CN111454862A - Lactobacillus paracasei freeze-dried powder with oral health function, preparation method and application - Google Patents
Lactobacillus paracasei freeze-dried powder with oral health function, preparation method and application Download PDFInfo
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- CN111454862A CN111454862A CN202010282419.6A CN202010282419A CN111454862A CN 111454862 A CN111454862 A CN 111454862A CN 202010282419 A CN202010282419 A CN 202010282419A CN 111454862 A CN111454862 A CN 111454862A
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- freeze
- lactobacillus paracasei
- dried powder
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- drying
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- 239000000843 powder Substances 0.000 title claims abstract description 57
- 241000186605 Lactobacillus paracasei Species 0.000 title claims abstract description 50
- 238000002360 preparation method Methods 0.000 title claims abstract description 24
- 230000036541 health Effects 0.000 title abstract description 14
- 206010006326 Breath odour Diseases 0.000 claims abstract description 8
- 208000032139 Halitosis Diseases 0.000 claims abstract description 8
- 230000002401 inhibitory effect Effects 0.000 claims abstract description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 25
- 238000004108 freeze drying Methods 0.000 claims description 24
- 241000894006 Bacteria Species 0.000 claims description 22
- 239000000047 product Substances 0.000 claims description 21
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- 238000000855 fermentation Methods 0.000 claims description 12
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- 238000009777 vacuum freeze-drying Methods 0.000 claims description 12
- 239000000126 substance Substances 0.000 claims description 11
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- 229940041514 candida albicans extract Drugs 0.000 claims description 8
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- 238000012216 screening Methods 0.000 abstract 1
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- 108020004465 16S ribosomal RNA Proteins 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 2
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 241001134770 Bifidobacterium animalis Species 0.000 description 2
- FERIUCNNQQJTOY-UHFFFAOYSA-N Butyric acid Chemical compound CCCC(O)=O FERIUCNNQQJTOY-UHFFFAOYSA-N 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- SIKJAQJRHWYJAI-UHFFFAOYSA-N Indole Chemical compound C1=CC=C2NC=CC2=C1 SIKJAQJRHWYJAI-UHFFFAOYSA-N 0.000 description 2
- JVTAAEKCZFNVCJ-REOHCLBHSA-N L-lactic acid Chemical compound C[C@H](O)C(O)=O JVTAAEKCZFNVCJ-REOHCLBHSA-N 0.000 description 2
- 244000199866 Lactobacillus casei Species 0.000 description 2
- 235000013958 Lactobacillus casei Nutrition 0.000 description 2
- 240000002605 Lactobacillus helveticus Species 0.000 description 2
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- 239000005913 Maltodextrin Substances 0.000 description 2
- 229920002774 Maltodextrin Polymers 0.000 description 2
- LSDPWZHWYPCBBB-UHFFFAOYSA-N Methanethiol Chemical compound SC LSDPWZHWYPCBBB-UHFFFAOYSA-N 0.000 description 2
- XBDQKXXYIPTUBI-UHFFFAOYSA-N Propionic acid Substances CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 2
- 241000194020 Streptococcus thermophilus Species 0.000 description 2
- 239000008272 agar Substances 0.000 description 2
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- 229940035034 maltodextrin Drugs 0.000 description 2
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- 238000000034 method Methods 0.000 description 2
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- 235000013618 yogurt Nutrition 0.000 description 2
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 description 1
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- 235000017166 Bambusa arundinacea Nutrition 0.000 description 1
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- 102000016938 Catalase Human genes 0.000 description 1
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- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 1
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241000204031 Mycoplasma Species 0.000 description 1
- 229910002651 NO3 Inorganic materials 0.000 description 1
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 description 1
- 208000025157 Oral disease Diseases 0.000 description 1
- 244000082204 Phyllostachys viridis Species 0.000 description 1
- 235000015334 Phyllostachys viridis Nutrition 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- KDYFGRWQOYBRFD-UHFFFAOYSA-N Succinic acid Natural products OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 description 1
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 1
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- 229930003268 Vitamin C Natural products 0.000 description 1
- 238000004220 aggregation Methods 0.000 description 1
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- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 1
- 239000011425 bamboo Substances 0.000 description 1
