Bifidobacterium metagen, bifidobacterium subgen and bifidobacterium pseudogen as well as preparation method and application thereof
By using inactivated Bifidobacterium posterior, parabiotic and biogenic preparations in animal breeding, the problems of poor stress resistance and storage resistance of probiotics in animal breeding are solved, and efficient and safe bacterial infection prevention and treatment effect are achieved , significantly better than traditional live bacteria preparations.
Patent Information
- Application Number
- CN202411081090.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-08-08
- Publication Date
- 2024-12-13
AI Technical Summary
The abuse of existing antibiotics in animal breeding leads to bacterial resistance, and probiotic preparations have problems such as poor stress resistance, intolerance to storage, low gastrointestinal survival rate, difficulty in colonization and virulence factors, and cannot effectively replace traditional antibiotics.
By fermenting Bifidobacterium in a specific culture medium, posterior, parabiotics and biomes were obtained. The posterior, inactivation was used to treat posterior, and the host intestine was prepared to prevent and treat animal bacterial infection without colonizing the host. And it has a preparation with high viable bacteria count, significant effects, small side effects and high safety.
It can significantly prevent and treat animal bacterial infections without relying on intestinal colonization, enhance the host's immune function and intestinal barrier function, reduce inflammatory response and mortality, and significantly better than traditional live bacterial preparations.
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Figure CN119120259A_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the technical field of biological fermentation, and in particular relates to a bifidobacterium postbiotic, parabiotic, quasibiotic, and a preparation method and application thereof. Background Art
[0002] The animal husbandry industry in my country has developed extremely rapidly and its scale is increasing. In the past, the use of antibiotics was a common method for preventing and treating bacterial infections in clinical practice, but the abuse of antibiotics has caused increasingly serious bacterial resistance. Since the implementation of a comprehensive ban on antibiotics in feed in my country on July 1, 2020, livestock and poultry farming has faced huge challenges, and people are in urgent need of safe, green and effective products to replace antibiotics.
[0003] Currently, there are more than 30 probiotic preparations approved for feed addition. While secreting peptides or proteins with antibacterial activity, probiotics can also provide nutrients to livestock and poultry to promote their healthy growth. As the main alternative to antibiotics, probiotic preparations have been widely used in animal production. They have many functions such as promoting the digestion and absorption of nutrients, improving the balance of gastrointestinal flora and enhancing intestinal immunity. They can significantly improve the growth performance of livestock and poultry and prevent and treat diseases. However, live probiotics also have certain problems, such as poor stress resistance, storage intolerance, low gastrointestinal survival rate, difficulty in colonization, and energy consumption for proliferation. In addition, some probiotics have virulence factors and certain drug resistance, and may even cause infection. Summary of the invention
[0004] The object of the present invention is to provide a bifidobacterium postbiotic, parabiotic, quasibiotic and its preparation method and application. The bifidobacterium postbiotic prepared by the preparation method provided by the present invention can prevent and treat bacterial infection in animals without colonization of the host intestine, and has high bacterial viability, significant effect, small side effects and high safety.
[0005] The present invention provides a method for preparing postbiotics, comprising the following steps: inoculating bifidobacteria into a culture solution for fermentation to obtain a fermentation solution, and inactivating the fermentation solution to obtain postbiotics; the fermentation temperature is 30-42° C., and the fermentation time is 30-40 hours;
[0006] Each liter of the culture solution includes the following raw materials: 8-12 g of peptone, 4-6 g of beef powder, 18-22 g of glucose, 3-5 g of yeast powder, 4-6 g of sodium acetate, 1-3 g of dipotassium hydrogen phosphate, 0.1-0.5 g of magnesium sulfate, 1-3 g of triammonium citrate, 0.01-0.1 g of manganese sulfate, 0.5-1.5 mL of Tween 80 and 0.5-1.5 g of L-cysteine hydrochloride.
[0007] Preferably, the bifidobacterium includes Bifidobacterium bifidum.
[0008] Preferably, the inactivation temperature is 60-85°C and the time is 20-80 minutes.
[0009] Preferably, the inactivation includes a first inactivation and a second inactivation; the first inactivation is carried out at a temperature of 60 to 65°C for 20 to 40 minutes; the second inactivation is carried out at a temperature of 80 to 85°C for 20 to 40 minutes.
