis Bb-12, available from Ke Hansen, Inc.
2. Lactobacillus acidophilus NCFM, available from Danisco.
3. Yolk globulin powder: adding deionized water into yolk to obtain yolk diluent, standing for 1-2 hr to obtain supernatant; adding a flocculating agent into the supernatant and uniformly stirring; separating to obtain precipitate and permeate, ultrafiltering the permeate to obtain crude IgY solution, filtering with microporous membrane to obtain concentrated IgY solution, and freeze drying to obtain yolk globulin powder.
3. Resveratrol: purchased from Nanjing Hegu Biotech Ltd.
4. Helicobacter pylori (ATCC 43504): purchased from biotechnology limited of baio bowei, beijing.
BALB/c mice: SPF grade, male, 6-8 weeks old, purchased from Kyork Biotech (Shanghai) Inc.
6. Trypticase soy peptone broth (TSB): purchased from Haibozbiol.
Example 1 an anti-helicobacter pylori composition
The number of living bacteria of Bifidobacterium animalis Bb-12 is 10 8 CFU/mL, viable count of Lactobacillus acidophilus NCFM is 10 7 CFU/mL, yolk globulin powder concentration of 1mg/L, resveratrol concentration of 12 mg/L.
Example 2 an anti-helicobacter pylori composition
The number of living bacteria of Bifidobacterium animalis Bb-12 is 10 8 CFU/mL, viable count of Lactobacillus acidophilus NCFM is 10 7 CFU/mL, yolk globulin powder concentration of 1 mg/L.
Example 3 an anti-helicobacter pylori composition
The number of living bacteria of Bifidobacterium animalis Bb-12 is 10 12 CFU/mL, viable count of Lactobacillus acidophilus NCFM is 10 10 CFU/mL, the concentration of egg yolk globulin powder is 1mg/L, and the concentration of resveratrol is 30 mg/L.
Comparative example 1 an anti-helicobacter pylori product
The concentration of resveratrol was 12 mg/L.
Experimental example 1 bacteriostatic detection of helicobacter pylori
(1) Preparing bacterial liquid: culturing helicobacter pylori to logarithmic phase, collecting 5mL bacterial solution, centrifuging for 5min at 5000g, resuspending the bacterial strain with TSB culture medium, and adjusting the bacterial solution concentration to 1 × 10 6 CFU/mL。
(2) A96-well plate was prepared, and 100. mu.L of the bacterial suspension of step (1) and 95. mu.L of TSB medium were added to each well, and 5. mu.L of the compositions of examples 1 to 3 and comparative example 1 were added to each well, and a blank was prepared by adding 5. mu.L of LDMSO solution, and each treatment was performed in 3 replicates. The 96-well plate was incubated at 37 ℃ with shaking at 150rpm for 48 h.
(3) The OD600 value reflects the concentration of the bacterial liquid, and the bacteriostasis rate (%) is calculated according to the following formula:
(OD600 control bacterial liquid-OD 600 treated bacterial liquid)/OD 600 control bacterial liquid-100%.
The results are shown in table 1 below.
TABLE 1
| Group of | Bacteriostatic ratio (%) |
| Example 1 | 72.5 |
| Example 2 | 57.9 |
| Example 3 | 73.3 |
| Comparative example 1 | 5.2 |
As can be seen from the above table, the anti-H.pylori efficacy can be better demonstrated with the composition described herein.
Experimental example 2.
(1) Preparation of the model
42 mice were selected and administered intragastric mixed antibiotic solutions (ampicillin 10mg/mL, gentamicin 1.2mg/mL, azithromycin 10mg/mL), 0.3 mL/mouse, 1 time/day, for a total of 3 days. After intragastric administration of mixed antibiotics for 7 days, mice were intragastric administered with 0.3mL of freshly cultured helicobacter pylori (1X 10) 9 CFU/mL), 1 time every other day, for a total of 5 times. Mice were fasted for 24h before gavage and for 2h after gavage. Randomly selecting 2 mice when the stomach is irrigated for 2 weeks at the last time, killing the mice, taking antral tissues, adding an RUT reagent, standing the mice at normal temperature, and determining that the reagent turns red within 12 hours to be positive, and determining that the reagent turns red to be negative otherwise. Meanwhile, antrum-stomach tissue is taken, fixed by 4% paraformaldehyde, embedded, sliced, and subjected to HE staining to judge whether infection is positive. The RUT and HE staining results showed positive results, indicating that the modeling was successful.