- HFACYLZERDEVSX-UHFFFAOYSA-N benzidine Chemical compound C1=CC(N)=CC=C1C1=CC=C(N)C=C1 HFACYLZERDEVSX-UHFFFAOYSA-N 0.000 description 1
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- KDYFGRWQOYBRFD-NUQCWPJISA-N butanedioic acid Chemical compound O[14C](=O)CC[14C](O)=O KDYFGRWQOYBRFD-NUQCWPJISA-N 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
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- PZOUSPYUWWUPPK-UHFFFAOYSA-N indole Natural products CC1=CC=CC2=C1C=CN2 PZOUSPYUWWUPPK-UHFFFAOYSA-N 0.000 description 1
- RKJUIXBNRJVNHR-UHFFFAOYSA-N indolenine Natural products C1=CC=C2CC=NC2=C1 RKJUIXBNRJVNHR-UHFFFAOYSA-N 0.000 description 1
- 239000000463 material Substances 0.000 description 1
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- 241001148471 unidentified anaerobic bacterium Species 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23G—COCOA; COCOA PRODUCTS, e.g. CHOCOLATE; SUBSTITUTES FOR COCOA OR COCOA PRODUCTS; CONFECTIONERY; CHEWING GUM; ICE-CREAM; PREPARATION THEREOF
- A23G4/00—Chewing gum
- A23G4/06—Chewing gum characterised by the composition containing organic or inorganic compounds
- A23G4/12—Chewing gum characterised by the composition containing organic or inorganic compounds containing microorganisms or enzymes; containing paramedical or dietetical agents, e.g. vitamins
- A23G4/123—Chewing gum characterised by the composition containing organic or inorganic compounds containing microorganisms or enzymes; containing paramedical or dietetical agents, e.g. vitamins containing microorganisms, enzymes
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/165—Paracasei
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- General Health & Medical Sciences (AREA)
- Virology (AREA)
- Medicinal Chemistry (AREA)
- Nutrition Science (AREA)
- Tropical Medicine & Parasitology (AREA)
- Polymers & Plastics (AREA)
- Food Science & Technology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Mycology (AREA)
- Inorganic Chemistry (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention discloses lactobacillus paracasei freeze-dried powder with an oral health care function, a preparation method and application thereof.A strain L PC48 is obtained by screening by an applicant, has a strong inhibiting effect on streptococcus mutans, has strong oral permanent planting capability and strong capability of inhibiting oral halitosis, can be used for oral health care, and greatly expands the application field of probiotics.
Description
Technical Field
The invention relates to lactobacillus paracasei freeze-dried powder with an oral health care function, a preparation method and application thereof. Belongs to the technical field of biological medicine processing.
Background
The oral cavity is a complex and complete microecological system, and the microorganisms planted in the oral cavity mainly comprise bacteria, fungi, mycoplasma, viruses, protozoa and the like, and the microorganisms inhabit, compete and antagonize at different parts of the oral cavity and form the oral cavity microecological system together with the inherent environment of the oral cavity. According to research, the microorganisms colonized in the oral cavity are mainly bacteria, and more than 700 species are found at present and are one of the most complex microorganism groups of the human body. At present, experts at home and abroad recognize that the adjustment of the balance of oral flora is the key to the prevention and treatment of various oral diseases.
The oral halitosis is mainly the result of the production of Volatile Sulfides (VSCs) by gram-negative anaerobes in the oral cavity through digestion of food residues, saliva, blood and other substances retained in the oral cavity, while hydrogen sulfide (H2S) and methyl mercaptan (CH3 SH) are the two most important types of VSCs, and research shows that probiotics can reduce the adhesion of oral pathogenic bacteria by competing with oral pathogenic bacteria for binding sites on the surface of oral mucosa or epithelial cells after entering the oral cavity environment. When the probiotic bacteria are integrated into the oral community, the extracellular polysaccharide produced by the probiotic bacteria and its strong aggregation capacity contribute to the formation of a biofilm in the oral environment. In such a complex ecological structure as a biofilm, probiotics can nonspecifically inhibit the growth of other microorganisms including oral pathogenic bacteria by secreting antibacterial substances such as hydrogen peroxide and organic acids. Meanwhile, when bacteria exist in a biological membrane system, the bacteria have strong resistance to external environmental changes, and are beneficial to propagation and survival in a complex oral environment. In addition, probiotics may exert their probiotic effects by modulating the immune system of the body. Therefore, it would be highly advantageous for oral health to screen suitable probiotics for the inhibition of oral-derived halitosis.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides lactobacillus paracasei freeze-dried powder with an oral health-care function, a preparation method and application thereof.