[0010] The present invention provides postbiotics prepared by the preparation method.
[0011] The present invention provides a method for preparing a parabiotic and a parabiotic, comprising the following steps: centrifuging the above-mentioned parabiotic to obtain a supernatant and a precipitate;
[0012] filtering the supernatant to obtain parabiotics;
[0013] resuspending the precipitate with culture medium to obtain biotin;
[0014] Each liter of the culture solution includes the following raw materials: 8-12 g of peptone, 4-6 g of beef powder, 18-22 g of glucose, 3-5 g of yeast powder, 4-6 g of sodium acetate, 1-3 g of dipotassium hydrogen phosphate, 0.1-0.5 g of magnesium sulfate, 1-3 g of triammonium citrate, 0.01-0.1 g of manganese sulfate, 0.5-1.5 mL of Tween 80 and 0.5-1.5 g of L-cysteine hydrochloride.
[0015] The invention provides a parabiotic prepared by the preparation method.
[0016] The present invention provides a quasi-biotin prepared by the above preparation method.
[0017] The present invention also provides a preparation for preventing and treating bacterial infection in animals, wherein the effective ingredients of the preparation include: one or more of the above-mentioned postbiotics, the above-mentioned parabiotics and the above-mentioned quasi-biotics.
[0018] Preferably, the bacteria include: Salmonella and / or Staphylococcus.
[0019] Beneficial effects: The present invention provides a method for preparing postbiotics, wherein bifidobacteria are fermented and cultured to obtain postbiotics, and the method is simple. In the present invention, L-cysteine hydrochloride is added to the culture solution, and the sugar content is increased, and the number of viable bacteria of the postbiotics obtained by fermentation reaches at least 1×10 9 CFU / mL. The present invention provides a method for preparing parabiotics and parabiotics using the above-mentioned postbiotics. The postbiotics, parabiotics and parabiotics prepared by the present invention have high safety, high content of viable bacteria, good stability, strong stress resistance and easy preservation.
[0020] The present invention provides postbiotics, parabiotics and parabiotics prepared by the above preparation method, which can directly act on the host intestinal cells and intestinal microorganisms without colonization in the host intestine, protect intestinal cells from bacterial infection by preventing bacterial adhesion and internalization, prevent the increase of paracellular permeability, restore cell tight junction function and membrane integrity, reduce bacterial colonization in intestinal cells, regulate cytokine gene expression, reduce inflammatory response, and play a role in preventing and treating bacterial infections in animals.
[0021] The present invention also provides a preparation for preventing and treating bacterial infection in animals using the above-mentioned postbiotics, parabiotics and parabiotics as effective ingredients. In the embodiment of the present invention, after the chicks take the preparation, the effective ingredients in the preparation regulate the immune function of the chicks, enhance the intestinal barrier function of the chicks, and improve the balance of intestinal flora. When the chicks are inoculated with Salmonella pullorum, the organic acids and bacteriocins in the preparation inhibit the activity of Salmonella pullorum and inhibit the adhesion of Salmonella pullorum to the intestine. The body's ability to resist Salmonella infection is enhanced, the inflammatory response is reduced, and intestinal damage is reduced. Thereby playing the effect of preventing and treating Salmonella pullorum. After mice were gavaged with the preparation, the effective ingredients in the preparation regulated the immune function of mice, enhanced the resistance of mice to Staphylococcus, thereby reducing the mortality rate, thereby playing the role of preventing and treating Staphylococcus infection in mice. BRIEF DESCRIPTION OF THE DRAWINGS
[0022] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required to be used in the embodiments are briefly introduced below.
[0023] Figure 1 This is a colony growth diagram of the first fermentation broth in Example 1;
[0024] Figure 2 This is a detection diagram showing that the second fermentation broth is completely inactivated in Example 1;
[0025] Figure 3 This is the inhibition zone determination diagram of the same preparation against Salmonella pullorum;
[0026] Figure 4 This is a curve chart showing the effects of different preparations on the average body weight of broilers. DETAILED DESCRIPTION
[0027] The present invention provides a method for preparing a postbiotic, comprising the following steps: inoculating bifidobacteria into a culture solution for fermentation to obtain a fermentation solution, and inactivating the fermentation solution to obtain a postbiotic.
