Bifidobacterium breve BBr37 with hypoglycemic ability, its application, product and method

Abstract

The present invention relates to a Bifidobacterium breve with blood sugar-lowering ability, its applications, products and methods, belonging to the field of microbial technology. The present invention provides a Bifidobacterium breve strain BBr37 with blood sugar-lowering ability, and its preservation number is CGMCC NO. 32423. The present invention also provides the use of the strain BBr37 in the preparation of hypoglycemic drugs, or health food with auxiliary blood sugar-lowering function, or bacteriostatic agents, or bacteriostasis for non-therapeutic purposes, as well as products and methods based on the strain BBr37. The strain BBr37 of the present invention has strong inhibitory effects on Escherichia coli, Staphylococcus aureus, Salmonella, Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, and Yersinia enterocolitica, and can effectively lower blood sugar and regulate the intestinal flora. Bifidobacterium breve ).

Description

Technical Field

[0001] The present invention belongs to the technical field of microbiology, and particularly relates to a Bifidobacterium breve BBr37 with hypoglycemic ability, and its application, product and method. Background Art

[0002] Type II diabetes is a complex chronic metabolic disease caused by insufficient insulin secretion or abnormal insulin secretion in insulin-sensitive tissues, leading to elevated blood sugar. It has currently become a serious global health problem, and exploring its treatment methods has also become the focus of attention in various countries.

[0003] Bifidobacterium breve ( Bifidobacterium breve ) is a class of Gram-positive bacilli, mainly present in breast milk and the feces of infants and young children, and its abundance gradually decreases or even disappears with age. Bifidobacterium in the human intestine plays an important role in human health. Bifidobacterium is a health marker of the human body and a beneficial bacterial flora with many benefits to the human body.

[0004] Therefore, there is a need in this field to develop Bifidobacterium breve with strong α-glucosidase inhibitory ability, hypoglycemic effect and broad-spectrum antibacterial and other functions. Summary of the Invention

[0005] Based on the above-mentioned needs existing in the prior art in this field, the present invention provides a Bifidobacterium breve BBr37 with hypoglycemic ability, and its application, product and method.

[0006] The technical solution of the present invention is as follows:

[0007] A Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with immunomodulatory function, and its preservation number is CGMCC NO. 32423.

[0008] Use of Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with preservation number CGMCC NO. 32423 in the preparation of functional foods for assisting in reducing blood sugar, and / or, functional foods for assisting in reducing blood lipids, and / or, functional foods for enhancing immune function, and / or, functional foods for regulating intestinal flora, and / or, bacteriostatic agents, and / or, bacteriostasis for non-therapeutic purposes.

[0009] The hypoglycemic effect includes: reducing blood sugar value, glycosylated serum protein, area under the oral glucose tolerance curve, insulin resistance index;

[0010] Preferably, the blood lipid reduction includes: reducing triglyceride, total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol;

[0011] Preferably, the immune enhancement includes: reducing the levels of lipopolysaccharide, tumor necrosis factor TNF-α, and interleukin-6, and / or increasing the level of interleukin-10;

[0012] Preferably, the regulation of the intestinal flora includes: increasing the abundances of the genera Blautia ), Allobaculum ), Bifidobacterium ), Akkermansia ), Faecalibaculum ), unclassified_Muribaculaceae ), and reducing the abundances of the genera Bacteroides ), Aerococcus );

[0013] Preferably, the antibacterial spectrum of the bacteriostasis includes: Escherichia coli, Salmonella, Staphylococcus aureus, Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, Yersinia enterocolitica.

[0014] A hypoglycemia-assisting functional food, comprising functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number of CGMCC NO. 32423.

[0015] A hypolipidemia-assisting functional food, comprising functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number of CGMCC NO. 32423.

[0016] An immune-enhancing functional food, comprising functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number of CGMCC NO. 32423.

[0017] A functional food for regulating intestinal flora, comprising functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number of CGMCC NO. 32423.

[0018] An antibacterial agent, comprising antibacterial active ingredients; the antibacterial active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number of CGMCC NO. 32423.

[0019] The antibacterial agent further includes: excipients.

[0020] An in vitro antibacterial method, using Bifidobacterium breve ( Bifidobacterium breve)BBr37 is used for antibacterial.

[0021] The beneficial effects of the present invention are as follows:

[0022] A strain of bacteria was isolated from infant fecal samples in the present invention, and it was found to have a relatively broad antibacterial ability, and has good inhibitory effects on many pathogenic bacteria including Escherichia coli, Salmonella, Staphylococcus aureus, Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, and Yersinia enterocolitica. After classifying and identifying this strain, it was known that it is a strain of Bifidobacterium breve ( Bifidobacterium breve ), so it was named BBr37. The present invention has proven through a large number of experiments that in addition to its broad antibacterial ability, the BBr37 strain also has excellent immune-enhancing and intestinal flora-regulating functions. The BBr37 strain can significantly reduce the levels of lipopolysaccharide, tumor necrosis factor TNF-α, and interleukin-6 and increase the level of interleukin-10; at the same time, the BBr37 strain can also significantly increase the abundances of the genera Akkermansia ( Blautia ), Allobaculum ( Allobaculum ), Bifidobacterium ( Bifidobacterium ), Akkermansia muciniphila ( Akkermansia ), Faecalibacterium prausnitzii ( Faecalibaculum ), and an unnamed Bacteroidetes ( unclassified_Muribaculaceae ), and reduce the abundances of the genera Bacteroides ( Bacteroides ), Aerococcus ( Aerococcus ). The BBr37 strain can assist in reducing blood sugar by reducing triglycerides, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol, and can assist in reducing blood lipids by reducing triglycerides, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol.