In order to achieve the purpose, the invention adopts the following technical scheme:
1. a lactobacillus paracasei is a lactobacillus paracasei (L actinobacillus paracasei) L PC48 which is preserved in China general microbiological culture Collection center (CGMCC) at 2-17.2020, with the preservation number of CGMCC No. 19422.
Preferably, the 16S rDNA sequence of the lactobacillus paracasei is shown as SEQ ID No. 1.
CAGGATGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAACGAGTTCTCGTT GATGATCGGTGCTTGCACCGAGATTCAACATGGAACGAGTGGCGGACGGGTGAGTAA CACGTGGGTAACCTGCCCTTAAGTGGGGGATAACATTTGGAAACAGATGCTAATACCG CATAGATCCAAGAACCGCATGGTTCTTGGCTGAAAGATGGCGTAAGCTATCGCTTTTG GATGGACCCGCGGCGTATTAGCTAGTTGGTGAGGTAATGGCTCACCAAGGCGATGATA CGTAGCCGAACTGAGAGGTTGATCGGCCACATTGGGACTGAGACACGGCCCAAACT CCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGCAAGTCTGATGGAGCAA CGCCGCGTGAGTGAAGAAGGCTTTCGGGTCGTAAAACTCTGTTGTTGGAGAAGAATG GTCGGCAGAGTAACTGTTGTCGGCGTGACGGTATCCAACCAGAAAGCCACGGCTAAC TACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTATCCGGATTTATTGGGC GTAAAGCGAGCGCAGGCGGTTTTTTAAGTCTGATGTGAAAGCCCTCGGCTTAACCGA GGAAGCGCATCGGAAACTGGGAAACTTGAGTGCAGAAGAGGACAGTGGAACTCCAT GTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACC CTGGTAGTCCATGCCGTAAACGATGAATGCTAGGTGTTGGAGGGTTTCCGCCCTTCAG TGCCGCAGCTAACGCATTAAGCATTCCGCCTGGGGAGTACGACCGCAAGGTTGAAAC TCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGC AACGCGAAGAACCTTACCAGGTCTTGACATCTTTTGATCACCTGAGAGATCAGGTTTC CCCTTCGGGGGCAAAATGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGA TGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATGACTAGTTGCCAGCATTTAGTT GGGCACTCTAGTAAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAA ATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGATGGTACAACGAG TTGCGAGACCGCGAGGTCAAGCTAATCTCTTAAAGCCATTCTCAGTTCGGACTGTAG GCTGCAACTCGCCTACACGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGC GGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTTTGTAA CACCCGAAGCCGGTGGCGTAACCCTTTTAGGGAGCGAGCCGTCTAAGGTGGGACAA A, as shown in SEQ ID NO. 1.
2. The application of the auxiliary cheese milk in preparing the oral health product for inhibiting the oral halitosis is provided.
Preferably, the oral health product comprises a daily chemical product and a food, wherein the daily chemical product is selected from an oral spray or a mouthwash, and the food is selected from a chewing gum or an oral lozenge.
3. The lactobacillus paracasei freeze-dried powder with the oral health care function is prepared by utilizing the lactobacillus paracasei.
4. The preparation method of the lactobacillus paracasei freeze-dried powder with the oral health care function comprises the steps of firstly inoculating the lactobacillus paracasei into a sterilized culture medium, fermenting for 24-72 hours at 35-44 ℃, centrifugally collecting precipitates, and carrying out vacuum freeze-drying to obtain the lactobacillus paracasei freeze-dried powder; wherein the medium comprises: 8-15 g of skimmed milk powder, 3-8 g of yeast extract, 15-25 g of glucose and K2HPO40.5～2g，CaCO31-3 g of distilled water (1000 m L).
Preferably, the specific steps are as follows:
(1) preparation of a culture medium: dissolving peptone, yeast extract, glucose and tween 80 in a formula amount in distilled water, and then sterilizing for 15-25 minutes at 120-125 ℃;
(2) fermenting by inoculating the Lactobacillus paracasei into the sterilized culture medium to obtain an original bacteria count of 1.0 × 104cfu/ml～1.0×105cfu/ml, fermentation to a bacterial count > 1.0 × 109cfu/ml；
(3) And (3) collecting thalli: collecting fermentation liquor, centrifuging for 4-12 minutes at the rotating speed of 4000-8000 r/min, and collecting bacterial precipitates by discarding supernatant;
(4) vacuum freeze drying: and (3) adding a freeze-drying protective agent and water into the precipitate collected in the step (3), then spreading the mixture in a pan for 1-3 cm, freezing the mixture for 12-15 hours at the temperature of-85 to-75 ℃, and freeze-drying the mixture for 18-30 hours by using a vacuum freeze-drying machine.
