CN112914103A - Probiotic composition for resisting helicobacter pylori infection and application thereof - Google Patents
Probiotic composition for resisting helicobacter pylori infection and application thereof Download PDFInfo
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- CN112914103A CN112914103A CN202110198819.3A CN202110198819A CN112914103A CN 112914103 A CN112914103 A CN 112914103A CN 202110198819 A CN202110198819 A CN 202110198819A CN 112914103 A CN112914103 A CN 112914103A
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Abstract
The invention provides a probiotic composition for resisting helicobacter pylori infection and application thereof, wherein the probiotic composition for resisting helicobacter pylori infection comprises: lactobacillus acidophilus, lactobacillus plantarum, bifidobacterium longum and prebiotics; wherein the Lactobacillus acidophilus is named as Lactobacillus acidophilus LA85, is preserved in the institute of microbiology (CGMCC) of China academy of sciences at 7-month and 20-month in 2020, and has the preservation number of CGMCC NO. 1.12735. The probiotic composition for resisting helicobacter pylori infection can effectively inhibit the proliferation of helicobacter pylori, reduce the planting amount of the helicobacter pylori in the stomach, relieve the inflammatory side reaction caused by helicobacter pylori infection in serum, and regulate the flora balance in the intestinal tract by the mutual matching of probiotics and prebiotics, and has wide application value.
Description
Technical Field
The invention belongs to the technical field of nutritional health-care food, and particularly relates to a helicobacter pylori infection resisting probiotic composition and application thereof.
Background
Helicobacter pylori is a unipolar, multifilamentary, helically-curved gram-negative bacterium, whose infection is closely associated with diseases such as functional dyspepsia, chronic gastritis, peptic ulcer, and gastric adenocarcinoma. In 1994 WHO has listed helicobacter pylori as the first causative agent of gastric cancer.
Antibiotic therapy is a standard scheme for clinically treating helicobacter pylori, and a triple method, namely a strategy of combining a Proton Pump Inhibitor (PPI) and two antibiotics or combining bismuth salt and the two antibiotics, is adopted for treatment, and main antibiotic medicaments comprise amoxicillin, clarithromycin, metronidazole and the like. However, with the continuous increase of the drug resistance rate of helicobacter pylori to antibacterial drugs, the increase of the side effects of antibiotics and other problems, the cure rate of the existing scheme is continuously reduced, even more serious adverse reactions are caused, such as antibiotic-associated enteritis, the occurrence of drug-resistant strains and the like, and the treatment repeatability is caused. Therefore, many researchers have recently explored new approaches to combat helicobacter pylori infection, such as the use of non-antibiotic agents, such as probiotics, antioxidants, and plant extracts (e.g., tea extracts, apple peel extracts) to improve or complement existing treatment regimens. Among them, probiotics are receiving wide attention in eradicating or alleviating helicobacter pylori infection because of their advantages of regulating microecology, reducing adverse effects of antibiotics, and the like.
There are several reports of the use of probiotics for the treatment or alleviation of helicobacter pylori. Analysis of the eradication rate of helicobacter pylori by Lexus et al (Lexus et al, Bifidobacterium tetragenous Living bacteria combination triple and quadruple therapy [ J ] Chinese and foreign medical treatment 2014, 33(1): 100-101.) the results show that the treatment effects of triple therapy, quadruple therapy and Bifidobacterium combination quadruple and triple therapy on helicobacter pylori can be eradicated by combining the traditional therapy with the bifidobacterium, the effect of the Bifidobacterium combination quadruple therapy is best, and the eradication rate can reach 94.7%. The probiotics can secrete helicobacter pylori competitive adhesion receptors and metabolize antibacterial substances, and the helicobacter pylori colonization on the gastric mucosa is prevented by stimulating the expression of adhesion proteins and stabilizing the gastric mucosa. The method has obvious effect, but antibiotics are used in the triple therapy and the quadruple therapy, which may increase the drug resistance of the helicobacter pylori and may cause adverse reactions such as gastrointestinal dysfunction, dysbacteriosis and the like. In addition, the therapeutic effect of using only bifidobacterium is limited, and if a plurality of probiotics are used in combination, the cure rate is increased.
