CN105725215A - Lactobacillus oral solution and preparation method thereof - Google Patents
Lactobacillus oral solution and preparation method thereof Download PDFInfo
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- CN105725215A CN105725215A CN201610184087.1A CN201610184087A CN105725215A CN 105725215 A CN105725215 A CN 105725215A CN 201610184087 A CN201610184087 A CN 201610184087A CN 105725215 A CN105725215 A CN 105725215A
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- lactobacillus
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- oral liquid
- lactobacillus plantarum
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- 241000186660 Lactobacillus Species 0.000 title claims abstract description 34
- 229940039696 lactobacillus Drugs 0.000 title claims abstract description 34
- 238000002360 preparation method Methods 0.000 title claims abstract description 15
- 229940100688 oral solution Drugs 0.000 title abstract 4
- 238000000855 fermentation Methods 0.000 claims abstract description 28
- 230000004151 fermentation Effects 0.000 claims abstract description 28
- 238000000034 method Methods 0.000 claims abstract description 11
- 240000006024 Lactobacillus plantarum Species 0.000 claims description 39
- 235000013965 Lactobacillus plantarum Nutrition 0.000 claims description 39
- 229940072205 lactobacillus plantarum Drugs 0.000 claims description 39
- 239000007788 liquid Substances 0.000 claims description 39
- 239000012530 fluid Substances 0.000 claims description 9
- 230000001954 sterilising effect Effects 0.000 claims description 7
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 5
- 229910000396 dipotassium phosphate Inorganic materials 0.000 claims description 5
- 239000008103 glucose Substances 0.000 claims description 5
- CSNNHWWHGAXBCP-UHFFFAOYSA-L magnesium sulphate Substances [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 5
- 239000007836 KH2PO4 Substances 0.000 claims description 4
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 4
- 235000013575 mashed potatoes Nutrition 0.000 claims description 4
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 4
- 239000000843 powder Substances 0.000 claims description 4
- 239000005862 Whey Substances 0.000 claims description 3
- 102000007544 Whey Proteins Human genes 0.000 claims description 3
- 108010046377 Whey Proteins Proteins 0.000 claims description 3
- 238000004806 packaging method and process Methods 0.000 claims description 3
- 238000012859 sterile filling Methods 0.000 claims description 3
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 abstract description 27
- 235000012000 cholesterol Nutrition 0.000 abstract description 13
- 230000000694 effects Effects 0.000 abstract description 12
- 239000003833 bile salt Substances 0.000 abstract description 11
- 210000004211 gastric acid Anatomy 0.000 abstract description 9
- 230000004083 survival effect Effects 0.000 abstract description 7
- 230000036039 immunity Effects 0.000 abstract description 4
- 230000001965 increasing effect Effects 0.000 abstract description 4
- 239000008280 blood Substances 0.000 abstract description 3
- 210000004369 blood Anatomy 0.000 abstract description 3
- 230000036541 health Effects 0.000 abstract description 3
- 210000000936 intestine Anatomy 0.000 abstract 2
- 241000186000 Bifidobacterium Species 0.000 abstract 1
- 244000005700 microbiome Species 0.000 abstract 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 40
- 241000894006 Bacteria Species 0.000 description 31
- 239000004310 lactic acid Substances 0.000 description 20
- 235000014655 lactic acid Nutrition 0.000 description 20
- 239000006228 supernatant Substances 0.000 description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 11
- 229940093761 bile salts Drugs 0.000 description 10
- 239000001963 growth medium Substances 0.000 description 10
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 description 9
- 239000008055 phosphate buffer solution Substances 0.000 description 9
- 150000003254 radicals Chemical class 0.000 description 9
- 230000000968 intestinal effect Effects 0.000 description 8
- 240000001046 Lactobacillus acidophilus Species 0.000 description 7
- 235000013956 Lactobacillus acidophilus Nutrition 0.000 description 7
- 230000001580 bacterial effect Effects 0.000 description 7
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 239000000284 extract Substances 0.000 description 6
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 6
- 238000001556 precipitation Methods 0.000 description 6
- 239000006041 probiotic Substances 0.000 description 6
- 235000018291 probiotics Nutrition 0.000 description 6
- 210000002784 stomach Anatomy 0.000 description 6
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 5
- 210000003608 fece Anatomy 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- 239000002244 precipitate Substances 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 239000000725 suspension Substances 0.000 description 5
- 241000186016 Bifidobacterium bifidum Species 0.000 description 4
- 238000002835 absorbance Methods 0.000 description 4
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 230000012010 growth Effects 0.000 description 4
- 230000000529 probiotic effect Effects 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- 235000013618 yogurt Nutrition 0.000 description 4
- 108020004465 16S ribosomal RNA Proteins 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 3
- 244000199866 Lactobacillus casei Species 0.000 description 3
- 235000013958 Lactobacillus casei Nutrition 0.000 description 3
- 230000004913 activation Effects 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 125000004122 cyclic group Chemical group 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 230000002496 gastric effect Effects 0.000 description 3