Further preferably, in the step (4), the composition of the lyoprotectant, water and the precipitate in percentage by weight is as follows: 85-90% of precipitate, 1.7-3.5% of freeze-drying protective agent and the balance of water.
Further preferably, in the step (4), the lyoprotectant is at least one selected from skim milk powder, trehalose, vitamin C and pectin.
Preferably, the vacuum freeze drying step is followed by a crushing operation to obtain powder, and then a diluent is added to obtain freeze-dried powder, and the freeze-dried powder is filled.
Further preferably, the diluent is maltodextrin or/and glucose, and the dosage of the diluent accounts for 70-90% of the weight of the freeze-dried powder.
5. The lactobacillus paracasei freeze-dried powder is applied to preparing oral health products for inhibiting oral halitosis.
Preferably, the oral health product comprises a daily chemical product and a food, wherein the daily chemical product is selected from an oral spray or a mouthwash, and the food is selected from a chewing gum or an oral lozenge.
Further preferably, in the oral spray or mouthwash, the content of the lactobacillus paracasei freeze-dried powder is 1-10% by weight.
Preferably, in the chewing gum or the buccal tablet, the weight content of the lactobacillus paracasei freeze-dried powder is 1-10%.
The applicant compares the bacteriostatic effect of the lactobacillus paracasei L PC48 of the present invention with that of other lactic acid bacteria on streptococcus mutans, and the results are shown in table 1, wherein lactobacillus casei 258, lactobacillus paracasei 213, lactobacillus plantarum L P28, lactobacillus helveticus L HR14, bifidobacterium animalis 342, and streptococcus thermophilus 912 are all purchased from bioscience, ltd (No. 66, north ridge, the gay region of luo bamboo, taiwan).
TABLE 1 comparison of the bacteriostatic effects on Streptococcus mutans
| Strain number | Bacteriostatic effect of streptococcus mutans |
| Lactobacillus casei 258 | ++ |
| Lactobacillus paracasei L PC48 | +++ |
| Lactobacillus paracasei 213 | ++ |
| Lactobacillus plantarum L P28 | ++ |
| Lactobacillus helveticus L HR14 | + |
| Bifidobacterium animalis 342 | - |
| Streptococcus thermophilus 912 | - |
Note: no inhibition; the diameter of the bacteriostatic circle is 7-10 mm (weak); a diameter of the antibacterial ring is 10-13 mm (good); bacteria inhibition circle diameter > 13mm (strong)
The invention has the beneficial effects that:
the applicant screens and obtains a lactobacillus paracasei L PC48, the strain has a strong inhibiting effect on streptococcus mutans, has strong oral permanent planting capability and strong capability of inhibiting oral halitosis, can be used for oral health care, and the application field of probiotics is greatly expanded.
Preservation information
And (3) classification and naming: lactobacillus paracasei
Name of Latin literature L actabacillus paracasei
The name of the depository: china general microbiological culture Collection center (CGMCC)
The address of the depository: xilu No.1 Hospital No. 3 of Beijing market facing Yang district
The preservation date is as follows: year 2020, 2 and 17
Detailed Description
The present invention will be further illustrated by the following examples, which are intended to be merely illustrative and not limitative.
Reference examples
The lactobacillus paracasei L PC48 of the invention is separated from dairy products and is obtained by the following method:
(1) preparation of MRS Medium
The MRS culture medium comprises the following specific components:
the preparation method comprises adding the above materials into 1000m L distilled water, heating to dissolve, adjusting pH, packaging, and autoclaving at 121 deg.C for 15-20 min.
(2) The lactobacillus paracasei L PC48 is separated from the yoghourt by the following separation method:
diluting yogurt sample in sterilized normal saline to a certain gradient, placing 0.1m L on MRS agar plate, coating surface with L bar, performing anaerobic culture at 37 deg.C for 48h, picking out characteristic colony on the plate with inoculating loop, and streaking and separating on new MRS agar plate.