CN109806222A discloses a toothpaste for inhibiting helicobacter pylori and a preparation method thereof, wherein the raw materials comprise sorbitol, deionized water, glycerol, silicon dioxide, sodium lauryl sulfate, essence, xanthan gum, sodium hydroxymethyl cellulose, tetrasodium pyrophosphate, saccharin sodium, methylparaben, frankincense, liquorice, evodia rutaecarpa, brucea javanica, ginseng, moringa oleifera, radix scutellariae extract, oral cavity probiotics, lactoferrin and lysozyme. The product can effectively remove helicobacter pylori in oral cavity, effectively prevent the oral transmission route of helicobacter pylori, but can not remove helicobacter pylori in stomach, and can not eradicate helicobacter pylori.
CN1021174450A discloses a Lactobacillus plantarum for resisting helicobacter pylori infection and application thereof, a Lactobacillus plantarum CGMCC No.4286 is obtained by breeding, has the inhibiting effect on the growth and urease activity of the helicobacter pylori, has stronger inhibiting effect on helicobacter pylori adhesion human gastric epithelial cell SGC7901, and has the effect of preventing or reducing the infection degree of mice infected with the helicobacter pylori. Side reactions such as inflammation, dysbacteriosis and the like are common after helicobacter pylori infection, and the scheme is not researched aiming at the side reactions; in the preparation process, the stability and other problems of the probiotics in the storage process are not fully considered in the scheme, so that the loss of the viable count is easy to cause, and the effect is weakened.
Currently, there are still improvements that can be made in the methods of inhibition and treatment of helicobacter pylori. It is an urgent problem to provide an effective method for inhibiting helicobacter pylori and alleviating side effects such as dysbacteriosis caused by helicobacter pylori infection.
Disclosure of Invention
Aiming at the defects and actual requirements of the prior art, the invention provides a probiotic composition for resisting helicobacter pylori infection and application thereof.
In order to achieve the purpose, the invention adopts the following technical scheme:
in a first aspect, the present invention provides a probiotic composition against helicobacter pylori infection, comprising: lactobacillus acidophilus, lactobacillus plantarum, bifidobacterium longum and prebiotics;
the Lactobacillus acidophilus is named Lactobacillus acidophilus LA85, is preserved in the institute of microbiology (CGMCC) at 7-20 days of 2020, has the address of No. 3 Xilu 1 Beijing Kogyo of the Chaoyang area, and has the preservation number of CGMCC NO. 1.12735.
In the invention, the probiotics have the functions of inhibiting helicobacter pylori infection, improving the immunity of the organism and maintaining the balance of intestinal flora; the prebiotics can stimulate the proliferation or activate the metabolism of beneficial bacteria in the intestinal tract, inhibit the growth of exogenous pathogenic bacteria and putrefying bacteria, improve the environment of the intestinal tract and promote the absorption of minerals by the intestinal tract. The probiotics and the prebiotics are matched with each other, so that the functions of the probiotics and the prebiotics can be better exerted, and the synergistic effect of inhibiting the helicobacter pylori is generated.
Preferably, the Lactobacillus plantarum is named Lactobacillus plantarum Lp90, and is deposited at the institute for microbiology (CGMCC) of China academy of sciences at 1 month and 27 days 2015, with the address of No. 3 of Xilu 1 of Beijing Korean district, zip code 100101 and the preservation number of CGMCC NO. 10453.
Preferably, the Bifidobacterium longum is named as Bifidobacterium longum BL21, which is preserved in the institute for microbiology (CGMCC) at 27 days 1 month 2015, Xilu 1 # 3 of the rising area of Beijing, Zijing, and the accession number is CGMCC NO. 10452.
According to the invention, the lactobacillus plantarum has strong metabolic capacity, can rapidly generate a large amount of organic acid, and the metabolic products of bifidobacterium longum also contain organic acid which can provide metabolic raw materials for lactobacillus acidophilus and promote the growth of lactobacillus acidophilus; the Lactobacillus acidophilus has strong protein decomposition capability, and can provide amino acids to stimulate proliferation of Lactobacillus plantarum and Bifidobacterium longum. The metabolites of the 3 probiotics have the effect of mutually promoting growth, can generate the effect of synergistic gain, has better inhibition effect on helicobacter pylori and stronger capability of regulating the balance of intestinal flora.
Preferably, the lactobacillus acidophilus comprises lactobacillus acidophilus lyophilized powder.
Preferably, the lactobacillus plantarum comprises lactobacillus plantarum freeze-dried powder.
Preferably, the bifidobacterium longum comprises bifidobacterium longum lyophilized powder.