[0023] The preservation information of the Bifidobacterium breve ( Bifidobacterium breve ) BBr37 of the present invention is as follows:

[0024] Preservation number: CGMCC NO. 32423;

[0025] Taxonomic name: Bifidobacterium breve Bifidobacterium breve ;

[0026] Preservation unit: China General Microbiological Culture Collection Center;

[0027] Address of the preservation unit: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing;

[0028] Preservation date: October 30, 2024. Description of the Drawings

[0029] Figure 1Line graph showing the effects of Bifidobacterium breve BBr37 in Experimental Example 5 of the present invention on the food intake and water intake of type II diabetic mice.

[0030] Figure 2 Line graph showing the effects of Bifidobacterium breve BBr37 in Experimental Example 5 of the present invention on the body weight of type II diabetic mice.

[0031] Figure 3 Line graph and bar graph showing the effects of Bifidobacterium breve BBr37 in Experimental Example 6 of the present invention on the blood glucose level and glycated serum protein level of type II diabetic mice.

[0032] Figure 4 Bar graph showing the effects of Bifidobacterium breve BBr37 in Experimental Example 6 of the present invention on the oral glucose tolerance of type II diabetic mice.

[0033] Figure 5 Bar graph showing the effects of Bifidobacterium breve BBr37 in Experimental Example 6 of the present invention on insulin resistance in type II diabetic mice.

[0034] Figure 6 Bar graph showing the effects of Bifidobacterium breve BBr37 in Experimental Example 8 of the present invention on inflammatory factors in type II diabetic mice; wherein, Figure a is the bar graph of lipopolysaccharide (LPS), Figure b is the bar graph of tumor necrosis factor (TNF-α), Figure c is the bar graph of interleukin-6 (IL-6), and Figure d is the bar graph of interleukin-10 (IL-10).

[0035] Figure 7 Bar graph showing the changes in the Chao1 index and Shannon index of the intestinal microbiota of mice in Experimental Example 9 of the present invention.

[0036] Figure 8 Analysis graph of β diversity of the intestinal flora of mice in Experimental Example 9 of the present invention.

[0037] Figure 9 Bar graph showing the abundances of various types of intestinal flora in mice in Experimental Example 9 of the present invention.

[0038] The markings in the figure are shown as follows:

[0039] Figures 1 - 9 The labels for each group in are: NC represents normal group mice, MC represents model group mice, MET represents mice in the drug treatment group, and BBr37 represents mice in the probiotic treatment group. Detailed implementation mode

[0040] The following further describes the specific content of the present invention in detail in combination with specific examples, experimental examples, and drawings, but does not limit the protection scope of the present invention thereto.

[0041] Source of biological material

[0042] I. The Escherichia coli, Staphylococcus aureus, and Salmonella used in Experimental Example 3 were purchased from the Guangdong Provincial Microbial Culture Collection Center; Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, and Yersinia enterocolitica were strains preserved in the laboratory.

[0043] II. The mice used in Experimental Examples 5 - 9 were commercially available.

[0044] "Above" and "below" in this article both include the base number.

[0045] "Bifidobacterium breve ( Bifidobacterium breve ) strain BBr37", "Bifidobacterium breve ( Bifidobacterium breve ) strain BBr37", "Bifidobacterium breve BBr37", "BBr37", "BBr37 strain", and "strain BBr37" in this article all refer to: Bifidobacterium breve ( Bifidobacterium breve ) strain BBr37 with the preservation number CGMCC NO. 32423.

[0046] The first - group examples, Bifidobacterium breve BBr37 of the present invention

[0047] This group of examples provides a Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with immunomodulatory function, and its preservation number is CGMCC NO. 32423.

[0048] Any act of culturing, propagating, fermenting, enriching, producing, preparing, using, inoculating, amplifying, transforming, modifying, reconstructing, selling, or offering to sell a strain of Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423, and / or, the act of combining a strain of Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 with other probiotics, and / or, the antagonism of a strain of Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 against various pathogenic bacteria, and / or, the preparation of bacteriostatic agents, and / or, the preparation of immune - enhancing functional foods, and / or, the preparation of intestinal flora - regulating functional foods all fall within the protection scope of the present invention.

