CN106883995A - Pediococcus acidilactici JQII-5 bacterial strains and application thereof - Google Patents
Pediococcus acidilactici JQII-5 bacterial strains and application thereof Download PDFInfo
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- CN106883995A CN106883995A CN201510936910.5A CN201510936910A CN106883995A CN 106883995 A CN106883995 A CN 106883995A CN 201510936910 A CN201510936910 A CN 201510936910A CN 106883995 A CN106883995 A CN 106883995A
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- 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
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- A—HUMAN NECESSITIES
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- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING OR TREATMENT THEREOF
- A23C11/00—Milk substitutes, e.g. coffee whitener compositions
- A23C11/02—Milk substitutes, e.g. coffee whitener compositions containing at least one non-milk component as source of fats or proteins
- A23C11/10—Milk substitutes, e.g. coffee whitener compositions containing at least one non-milk component as source of fats or proteins containing or not lactose but no other milk components as source of fats, carbohydrates or proteins
- A23C11/103—Milk substitutes, e.g. coffee whitener compositions containing at least one non-milk component as source of fats or proteins containing or not lactose but no other milk components as source of fats, carbohydrates or proteins containing only proteins from pulses, oilseeds or nuts, e.g. nut milk
- A23C11/106—Addition of, or treatment with, microorganisms
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
- A61K35/741—Probiotics
- A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
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- 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
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Abstract
The invention discloses a kind of Pediococcus acidilactici (Pediococcus acidilactici) JQII-5 and application thereof, specifically disclose Pediococcus acidilactici and its purposes in food or medicine is prepared, food and medicine, wherein the Pediococcus acidilactici has carried out preservation on 2 2nd, 2015 in China Committee for Culture Collection of Microorganisms's common micro-organisms center, and preserving number is CGMCC No.10512.Pediococcus acidilactici JQII-5 of the invention has excellent prebiotic effect, and acidproof and bile tolerance ability can be preserved for a long time, and can be effective for reducing Blood Cholesterol and triglycerides.
Description
Technical field
The present invention relates to microorganism field, in particular it relates to Pediococcus acidilactici and application thereof, more specifically, is related to lactic acid sheet
Coccus and application thereof, purposes, food and medicine more particularly, to Pediococcus acidilactici and its in food or medicine is prepared.
Background technology
Hypercholesterolemia is also referred to as high low density lipoprotein cholesterolemia, is to cause atherosclerotic cardiovascular disease
Primary hazards.Current research shows that various cardiovascular risk crowd low-density lipoproteins often reduce 1mmol/L, main
Wanting vascular events can reduce by 21%.Shown according to the statistics of 2013, China there are about 200,000,000 critical hypercholesterolemia crowds and
88000000 hypercholesterolemiapatients patients.And in China adult the awareness of high cholesterol, treatment rate, control rate be only 11%,
5.1% and 2.8%.In the 4th Chinese chronic diseases management conference cardiovascular disease prevention forum, multidigit expert call, policy
Maker, medical worker and the public will be appreciated that important function of the control blood fat in cardiovascular disease prevention, really
Accomplish the integrated management of hazards.
Prevention of cardiovascular disease is currently directed to, the meals principle in accordance with low fat or low saturated fat food is advocated.It is substantial amounts of
It is demonstrated experimentally that low-fat diet can effectively reduce the content of cholesterol in serum, but the taste and flavor of low-fat diet is fairly simple,
This diet suggestion is not accepted by the public or adheres to for a long time yet, therefore crowd's serum cholesterol level is in increase trend year by year.By
In the low acceptance that LF is advocated, people's hope develops other can be with reduce serum cholesterol and the meal supplement of blood fat
Product.
Find that lactic acid bacteria can reduce the discovery that body inner cholesterol content comes from last century the seventies Mann earliest, he
It was found that drinks fermented dairy product people has relatively low cholesterol level in vivo, in 1985, researcher Gilliland etc. carried out
Lactobacillus acidophilus carries out the research of external norcholesterol, as a result find the bacterium can reduce zymotic fluid in cholesterol level.