In the invention, the preparation method of the freeze-dried powder comprises the following steps:
respectively inoculating lactobacillus acidophilus, lactobacillus plantarum and bifidobacterium longum in an MRS liquid culture medium according to the inoculation amount of 5-10%, and performing static culture at 36-38 ℃ for 23-24 hours to obtain a first-grade seed solution;
respectively inoculating the primary seed liquid into an MRS liquid culture medium according to the inoculation amount of 3-5%, and carrying out anaerobic culture at 36-38 ℃ for 23-24 h to obtain a secondary seed liquid;
and respectively inoculating the secondary seed liquid into an MRS liquid culture medium according to the inoculation amount of 3-5%, performing high-density culture, performing anaerobic culture at 36-38 ℃ for 12-18 h, respectively centrifugally collecting thalli, adding protective agents (skim milk powder 20%, trehalose 50% and water 30%), performing vacuum freeze drying, controlling the water content to be below 4% and the water activity to be below 0.1, and thus obtaining the lactobacillus acidophilus freeze-dried powder, the lactobacillus plantarum freeze-dried powder and the bifidobacterium longum freeze-dried powder.
Preferably, the number of viable bacteria in the lactobacillus acidophilus freeze-dried powder, the lactobacillus plantarum freeze-dried powder and the bifidobacterium longum freeze-dried powder is 108～1010CFU/g, for example, may be 1X 108CFU/g、5×108CFU/g、1×109CFU/g、5×109CFU/g or 1X 1010CFU/g。
Preferably, the water content in the lactobacillus acidophilus freeze-dried powder, the lactobacillus plantarum freeze-dried powder and the bifidobacterium longum freeze-dried powder is not higher than 4 percent, and can be 1 percent, 2 percent, 3 percent or 4 percent; the water activity content is not higher than 0.1, and may be, for example, 0.02, 0.04, 0.06, 0.08, or 0.1.
Preferably, the prebiotic comprises any one of, or a combination of at least two of, inulin, fructo-oligosaccharide or maltodextrin, for example inulin or a combination of fructo-oligosaccharide and maltodextrin.
In the invention, the prebiotics are fermented and absorbed by intestinal flora when passing through the intestinal tract, and the fermentation product promotes the growth of the prebiotics, reduces the pH value of the intestinal tract, promotes the gastrointestinal motility, accelerates the discharge of harmful bacteria and enhances the immune function of the intestinal tract.
Preferably, the lactobacillus acidophilus freeze-dried powder accounts for 1-5 parts by weight of the helicobacter pylori infection resisting probiotic composition, for example, 1 part, 2 parts, 3 parts, 4 parts or 5 parts, preferably 3 parts.
Preferably, the weight part of the lactobacillus plantarum freeze-dried powder in the helicobacter pylori infection resisting probiotic composition is 2-8 parts, such as 2 parts, 3 parts, 4 parts, 5 parts, 6 parts, 7 parts or 8 parts, and preferably 5 parts.
Preferably, the weight part of the bifidobacterium longum freeze-dried powder in the helicobacter pylori infection resisting probiotic composition is 2-6 parts, such as 2 parts, 3 parts, 4 parts, 5 parts or 6 parts, and preferably 4 parts.
Preferably, the weight part of the inulin in the helicobacter pylori infection resisting probiotic composition is 30-50 parts, such as 30 parts, 31 parts, 32 parts, 33 parts, 34 parts, 35 parts, 36 parts, 37 parts, 38 parts, 39 parts, 40 parts, 41 parts, 42 parts, 43 parts, 44 parts, 45 parts, 46 parts, 47 parts, 48 parts, 49 parts or 50 parts, preferably 40 parts.
Preferably, the weight part of the fructo-oligosaccharide in the probiotic composition for resisting helicobacter pylori infection is 20-40 parts, such as 20 parts, 21 parts, 22 parts, 23 parts, 24 parts, 25 parts, 26 parts, 27 parts, 28 parts, 29 parts, 30 parts, 31 parts, 32 parts, 33 parts, 34 parts, 35 parts, 36 parts, 37 parts, 38 parts, 39 parts or 40 parts, preferably 25 parts.
Preferably, the weight part of the maltodextrin in the probiotic composition for resisting helicobacter pylori infection is 20-30 parts, such as 20 parts, 21 parts, 22 parts, 23 parts, 24 parts, 25 parts, 26 parts, 27 parts, 28 parts, 29 parts or 30 parts, and preferably 23 parts.
Preferably, the probiotic composition for resisting helicobacter pylori infection comprises, by weight, 1-5 parts of lactobacillus acidophilus freeze-dried powder, 2-8 parts of lactobacillus plantarum freeze-dried powder, 2-6 parts of bifidobacterium longum freeze-dried powder, 30-50 parts of inulin, 20-40 parts of fructo-oligosaccharide and 20-30 parts of maltodextrin.