- 229940017800 lactobacillus casei Drugs 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 235000013336 milk Nutrition 0.000 description 3
- 239000008267 milk Substances 0.000 description 3
- 210000004080 milk Anatomy 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- 230000003647 oxidation Effects 0.000 description 3
- 238000007254 oxidation reaction Methods 0.000 description 3
- 239000001301 oxygen Substances 0.000 description 3
- 229910052760 oxygen Inorganic materials 0.000 description 3
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical compound [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 description 2
- 230000003078 antioxidant effect Effects 0.000 description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 2
- 235000013361 beverage Nutrition 0.000 description 2
- 238000009395 breeding Methods 0.000 description 2
- 230000001488 breeding effect Effects 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 229960000935 dehydrated alcohol Drugs 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 239000012153 distilled water Substances 0.000 description 2
- 230000002708 enhancing effect Effects 0.000 description 2
- 230000003203 everyday effect Effects 0.000 description 2
- 230000006870 function Effects 0.000 description 2
- 210000000232 gallbladder Anatomy 0.000 description 2
- 230000007413 intestinal health Effects 0.000 description 2
- 229940039695 lactobacillus acidophilus Drugs 0.000 description 2
- 230000035790 physiological processes and functions Effects 0.000 description 2
- 238000012797 qualification Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 102000057297 Pepsin A Human genes 0.000 description 1
- 108090000284 Pepsin A Proteins 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- -1 alkane free radical Chemical class 0.000 description 1
- 229940040526 anhydrous sodium acetate Drugs 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
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- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229940099352 cholate Drugs 0.000 description 1
- BHQCQFFYRZLCQQ-OELDTZBJSA-N cholic acid Chemical compound C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)[C@@H](O)C1 BHQCQFFYRZLCQQ-OELDTZBJSA-N 0.000 description 1
- 230000009514 concussion Effects 0.000 description 1
- 239000006071 cream Substances 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 235000019797 dipotassium phosphate Nutrition 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 235000013345 egg yolk Nutrition 0.000 description 1
- 210000002969 egg yolk Anatomy 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 235000019634 flavors Nutrition 0.000 description 1
- 238000009920 food preservation Methods 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 238000009630 liquid culture Methods 0.000 description 1
- 230000033001 locomotion Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 235000013622 meat product Nutrition 0.000 description 1
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- 239000000203 mixture Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000003715 nutritional status Nutrition 0.000 description 1
- 230000000050 nutritive effect Effects 0.000 description 1
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- 229940111202 pepsin Drugs 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 238000011056 performance test Methods 0.000 description 1
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 1
- 229920000053 polysorbate 80 Polymers 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
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- 239000007787 solid Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000010025 steaming Methods 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 229930193551 sterin Natural products 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000025366 tissue development Effects 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 230000001228 trophic effect Effects 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 239000000273 veterinary drug Substances 0.000 description 1
- 239000012138 yeast extract 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
- 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
- A23V2400/169—Plantarum
-
- 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
- C12R2001/25—Lactobacillus plantarum
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Tropical Medicine & Parasitology (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to the technical field of application of microorganisms and particularly relates to a lactobacillus oral solution and a preparation method thereof. The oral solution is prepared through a plant lactobacillus fermentation method, and the viable count reaches 1-3 billion/mL. The collection number of adopted plant lactobacillus DY-1 is CCTCC (China Center For Type Culture Collection) NO:M2016137, the plant lactobacillus has very strong gastric acid resisting capability and bile salt resisting capability, and particularly, the survival rate in gastric acid reaches 165.41%; the strain can be used for effectively removing cholesterol and free radicals, and the removing rates reach 88.5% and 96.9% respectively; and the unexpected technical effect is obtained. When the lactobacillus oral solution provided by the invention is taken, the quantity of the lactobacillus and bifidobacterium in intestines of human bodies can be obviously increased, and the intestine health can be easily maintained; and the content of the cholesterol in blood of the human bodies can also be effectively reduced and the free radicals are eliminated, so that the immunity of organisms is enhanced and the market prospect is wide.