[0028] Unless otherwise specified, the present invention has no special requirements for the preparation raw materials, and commercially available products known to those skilled in the art can be used.
[0029] The bifidobacterium of the present invention preferably includes Bifidobacterium bifidum. In the embodiment of the present invention, the selected Bifidobacterium bifidum is a conventional commercially available product, and the strain number is BNCC 186304. The present invention preferably inoculates the above-mentioned bifidobacterium into the culture solution; during inoculation, the volume ratio of the bifidobacterium strain to the culture solution is preferably 0.5-1.5:40-60, more preferably 0.8-1.2:45-55, and most preferably 1:50.
[0030] In the present invention, each L of the culture solution includes the following raw materials: 8-12 g of peptone, 4-6 g of beef powder, 18-22 g of glucose, 3-5 g of yeast powder, 4-6 g of sodium acetate, 1-3 g of dipotassium hydrogen phosphate, 0.1-0.5 g of magnesium sulfate, 1-3 g of triammonium citrate, 0.01-0.1 g of manganese sulfate, 0.5-1.5 mL of Tween 80 and 0.5-1.5 g of L-cysteine hydrochloride.
[0031] The culture medium of the present invention comprises 8-12g of peptone per L, more preferably 9-11g, and most preferably 10g; the culture medium comprises 4-6g of beef powder per L, more preferably 4.5-5.5g, and most preferably 5g; the culture medium comprises 18-22g of glucose per L, more preferably 19-21g, and most preferably 20g; the culture medium comprises 3-5g of yeast powder per L, more preferably 3.5-4.5g, and most preferably 4g; the culture medium comprises 4-6g of sodium acetate per L, more preferably 4.5-5.5g, and most preferably 5g; each liter of the culture solution includes 1-3g of dipotassium hydrogen phosphate, more preferably 1.5-2.5g, and most preferably 2g; each liter of the culture solution includes 0.1-0.5g of magnesium sulfate, more preferably 0.2-0.4g, and most preferably 0.3g; each liter of the culture solution includes 1-3g of triammonium citrate, more preferably 1.5-2.5g, and most preferably 2g; each liter of the culture solution includes 0.01-0.1g of manganese sulfate, more preferably 0.03-0.07g, and most preferably 0.05g; each liter of the culture solution includes 0.5-1.5mL of Tween 80, more preferably 0.8-1.2mL, and most preferably 1mL; each liter of the culture solution includes 0.5-1.5g of L-cysteine hydrochloride, more preferably 0.8-1.2g, and most preferably 1g. As a preferred embodiment of the present invention, each L of the culture solution comprises the following raw materials: 10 g of peptone, 5 g of beef powder, 20 g of glucose, 4 g of yeast powder, 5 g of sodium acetate, 2 g of dipotassium hydrogen phosphate, 0.3 g of magnesium sulfate, 2 g of triammonium citrate, 0.05 g of manganese sulfate, 1 mL of Tween 80 and 1 g of L-cysteine hydrochloride.
[0032] The present invention preferably inoculates the above-mentioned bifidobacterium into the above-mentioned culture solution for fermentation culture to obtain a fermentation solution. The fermentation temperature of the present invention is 30-42°C, preferably 35-40°C, more preferably 37°C; the fermentation time is 30-40h, more preferably 34-38h, and most preferably 36h. The fermentation of the present invention is preferably anaerobic fermentation, and the purpose of extending the fermentation time is to obtain more metabolites of Bifidobacterium bifidum.
[0033] The present invention preferably inactivates the fermentation broth obtained above to obtain postbiotics. The inactivation temperature of the present invention is preferably 60-85°C, more preferably 62-82°C; the inactivation time is preferably 20-80min, more preferably 30-60min. The inactivation of the present invention preferably includes a first inactivation and a second inactivation; the temperature of the first inactivation is preferably 60-65°C, more preferably 62°C; the time of the first inactivation is preferably 20-40min, more preferably 30min; the temperature of the second inactivation is preferably 80-85°C, more preferably 82°C; the time is preferably 20-40min, more preferably 30min. The present invention is inactivated twice, the inactivation is more sufficient, the obtained postbiotics are more stable, and the inactivation temperature is controlled within a certain range, even if the time is increased, it will not affect the active ingredients of the postbiotics.