Grill etc. takes lactobacillus acidophilus and galactenzyme to the higher patient of serum total cholesterol level, after 8 months
It was found that tested crowd's serum cholesterol content is remarkably decreased.The country is directed to more deep two of norcholesterol functional study at present
Individual bacterial strain belongs to Lactobacillus plantarum, respectively L.plantarum P8 and Lactobacillus plantarum ST-III.L.plantarum P8
The team of the peace leader from Agricultural University of the Inner Mongol, animal experiment shows that L.plantarum P8 have significant drop
Blood fat.Zeng Ren Northeast Agricultural Universities Foodstuffs Academy vice-president, incumbent Shanghai Bright Dairy ＆ Food Co., Ltd. general manager Guo Benheng
The team of leader and the Lactobacillus plantarum ST-III of Southern Yangtze University's R ＆ D Cooperation, on the basis of a large amount of basic research, successfully lead to
Cross the effect for confirming its norcholesterol of zoopery and crowd's clinical test, and industrialization, one as bright company
Probiotic products with independent intellectual property right.This also fully indicates the lactic acid bacteria with norcholesterol in fermented food field
It is with a wide range of applications.
The content of the invention
The purpose of the present invention is to alleviate the growing severe trend of domestic people with hyperlipidemia, there is provided there is one kind drop courage to consolidate
The Pediococcus acidilactici JQII-5 of alcohol, triglyceride reducing function, and its preparing the purposes of the aspects such as food or medicine.
The first aspect of the present invention, the invention discloses a kind of Pediococcus acidilactici, during it was preserved on 2 2nd, 2015
State's Microbiological Culture Collection administration committee common micro-organisms center, the address of depositary institution is BeiChen West Road, Chaoyang District, BeiJing City
No. 3 Institute of Microorganism, Academia Sinica of No. 1 institute, preserving number is CGMCC No.10512, the entitled Pediococcus acidilactici of preservation
(Pediococcus acidilactici)JQII-5.Embodiments in accordance with the present invention, Pediococcus acidilactici JQII-5 of the invention
Not only there is excellent prebiotic effect, also acidproof and bile tolerance ability can be preserved for a long time, and can be effective for reducing blood
Fat, specifically, can effectively reduce Blood Cholesterol and triglycerides.
The second aspect of the present invention, the invention provides purposes of the Pediococcus acidilactici in food or medicine is prepared.
Embodiments in accordance with the present invention, Pediococcus acidilactici of the invention is provided to animal, being capable of effectively reduction blood fat, especially cholesterol
And triglycerides.Thus, the food or medicine are used for reduction blood fat.
The third aspect of the present invention, the present invention provides a kind of composition.The composition includes lactic acid sheet described above
Coccus, being capable of effectively reduction blood fat, especially cholesterol and triglycerides.Thus, the composition is used for reduction blood fat.
The fourth aspect of the present invention, the present invention provides a kind of medicine.The medicine is included：Foregoing lactic acid sheet ball
Bacterium；And pharmaceutical non-toxic carrier.Embodiments in accordance with the present invention, it is described to be not particularly limited with non-toxic carrier, as long as energy
The medicine is enough set to form the formulation of easy administration, those skilled in the art can flexibly select as the case may be.Root
According to some specific examples of the invention, the pharmaceutical non-toxic carrier be selected from glucose, lactose, sucrose, starch, mannitol,
Dextrin, fatty glyceride, polyethylene glycol, HES, ethylene glycol, polyoxyethylene sorbitan fatty acid ester, amino
Acid, gelatin, albumin, water, at least one of normal saline solution.Thus, medicine of the invention can be effectively formed easily administration
Formulation, be easy to administration.
Embodiments in accordance with the present invention, medicine of the invention is further included：Stabilizer, wetting agent, emulsifying agent, bonding
Agent, at least one of isotonic agent.Specifically, the medicine is compound probiotic piece.The compound probiotic piece includes：It is noted earlier
Pediococcus acidilactici, streptococcus thermophilus, Lactobacillus rhamnosus, Lactobacillus casei, lactobacillus acidophilus, Lactobacillus plantarum, pentose piece
Coccus, inulin, dietary fiber (resistant dextrin), D-sorbite, microcrystalline cellulose, magnesium stearate, after being mixed uniformly, compressing tablet
It is made tablet.
Embodiments in accordance with the present invention, the dosage of the medicine is not particularly limited, in practical application, can basis
The health status of administration object is flexibly selected.Some embodiments of the invention, by the medicine with 2.0 × 109CFU lives
The dosage in bacterium/day is administered to animal.Thus, the cholesterol level and content of triglyceride for being administered animal are substantially reduced.