Preferably, the probiotic composition for resisting helicobacter pylori infection comprises 3 parts by weight of lactobacillus acidophilus freeze-dried powder, 5 parts by weight of lactobacillus plantarum freeze-dried powder, 4 parts by weight of bifidobacterium longum freeze-dried powder, 40 parts by weight of inulin, 25 parts by weight of fructo-oligosaccharide and 23 parts by weight of maltodextrin.
In a second aspect, the invention provides the use of the probiotic composition against helicobacter pylori infection of the first aspect in the preparation of a food and/or health care product against helicobacter pylori infection.
According to the invention, the probiotics and the prebiotics are scientifically matched and have a synergistic effect, so that the function of inhibiting the propagation of harmful bacteria such as helicobacter pylori and the like is realized, the intestinal flora is improved, the side reaction of inflammation is relieved, the occurrence probability of diseases such as functional dyspepsia, chronic gastritis, peptic ulcer, gastric adenocarcinoma and the like is further reduced, the prebiotics is added into food and/or health care products, the prebiotics is more convenient to use, and the popularization and use of the product are promoted.
Compared with the prior art, the invention has the following beneficial effects:
(1) the probiotics freeze-dried powder and the prebiotics are matched with each other, so that the intestinal microenvironment is effectively improved, the helicobacter pylori is more strongly inhibited by reasonable proportioning of the raw materials, the planting amount of the helicobacter pylori in stomach is obviously reduced, and the log value of the planting amount of the helicobacter pylori in each gram of stomach tissue is not higher than 5; the concentration of antibodies generated by helicobacter pylori infection in serum is reduced, the absorbance of enzyme-linked immune reaction products of serum IgG at 450nm is not higher than 0.13, and the side reaction of inflammation is relieved; the intestinal flora change can be adjusted, the growth of beneficial bacteria such as bifidobacteria and lactic acid bacteria is promoted, the log value of the number of bacteria in per gram of excrement is not lower than 7, the proliferation of harmful bacteria such as enterobacteria, enterococcus and clostridium perfringens is inhibited, the log value of the number of bacteria in per gram of excrement is not higher than 7, the flora balance is maintained, and the side reactions such as flora imbalance and the like caused by helicobacter pylori infection are obviously improved;
(2) according to the invention, by selecting the appropriate probiotic freeze-dried powder and prebiotics and reasonably matching, compared with the use of single probiotic freeze-dried powder or single prebiotics, the effect is remarkably improved, and the effect of 1+1>2 is achieved;
(3) the preparation method of the probiotic composition for resisting helicobacter pylori infection is simple and easy to implement, does not relate to a complex preparation process, does not need strict production conditions, and has low cost, low energy consumption, high preparation efficiency and wide application value.
Drawings
FIG. 1 is a graph showing the results of the measurement of the amount of colonization of helicobacter pylori in the present invention;
FIG. 2 is a graph showing the results of the measurement of anti-H.pylori antibody in mouse serum according to the present invention;
FIG. 3 is a graph showing the results of the measurement of changes in the intestinal flora of mice according to the present invention.
Detailed Description
To further illustrate the technical means adopted by the present invention and the effects thereof, the present invention is further described below with reference to the embodiments and the accompanying drawings. It is to be understood that the specific embodiments described herein are merely illustrative of the invention and are not limiting of the invention.
The examples do not show the specific techniques or conditions, according to the technical or conditions described in the literature in the field, or according to the product specifications. The reagents or apparatus used are conventional products commercially available from normal sources, not indicated by the manufacturer.