[0049] The other probiotics include, but are not limited to: Lactobacillus plantarum, Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus delbrueckii subsp. lactis, Lactobacillus helveticus, Lactobacillus casei, Lactobacillus crispatus, Lactobacillus fermentum, Lactobacillus gasseri, Lactobacillus johnsonii, Lactobacillus paracasei, Lactobacillus rhamnosus, Lactobacillus salivarius, Saccharomyces cerevisiae, Torulaspora delbrueckii, Candida spp., Wickerhamomyces spp., Pichia spp., Saccharomyces boulardii, Trichosporon beigelii, Schizosaccharomyces pombe, Rhodotorula rubra, Schizosaccharomyces pombe, Bacillus cereus, Brevibacillus laterosporus, Bacillus megaterium, Bacillus mucilaginosus, Bacillus azotofixans, Bacillus sphaericus, Clostridium butyricum, Bifidobacterium adolescentis, Bifidobacterium angulatum, Bifidobacterium animalis, Bifidobacterium asteroides, Bifidobacterium bifidum, Bifidobacterium bovis, Bifidobacterium breve, Bifidobacterium dentium, Bifidobacterium infantis (i.e., Bifidobacterium longum subsp. infantis), Bifidobacterium lactis (i.e., Bifidobacterium animalis subsp. lactis), Bifidobacterium longum, Bifidobacterium pseudocatenulatum, Bifidobacterium thermophilum, and Bifidobacterium thermophilum acidophilus.

[0050] Those skilled in the art can, according to the actual production needs, combine the conventional technical means or basic common sense of the production process in the pharmaceutical field (for example, "Encyclopedia of Pharmaceutical Technology", "Encyclopedia of Food and Food Production", "Lactic Acid Bacteria and Their Fermented Foods", "Research and Application of Microbial Inoculant Technology", etc.) to make routine selections or adjustments of pharmaceutical excipients, and then prepare products of different dosage forms, different storage conditions, and different shelf lives from a strain of Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423. This poses no technical obstacle to those skilled in the art and can be easily achieved.

[0051] Group 2 of the examples, uses of the Bifidobacterium breve BBr37 of the present invention

[0052] This group of examples provides the use of Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number CGMCC NO. 32423 in the preparation of functional foods for assisting in reducing blood sugar, and / or, functional foods for assisting in reducing blood lipid, and / or, functional foods for enhancing immune function, and / or, functional foods for regulating intestinal flora, and / or, bacteriostatic agents, and / or, bacteriostasis for non-therapeutic purposes.

[0053] In specific examples, the reduction of blood sugar includes: reducing blood sugar value, glycated serum protein, area under the oral glucose tolerance curve, insulin resistance index;

[0054] Preferably, the reduction of blood lipid includes: reducing triglyceride, total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol;

[0055] Preferably, the enhancing immunity includes: reducing the levels of lipopolysaccharide, tumor necrosis factor TNF-α, and interleukin-6, and / or increasing the level of interleukin-10;

[0056] Preferably, the regulating intestinal flora includes: increasing the abundances of the genera Blautia ), Allobaculum ), Bifidobacterium ), Akkermansia ), Faecalibaculum ), unclassified_Muribaculaceae ), and reducing the abundances of the genera Bacteroides ), Aerococcus );

[0057] Preferably, the antibacterial spectrum of the antibacterial includes: Escherichia coli, Salmonella, Staphylococcus aureus, Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, Yersinia enterocolitica.

[0058] Group 3 Examples, the hypoglycemic function food of the present invention

[0059] This group of examples provides a hypoglycemic function food. All examples in this group have the following common features: The hypoglycemic function food includes functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number of CGMCC NO. 32423.

[0060] In a further example, the hypoglycemic function food further includes: excipients.

[0061] In a more specific example, the excipients are selected from: solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, antiadhesives, chelating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, clathrates, humectants, absorbents, diluents, flocculants, deflocculants, filter aids, release retardants, etc.

[0062] According to the content of the present invention, for different requirements in actual production applications, combined with the conventional technical means in the field of preparation (for example, "Encyclopedia of Preparation Technology", "Encyclopedia of Food and Food Production", "Lactic Acid Bacteria and Their Fermented Foods", "Research and Application of Microbial Bacterial Agents", etc.), those skilled in the art can select and formulate the above excipients, and prepare Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 into different dosage forms, such as powder, tablet, oral liquid, gel, patch, spray, lotion, granule, etc.

[0063] In a specific embodiment, the dosage form of the functional food for assisting in reducing blood sugar is selected from one or more of the following: powder, tablet, liquid, capsule.

[0064] Group 4 of the embodiments, the functional food for assisting in reducing blood lipid of the present invention

[0065] This group of embodiments provides a functional food for assisting in reducing blood lipid. All embodiments in this group have the following common characteristics: the functional food for assisting in reducing blood lipid includes functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number CGMCC NO. 32423.

[0066] In a further embodiment, the functional food for assisting in reducing blood lipid further includes: excipients.

[0067] In a more specific embodiment, the excipients are selected from: solvent, propellant, solubilizer, cosolvent, emulsifier, colorant, binder, disintegrant, filler, lubricant, wetting agent, osmotic pressure regulator, stabilizer, glidant, flavoring agent, preservative, suspending agent, coating material, fragrance, antiadhesive, chelating agent, penetration enhancer, pH regulator, buffer, plasticizer, surfactant, foaming agent, defoaming agent, thickening agent, clathrate, humectant, absorbent, diluent, flocculant, deflocculant, filter aid, release retarder, etc.

