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CN-122012289-A - Preparation method of probiotic composition, preparation and application thereof

CN122012289ACN 122012289 ACN122012289 ACN 122012289ACN-122012289-A

Abstract

The application provides a preparation method of a probiotic composition and a preparation and application thereof, and the preparation comprises the following steps of respectively placing lactobacillus plantarum GOLDGUT-LP1024 strain, lactobacillus paracasei GOLDGUT-LC12345 strain, lactobacillus reuteri GOLDGUT-LR99 strain, pediococcus acidilactici GOLDGUT-PA0755 strain and lactobacillus rhamnosus GOLDGUT-L818 strain in a sterile MRS culture solution for activation, continuously passaging for a plurality of times to enable the strain to enter a logarithmic growth phase and expand and culture, centrifugally collecting bacterial solutions to obtain bacterial sludge of each strain, adding a protective agent into the bacterial sludge of each strain, freeze-drying, crushing and sieving to obtain freeze-dried powder of each strain, and mixing the freeze-dried powder of each strain with first lactobacillus compound powder and second lactobacillus compound powder according to parts by weight to obtain the probiotic composition with the effect of relieving diarrhea related to antibiotics.

Inventors

  • ZHONG SHUYING
  • ZHOU XIAOLI
  • ZHANG SILU
  • XIAO GUOXUN

Assignees

  • 深圳保时健生物工程有限公司

Dates

Publication Date
20260512
Application Date
20260408

Claims (10)