Raw materials:
the Lactobacillus acidophilus LA85 is derived from strain stock of Microkang Probiotics (Suzhou) GmbH, is named as Lactobacillus acidophilus LA85, is preserved in the institute of microbiology (CGMCC) at 7-20 days 2020, and has the address of No. 3 of Xilu 1 of Beijing area sunward, Beijing, and the preservation number of 100101, and is CGMCC NO. 1.12735;
the Lactobacillus plantarum Lp90 is from strain stock of the limited micro-health probiotic (Suzhou) and named as Lactobacillus plantarum Lp90, which is preserved in the institute for microbiology (CGMCC) of China academy of sciences (CGMCC) 1 month 27 days 2015, with the address of No. 3 of Xilu 1 of Beijing north morning area of the rising area, the postal code 100101 and the preservation number of CGMCC NO. 10453;
bifidobacterium longum BL21 is derived from strain stock of Mikang Probiotics (Suzhou) GmbH, named Bifidobacterium longum BL21, and is preserved in the institute of microbiology (CGMCC) 1/27 th of China academy of sciences 2015 at the address of No. 3 of No.1 North road of West Chen of the sunward area in Beijing, the postal code 100101, and the preservation number of CGMCC NO. 10452;
in the invention, the lactobacillus acidophilus freeze-dried powder, the lactobacillus plantarum freeze-dried powder and the bifidobacterium longum freeze-dried powder are prepared by the following methods:
respectively inoculating lactobacillus acidophilus, lactobacillus plantarum and bifidobacterium longum in an MRS liquid culture medium according to the inoculation amount of 10%, and performing static culture at 37 ℃ for 24 hours to obtain a first-grade seed solution;
respectively inoculating the primary seed liquid into an MRS liquid culture medium according to the inoculation amount of 5%, and performing anaerobic culture at 37 ℃ for 24 hours to obtain a secondary seed liquid;
and respectively inoculating the secondary seed liquid into an MRS liquid culture medium according to the inoculation amount of 5%, performing high-density culture, performing anaerobic culture at 37 ℃ for 16h, respectively centrifugally collecting thalli, adding protective agents (skim milk powder 20%, trehalose 50% and water 30%), performing vacuum freeze drying, controlling the water content to be 3% and the water activity to be 0.05, and thus obtaining the lactobacillus acidophilus freeze-dried powder, the lactobacillus plantarum freeze-dried powder and the bifidobacterium longum freeze-dried powder.
MRS medium was purchased from Qingdao Haibo Biotech, Inc.;
the skim milk powder was purchased from Hengtian Natural Co Ltd;
trehalose was purchased from sienna gevin biotechnology limited;
inulin was purchased from Fengning safety high-tech practice Co., Ltd;
fructooligosaccharides were purchased from quantum high-tech (china) bio-resources limited;
maltodextrin was purchased from underwriter bio-ltd;
SPF mice were purchased from shanghai slek experimental animals center;
helicobacter pylori was purchased from ATCC, strain No. ATCC 43504;
GSSA culture medium, BBL agar culture medium, Lbs agar culture medium, EMB agar culture medium, sodium azide-crystal violet-esculin agar culture medium and TSC agar culture medium were purchased from Qingdao Haibo biotechnology, Inc.;
the Cusabio kit is purchased from Shanghai enzyme-linked Biotechnology, Inc.
Example 1
This example provides a probiotic composition against helicobacter pylori infection, comprising: 5 parts of lactobacillus acidophilus freeze-dried powder, 2 parts of lactobacillus plantarum freeze-dried powder, 5 parts of bifidobacterium longum freeze-dried powder, 30 parts of inulin, 38 parts of fructo-oligosaccharide and 20 parts of maltodextrin.
Example 2
This example provides a probiotic composition against helicobacter pylori infection, comprising: 3 parts of lactobacillus acidophilus freeze-dried powder, 5 parts of lactobacillus plantarum freeze-dried powder, 4 parts of bifidobacterium longum freeze-dried powder, 40 parts of inulin, 25 parts of fructo-oligosaccharide and 23 parts of maltodextrin.
Example 3
This example provides a probiotic composition against helicobacter pylori infection, comprising: 2 parts of lactobacillus acidophilus freeze-dried powder, 8 parts of lactobacillus plantarum freeze-dried powder, 2 parts of bifidobacterium longum freeze-dried powder, 48 parts of inulin, 20 parts of fructo-oligosaccharide and 20 parts of maltodextrin.
Example 4
This example provides a probiotic composition against helicobacter pylori infection, comprising: 2 parts of lactobacillus acidophilus freeze-dried powder, 4 parts of lactobacillus plantarum freeze-dried powder, 6 parts of bifidobacterium longum freeze-dried powder, 50 parts of inulin, 20 parts of fructo-oligosaccharide and 20 parts of maltodextrin.
Example 5
This example provides a probiotic composition against helicobacter pylori infection, comprising: 1 part of lactobacillus acidophilus freeze-dried powder, 6 parts of lactobacillus plantarum freeze-dried powder, 5 parts of bifidobacterium longum freeze-dried powder, 30 parts of inulin, 40 parts of fructo-oligosaccharide and 30 parts of maltodextrin.