[0034] The present invention provides a postbiotic prepared by the above preparation method. The viable count of the postbiotic obtained by the present invention can reach 1×10 9 CFU / mL, and have a long shelf life, higher safety and greater stability, which can avoid many disadvantages of live bacteria.
[0035] The present invention provides a method for preparing parabiotics and quasi-bionts, comprising the following steps: centrifuging the above-mentioned postbiotics to obtain a supernatant and a precipitate; filtering the supernatant to obtain parabiotics; and resuspending the precipitate with a culture medium to obtain quasi-bionts.
[0036] The present invention preferably first centrifuges the above-mentioned postbiotics, and the centrifugation is preferably ultra-low temperature centrifugation; the speed of the ultra-low temperature centrifugation is preferably 7000-9000r / min, more preferably 8000r / min; the time of the ultra-low temperature centrifugation is preferably 5-15min, more preferably 10min; the temperature of the ultra-low temperature centrifugation is 2-6°C, more preferably 4°C. The present invention preferably filters the supernatant after centrifugation to obtain a parabiotic; the filtration is preferably membrane filtration, and the filter membrane is preferably a 0.22μm filter membrane. The present invention preferably resuspends the above-mentioned centrifuged precipitate with the above-mentioned culture solution to obtain a quasi-biont; when resuspended, the volume ratio of the precipitate to the culture solution is preferably 0.5-1.5:05-1.5, more preferably 1:1. The culture solution of the present invention is the same as the culture solution inoculated with the above-mentioned bifidobacterium, which will not be repeated here.
[0037] The present invention provides a parabiotic prepared by the above-mentioned preparation method. The present invention provides a parabiotic prepared by the above-mentioned preparation method. The parabiotic and parabiotic prepared by the present invention contain metabolites and inactivated bacteria in the fermentation and cultivation process of bifidobacteria.
[0038] The present invention also provides a preparation for preventing and treating bacterial infection in animals, wherein the active ingredients of the preparation include: one or more of the above-mentioned postbiotics, the above-mentioned parabiotics and the above-mentioned quasi-biotics. The preparation of the present invention can directly act on the host intestinal cells and intestinal microorganisms, protect the intestinal cells from bacterial infection by preventing the adhesion and internalization of bacteria, regulate cytokine gene expression, reduce inflammatory response, and play a role in preventing and treating bacterial infection in animals. The bacteria of the present invention preferably include: Salmonella and / or Staphylococcus; the Salmonella is more preferably Salmonella pullorum.
[0039] In order to further illustrate the present invention, a bifidobacterium postbiotic, parabiotic, quasibiotic, and preparation method and application thereof provided by the present invention are described in detail below with reference to the accompanying drawings and examples, but they should not be construed as limiting the scope of protection of the present invention.
[0040] Unless otherwise specified, the reagents used in the examples of the present invention are conventional commercially available products in the art. The 0.22 μm Syringe Filter (water system) was purchased from Biosharp Biotechnology Co., Ltd., and the culture medium raw materials were purchased from Beijing Solebow Technology Co., Ltd.
[0041] The chicks used in the examples of the present invention were purchased from Qiusheng Poultry, the feed used was Zhengda chick pellets, and the mice and feed were purchased from Hunan Slake Jingda. The experimental process complied with the animal welfare policy.
[0042] Example 1
[0043] 1. Fermented Bifidobacterium bifidum
[0044] Culture medium: Each 1000 mL contains peptone 10 g, beef powder 5 g, glucose 20 g, yeast powder 4 g, sodium acetate 5 g, dipotassium hydrogen phosphate 2 g, magnesium sulfate 0.3 g, triammonium citrate 2 g, manganese sulfate 0.05 g, Tween 80 1 mL, and L-cysteine hydrochloride 1 g.
[0045] Bifidobacterium bifidum (strain number: BNCC186304) was inoculated into the culture medium at a volume ratio of 1:50, and a 50 mL centrifuge tube was filled as much as possible. The first fermentation liquid was harvested after shaking at 37° C. for 36 h.