[0068] According to the content of the present invention, for different requirements in actual production applications, combined with the conventional technical means in the field of preparation (for example, "Encyclopedia of Preparation Technology", "Encyclopedia of Food and Food Production", "Lactic Acid Bacteria and Their Fermented Foods", "Research and Application of Microbial Bacterial Agents", etc.), those skilled in the art can select and formulate the above excipients, and prepare Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 into different dosage forms, such as powder, tablet, oral liquid, gel, patch, spray, lotion, granule, etc.

[0069] In a specific embodiment, the dosage form of the auxiliary lipid-lowering functional food is selected from one or more of: powder, tablet, liquid, capsule.

[0070] Group 5 embodiments, the immune-enhancing functional food of the present invention

[0071] This group of embodiments provides an immune-enhancing functional food. All embodiments in this group have the following common features: The immune-enhancing functional food includes functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number CGMCC NO. 32423.

[0072] In a further embodiment, the immune-enhancing functional food further includes: excipients.

[0073] In a more specific embodiment, the excipients are selected from: solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, antiadhesives, chelating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, clathrates, humectants, absorbents, diluents, flocculants, deflocculants, filter aids, release retardants, etc.

[0074] According to the content of the present invention, for different requirements in actual production applications, combined with conventional technical means in the field of preparation (for example, "Encyclopedia of Pharmaceutical Technology", "Encyclopedia of Food and Food Production", "Lactic Acid Bacteria and Their Fermented Foods", "Research and Application of Microbial Inoculant Technology", etc.), those skilled in the art can select and formulate the above excipients, and make Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 into different dosage forms, such as powder, tablet, oral liquid, gel, patch, spray, lotion, granule, etc.

[0075] In a specific embodiment, the dosage form of the immune-enhancing functional food is selected from one or more of: powder, tablet, liquid, capsule.

[0076] Group 6 embodiments, the intestinal flora-regulating functional food of the present invention

[0077] This group of embodiments provides an intestinal flora-regulating functional food. All embodiments in this group have the following common features: The intestinal flora-regulating functional food includes functional active ingredients; the functional active ingredients include: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number CGMCC NO. 32423.

[0078] In a further embodiment, the functional food for regulating intestinal flora further comprises: excipients.

[0079] In a more specific embodiment, the excipients are selected from: solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, antiadhesives, chelating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, clathrates, humectants, absorbents, diluents, flocculants, deflocculants, filter aids, release retardants, etc.

[0080] According to the content of the present invention, for different requirements in actual production applications, combined with conventional technical means in the field of preparation (for example, "Encyclopedia of Pharmaceutical Technology", "Encyclopedia of Food and Food Production", "Lactic Acid Bacteria and Their Fermented Foods", "Research and Application of Microbial Bacterial Agents", etc.), those skilled in the art can select and formulate the above excipients, and make Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 into different dosage forms, such as powder, tablet, oral liquid, gel, patch, spray, lotion, granule, etc.

[0081] In a specific embodiment, the dosage form of the functional food for regulating intestinal flora is selected from one or more of powder, tablet, liquid, capsule.

[0082] Group 7 embodiments, the bacteriostatic agent of the present invention

[0083] This group of embodiments provides a bacteriostatic agent. All embodiments in this group have the following common features: the bacteriostatic agent includes a bacteriostatic active ingredient; the bacteriostatic active ingredient includes: Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number CGMCC NO. 32423.

[0084] In a further embodiment, the bacteriostatic agent further comprises: excipients.

[0085] In a more specific embodiment, the excipients are selected from: solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, antiadhesives, chelating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, clathrates, humectants, absorbents, diluents, flocculants, deflocculants, filter aids, release retardants, etc.

[0086] According to the content of the present invention, for different requirements in actual production applications, combined with conventional technical means in the field of preparation preparation (for example, "Encyclopedia of Preparation Technology", "Encyclopedia of Food and Food Production", "Lactic Acid Bacteria and Their Fermented Foods", "Research and Application of Microbial Bacterial Agent Technology", etc.), those skilled in the art can select and formulate the above excipients, and prepare Bifidobacterium breve BBr37 with the preservation number CGMCC NO. 32423 into different dosage forms, such as powder, tablet, oral liquid, gel, patch, spray, lotion, granule, etc.

[0087] In a specific embodiment, the dosage form of the bacteriostatic agent is selected from one or more of the following: powder, tablet, liquid agent, capsule.

[0088] Group 8 Examples, in vitro bacteriostatic method of the present invention

[0089] This group of examples provides an in vitro bacteriostatic method. All examples in this group have the following common characteristics: using Bifidobacterium breve ( Bifidobacterium breve ) BBr37 with the preservation number CGMCC NO. 32423 for bacteriostasis.