Description
Technical field
The present invention relates to technical field of microbe application, be specifically related to a kind of lactobacillus oral liquid and preparation method thereof.
Background technology
Lactic acid bacteria (lactic acid bacteria, LAB) is that a class can utilize fermentable carbohydrate to produce in a large number
The common name of the antibacterial of lactic acid, extremely wide in distributed in nature, there is abundant species diversity.Lactic acid bacteria is also widely present in
In human body and animal intestinal, numerous food product, material and minority clinical sample, it is possible not only to improve nutritive value of food, improves
Flavour of food products, improves food preservation and added value, and the special physiological of lactic acid bacteria is active and trophic function, the most day by day causes
The attention of people.Numerous studies show, lactic acid bacteria can regulate body gastrointestinal tract normal flora, keep microecological balance, improves
Food digestion rate and biological value, reduce serum cholesterol, controls endotoxin, and in suppression intestinal, putrefaction bacteria growth and breeding and corruption are produced
The generation of thing, manufactures nutrient substance, stimulates tissue development, thus to the nutritional status of body, physiological function, cell infection, medicine
The generation effects such as thing effect, toxic reaction, immunoreation, tumor generation, aging course and unexpected emergency reaction.Thus may be used
Seeing, the physiological function of lactic acid bacteria is closely bound up with the vital movement of body.
Lactic acid bacteria is mainly used in the fields such as yogurt, lactobacillus beverage, fermented type meat products, Medicines and Health Product.
Lactic acid bacteria industry has become one of health food and field of medicaments industry with fastest developing speed, and world market scale is more than 300,000,000,000
Dollar.The total scale of China's lactic acid bacteria industry has been over 16,000,000,000 yuan, and average year increasing degree is about 20%.Along with city
Popularizing of the most ripe and probiotic bacteria concept of field, the same with developed country, the probiotics fermention milk product of China will become
For the product that growth rate in dairy industry is the fastest.China's probiotic bacteria milk product year gross sales amount is more than 1,000,000,000 yuan of people at present
Coin, the yield of Yoghourt with 40% speed increment, the increasing degree of probiotics yogurt and lactobacillus beverage is higher than the growth of Yoghourt.
The performance of lactic acid bacteria directly determines the quality of its fermented product, therefore, sets up lactic acid bacteria culturers resources bank, and
Screen and develop probiotic lactobacillus strain on the basis of this to be particularly important.
Summary of the invention
It is an object of the invention to provide a kind of lactobacillus oral liquid and preparation method thereof.Described lactobacillus oral liquid is to pass through
Lactobacillus plantarum (Lactobacillus plantarum) fermentation prepares, and gastric acid and bile salts are had very by this bacterial strain
Strong toleration, and can effectively Cholesterol removal and free radical, oxidation resistance is strong, is conducive to safeguarding intestinal health, improves machine
Body immunity.
One aspect of the present invention provides a kind of lactobacillus oral liquid, and described lactobacillus oral liquid is by by Lactobacillus plantarum
(Lactobacillus plantarum) fermentation prepares.
Described Lactobacillus plantarum is Lactobacillus plantarum DY-1(Lactobacillus plantarum DY-1), in 2016
The China typical culture collection center that on March 22, in is preserved in Wuhan, China Wuhan University, preserving number is CCTCC NO:M
2016137。
In described lactobacillus oral liquid, viable count is 10-30 hundred million/mL.
Present invention also offers the preparation method of above-mentioned lactobacillus oral liquid, concrete steps include:
(1) preparation liquid fermentation medium, sterilizing；
(2) by the 3-5% of amount of fermentation, Lactobacillus plantarum is seeded in liquid fermentation medium；
(3) fermentation temperature 37～40 DEG C, fermentation time is 10～24 hours；
(4) fermentation liquid is carried out sterile filling, packaging, obtain lactobacillus oral liquid.
In described liquid fermentation medium, each component and mass volume ratio thereof are respectively as follows: brown sugar 6%, glucose 5%, milk surum
Powder 0.07%, mashed potatoes 0.04%, K2HPO4 0.03%、KH2PO4 0.02%、MgSO4 0.03%。
The original ph of described fluid medium is 5.5-6.5.