[0046] Use sterilized PBS to dilute the bifidobacterium fermentation broth in a ten-fold gradient, mix well after dilution, and dilute to 10 -7 , take 10 -5 , 10-6 , 10 -7 100 μL of the dilution solution was spread on the MRS plate. Two replicates were made for each treatment, sealed in anaerobic bags, added to anaerobic gas-generating bags, and cultured in a 37°C incubator for 48 hours. The growth of the colonies after culture was as follows: Figure 1 As shown in Figure 2, the number of viable bacteria in the first fermentation broth obtained by colony counting was 1.84×10 9 CFU / mL.
[0047] 2. Preparation and inactivation of postbiotics, parabiotics and parabiotics
[0048] The first fermentation liquid was inactivated at 62° C. for 30 min, and then inactivated at 82° C. for 30 min. The first fermentation liquid was inactivated to obtain a second fermentation liquid, namely, postbiotics.
[0049] The second fermentation liquid was applied to MRS solid culture medium and sealed with anaerobic bags, and placed in an anaerobic gas production bag in a 37°C incubator for 48 hours. After testing, it was completely inactivated. Figure 2 shown.
[0050] The second fermentation liquid was centrifuged at 4°C ultra-low temperature and 8000r / min for 10min. The supernatant after centrifugation was filtered using a 0.22μm filter membrane to obtain the parabiotics. The precipitate after centrifugation was added with an equal volume of fermentation liquid and resuspended to obtain the parabiotics.
[0051] Comparative Example 1
[0052] The method of Example 1 is followed, except that Bifidobacterium bifidum is fermented and cultured using common MRS medium to obtain the first fermentation liquid.
[0053] Comparative Example 2
[0054] The method of Example 1 is followed, except that after obtaining the first fermentation liquid, it is inactivated only once at 100° C. to obtain the postbiotics.
[0055] Experimental Example 1: Resuscitation of Salmonella pullorum and determination of postbiotic inhibition zone by Oxford cup method
[0056] Inoculate Salmonella pullorum in LB liquid culture medium at a ratio of 1:50 (volume ratio) and culture at 37°C and 180r / min for 16h. Prepare 100mL LB solid culture medium with 2% agar, sterilize by high pressure and cool to 45°C, add 10mL of revived Salmonella pullorum solution, shake well, and pour into a culture dish with two sterile Oxford cups, with 20mL of culture medium poured into each dish. Cool to room temperature for 20min, take out the Oxford cup after the culture medium solidifies, add 200μL of postbiotics, parabiotics and parabiotics of Example 1 to each well, and obtain postbiotic group, parabiotic group and parabiotic group, respectively, and add 200μL of live Bifidobacterium (live bacteria count 1.84×10 9CFU / mL) as the control group, and 200 μL of culture solution as the blank control group, and the culture solution was allowed to diffuse into LB agar at rest and cultured at 37°C for 12 to 14 h.
[0057] The outer diameter of the inhibition zone and the inner diameter of the Oxford cup were measured, and the value was obtained by subtracting the inner area from the outer area. The inhibition zone of the postbiotics in Example 1 was 17.5 mm, the inhibition zone of the parabiotics was 17 mm, the inhibition zone of the live bacteria control was 17 mm, the parabiotics had no inhibition zone, and the culture medium blank control had no inhibition zone. The results are as follows: Figure 3 shown.
[0058] The number of viable bacteria in the first fermentation liquid of Comparative Example 1 was 4.9×10 8 CFU / mL.
[0059] The postbiotics of Example 2 were subjected to the Oxford Cup inhibition zone test, and the inhibition zone was only 13 mm.
[0060] Test Example 2 Salmonella pullorum challenge protection experiment
[0061] 72 SPF1 one-day-old hemp-feathered broilers were randomly divided into 6 groups, each with 12 chicks, namely the postbiotic group, parabiotic group, parabiotic group, live bacteria group, blank control group, and anti-virus control group. Each group had a cage, equipped with a warm lamp, a kettle, and a feed trough. After the chicks opened their mouths on the first day, 200mL of warm water was added with 10g of glucose and 0.1g of 10% veterinary enrofloxacin for each group for three consecutive days. During this period, the normal basic diet was fed two hours after the medicine was fed every day, 36g per group. For chicks under 10 days of age, the daily feed intake for each chicken was (day age + 2)g, and for chicks aged 11 to 20 days, the daily feed intake for each chicken was (day age + 1)g.