[0090] Experimental Example 1, isolation, screening and identification of Bifidobacterium breve

[0091] (1) Isolation and screening of Bifidobacterium breve

[0092] The collected infant feces were placed in a sterile sampling tube and transported in an ice box. Under sterile conditions, it was serially diluted with 0.85% saline. An appropriate dilution gradient was selected and spread on an LMRS + lithium salt agar plate, and cultured at 37°C for 48 - 72 hours. Suspected single colonies were picked by observing their colony morphology with the naked eye, and they were examined under a microscope and subjected to preliminary screening and purification culture. After purification, it was cultured in an LMRS liquid anaerobic tube at 37°C for 8 - 12 hours. After centrifugation to remove the supernatant, it was resuspended in a sterile 25% glycerol aqueous solution and stored in the strain bank of Wuhan Microkang Probiotics Research Institute.

[0093] (2) Identification of Bifidobacterium breve:

[0094] The target strain screened was cultured in liquid for expansion, the bacterial cells were collected, genomic DNA was extracted, and the 16S rDNA fragment was amplified using the universal primers 27F and 1492R described in paragraph 58 of Chinese invention patent ZL202210478937.4. The PCR amplification product was detected by agarose gel electrophoresis, and the PCR product was sequenced. Among them, the PCR reaction system: 10×buffer 10μL, 10mM dNTP 2μL, 1μL of each of the upstream and downstream primers, 2μL of DNA template, 0.5μL of Taq enzyme, ddH 234 μL of O. Pre-denature at 95 °C for 10 min; then perform 35 cycles of 94 °C for 30 s, 60 °C for 30 s, and 72 °C for 1 min. After completion, extend at 72 °C for 5 min. The PCR products were detected by gel electrophoresis and then sent to Wuhan Kingcare Bioengineering Co., Ltd. for sequencing. The identified gene sequences were aligned in the NCBI database using the BLAST tool. According to the results of molecular biological identification, the Latin name of the strain category is Bifidobacterium breve , and it was determined that the strain is Bifidobacterium breve. The strain was named Bifidobacterium breve BBr37 and sent for preservation. The preservation information is as follows:

[0095] Preservation number: CGMCC NO. 32423;

[0096] Taxonomic name: Bifidobacterium breve Bifidobacterium breve ;

[0097] Preservation unit: China General Microbiological Culture Collection Center;

[0098] Address of the preservation unit: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing;

[0099] Preservation date: October 30, 2024.

[0100] Experimental Example 2. Tolerance experiments in artificial gastric juice and intestinal juice

[0101] Preparation of artificial gastric juice: Prepare 0.5% sodium chloride solution, adjust the pH to 3 with 1 mol / L HCl, add 0.3% pepsin, and after fully dissolving, filter and sterilize with a 0.22 μm microporous membrane for standby.

[0102] Preparation of artificial intestinal juice: Prepare 0.5% NaCl solution, adjust the pH value to 8.0 with 0.1 mol / L NaOH, add 0.1% trypsin, and after fully dissolving, filter and sterilize with a 0.22 μm microporous membrane for standby.

[0103] The test strain was activated for 2 generations and then the bacterial liquid concentration was adjusted to 10 8 CFU / mL. Take 1 mL of the bacterial suspension, centrifuge to collect the bacteria, and separately inoculate into 1 mL of the prepared artificial gastric juice and intestinal juice, mix well, digest at 37 °C, and at the same time, take the digestive juices at 0 h and 3 h to detect the viable bacteria count and calculate the survival rate. The results are shown in Table 1. Among them, the strain survival rate (%) = Nt / N0 × 100%, where N0 represents the viable bacteria count of the strain at 0 h, CFU / mL, and Nt represents the viable bacteria count of the strain at 3 h, CFU / mL.

[0104] Table 1. Tolerance survival rate of Bifidobacterium breve BBr37 to artificial gastric juice and intestinal juice

[0105]

[0106] The experimental results showed that after 3 hours of gastric juice digestion, the survival rate of Bifidobacterium breve BBr37 was 87.96%. After 3 hours of intestinal juice digestion, the survival rate of Bifidobacterium breve BBr37 was 96.83%. This indicated that Bifidobacterium breve BBr37 had good abilities to tolerate gastric acid and intestinal juice.

[0107] Experimental Example 3: Inhibitory ability of Bifidobacterium breve against pathogenic bacteria

[0108] Cool the MRS broth medium containing 1.5% agar to about 55 °C, mix it with the bacterial suspensions of Escherichia coli, Salmonella, Staphylococcus aureus, Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, and Yersinia enterocolitica in a certain proportion, so that the viable count of the indicator bacteria was at the order of 10 6 CFU / mL, and then quickly pour it into the plate pre-placed with Oxford cups. After the medium cooled and solidified, take out the Oxford cups, inject 200 μL of the fermentation broth of Bifidobacterium breve BBr37 strain into each well. After overnight culture at 37 °C, measure the diameter of the inhibition zone.

[0109] Table 2. Inhibition zone diameter (mm) of Bifidobacterium breve against pathogenic bacteria

[0110]

[0111] As can be seen from the above table, Bifidobacterium breve could effectively inhibit intestinal pathogenic bacteria, especially Campylobacter jejuni and Escherichia coli, and the inhibition zone diameters were 28.74 mm and 19.39 mm respectively, showing good inhibitory abilities.