It is further preferred that described fermentation temperature is 37 DEG C.
The method that the present invention uses Lactobacillus plantarum to ferment prepares high-activity lactic acid bacteria oral liquid, and viable count is up to
10-30 hundred million/mL.The Lactobacillus plantarum DY-1 screening wherein used from the feces of normal adults, have the strongest stomach juice-resistant,
Resistance to bile salts ability, especially its survival rate in gastric acid are up to 165.41%；And this bacterial strain can effectively Cholesterol removal and
Free radical, clearance is up to 88.5% and 96.9% respectively, achieves unexpected technique effect.The breast that the edible present invention provides
Acid fungus oral liquid can substantially increase lactic acid bacteria and the quantity of bacillus bifidus in human body intestinal canal, is conducive to safeguarding intestinal health, moreover it is possible to
Effectively reduce the content of cholesterol in blood of human body, remove free radical, the immunity of enhancing body, wide market.
Detailed description of the invention
In addition to the bacterial strain that the present invention filters out, implement various raw materials used by the present invention, production equipment is all selected from city
Sell any one, the most special restriction.
Following example are in order to elaboration present invention is better described, and those skill in the art related can be by
Embodiment is more fully understood that and grasps the present invention.But, the protection of the present invention and right are not limited to provided case
Example.
The screening of embodiment 1 Lactobacillus plantarum and qualification
1, screening
(1) take several normal adults fresh excreta in the sterilizing triangular flask being built-in with bead, add 10 times of weight PBS and delay
Rush liquid and be placed in shaking table concussion 30min, then collect 2000g in centrifuge tube and be centrifuged 5min collection supernatant；
(2) above-mentioned supernatant 5000g is centrifuged 10min and collects precipitation, precipitate with twice collection of PBS cyclic washing；
(3) in the centrifuge tube that above-mentioned collection precipitates, add the isopyknic simulated gastric fluid of former supernatant fully suspend precipitation, 37 DEG C of water
Bath 30min；
(4) above-mentioned suspension 5000g is centrifuged 10min and collects precipitation, precipitate with twice collection of PBS cyclic washing；
(5) in the centrifuge tube that above-mentioned collection precipitates, add the isopyknic simulated intestinal fluid of former supernatant fully suspend precipitation, 37 DEG C of water
Bath 30min；
(6) above-mentioned suspension 5000g is centrifuged 10min and collects precipitation, precipitate with twice collection of PBS cyclic washing, finally
Fully suspend precipitation with a small amount of PBS；
(7) (glucose 20g, Carnis Bovis seu Bubali cream 10g, yeast extract 5g, peptone in MRS culture medium will be coated after above-mentioned suspension
10g, anhydrous sodium acetate 5g, calcium carbonate 1g, tween 80 1mL, K2HPO4 2g, agar 15-20g, saline solution A 5mL, distilled water
1000mL, pH6.5), cultivate 24～48 hours for 37 DEG C, with the bacterium colony respectively streak inoculation of transparent circle to MRS on picking flat board
In culture medium, cultivating 24～48 hours for 37 DEG C, the pure culture that repetitive operation obtains several times preserves.
2, identify
The lactic acid bacteria of above-mentioned screening is carried out stomach juice-resistant, the detection of resistance to bile salts ability, respectively to gastric acid, bile salts toleration all phases
Stronger bacterial strain is carried out molecular biology method qualification, records its 16S rDNA sequence.Then will be resistance to gastric acid and bile salts
Being compared by NCBI BLAST by the 16S rDNA sequence of the strongest bacterial strain DY-1 of property, result shows this sequence and plants
The 16S rDNA sequence homology of thing lactobacillus (Lactobacillus plantarum) is more than 99%, and therefore the present invention screens
DY-1 bacterial strain be Lactobacillus plantarum.