The characteristics of the lactobacillus paracasei L PC48 obtained by the above method are as follows:
gram-positive, acid-fast, spore-forming, immotile obligate anaerobic bacteria, ferment glucose mainly to produce acetic acid and L (+) lactic acid in a 3:2 molar ratio, and also produce small amounts of formic acid and succinic acid, without producing CO2Propionic acid and butyric acid. Catalase negative, benzidine reaction and indole reaction negative, nitrate is not reduced. The molar content of G + C is between 57.2 and 64.5 percent.
The obtained 16S rDNA sequence of the lactobacillus paracasei L PC48 has a sequence structure shown in SEQ ID No.1, and the specific structure of the SEQ ID No.1 is as follows:
CAGGATGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAACGAGTTCTCGTT GATGATCGGTGCTTGCACCGAGATTCAACATGGAACGAGTGGCGGACGGGTGAGTAA CACGTGGGTAACCTGCCCTTAAGTGGGGGATAACATTTGGAAACAGATGCTAATACCG CATAGATCCAAGAACCGCATGGTTCTTGGCTGAAAGATGGCGTAAGCTATCGCTTTTG GATGGACCCGCGGCGTATTAGCTAGTTGGTGAGGTAATGGCTCACCAAGGCGATGATA CGTAGCCGAACTGAGAGGTTGATCGGCCACATTGGGACTGAGACACGGCCCAAACT CCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGCAAGTCTGATGGAGCAA CGCCGCGTGAGTGAAGAAGGCTTTCGGGTCGTAAAACTCTGTTGTTGGAGAAGAATG GTCGGCAGAGTAACTGTTGTCGGCGTGACGGTATCCAACCAGAAAGCCACGGCTAAC TACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTATCCGGATTTATTGGGC GTAAAGCGAGCGCAGGCGGTTTTTTAAGTCTGATGTGAAAGCCCTCGGCTTAACCGA GGAAGCGCATCGGAAACTGGGAAACTTGAGTGCAGAAGAGGACAGTGGAACTCCAT GTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACC CTGGTAGTCCATGCCGTAAACGATGAATGCTAGGTGTTGGAGGGTTTCCGCCCTTCAG TGCCGCAGCTAACGCATTAAGCATTCCGCCTGGGGAGTACGACCGCAAGGTTGAAAC TCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGC AACGCGAAGAACCTTACCAGGTCTTGACATCTTTTGATCACCTGAGAGATCAGGTTTC CCCTTCGGGGGCAAAATGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGA TGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATGACTAGTTGCCAGCATTTAGTT GGGCACTCTAGTAAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAA ATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGATGGTACAACGAG TTGCGAGACCGCGAGGTCAAGCTAATCTCTTAAAGCCATTCTCAGTTCGGACTGTAG GCTGCAACTCGCCTACACGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGC GGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTTTGTAA CACCCGAAGCCGGTGGCGTAACCCTTTTAGGGAGCGAGCCGTCTAAGGTGGGACAA A。
example 1:
(1) preparation of a culture medium:
mixing 15g of skimmed milk powder, 3g of yeast extract,glucose 25g, K2HPO42g，CaCO33g of the extract was dissolved in 1000m of L distilled water and sterilized at 125 ℃ for 25 minutes;
(2) high-density fermentation of lactic acid bacteria:
the culture medium after the sterilization was inoculated with the Lactobacillus paracasei L PC48 obtained in the reference example so that the number of original bacteria was 1.0 × 104cfu/ml, fermenting at 35 deg.C for 72 hr to make the number of bacteria > 1.0 × 109cfu/ml；
(3) And (3) collecting thalli: collecting lactobacillus fermentation liquor, centrifuging for 12 minutes at the rotating speed of 800r/min, discarding supernatant, and collecting bacterial precipitate;
(4) adding freeze-drying protective agent and water into the precipitate collected in the step (3), wherein the freeze-drying protective agent comprises 90% of bacterial sludge, 0.3% of VC, 0.8% of soybean phospholipid and 8.9% of water in percentage by weight,
(5) freeze drying of bacterial mud: adding a freeze-drying protective agent and water into the precipitate collected in the step (4), then spreading for 3cm, freezing at the temperature of a cold trap of-75 ℃ overnight, and freeze-drying for 30h by using a vacuum freeze-drying machine with the vacuum degree of 1 Pa;
(6) crushing and mixing: crushing the semi-finished freeze-dried powder obtained in the step (5) into powder, adding a certain amount of diluent, and canning to obtain a finished product; the freeze-dried powder comprises, by weight, 20% of a semi-finished product of the freeze-dried powder, 1.5% of maltodextrin and 78.5% of glucose.