  1. 1. A method of preparing a probiotic composition, the probiotic composition being prepared by the method of: s1, respectively placing lactobacillus plantarum GOLDGUT-LP1024 strain, lactobacillus paracasei GOLDGUT-LC12345 strain, lactobacillus reuteri GOLDGUT-LR99 strain, pediococcus acidilactici GOLDGUT-PA0755 strain and lactobacillus rhamnosus GOLDGUT-L818 strain in a 37 ℃ sterile MRS culture solution for activation for 18-24 hours, continuously carrying out 2-3 times to enable the strains to enter a logarithmic growth phase, and carrying out expanded culture; S2, at 2-8 ℃ and 6000 DEG C Centrifuging the bacterial liquid at 12000rpm for 15 Adding protective agent into the bacterial sludge of each strain, freeze-drying, pulverizing and sieving to obtain freeze-dried powder of each strain; S3, selecting 0.8-1 part of lactobacillus plantarum GOLDGUT-LP1024 powder, 0.2-0.3 part of lactobacillus paracasei GOLDGUT-LC12345 powder, 1.5-2 parts of lactobacillus reuteri GOLDGUT-LR99 powder, 0.1-0.2 part of pediococcus acidilactici GOLDGUT-PA0755 powder and 0.3-0.4 part of lactobacillus rhamnosus GOLDGUT-L818 powder, fully mixing with 2-3 parts of first lactobacillus compound powder and 3-4 parts of second lactobacillus compound powder to obtain the probiotic composition, The first lactobacillus compound powder is obtained by mixing lactobacillus acidophilus NCFM powder, bifidobacterium animalis subspecies B1-04 powder, lactobacillus plantarum Lp116 powder, bifidobacterium bifidum Bb-06 powder and bifidobacterium longum subspecies infant Bi-26 powder; The second lactobacillus compound powder is obtained by mixing bifidobacterium longum subspecies BORI powder, lactobacillus acidophilus AD031 powder, bifidobacterium bifidum BGN4 powder, bifidobacterium animalis subspecies lactis AD011 powder and bifidobacterium longum subspecies infantis IBS007 powder.
  2. 2. The method of claim 1, wherein the probiotic composition has a viable count of greater than or equal to 2.5X1011 CFU/g.
  3. 3. The preparation method of the compound bacteria of the invention according to claim 1, wherein the expanding culture in the step S1 comprises the steps of respectively inoculating bacterial liquid into a fermentation tank according to the inoculation amount of 3% -8%, continuously culturing and fermenting for 20-40h, and rapidly cooling the bacterial liquid to 15-25 ℃ after the fermentation is finished; The pH value of the bacterial liquid in the fermentation tank is 6.0-7.5 in the culture fermentation process, and the culture temperature is 35-39 ℃.
  4. 4. The method of claim 1, wherein the activating in step S1 comprises adding a supplement to the sterile MRS broth, respectively, the supplement comprising one or more of a nitrogen source supplement, a carbon source supplement, and a buffer, the supplement being present in the sterile MRS broth at a mass volume ratio of no more than 1%; The nitrogen source supplement comprises one or more of yeast extract, beef extract, fish peptone, soybean peptone and corn steep liquor dry powder, the carbon source supplement comprises one or more of trehalose and glucose, and the buffer comprises one or more of tri-ammonium citrate and sodium acetate.
  5. 5. The preparation method of the bacterial sludge of each bacterial strain according to claim 1, wherein the collection in the step S2 comprises the steps of centrifuging the bacterial liquid, discarding supernatant, adding sterile PBS buffer solution into the precipitate, gently stirring and washing, centrifuging again under the same centrifugal condition, repeatedly washing for 1-3 times, and collecting bacterial sludge of each bacterial strain, wherein the pH value of the sterile PBS buffer solution is 7.0-7.4.
  6. 6. The preparation method according to claim 1, wherein in the step S2, the mixture is stirred at 20 ℃ to 30 ℃ for 25 to 40 minutes after the protective agent is added, and the mass ratio of the bacterial sludge of each strain to the protective agent is 1:2 to 3; the protective agent comprises one or more of a carbon source substance, a nitrogen source substance, a pH regulator, a stabilizer, a prebiotic, an osmotic pressure regulator, an amino acid substance and sterile water.
  7. 7. The preparation method of the probiotic composition according to claim 1, wherein the mesh number of the sieving in the step S2 is 60-120 mesh, and the bifidobacterium animalis milk subspecies B1-04 bacterial powder, bifidobacterium bifidum Bb-06 bacterial powder, bifidobacterium longum subspecies infant Bi-26 bacterial powder, bifidobacterium longum subspecies BORI bacterial powder, bifidobacterium bifidum BGN4 bacterial powder, bifidobacterium animalis milk subspecies AD011 bacterial powder and bifidobacterium longum subspecies infant IBS007 bacterial powder are obtained by 60-80 mesh sieving; The lactobacillus acidophilus NCFM powder, lactobacillus plantarum Lp116 powder, lactobacillus acidophilus AD031 powder, lactobacillus plantarum GOLDGUT-LP1024 powder, lactobacillus paracasei GOLDGUT-LC12345 powder, lactobacillus reuteri GOLDGUT-LR99 powder and lactobacillus rhamnosus GOLDGUT-L818 powder are obtained by 60-100 mesh screening and crushing; The pediococcus acidilactici GOLDGUT-PA0755 bacterial powder is obtained by crushing through a 60-120 mesh sieve, wherein, And (3) sieving the bacterial powder of each bacterial strain in the first lactobacillus compound powder and the second lactobacillus compound powder according to the sieving mesh number in advance, and then mixing and compounding.
  8. 8. The method of claim 1, wherein said thoroughly mixing in step S3 results in said probiotic composition comprising: Selecting 2-3 parts of first lactobacillus compound powder and 3-4 parts of second lactobacillus compound powder, mixing, and Adding lactobacillus reuteri GOLDGUT-LR99 powder 1.5-2 parts, lactobacillus plantarum GOLDGUT-LP1024 powder 0.8-1 part and lactobacillus rhamnosus GOLDGUT-L818 powder 0.3-0.4 part, mixing, and mixing Adding 0.2-0.3 part of Lactobacillus paracasei GOLDGUT-LC12345 powder and 0.1-0.2 part of Pediococcus acidilactici GOLDGUT-PA0755 powder, and mixing for 5 times And (5) 15min to obtain the probiotic composition.
  9. 9. A probiotic preparation, characterized in that it comprises a probiotic composition prepared by the preparation method according to any one of claims 1 to 8, and pharmaceutically or food acceptable excipients.
  10. 10. Use of a probiotic formulation according to claim 9 for the preparation of a product for alleviating antibiotic-associated diarrhea, wherein said product comprises a pharmaceutical product, a nutraceutical or a food product.