This Strain Designation is Lactobacillus plantarum DY-1(Lactobacillus plantarum DY-1 by applicant).And
In the China typical culture collection center that on March 22nd, 2016 is preserved in Wuhan, China Wuhan University, preserving number is CCTCC
NO:M2016137。
Embodiment 2 Lactobacillus plantarum DY-1 is to gastric acid and the toleration of bile salts
1, stomach juice-resistant method for testing performance:
Taking the Lactobacillus plantarum DY-1 bacteria suspension 40ml after activation and add in 50ml centrifuge tube, centrifugal (5000g, 5min) collects bacterium
Body, (compound method uses for reference the People's Republic of China (PRC) to wash twice addition 40ml simulated gastric fluid in backward centrifuge tube with PBS
The method of veterinary drug allusion quotation, takes dilute hydrochloric acid 16.4ml and adds water to shake up and be diluted to 1000ml, adjusts pH with the NaOH of concentrated hydrochloric acid or 10% and is
3.0, in 121 DEG C of sterilizing 30min, add pepsin in sterilizing room with 1g/100ml ratio), 2h is cultivated in 37 DEG C of water-baths, every
30min shakes up once, makees count plate in 0h and 2h is separately sampled, with 0h sample for comparison, calculates the survival of 2h sample thalline
Rate, survival rate=(viable count during viable count/0h during 2h) × 100%.
2, the method for testing performance of resistance to bile salts:
Taking the Lactobacillus plantarum DY-1 bacteria suspension 40ml after activation and add in 50ml centrifuge tube, centrifugal (5000g, 5min) collects bacterium
Body, washes twice addition 40ml simulated intestinal fluid (compound method: take KH in backward centrifuge tube with PBS2PO46.8g, adds steaming
Distilled water 500ml dissolve, with 0.4%(w/v) NaOH solution regulation pH value to 6.8, add water to 1000ml, every 100ml solution add
0.3g fowl cholate, after fully dissolving, in 115 DEG C of sterilizing 15min.), 2h is cultivated in 37 DEG C of water-baths, shakes up once every 30min, in
0h, 2h are separately sampled makees count plate, with 0h sample for comparison, and the survival rate of calculating 2h sample thalline, survival rate=(during 2h
Viable count during viable count/0h) × 100%.
Meanwhile, with bacillus acidophilus (Lactobacillus acidophilus) CGMCC1.1854 and lactobacillus casei
(Lactobacillus casei) CGMCC1.2435, as comparison, carries out above-mentioned stomach juice-resistant and the detection of resistance to bile salts performance.Tool
Body the results are shown in Table 1.
Table 1 stomach juice-resistant, resistance to bile salts performance test results
Data from table 1 it can be seen that with bacillus acidophilus CGMCC:1.1854 and lactobacillus casei CGMCC:1.2435 phase
Ratio, the Lactobacillus plantarum DY-1 that the present invention screens has the strongest stomach juice-resistant, resistance to bile salts ability, and especially it is at gastric acid
In survival rate be up to 165.41%, hence it is evident that higher than control strain, illustrate that Lactobacillus plantarum DY-1 is not only able to deposit in gastric acid
Live, the most a certain degree of breeding.
The embodiment 3 Lactobacillus plantarum DY-1 removal effect to cholesterol
This experiment is with reference to the method for (1998) such as Brashears, and slightly improves.
Being inoculated in MRS culture medium after being activated by Lactobacillus plantarum DY-1 bacterial strain, 37 DEG C of overnight incubation, as seed liquor.
Preparation is containing 5%(v/v) the MRS culture medium of egg yolk liquid, take wherein 2 ml culture medium, 5000 r/min are centrifuged 7min,
Taking 1ml supernatant to be placed in centrifuge tube, add 9ml dehydrated alcohol, oscillation treatment 5 min, 10000 r/min is centrifuged 10min,
Take 2 ml supernatant and add 2 ml P-Fe-S reagent, ice bath mixing reaction 30 min, survey OD550, calculate gallbladder in culture medium solid
The initial content of alcohol.
Then, the seed liquor of Lactobacillus plantarum DY-1 is seeded in above-mentioned MRS culture medium, under the conditions of 37 DEG C, cultivates 24
After h, again measure the final content of cholesterol in culture medium, calculate the Lactobacillus plantarum DY-1 clearance to cholesterol.Simultaneously
Using bacillus acidophilus (Lactobacillus acidophilus) CGMCC:1.1854 as control strain, use above-mentioned equally
Operation, calculate bacillus acidophilus's clearance to cholesterol.
Removal rate of cholesterol=(initial content-final content)/initial content × 100%
Result shows: the clearance of cholesterol in MRS culture medium is reached by the Lactobacillus plantarum DY-1 that the present invention screens
88.5%；And control strain bacillus acidophilus is only 19.8% to removal rate of cholesterol.Thus illustrate that Lactobacillus plantarum DY-1 is to gallbladder
The removal effect of sterin is preferable.