(7) The lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) is applied to preparing foods such as chewing gum, tablets and the like, and the dosage of the freeze-dried powder is 10 percent of the total amount of the chewing gum.
Example 2:
(1) preparation of a culture medium:
mixing skimmed milk powder 15g, yeast extract 3g, glucose 25g, and K2HPO40.5g of CaCO 33 g was dissolved in 1000m of L distilled water and sterilized at 125 ℃ for 25 minutes;
(2) high-density fermentation of lactic acid bacteria:
inoculating Lactobacillus paracasei L PC48 into the sterilized culture medium, so that the original bacteria number is 1.0 × 104cfu/ml, fermenting at 35 deg.C for 24 hr to make the number of bacteria > 1.0 × 109cfu/ml；
(3) And (3) collecting thalli: collecting lactobacillus fermentation liquor, centrifuging for 4 minutes at the rotating speed of 800r/min, discarding the supernatant, and collecting the bacterial precipitate;
(4) adding freeze-drying protective agent and water into the precipitate collected in the step (3), wherein the mixture comprises 90% of bacterial sludge, 0.3% of VC, 0.8% of soybean lecithin and 8.9% of water,
(5) freeze drying of bacterial mud: and (3) adding a freeze-drying protective agent and water into the precipitate collected in the step (4), then spreading the mixture in a pan for 1.0-3 cm, freezing the mixture overnight at the temperature of-85 to-75 ℃, and freeze-drying the mixture for 18-30 h by using a vacuum freeze-drying machine. The temperature of the cold trap is-50 ℃, and the vacuum degree is 1 Pa;
(6) crushing and mixing: crushing the semi-finished freeze-dried powder obtained in the step (5) into powder, adding a certain amount of diluent, and canning to obtain a finished product; 20% of semi-finished freeze-dried powder, 1.5% of sorbitol and 78.5% of glucose.
(7) And (3) applying the lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) to preparation of foods such as chewing gum, tablets and the like, wherein the dosage of the freeze-dried powder is 1% of the total amount of the chewing gum.
(8) And (3) applying the lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) to preparation of daily chemical products such as mouth wash for oral cavity, wherein the dosage of the freeze-dried powder is 1% of the total amount of the fat food.
Example 3:
(1) preparation of a culture medium:
mixing skimmed milk powder 8g, yeast extract 8g, glucose 15g, and K2HPO42g，CaCO31g of the extract was dissolved in 1000m of L distilled water and sterilized at 125 ℃ for 15 minutes;
(2) high-density fermentation of lactic acid bacteria:
inoculating Lactobacillus paracasei L PC48 into the sterilized culture medium, so that the original bacteria number is 1.0 × 105cfu/ml, fermenting at 35 deg.C for 72 hr to make the number of bacteria > 1.0 × 109cfu/ml；
(3) And (3) collecting thalli: collecting lactobacillus fermentation liquor, centrifuging for 12 minutes at the rotating speed of 4000r/min, discarding supernatant, and collecting bacterial precipitate;
(4) adding freeze-drying protective agent and water into the precipitate collected in the step (3), wherein the freeze-drying protective agent comprises 90% of bacterial sludge, 0.3% of VC, 0.8% of soybean phospholipid and 8.9% of water in percentage by weight,
(5) freeze drying of bacterial mud: and (4) adding a freeze-drying protective agent and water into the precipitate collected in the step (4), then spreading the mixture on a tray for 1.0cm, freezing the mixture at the temperature of-75 ℃ overnight, and freeze-drying the mixture for 18-30 h by using a vacuum freeze-drying machine. The temperature of the cold trap is-50 ℃, and the vacuum degree is 1 Pa;
(6) crushing and mixing: crushing the semi-finished freeze-dried powder obtained in the step (5) into powder, adding a certain amount of diluent, and canning to obtain a finished product; the freeze-dried powder comprises, by weight, 20% of a semi-finished product of the freeze-dried powder, 1.5% of sorbitol and 78.5% of glucose.
(7) And (3) applying the lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) to preparation of foods such as chewing gum, tablets and the like, wherein the dosage of the freeze-dried powder is 1% of the total amount of the chewing gum.
And (3) applying the lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) to preparation of daily chemical products such as mouth wash for oral cavity, wherein the dosage of the freeze-dried powder is 10% of the total amount of the fat food.