Description

Preparation method of probiotic composition, preparation and application thereof Technical Field The application belongs to the technical field of microorganisms, and particularly relates to a preparation method of a probiotic composition, a preparation and application thereof. Background Antibiotics are a core means for treating infectious diseases in modern medicine, greatly improve the health level and average life expectancy of human beings, and are widely applied to the fields of clinical diagnosis and treatment, livestock breeding and the like, and the use amount of the antibiotics in the world is continuously rising. The intestinal microbiota of human body is a micro-ecological system with complex structure, and deeply participates in a plurality of physiological processes such as host immune development, metabolism regulation, intestinal barrier maintenance, pathogen resistance and the like, but broad-spectrum antibiotics do not have accurate targeting, and can generate nonselective inhibition or killing effect on symbiotic beneficial microorganisms in intestinal tracts while killing pathogenic bacteria, so that the caused imbalance of intestinal microbiota becomes a common problem after the use of antibiotics, and the diarrhea related to antibiotics is one of the most typical adverse reactions, and the intestinal micro-ecological intervention strategy based on probiotics also becomes a research hot spot in the field of gastrointestinal disease prevention and treatment. The use of antibiotics can cause the decrease of intestinal microorganism diversity and the unbalance of flora structure, and the interference to intestinal microecology has certain persistence, and the dominant intestinal flora is difficult to quickly restore to the original state after stopping the medicine, thereby causing the problems of intestinal barrier function injury, inflammatory reaction and the like, becoming the core cause of occurrence of antibiotic-related diarrhea, and bringing a plurality of challenges to clinical intervention. At present, the intervention means for the antibiotic-associated diarrhea still has obvious limitation, the traditional antidiarrheal drug can only relieve diarrhea manifestation from the symptom level, the unbalance of intestinal flora and the damage of mucous membrane barrier caused by antibiotics cannot be repaired from the root, while probiotics are proved to have potential for regulating intestinal microecology, but the traditional probiotics preparation is mostly single strain or simple compound strain, has limited suitability and colonisation capability, and the traditional culture and preservation process cannot fully consider the physiological characteristic difference of different probiotics, so that the activity of the strain is difficult to be effectively maintained, and the effect stability is insufficient when the antibiotic-associated diarrhea is relieved. Disclosure of Invention The application provides a preparation method of a probiotic composition and a preparation and application thereof, which are characterized in that differential special supplements are added to different probiotic bacterial strain characteristics and personalized culture condition regulation and control are carried out, the protective agent treatment and low-temperature freeze-drying process of the strain types are matched, the probiotic composition is sieved and crushed according to the classification of the bacterial morphology and characteristics, the stepwise accurate compounding of the bacterial ratio, the particle size and the fluidity is combined, the bacterial mud preparation means of low-temperature centrifugal purification is assisted, the living bacterial activity and the biological function of each bacterial strain are reserved to the maximum extent, the uniformity, the stability and the field planting capability of the probiotic composition are ensured, and finally, the purposes of effectively regulating the host intestinal flora structure, improving the intestinal flora diversity, repairing the intestinal mucosa barrier and regulating the organism inflammatory reaction are realized, so that the diarrhea related to antibiotics is effectively relieved, and meanwhile, the standardized and repeatable preparation process support is provided for the application of the probiotic composition in the fields of medicines, health products, foods and the like. In a first aspect, embodiments of the present application provide a method for preparing a probiotic composition, the probiotic composition being prepared by: s1, respectively placing lactobacillus plantarum GOLDGUT-LP1024 strain, lactobacillus paracasei GOLDGUT-LC12345 strain, lactobacillus reuteri GOLDGUT-LR99 strain, pediococcus acidilactici GOLDGUT-PA0755 strain and lactobacillus rhamnosus GOLDGUT-L818 strain in a 37 ℃ sterile MRS culture solution for activation for 18-24 hours, continuously carrying out 2-3 times to enable the strains to ent