[0062] During the period, starting from the 3rd day of age, after cleaning the feed trough and water trough chicken cage at 2 pm, 12 mL of the corresponding group's preparation was added to 200 mL of water in the postbiotic group, parabiotic group, parabiotic group and live bacteria group chickens, and 12 mL of normal saline was added to the blank group. All animals were fed a safe basic diet without Salmonella according to their age. After weighing on an empty stomach every morning, ultrapure water was added to each group. After feeding for 7 days, on the 8th day, 0.1 mL of the above-mentioned attacking bacterial solution was administered orally to each chicken. Feeding and watering were stopped for 16 hours and 0.5 hours before exposure to the virus, and drinking water was resumed 0.5 hours after exposure to the virus. Feeding and watering were stopped for 0.5 hours before the second exposure to the virus, and drinking water was resumed 0.5 hours after exposure to the virus. The two exposures were completed within 24 hours (each chicken was exposed to 1.0×10 8 CFU / mL of Salmonella pullorum). The chicks were fed for 7 days after infection, and the corresponding group of preparations were added to 200mL of water at 2 pm every day until the end of the experiment. The status and mortality of the chicks in each group were observed after infection, and the morbidity and mortality results were recorded and calculated. The results are shown in Tables 1, 2 and Figure 4 As shown (the result is rounded to two decimal places).
[0063] Table 1 Comparison of morbidity and mortality in Salmonella pullorum challenge protection experiment
[0064] Grouping Blank control group The challenge control group Epigenetic tuple Biotype Parabiotic Group Live bacteria control group Number of cases (only) 0 12 8 9 10 10 Incidence 0％ 100％ 66.66％ 75％ 83.33％ 83.33％ Number of deaths (individuals) 0 5 1 2 3 2 Case fatality rate 0％ 41.66％ 8.33％ 16.66％ 25％ 16.66％
[0065] Table 2 Comparison of average body weight in Salmonella pullorum challenge protection experiment
[0066]
[0067] From Table 1, Table 2 and Figure 4 It can be seen that the antiviral effects of postbiotics, parabiotics and parabiotic preparations are obvious and significantly better than live bacteria; and the weight of the test subjects fed with postbiotics, parabiotics and parabiotic preparations has steadily increased. It can be seen that after the chicks take the preparation, the effective ingredients in the preparation regulate the immune function of the chicks, enhance the intestinal barrier function of the chicks, and improve the balance of intestinal flora. When the chicks are inoculated with Salmonella pullorum, the organic acids and bacteriocins in the preparation inhibit the activity of Salmonella pullorum and inhibit the adhesion of Salmonella pullorum to the intestine. The body's ability to resist Salmonella infection is enhanced, the inflammatory response is reduced, and intestinal damage is reduced, thereby playing a role in preventing and treating Salmonella pullorum.
[0068] Experimental Example 3: Prevention and Treatment of Staphylococcus Infection in Mice
[0069] Seventy-two 8-week-old Kunming mice were divided into 6 groups, each with 12 mice, namely, postbiotic group, parabiotic group, parabiotic group, live bacteria group, blank control group, and challenge control group. Among them, each mouse in the postbiotic group, parabiotic group, parabiotic group, and live bacteria group was gavaged with 1 mL of each preparation of the corresponding group, the blank group was gavaged with 1 mL of normal saline, and the challenge control group was gavaged with 1 mL of normal saline. After gavage for 7 days, the intraperitoneal inoculation concentration of mice except the blank group was 2.4×10 8 CFU / mL of Staphylococcus aureus was 0.2mL. The mortality was observed within 24 hours after inoculation, and the morbidity and mortality rates were recorded and calculated. The results are shown in Table 3.