Example 6
This example provides a probiotic composition against helicobacter pylori infection, comprising: 0.5 part of lactobacillus acidophilus freeze-dried powder, 10.5 parts of lactobacillus plantarum freeze-dried powder, 1 part of bifidobacterium longum freeze-dried powder, 25 parts of inulin, 48 parts of fructo-oligosaccharide and 15 parts of maltodextrin.
Example 7
This example provides a probiotic composition against helicobacter pylori infection, comprising: 10 parts of lactobacillus acidophilus freeze-dried powder, 1 part of lactobacillus plantarum freeze-dried powder, 1 part of bifidobacterium longum freeze-dried powder, 55 parts of inulin, 15 parts of fructo-oligosaccharide and 18 parts of maltodextrin.
Example 8
The only difference from example 2 is that in this example, fructooligosaccharides and maltodextrin are not added, the missing parts by weight are made up of inulin, and the remaining raw materials are the same as in example 2.
Comparative example 1
The difference from the example 2 is only that no lactobacillus acidophilus freeze-dried powder and no bifidobacterium longum freeze-dried powder are added in the comparative example, the missing parts by weight are complemented by lactobacillus plantarum freeze-dried powder, and the rest raw materials are the same as the example 2.
Detection of anti-helicobacter pylori infection
110 SPF-class mice aged 4 weeks were taken, and after being fed normally for 1 week, the mice were first gavaged with 0.5mL of mixed antibiotics (10mg/mL ampicillin, 2mg/mL gentamicin, and 25mg/mL azithromycin), and after 3 days, the mice were randomly divided into 11 groups, i.e., a control group, an infected group, examples 1-8 groups, and comparative example 1 group, each of which was 10 mice. Control mice were first treated with 0.2mol/L NaHCO3Performing intragastric administration, wherein after each mouse is intragastric administered with 0.25mL every day for 1h, the mice are intragastric administered with 0.3mL of physiological saline once every 1 day for 5 times; the infection group was first treated with 0.2mol/L NaHCO3Gavage, each mouse gavage 0.25mL daily, after 1h, 0.3mL concentration of 1 × 109CFU/Carrying out intragastric administration on mL helicobacter pylori bacterial liquid once every 1 day for 5 times; after the groups 1-8 and the group 1 are gavaged by the same method as the infected group, the probiotic composition for resisting helicobacter pylori infection of the corresponding group is used for intervening for 4 weeks, and the normal feeding is carried out for 1 month.
Determination of the amount of helicobacter pylori colonized
The homogenate of the stomach tissue was taken, after gradient dilution, inoculated onto a Glax Selective Supplement A (GSSA) plate (containing 7% sheep blood, 2.5. mu.g/mL amphotericin, 200. mu.g/mL bacitracin, 6. mu.g/mL vancomycin, 2. mu.g/mL nalidixic acid and 0.5. mu.g/mL polymyxin), cultured under anaerobic conditions for 48h, and the number of H.pylori was counted, as shown in FIG. 1.
As can be seen from FIG. 1, in examples 1-5, the amount of helicobacter pylori colonized in the stomach of the mouse is small, and the log value of the amount of helicobacter pylori per gram of tissue is not higher than 5;
compared with the examples 1-5, the amount of helicobacter pylori colonized in the examples 6-8 and the comparative example 1 is larger, wherein the content of each raw material in the examples 6-7 is not in an optimal range, which shows that the addition amount of each raw material can influence the helicobacter pylori resistance effect of the probiotic composition, and when the content is not in the optimal range, the helicobacter pylori resistance effect is not obvious;
in the embodiment 8, only inulin is added, and only lactobacillus plantarum is added in the comparative example 1, so that the antibacterial ability is influenced by different degrees, which shows that the formula effect of the composite probiotic and prebiotics is obviously better than that of the single prebiotic formula or the single probiotic formula, and shows that the helicobacter pylori infection resisting probiotic composition of different probiotics and prebiotics can effectively eliminate helicobacter pylori under the same intake;
further comparing the effects of examples 1 to 5, it can be found that example 2 is the most effective in eliminating helicobacter pylori.
Mouse serum anti-helicobacter pylori antibody assay
The antibody content in mouse serum was determined by enzyme-linked immunoassay, and detection was performed using the Cusabio kit.