Example 4:
(1) preparation of a culture medium:
mixing skimmed milk powder 15g, yeast extract 3g, glucose 25g, and K2HPO40.5g，CaCO33g of the extract was dissolved in 1000m of L distilled water and sterilized at 120 ℃ for 25 minutes;
(2) high-density fermentation of lactic acid bacteria:
inoculating Lactobacillus paracasei L PC48 into the sterilized culture medium, so that the original bacteria number is 1.0 × 104cfu/mlcfu/ml, fermenting at 44 deg.C for 24 hr to make the number of bacteria > 1.0 × 109cfu/ml；
(3) And (3) collecting thalli: collecting lactobacillus fermentation liquor, centrifuging for 4 minutes at the rotating speed of 8000r/min, discarding the supernatant, and collecting the bacterial precipitate;
(4) adding freeze-drying protective agent and water into the precipitate collected in the step (3), wherein the freeze-drying protective agent comprises 90% of bacterial sludge, 0.3% of VC, 0.8% of soybean phospholipid and 8.9% of water in percentage by weight,
(5) freeze drying of bacterial mud: and (4) adding a freeze-drying protective agent and water into the precipitate collected in the step (4), spreading for 3cm, freezing overnight at-85 ℃, and freeze-drying for 18-30 h by using a vacuum freeze-drying machine. The temperature of the cold trap is-50 ℃, and the vacuum degree is 1 Pa;
(6) crushing and mixing: crushing the semi-finished freeze-dried powder obtained in the step (5) into powder, adding a certain amount of diluent, and canning to obtain a finished product; the freeze-dried powder comprises, by weight, 20% of a semi-finished product of the freeze-dried powder, 1.5% of sorbitol and 78.5% of glucose.
(7) The lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) is applied to preparing foods such as chewing gum, tablets and the like, and the dosage of the freeze-dried powder is 10 percent of the total amount of the chewing gum.
(8) And (3) applying the lactobacillus paracasei L PC48 freeze-dried powder prepared in the step (6) to preparation of daily chemical products such as mouth wash for oral cavity, wherein the dosage of the freeze-dried powder is 1% of the total amount of the fat food.
Although the present invention has been described with reference to the specific embodiments, it is not intended to limit the scope of the present invention, and various modifications and variations can be made by those skilled in the art without inventive changes based on the technical solution of the present invention.
Sequence listing
<110> Biotechnology (Yangzhou) Ltd
<120> lactobacillus paracasei freeze-dried powder with oral health care function, preparation method and application
<141>2020-04-07
<160>1
<170>SIPOSequenceListing 1.0
<210>1
<211>1484
<212>DNA
<213>Artificial Sequence
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caggatgaac gctggcggcg tgcctaatac atgcaagtcg aacgagttct cgttgatgat 60
cggtgcttgc accgagattc aacatggaac gagtggcgga cgggtgagta acacgtgggt 120
aacctgccct taagtggggg ataacatttg gaaacagatg ctaataccgc atagatccaa 180
gaaccgcatg gttcttggct gaaagatggc gtaagctatc gcttttggat ggacccgcgg 240
cgtattagct agttggtgag gtaatggctc accaaggcga tgatacgtag ccgaactgag 300
aggttgatcg gccacattgg gactgagaca cggcccaaac tcctacggga ggcagcagta 360
gggaatcttc cacaatggac gcaagtctga tggagcaacg ccgcgtgagt gaagaaggct 420
ttcgggtcgt aaaactctgt tgttggagaa gaatggtcgg cagagtaact gttgtcggcg 480
tgacggtatc caaccagaaa gccacggcta actacgtgcc agcagccgcg gtaatacgta 540
ggtggcaagc gttatccgga tttattgggc gtaaagcgag cgcaggcggt tttttaagtc 600
tgatgtgaaa gccctcggct taaccgagga agcgcatcgg aaactgggaa acttgagtgc 660
agaagaggac agtggaactc catgtgtagc ggtgaaatgc gtagatatat ggaagaacac 720
cagtggcgaa ggcggctgtc tggtctgtaa ctgacgctga ggctcgaaag catgggtagc 780
gaacaggatt agataccctg gtagtccatg ccgtaaacga tgaatgctag gtgttggagg 840
gtttccgccc ttcagtgccg cagctaacgc attaagcatt ccgcctgggg agtacgaccg 900
caaggttgaa actcaaagga attgacgggg gcccgcacaa gcggtggagc atgtggttta 960
attcgaagca acgcgaagaa ccttaccagg tcttgacatc ttttgatcac ctgagagatc 1020