[0112] Experimental Example 4: Inhibitory ability of Bifidobacterium breve against α-glucosidase

[0113] Inoculate Bifidobacterium breve in liquid L-MRS medium, culture it overnight at 37 °C, transfer it to the second generation and culture for 8 hours, then transfer it to the third generation and ferment for 14 hours. Take the fermentation broth for centrifugation, discard the supernatant, resuspend it with physiological saline, and adjust the bacterial suspension concentration to 10 8 CFU / mL for standby. Add 150 μL of PBS solution to 75 μL of p-nitrophenyl-α-D-glucopyranoside (PNPG) solution and 25 μL of the sample to be tested. Place the mixture on ice for 3 minutes, add 50 μL of α-glucosidase solution at 0.2 U / mL and react in a water bath at 37 °C for 15 minutes, then add 1 mL of a concentration of 1 mol / L Na 2 CO 3 as the reaction termination solution, and measure the absorbance value of the reaction solution at 405 nm:

[0114] A: Add sample and α-glucosidase;

[0115] B: Add sample without adding α-glucosidase;

[0116] C: Do not add sample but add α-glucosidase;

[0117] D: Do not add sample and do not add α-glucosidase

[0118] Table 3. Inhibition rate of Bifidobacterium breve BBr37 on α-glucosidase

[0119]

[0120] As can be seen from the above table, Bifidobacterium breve has the ability to inhibit α-glucosidase, indicating that the strain has potential hypoglycemic ability.

[0121] Experimental Example 5. Effects of Bifidobacterium breve on food intake, water intake, and body weight of type 2 diabetic mice

[0122] Male C57BL / 6j mice (5 - 6 weeks old; 19 ± 1 g) were housed in the animal house of Hubei Center for Disease Control and Prevention. The experimental animal use license number is: SCXK (E) 2020 - 0019. The animal experiment was maintained at 20 - 22 °C, humidity 40 - 60%, with a 12h light / 12h dark cycle. Mice had free access to food and water. After one week of adaptation, according to the reference F. Yan, N. Li, J. Shi, H. Li, Y. Yue, W. Jiao, N. Wang, Y. Song, G. Huo and B. Li, Bifidobacterium breve alleviates type 2 diabetes by regulating 2 hepaticglucose and lipid metabolism and gut microbiota in mice, Food Funct, 2019, the mice were induced to develop type 2 diabetes. The mice were divided into 4 groups (n = 12): normal group (NC group), model group (MC group), probiotic group (BBr37 group, 6*10 9 CFU / d), and drug treatment group (MET, 0.2 g / kg / d). The normal group and the model group were gavaged with sterile normal saline, the probiotic group was gavaged with Bifidobacterium breve BBr37 once a day for 35 consecutive days, and the drug treatment group (MET group) was gavaged with metformin hydrochloride once a day for 35 consecutive days. The daily food intake, daily water intake, and weekly body weight of the mice were monitored.

[0123] The typical characteristics associated with type 2 diabetic mice are mainly manifested as increased daily food intake, increased water intake, increased urine output, and decreased body weight. After five weeks of gavage treatment in the probiotic treatment group, the overall symptoms of diabetes were alleviated and improved, and the results are as Figure 1As shown. The therapeutically effective strain controlled the increase in the food and water intake of mice. During daily monitoring of the bedding humidity, the urine volume in the cages of the treatment group showed a tendency to return to normal. From Figure 2 The change in the body weight value of mice shows that the intervention of probiotics has a good stabilizing or rising effect on the decrease in the body weight value of mice, effectively improving the nutrient absorption of type II diabetic mice, and overall showing a tendency to approach the healthy group.

[0124] Experimental Example 6. Effect of Bifidobacterium breve on glucose metabolism in type II diabetic mice

[0125] Before treatment and during the five-week treatment period, tail vein blood was collected weekly to measure the fasting blood glucose value of mice. The measurement results of the fasting blood glucose value (FBG) are as Figure 3 shown on the left. After five weeks of treatment, the glycated serum protein concentration and oral glucose tolerance of mice were measured. Glucose was orally administered to each group of mice, and the blood glucose values were measured at 0 min, 15 min, 30 min, 60 min, 90 min, and 120 min after oral glucose administration. The area under the oral glucose tolerance curve in the oral glucose tolerance test. At the end of the experiment, the serum was taken to measure the serum insulin concentration of mice.

[0126] Experimental steps for oral glucose tolerance: The amount of glucose used in the glucose tolerance test for mice is 2 g per kilogram of body weight (2 g / kg). A glucose solution with an appropriate concentration is prepared according to the experimental requirements, and a 20% glucose solution is prepared with physiological saline. Mice are fasted for 16 hours, and during this period, mice maintain normal drinking water; before starting the glucose tolerance test, the weight of each mouse is weighed. Blood is taken from the tip of the mouse's tail, and the fasting blood glucose is measured with a blood glucose meter. The measured value is regarded as the blood glucose value at 0 min. Glucose is administered by gavage and timing starts from the moment it is completed. The blood glucose values of each mouse are measured at 15, 30, 60, 90, and 120 min.