(2) Grouping and administration of drugs
Mice successfully modeled were randomly divided into 4 groups, which were a model group, an example 1 group, an example 2 group, and a comparative example 1 group, each of which was 10 mice. Unmodeled mice were a blank control group (10). The administration modes of each group are as follows:
blank control group: no intervention is taken.
Model group: the normal saline solution for stomach irrigation is 100 mg/(kg. d) for 4 weeks.
Example 1 group: gavage the composition described in example 1, 100 mg/(kg. d), for 4 consecutive weeks.
Example 2 group: gavage the composition described in example 2, 100 mg/(kg. d), for 4 consecutive weeks.
Comparative example 1 group: gavage the composition described in comparative example 1, 100 mg/(kg. d), for 4 consecutive weeks.
(3) Therapeutic results with helicobacter pylori
After 4 weeks of administration as described above, the mice were sacrificed, the antral tissues of the stomach were taken, stained with RUT reagent, and left to stand at room temperature for 12 hours, where the conversion of the reagent to red color was positive, and negative was negative, i.e., the helicobacter pylori was completely eradicated. The results are shown in Table 2 below.
TABLE 2
| Group of | Total number (only) | Negative animal number (only) | Eradication Rate (%) |
| Blank control group | 10 | 10 | - |
| Model set | 10 | 0 | - |
| Example 1 | 10 | 10 | 100 |
| Example 2 | 10 | 8 | 80 |
| Comparative example 1 | 10 | 0 | 0 |
As can be seen from the content in table 2 above, the composition of the present invention also has a significant inhibitory effect on helicobacter pylori in vivo, and can effectively eliminate helicobacter pylori and reduce the occurrence of gastric cancer.
Experimental example 3 human body experiment
To verify that the composition of the present application was indeed effective in patients with helicobacter pylori infection, the composition of example 1 was prepared into tablets by adding strawberry powder 1mg/mL, fucoidan 0.03mg/mL, dextrin 5mg/mL, mannitol 0.2mg/mL, starch 0.2mg/mL, and magnesium stearate 0.5mg/mL, and then administered orally at 2 g/person/day. The patient's own informed and consented experience was obtained in all experiments.
Selecting 60 cases of urea through hospital 14 C]And (5) detecting through an expiration test, and determining that the patient is positive for the helicobacter pylori. The age is 30-70, and the ratio of male to female is 34: 26. Randomly dividing the patients into two groups, wherein one group is a test group, and orally taking the composition tablets; the other group is' anPlacebo group, orally administered the same drug as the tablet described herein, but without the composition described in example 1 herein. After oral administration for 10-15d, urea [ 2 ] 14 C]And (4) detecting in a breath test.
The test group orally administered the composition described herein, then with urea 14 C]Breath test tests all showed negative, while placebo group remained positive. The compositions described herein are indicated for use in the treatment of helicobacter pylori infection.
The following is a case specific: the patients: yuci clinic department of Jinshui Yuci, Zheng Zhou, 40 years old, Zheng Zhou City 14 C]A positive Hp (+) test wherein the test result in the breath test C80 is that after the above tablet 10d is administered, urea [ 2 ], [ 14 C]Breath test detected C38, and diagnosed as positive Hp (-).
The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the present invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.
Claims (10)
1. A composition characterized by: the composition comprises: bifidobacterium animalis Bb-12, Lactobacillus acidophilus NCFM and egg yolk globulin powder.
2. The composition of claim 1, wherein: the composition also comprises resveratrol.
3. The composition of claim 2, wherein: the viable count of the animal bifidobacterium Bb-12 is 10 6 -10 10 CFU/mL。
4. The composition of claim 2, wherein: the lactobacillus acidophilus NCFM activityThe number of bacteria is 10 5 -10 9 CFU/mL。
5. The composition of claim 2, wherein: the concentration of the egg yolk globulin powder is 0.1-10mg/L, preferably 1 mg/L.
6. The composition of claim 2, wherein: the concentration of the resveratrol is 5-20mg/L, preferably 10-15 mg/L.
7. Use of a composition according to any one of claims 1 to 6 for the manufacture of a product against helicobacter pylori.
8. Use according to claim 7, characterized in that: the product is a health product, food or medicine.
9. An anti-helicobacter pylori product, characterized in that: said product comprising a composition according to any one of claims 1 to 6.
10. The product of claim 9, wherein: the product is a health product, food or medicine.
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