The antioxidant effect of embodiment 4 Lactobacillus plantarum DY-1
DPPH is the most stable a kind of free radical centered by nitrogen, if tested material can remove it, then it represents that tested material has
Reducing the ability of the free radicals such as hydroxy radical, alkane free radical or superoxide radical, antioxidant effect is notable.
After carrying out passing on activation by Lactobacillus plantarum DY-1, with 5%(mass ratio) inoculum concentration is inoculated in MRS liquid culture
Base, 37 DEG C of quiescent culture 18 h, 3 000 r/min, centrifugal 15 min, collect fermentation supernatant and thalline respectively.
Prepare cell-free extract: wash thalline 3 times with the phosphate buffer solution (PBS) that pH value is 7.4, again hang
Float in phosphate buffer solution, adjust bacterium number to 109 cfu /ml；Then ultrasound wave ice bath crushes thalline, 10000 r/min
Centrifugal 10 min, supernatant is cell-free extract.
The assay method of DPPH: first DPPH is dissolved in dehydrated alcohol, is prepared as the DPPH solution that concentration is 0.2M；
Respectively take 1 ml fermented supernatant fluid and cell-free extract, be separately added into the DPPH that 1 ml concentration is 0.2 mmol/L, mixing,
After left at room temperature 30min, at 517 nm, survey absorbance change.
The clearance rate (%) of DPPH=［ 1-(A1-A2)/A3 ］ × 100
In formula: A1 represents the original absorbance of the DPPH solution not adding sample;
A2 represents the sample absorbance when measuring wavelength own;
A3 represents the absorbance of DPPH solution after sample-adding.
Result shows: the clearance rate of DPPH is divided by fermented supernatant fluid and the cell-free extract of Lactobacillus plantarum DY-1
Not Gao Da 96.9% and 84.4%, thus the fermented liquid supernatant liquid of the Lactobacillus plantarum DY-1 that the present invention provides is described and acellular carries
Take thing and be respectively provided with stronger oxidation resistance, can effectively remove free radical, and the elimination effect of this strain fermentation supernatant is wanted
Apparently higher than cell-free extract, the removing of free radical is played by the metabolite during this explanation Lactobacillus plantarum DY-1 growth
Prior effect, oxidation resistance is higher.
Additionally, the Lactobacillus plantarum DY-1 fermented supernatant fluid of present invention offer and cell-free extract are to superoxide anion certainly
By base (O-2) also there is good removal effect, clearance respectively reaches 83.4% and 70.1%.Ultra-oxygen anion free radical is
First oxygen-derived free radicals, can generate other oxygen-derived free radicals by series reaction, has important biological function, and with multiple
Disease is closely related.Lactobacillus plantarum DY-1 of the present invention is to safeguarding that health will produce important effect.
The preparation method of embodiment 5 lactobacillus oral liquid
Preparation liquid fermentation medium: in fermentation tank add suitable quantity of water, respectively by brown sugar, glucose, whey powder, mashed potatoes,
K2HPO4、KH2PO4、MgSO4Join in tank, constant volume after stirring and dissolving.Utilize high-temperature steam control fermentation pressure inside the tank 0.09～
0.10MPa, makes temperature control at 115 DEG C～121 DEG C, sterilization 25～45min, cools to 37～40 DEG C, wherein after terminating
The mass volume ratio of each composition is finally respectively as follows: brown sugar 6%, glucose 5%, whey powder 0.07%, mashed potatoes 0.04%, K2HPO4
0.03%、KH2PO4 0.02%、MgSO40.03%, the original ph of culture medium is 5.5-6.5.
By amount of fermentation 3-5% inoculate above-mentioned Lactobacillus plantarum DY-1(CCTCC NO:M2016137) seed liquor in fermentation
In tank, stir 15min, fully close stirring after mixing；Controlling fermentation temperature is 37～40 DEG C, and fermentation time is 10～24 little
Time, cold water is cooled to rapidly less than 10 DEG C, carries out sterile filling, packaging, i.e. obtains lactobacillus oral liquid, wherein Lactobacillus plantarum
DY-1 viable count is up to 10-30 hundred million/mL.
The effect assessment of embodiment 6 lactobacillus oral liquid
From healthy population, random choose is grown up each 20 of men and women, collects its fresh excreta and detects lactic acid bacteria therein and bifid bar
Bacterium number average out to 2.2 × 108CFU/g and 8.1 × 109CFU/g.These 40 normal adults eat above-mentioned lactic acid bacteria for each person every day
Oral liquid 50 ml, detects lactic acid bacteria and bacillus bifidus number in its fresh excreta after one week.