[0127] The detection results of the weekly blood glucose value of mice are as Figure 3 shown. It can be concluded that the strain can significantly reduce the blood glucose value of diabetic mice during the five-week intervention. The Bifidobacterium breve treatment group showed an effect of reducing blood glucose in the third week, and as the treatment time extended, the blood glucose-lowering effect of mice became more obvious. At the fifth week of treatment, there was a highly significant difference between the fasting blood glucose value of the probiotic group and the model group, and there was no significant difference compared with the healthy group. It shows that the BBr37 strain can effectively reduce the blood glucose value of mice and approach the healthy group. The glycated serum protein index reflects the average blood glucose level in the past 1-3 weeks. As Figure 3 shown on the right. It can be concluded that probiotic treatment effectively reduces the blood glucose value of mice. After five weeks of treatment, the indicators of oral glucose tolerance of mice were measured. According to Figure 4As shown, the area under the oral glucose tolerance curve in the BBr37 treatment group and the model group reached a highly significant level. The experimental results showed that while reducing blood glucose in mice, the BBr37 strain also had a good effect on improving the glucose absorption and metabolism function of diabetic mice.

[0128] The insulin resistance index (HOMA-IR) is a value calculated according to the serum insulin level and fasting blood glucose value, and is an indicator used to judge the degree of insulin resistance. According to Figure 5 it can be concluded that the degree of insulin resistance in the BBr37 strain treatment group and the drug treatment group was significantly improved and relieved, and approached the healthy group.

[0129] The above results indicate that the treatment with the BBr37 strain can effectively improve and enhance the glucose metabolism ability of mice, reduce the blood glucose value of mice, and thus control the development of diabetes in mice.

[0130] Experimental Example 7. Effect of Bifidobacterium breve on blood lipid levels in type 2 diabetic mice

[0131] Index determination was performed on the serum of mice: determination of triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C), as shown in Table 4:

[0132] Table 4. Detection results of total cholesterol, total triglyceride, high-density lipoprotein, and low-density lipoprotein in serum

[0133]

[0134] As can be seen from Table 4, gavage with the BBr37 strain had a certain effect on reducing the blood lipid levels of type 2 diabetic mice. The indicators of total cholesterol and low-density lipoprotein showed good treatment effects and reached a significant difference (p < 0.05) compared with the model group (MC). The BBr37 strain was particularly significant in reducing total cholesterol and low-density lipoprotein in the serum during the treatment of diabetic dyslipidemia.

[0135] The above results can show that the BBr37 strain can effectively reduce the blood lipid levels of type 2 diabetic mice and has a good effect.

[0136] Experimental Example 8. Effect of Bifidobacterium breve on inflammatory factors in type 2 diabetic mice

[0137] The onset of diabetes can cause systemic inflammation in the body. The occurrence of chronic inflammatory diseases can lead to an increase in lipopolysaccharide toxins in the serum of patients, and at the same time promote the production of tumor necrosis factor and interleukin-6. At the end of the experiment, the whole blood of the mice was centrifuged to obtain serum, and an ELISA kit was used to measure the inflammatory factors in the mouse serum, including the concentration levels of lipopolysaccharide (LPS), tumor necrosis factor (TNF-α), interleukin-6 (IL-6), and interleukin-10 (IL-10). The results are shown respectively as Figure 6 shown in a, b, c, and d in Figure 6 . As shown in d in

[0138] , gavage with BBr37 strain reduced the inflammation in the mice and slowed down the occurrence of body inflammation compared with the model group (MC). As an anti-inflammatory factor in the body, interleukin-10 participates in various regulations of immune cells, inflammatory cells, etc.

[0139] Experimental Example 9. Effect of Bifidobacterium breve on the intestinal flora of type 2 diabetic mice

[0140] After the experiment, the mice were sacrificed. After abdominal disinfection, they were dissected, and the cecal contents of each group of mice were taken for microbial diversity analysis. The 16S rDNA V3-V4 hypervariable regions of all bacteria in the samples were sequenced on the Mi Seq Illumina sequencing platform to determine the characteristics of the intestinal microbiota.

[0141] α-diversity reflects the species richness and species diversity of the sample. The Chao1 index and the Shannon index are commonly used judgment indicators for the diversity and abundance of intestinal microbial colonies. As Figure 7 shown, the abundance and diversity of the intestinal flora of type 2 diabetic mice decreased extremely significantly compared with those of the healthy group mice (p < 0.01). According to the Chao1 index analysis, the treatment with BBr37 strain could increase and improve the abundance of the intestinal flora, reaching a significant difference level compared with the model group. There was no significant difference in the effect compared with the drug treatment group (p > 0.05). According to the Shannon index analysis, gavage with BBr37 strain could extremely significantly increase the diversity of the intestinal flora of type 2 diabetic mice and approach the healthy group. The results showed that five weeks of gavage treatment with BBr37 strain could increase and improve the abundance and diversity of the intestinal flora of type 2 diabetic mice.