Centrifuging the blood of the mouse at 1000g for 20min, and taking the supernatant; diluting the supernatant with diluent, adding 100 μ L diluted liquid into an enzyme label plate, reacting at 37 deg.C for 30min, washing the plate for 3 times, and spin-drying; adding 100 μ L of horse radish peroxidase-labeled anti-mouse IgG working solution, reacting at 37 deg.C for 30min, washing the plate for 3 times, and spin-drying; adding 90 μ L of color-developing agent TMB, and reacting at 37 deg.C for 15 min; finally, 50. mu.L of stop solution is added, and the product is detected by an enzyme-linked immunosorbent assay (OD)450The results are shown in FIG. 2.
After helicobacter pylori infection, corresponding antibodies are generated in serum, and when the content of the antibodies is too high, inflammation is easily caused, and repeated treatment effects or other side effects are caused. As can be seen from FIG. 2, in examples 1 to 5, the content of IgG antibody in the mouse serum is low, and the absorbance at 450nm is not higher than 0.13, which indicates that the relieving effect on the inflammatory reaction is good;
compared with the examples 1 to 5, the examples 6 to 8 and the comparative example 1 are poor in effect, wherein the content of each raw material in the examples 6 to 7 is not in an optimal range, and the obtained probiotic composition has an insignificant relieving effect on the inflammatory side reaction caused by helicobacter pylori infection when the addition amount of the probiotic is not in the optimal range;
in addition, compared with the single prebiotic formulation (example 8) and the single probiotic formulation (comparative example 1), the formulation using the composite probiotics and prebiotics has a significantly better effect of relieving inflammation than the single prebiotic formulation or the single probiotic formulation, which indicates that the probiotic composition using different probiotics and prebiotics has a better effect of relieving the side effect of inflammation caused by helicobacter pylori at the same intake;
further comparing the effects of examples 1 to 5, it was found that example 2 is the best effect for relieving the side effects of inflammation caused by helicobacter pylori infection.
Determination of changes in intestinal flora in mice
Collecting fresh excrement of a mouse, diluting the excrement by 10 times in a gradient manner, inoculating the diluted excrement into a selective culture medium, identifying and counting colonies according to colony morphology, gram-staphyloscopy and biochemical reaction, and calculating the number of bifidobacteria, lactobacilli, enterobacteria, enterococci and clostridium perfringens in each gram of wet excrement. The culture media and culture conditions used for the different species are shown in table 1, and the results of the intestinal flora changes are shown in fig. 3.
TABLE 1
Among the above-mentioned strains, bifidobacteria and lactic acid bacteria are microorganisms beneficial to the intestinal tract and help to maintain the intestinal microecological balance. On the contrary, enterobacteria, enterococci and clostridium perfringens are harmful bacteria and easily produce toxins, etc. As can be seen from FIG. 3, in examples 1-5, the content of beneficial microorganisms in intestinal tracts of mice is increased, the log value of the number of bacteria in each gram of feces is not less than 7, the content of harmful microorganisms is decreased, and the log value of the number of bacteria in each gram of feces is not more than 7;
the anti-helicobacter pylori infection probiotic compositions prepared in examples 6 to 8 and comparative example 1 have slightly poor intestinal flora regulating ability compared with examples 1 to 5, wherein the addition amount of each component in examples 6 to 7 is not in a preferred range, which shows that the addition amount of the substance affects the efficacy of the probiotic composition;
in addition, compared with the single prebiotic formulation (example 8) and the single probiotic formulation (comparative example 1), the formulation using the composite probiotic and prebiotic has poor promoting effect on beneficial microorganisms and poor inhibiting effect on harmful microorganisms, which indicates that the probiotic composition using different probiotics and prebiotics has better effect on relieving the side effect of flora imbalance caused by helicobacter pylori at the same intake;
further, the effects of comparative examples 1 to 5 were found to be the best in the regulation of intestinal flora in example 2.
In conclusion, the probiotic composition for resisting helicobacter pylori infection can inhibit the proliferation of helicobacter pylori in vivo and reduce the colonization amount in vivo; simultaneously, the quantity of anti-helicobacter pylori antibodies in serum can be reduced, and inflammatory side effects can be relieved; the quantity of beneficial microorganisms in the intestinal tract can be effectively increased, the quantity of harmful microorganisms can be reduced, the flora balance in the intestinal tract can be maintained, and the environment in the intestinal tract can be adjusted; the probiotic composition for resisting helicobacter pylori infection is convenient to use, low in cost and energy consumption and wide in application value.
The applicant states that the present invention is illustrated in detail by the above examples, but the present invention is not limited to the above detailed methods, i.e. it is not meant that the present invention must rely on the above detailed methods for its implementation. It should be understood by those skilled in the art that any modification of the present invention, equivalent substitutions of the raw materials of the product of the present invention, addition of auxiliary components, selection of specific modes, etc., are within the scope and disclosure of the present invention.