aggtttcccc ttcgggggca aaatgacagg tggtgcatgg ttgtcgtcag ctcgtgtcgt 1080
gagatgttgg gttaagtccc gcaacgagcg caacccttat gactagttgc cagcatttag 1140
ttgggcactc tagtaagact gccggtgaca aaccggagga aggtggggat gacgtcaaat 1200
catcatgccc cttatgacct gggctacaca cgtgctacaa tggatggtac aacgagttgc 1260
gagaccgcga ggtcaagcta atctcttaaa gccattctca gttcggactg taggctgcaa 1320
ctcgcctaca cgaagtcgga atcgctagtaatcgcggatc agcacgccgc ggtgaatacg 1380
ttcccgggcc ttgtacacac cgcccgtcac accatgagag tttgtaacac ccgaagccgg 1440
tggcgtaacc cttttaggga gcgagccgtc taaggtggga caaa 1484
Claims (10)
1. The lactobacillus paracasei is characterized in that the strain is lactobacillus paracasei (L actinobacillus paracasei) L PC48, which is preserved in China general microbiological culture Collection center (CGMCC NO. 19422) at 17.2.2020.
2. The lactobacillus paracasei according to claim 1, wherein the 16SrDNA sequence of the lactobacillus paracasei is shown as SEQ ID No. 1.
3. Use of a cheese milk of claim 1 or 2 for the manufacture of an oral care product for inhibiting halitosis of oral origin.
4. The use according to claim 3, wherein the oral care product comprises a daily chemical product selected from a mouth spray or a mouthwash and a food product selected from a chewing gum or a buccal tablet.
5. A freeze-dried powder of Lactobacillus paracasei prepared by using the Lactobacillus paracasei of claim 1 or 2.
6. The preparation method of the lactobacillus paracasei freeze-dried powder, according to claim 5, is characterized in that the lactobacillus paracasei is inoculated into a sterilized culture medium, fermented for 24-72 hours at 35-44 ℃, centrifuged to collect precipitates, and subjected to vacuum freeze drying to obtain the lactobacillus paracasei freeze-dried powder; wherein the medium comprises: 8-15 g of skimmed milk powder, 3-8 g of yeast extract, 15-25 g of glucose and K2HPO40.5～2g，CaCO31-3 g of distilled water (1000 m L).
7. The preparation method according to claim 6, comprising the following steps:
(1) preparation of a culture medium: dissolving peptone, yeast extract, glucose and tween 80 in a formula amount in distilled water, and then sterilizing for 15-25 minutes at 120-125 ℃;
(2) fermenting by inoculating the Lactobacillus paracasei into the sterilized culture medium to obtain an original bacteria count of 1.0 × 104cfu/ml～1.0×105cfu/ml, fermentation to a bacterial count > 1.0 × 109cfu/ml；
(3) And (3) collecting thalli: collecting fermentation liquor, centrifuging for 4-12 minutes at the rotating speed of 4000-7500 r/min, and collecting bacterial sediments by discarding supernatant;
(4) vacuum freeze drying: and (3) adding a freeze-drying protective agent and water into the precipitate collected in the step (3), then spreading the mixture in a pan for 1-3 cm, freezing the mixture for 12-15 hours at the temperature of-85 to-75 ℃, and freeze-drying the mixture for 18-30 hours by using a vacuum freeze-drying machine.
8. The preparation method according to claim 7, wherein the vacuum freeze-drying step is followed by a pulverization operation to obtain a powder, and the diluent is added to obtain a lyophilized powder, which is filled.
9. Use of a freeze-dried powder of lactobacillus paracasei according to claim 5 for the preparation of an oral care product for inhibiting halitosis of mouth origin.
10. The use according to claim 9, wherein the oral care product comprises a daily chemical product selected from a mouth spray or a mouthwash and a food product selected from a chewing gum or a buccal tablet.
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| CN115812882A (en) * | 2022-11-19 | 2023-03-21 | 西华大学 | Preparation method of kiwi fruit fermented beverage rich in protocatechuic acid and GABA |
| CN115812882B (en) * | 2022-11-19 | 2023-12-12 | 西华大学 | Preparation method of kiwi fruit fermented beverage rich in protocatechuic acid and GABA |
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