[0142] β-diversity is used to compare the similarity in species diversity among different samples. The PCA method is a common analysis method that can reflect the differences and distances between samples. PCA uses variance decomposition to reflect the differences in multiple sets of data on a two-dimensional coordinate graph, and the coordinate axes take the two eigenvalues that can best reflect the variance. The closer the distance between two samples, the more similar the composition of these two samples. As Figure 8 shown, both the treatment with probiotics and drugs can gradually show differences between the intestinal flora of mice and the model group and move closer to the healthy group, indicating that the strain can effectively improve the intestinal flora of type II diabetic mice and tend to be normal.

[0143] Metformin, as a first-line drug for the treatment of type II diabetes, its mechanism of action has been continuously studied and revealed. Many studies have shown that metformin can act on type II diabetic mice by changing the intestinal flora. As Figure 9 shown in the results analysis, the genera Mucilaginibacter ( Blautia ) and Allobaculum ( Allobaculum ) were significantly increased in the intestinal flora of mice gavaged with BBr37. Literature reports that Mucilaginibacter can maintain glucose homeostasis and have anti-inflammatory effects. Allobaculum ( Allobaculum ) is a bacterium that can utilize carbohydrates to produce butyric acid, and butyric acid can upregulate intestinal tight junction proteins, which is closely related to human health. As shown in the figure, the gavage treatment for five weeks significantly increased the abundance of the genus Bifidobacterium ( Bifidobacterium ) in mice, indicating that BBr37 can achieve good therapeutic effects by colonizing and metabolizing in the intestine. In addition, the beneficial intestinal flora, Akkermansia muciniphila ( Akkermansia ), Faecalibacterium prausnitzii ( Faecalibaculum ), and unclassified Bacteroidetes ( unclassified_Muribaculaceae ) showed significant increases in abundance in the treatment group of BBr37 strain (p < 0.05). The harmful intestinal flora, Bacteroides ( Bacteroides ), Aerococcus ( Aerococcus ), etc., showed significant decreases in the BBr37 treatment group and the drug treatment group (p < 0.01).

[0144] The above results indicate that: There is intestinal flora disorder in type II diabetic mice. The treatment with Bifidobacterium breve BBr37 can effectively colonize in the mouse intestine, regulate the intestinal flora of type II diabetic mice, significantly reduce the abundance of harmful flora, significantly increase the abundance of beneficial intestinal flora, and improve the species diversity and abundance of intestinal flora. It can effectively control and improve the glucose metabolism, inflammation, and blood lipid levels of type II diabetic mice.

Claims

1. A Bifidobacterium breve BBr37 having an immunomodulatory function, characterized in that: Its deposit number is CGMCC NO. 32423.

2. Use of Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423 in the preparation of functional foods for assisting in lowering blood sugar, and / or functional foods for assisting in lowering blood lipids, and / or functional foods for enhancing immunity, and / or functional foods for regulating intestinal flora, and / or antibacterial agents, and / or antibacterial agents for non-therapeutic purposes.

3. The use of Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423 according to claim 2 in the preparation of functional foods for assisting in lowering blood sugar, and / or functional foods for assisting in lowering blood lipids, and / or functional foods for enhancing immunity, and / or functional foods for regulating intestinal flora, and / or antibacterial agents, and / or antibacterial agents for non-therapeutic purposes, characterized in that: The blood sugar reduction includes: reducing blood sugar level, glycosylated serum protein, area below the oral glucose tolerance line, and insulin resistance index; And / or, the lipid-lowering includes: lowering triglycerides, total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol; And / or, the immunity enhancement includes: reducing the level of lipopolysaccharide, tumor necrosis factor TNF-α, interleukin-6, and / or increasing the level of interleukin-10; And / or, the regulating intestinal flora includes: increasing the abundance of Blautia, Allobaculum, Bifidobacterium, Akkermansia, Faecalibaculum, and unclassified Muribacula, and reducing the abundance of Bacteroides and Aerococcus; And / or, the antibacterial spectrum includes: Escherichia coli, Salmonella, Staphylococcus aureus, Haemophilus parasuis, Haemophilus parainfluenzae, Campylobacter jejuni, and Yersinia enterocolitica.

4. A functional food for assisting in lowering blood sugar, comprising functional active ingredients; characterized in that: The functional active ingredients include: Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423.

5. A functional food for assisting in lowering blood lipids, comprising functional active ingredients; characterized in that: The functional active ingredients include: Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423.

6. A food for enhancing immune function, comprising functional active ingredients; characterized in that: The functional active ingredients include: Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423.

7. A functional food for regulating intestinal flora, comprising functional active ingredients; characterized in that: The functional active ingredients include: Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423.

8. An antibacterial agent, comprising an antibacterial active ingredient; characterized in that: The antibacterial active ingredients include: Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423.

9. An antibacterial agent according to claim 8, characterized in that: Also includes: Auxiliary materials.

10. An in vitro antibacterial method for non-therapeutic purposes, characterized in that: Bifidobacterium breve BBr37 with a preservation number of CGMCC NO. 32423 was used for bacteriostasis.

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