Result shows: after edible lactobacillus oral liquid, lactic acid bacteria and bacillus bifidus in 40 normal adults fresh excretas
Number averagely reaches 9.2 × 109CFU/g and 8.9 × 1011CFU/g, is significantly larger than the bacterium number before edible lactobacillus oral liquid.From
And illustrate, the lactobacillus oral liquid that the edible present invention provides can substantially increase lactic acid bacteria and the number of bacillus bifidus in human body intestinal canal
Amount, is conducive to safeguarding that human body intestinal canal flora is healthy.
Additionally, study for a long period of time, data show, drink the lactobacillus oral liquid that the present invention provides every day, can not only be significantly increased
The quantity of probiotic bacteria in intestinal, improves gastrointestinal digestion and absorption function, moreover it is possible to effectively reduce cholesterol in blood of human body
Content, removes free radical, the immunity of enhancing body, safeguards healthy.
Claims (7)
1. a lactobacillus oral liquid, it is characterised in that described lactobacillus oral liquid is by by Lactobacillus plantarum
(Lactobacillus plantarum) carries out fermenting and prepares.
2. lactobacillus oral liquid as claimed in claim 1, it is characterised in that the preserving number of described Lactobacillus plantarum is
CCTCCNO:M 2016137。
3. lactobacillus oral liquid as claimed in claim 1 or 2, it is characterised in that viable count in described lactobacillus oral liquid
For 10-30 hundred million/mL.
4. the preparation method of a lactobacillus oral liquid, it is characterised in that described method comprises the steps:
(1) preparation liquid fermentation medium, sterilizing；
(2) by the 3-5% of amount of fermentation, Lactobacillus plantarum is seeded in liquid fermentation medium；
(3) fermentation temperature 37～40 DEG C, fermentation time 10～24 hours；
(4) fermentation liquid is carried out sterile filling, packaging, obtain lactobacillus oral liquid.
5. preparation method as claimed in claim 4, it is characterised in that each component and matter thereof in described liquid fermentation medium
Amount volume ratio is respectively as follows: brown sugar 6%, glucose 5%, whey powder 0.07%, mashed potatoes 0.04%, K2HPO40.03%,
KH2PO40.02%, MgSO40.03%.
6. preparation method as claimed in claim 5, it is characterised in that the original ph of described fluid medium is 5.5-
6.5。
7. the preparation method as described in claim 5 or 6, it is characterised in that described fermentation temperature is 37 DEG C.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107227278A (en) * | 2017-07-17 | 2017-10-03 | 浙江鸣食品股份有限公司 | A kind of Lactobacillus plantarum A11 and its application |
| CN107227277A (en) * | 2017-07-17 | 2017-10-03 | 浙江鸣食品股份有限公司 | A kind of Lactobacillus plantarum E680 and its application |
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| JP2007189973A (en) * | 2006-01-20 | 2007-08-02 | Nissin Food Prod Co Ltd | New lactic acid bacterium having blood cholesterol reducing action |
| CN102732470A (en) * | 2012-07-20 | 2012-10-17 | 光明乳业股份有限公司 | Purely natural and low-cost lactobacillus (Lb.) plantarum ST-III culture method and products thereof, and application of products |
| CN103636779A (en) * | 2013-12-17 | 2014-03-19 | 山东兴牛乳业有限公司 | Functional fermented milk and preparation method thereof |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2007189973A (en) * | 2006-01-20 | 2007-08-02 | Nissin Food Prod Co Ltd | New lactic acid bacterium having blood cholesterol reducing action |
| CN102732470A (en) * | 2012-07-20 | 2012-10-17 | 光明乳业股份有限公司 | Purely natural and low-cost lactobacillus (Lb.) plantarum ST-III culture method and products thereof, and application of products |
| CN103636779A (en) * | 2013-12-17 | 2014-03-19 | 山东兴牛乳业有限公司 | Functional fermented milk and preparation method thereof |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107227278A (en) * | 2017-07-17 | 2017-10-03 | 浙江鸣食品股份有限公司 | A kind of Lactobacillus plantarum A11 and its application |
| CN107227277A (en) * | 2017-07-17 | 2017-10-03 | 浙江鸣食品股份有限公司 | A kind of Lactobacillus plantarum E680 and its application |
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