[0070] Table 3 Staphylococcus aureus protection experiment
[0071] Grouping Blank control group The challenge control group Epigenetic tuple Biological element group Parabiotic Group Live bacteria control group Incidence 100％ 100％ 100％ 100％ 100％ 100％ Case fatality rate 0％ 100％ 41.66％ 66.66％ 58.33％ 58.33％
[0072] As can be seen from Table 3, the postbiotic, parabiotic and parabiotic preparations have obvious effects in preventing and treating bacterial infections in animals. After mice were gavaged with the preparations, the active ingredients in the preparations regulated the immune function of the mice, enhanced the resistance of the mice to staphylococci, thereby reducing the mortality rate, and thus played a role in preventing and treating staphylococcal infection in mice.
[0073] It can be seen that the postbiotics, parabiotics and quasi-biotics prepared by the method of the present invention include metabolites and inactivated bacteria in the fermentation and cultivation process of bifidobacteria, which are different from traditional probiotics that need to colonize in the intestine for a period of time before they can take effect. Inactivated probiotics represent metabolites or bacterial components of probiotics after heat sealing treatment. Because of their small molecules, they can quickly enter the body through the intestinal mucosa and produce a number of benefits that help health, including enhancing immunity, balancing intestinal flora, regulating physiological functions, etc. The postbiotics, parabiotics and quasi-biotic preparations described in the present invention play a better role than live probiotics, and because of their clear chemical structure, long shelf life, higher safety and stronger stability, they can avoid many shortcomings of live bacteria and have a significant effect on preventing and treating pathogenic bacteria from infecting animals.
[0074] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.
Claims
1. A method for preparing postbiotics, characterized in that: The following steps are involved: Inoculating bifidobacteria into a culture medium for fermentation to obtain a fermentation liquid, and inactivating the fermentation liquid to obtain postbiotics; the fermentation temperature is 30 to 42° C., and the fermentation time is 30 to 40 hours; Each liter of the culture solution includes the following raw materials: 8-12 g of peptone, 4-6 g of beef powder, 18-22 g of glucose, 3-5 g of yeast powder, 4-6 g of sodium acetate, 1-3 g of dipotassium hydrogen phosphate, 0.1-0.5 g of magnesium sulfate, 1-3 g of triammonium citrate, 0.01-0.1 g of manganese sulfate, 0.5-1.5 mL of Tween 80 and 0.5-1.5 g of L-cysteine hydrochloride.
2. The preparation method according to claim 1, characterized in that: The bifidobacteria include Bifidobacterium bifidum.
3. The preparation method according to claim 1, characterized in that: The inactivation temperature is 60-85° C. and the inactivation time is 20-80 min.
4. The preparation method according to claim 3, characterized in that: The inactivation includes a first inactivation and a second inactivation; the first inactivation is performed at a temperature of 60 to 65° C. and a time of 20 to 40 minutes; the second inactivation is performed at a temperature of 80 to 85° C. and a time of 20 to 40 minutes.
5. Postbiotics prepared by the preparation method according to any one of claims 1 to 4.
6. A method for preparing parabiotics and parabiotics, characterized in that: The following steps are involved: Centrifuging the postbiotics according to claim 5 to obtain a supernatant and a precipitate; filtering the supernatant to obtain parabiotics; resuspending the precipitate with culture medium to obtain biotin; Each liter of the culture solution includes the following raw materials: 8-12 g of peptone, 4-6 g of beef powder, 18-22 g of glucose, 3-5 g of yeast powder, 4-6 g of sodium acetate, 1-3 g of dipotassium hydrogen phosphate, 0.1-0.5 g of magnesium sulfate, 1-3 g of triammonium citrate, 0.01-0.1 g of manganese sulfate, 0.5-1.5 mL of Tween 80 and 0.5-1.5 g of L-cysteine hydrochloride.
7. A parabiotic prepared by the preparation method according to claim 6.
8. The quasi-biotics prepared by the preparation method according to claim 6.
9. A preparation for preventing and treating bacterial infection in animals, characterized in that: The active ingredients of the preparation include: one or more of the postbiotics described in claim 5, the parabiotics described in claim 7, and the parabiotics described in claim 8.
10. The preparation according to claim 9, characterized in that The bacteria include: Salmonella and / or Staphylococcus.