Claims (10)
1. A probiotic composition against helicobacter pylori infection, wherein said probiotic composition against helicobacter pylori infection comprises: lactobacillus acidophilus, lactobacillus plantarum, bifidobacterium longum and prebiotics;
wherein the Lactobacillus acidophilus is named as Lactobacillus acidophilus LA85, is preserved in the institute of microbiology (CGMCC) of China academy of sciences at 7-month and 20-month in 2020, and has the preservation number of CGMCC NO. 1.12735.
2. The probiotic composition for resisting helicobacter pylori infection according to claim 1, wherein the Lactobacillus plantarum is named Lactobacillus plantarum Lp90 and is deposited at the institute for microbiology (CGMCC) of china academy of sciences (CGMCC) at 1 month and 27 days 2015 with the collection number of CGMCC No. 10453.
3. The probiotic composition for the treatment of helicobacter pylori infection according to claim 1 or 2, wherein the Bifidobacterium longum is named Bifidobacterium longum BL21 and deposited at the institute for microbiology (CGMCC) of the institute of sciences china (CGMCC) at 1 month and 27 days 2015 with the collection number of CGMCC No. 10452.
4. The probiotic composition for resisting helicobacter pylori infection according to any one of claims 1 to 3, wherein the lactobacillus acidophilus comprises lactobacillus acidophilus freeze-dried powder;
preferably, the lactobacillus plantarum comprises lactobacillus plantarum lyophilized powder;
preferably, the bifidobacterium longum comprises bifidobacterium longum lyophilized powder.
5. The probiotic composition for resisting helicobacter pylori infection according to claim 4, wherein the viable count of the lactobacillus acidophilus lyophilized powder, the lactobacillus plantarum lyophilized powder and the bifidobacterium longum lyophilized powder is 108～1010CFU/g。
6. The probiotic composition for resisting helicobacter pylori infection according to any one of claims 1 to 5, wherein the prebiotics comprise any one or a combination of at least two of inulin, fructo-oligosaccharide or maltodextrin.
7. The probiotic composition for resisting helicobacter pylori infection according to any one of claims 1 to 6, wherein the lactobacillus acidophilus lyophilized powder is 1 to 5 parts by weight, preferably 3 parts by weight of the probiotic composition for resisting helicobacter pylori infection;
preferably, the lactobacillus plantarum freeze-dried powder accounts for 2-8 parts by weight, preferably 5 parts by weight, of the helicobacter pylori infection resisting probiotic composition;
preferably, the weight part of the bifidobacterium longum freeze-dried powder in the helicobacter pylori infection resisting probiotic composition is 2-6 parts, and preferably 4 parts.
8. The helicobacter pylori infection-resisting probiotic composition according to any one of claims 1 to 7, wherein the inulin is contained in the helicobacter pylori infection-resisting probiotic composition in an amount of 30 to 50 parts by weight, preferably 40 parts by weight;
preferably, the weight part of the fructo-oligosaccharide in the helicobacter pylori infection resisting probiotic composition is 20-40 parts, preferably 25 parts;
preferably, the weight part of the maltodextrin in the probiotic composition for resisting helicobacter pylori infection is 20-30 parts, and 23 parts is preferred.
9. The probiotic composition for resisting helicobacter pylori infection according to any one of claims 1 to 8, wherein the probiotic composition for resisting helicobacter pylori infection comprises 1 to 5 parts by weight of lactobacillus acidophilus lyophilized powder, 2 to 8 parts by weight of lactobacillus plantarum lyophilized powder, 2 to 6 parts by weight of bifidobacterium longum lyophilized powder, 30 to 50 parts by weight of inulin, 20 to 40 parts by weight of fructo-oligosaccharide and 20 to 30 parts by weight of maltodextrin;
preferably, the probiotic composition for resisting helicobacter pylori infection comprises 3 parts by weight of lactobacillus acidophilus freeze-dried powder, 5 parts by weight of lactobacillus plantarum freeze-dried powder, 4 parts by weight of bifidobacterium longum freeze-dried powder, 40 parts by weight of inulin, 25 parts by weight of fructo-oligosaccharide and 23 parts by weight of maltodextrin.
10. Use of the probiotic composition for resisting helicobacter pylori infection according to any one of claims 1 to 9 in the preparation of food and/or health care products for resisting helicobacter pylori infection.
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