The fifth aspect of the present invention, additionally provides a kind of food.Embodiments in accordance with the present invention, the food is included：Before
Pediococcus acidilactici described in face；And acceptable additive in bromatology.Specifically, the food is acid bean milk drink.Institute
The acid bean milk drink stated is included：Bean powder, inulin, white granulated sugar, lactobacillus bulgaricus and lactobacillus thermophilus, ferment 10h at 42 DEG C,
It is subsequently adding 0.1% foregoing Pediococcus acidilactici and 0.1% Pediococcus pentosaceus.With by the effect of low blood fat, especially
It is to reduce cholesterol and triglycerides.
It should be noted that term " food " used herein should broadly understood, it can any can be eaten
Form, i.e., in addition to conventional food form, food of the invention can also be health products, drink, fermented food etc., and
And, fermented food also includes the feed of animal.
Embodiments in accordance with the present invention, the intake dosage of the food is not particularly limited, in practical application, can basis
Actual state is flexibly selected.Some embodiments of the invention, by the food with 2.0 × 109The dosage in CFU viable bacterias/day
It is supplied to animal.Thus, the cholesterol level and content of triglyceride for taking the animal of food of the present invention are substantially reduced.
In addition, it is necessary to explanation, Pediococcus acidilactici of the invention can be with the food material used in common food
Material is matched.For example, cereal and potato class：Cereal includes rice, face, coarse cereals, and potato class include potato, Ipomoea batatas etc.；Animality is eaten
Thing, including meat, fowl, fish, milk, egg etc.；Beans and its product, including soybean and other dry beans；Vegetable and fruit class, including it is fresh
Beans, rhizome, leaf vegetables, solanberry etc.；Pure heat energy food, including vegetable and animals oils, starch, table sugar and drinks etc..
Additional aspect of the invention and advantage will be set forth in part in the description, and will partly become from the following description
Obtain substantially, or recognized by practice of the invention.
Brief description of the drawings
Of the invention above-mentioned and/or additional aspect and advantage will become from description of the accompanying drawings below to embodiment is combined
Obtain easier to understand：
Fig. 1：Aspect graph under the light microscopic of Pediococcus acidilactici JQII-5；
Fig. 2：Aspect graph under 10000 times of Electronic Speculum of Pediococcus acidilactici JQII-5；
Fig. 3：The result figure of the external Cholesterol removals of Pediococcus acidilactici JQII-5.Wherein：Ordinate represents cholesterol degradation
Rate, abscissa represents group.
Fig. 4：The external acid resistance results figures of Pediococcus acidilactici JQII-5.
Fig. 5：The external cholate resistance results figures of Pediococcus acidilactici JQII-5.
Fig. 6：Pediococcus acidilactici JQII-5 triglyceride reducing (Triglyceride, TG) result figures in animal body.Wherein：
Ordinate represents the content (mmol/L) of triglycerides (TG)；Abscissa represents different groups.
Fig. 7：Pediococcus acidilactici JQII-5 norcholesterol (total cholesterol, TC) result figures in animal body.Its
In：Ordinate represents the content (mmol/L) of cholesterol (TC)；Abscissa represents different groups.
Fig. 8：Soy yogurt feeding hyperlipidemia model rat, the changes of contents figure of cholesterol (TC) in rat body.
Fig. 9：Soy yogurt feeding hyperlipidemia model rat, the changes of contents figure of triglycerides (TG) in rat body.
Specific embodiment
Probiotics is colonized in human body intestinal canal, genital tract, can be produced definite health efficacy and be put down so as to improve host's Tiny ecosystem
Weighing apparatus, performance beneficial effect.Probiotics in human body, animal body mainly has：Lactic acid bacteria, Bifidobacterium, lactobacillus acidophilus, butyric acid shuttle
Bacterium, actinomyces, saccharomycete etc..At present the most powerful product of the function studied in the world be mainly all kinds of microorganism groups of the above into
Composite reactive probiotics, it is widely used in bioengineering, industrial or agricultural, food security and life and health field.
Pediococcus acidilactici JQII-5 of the invention is a kind of lactic acid bacteria, can effectively reduce cholesterol and triglycerides,
Human body hyperlipidemic conditions can be improved as food and the composition of medicine.
New strains Pediococcus acidilactici (Pediococcus acidilactici) JQII-5 of the invention is in 2015 2
The moon has carried out preservation on 2nd in China Committee for Culture Collection of Microorganisms's common micro-organisms center, and the address of depositary institution is
Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 Institute of Microorganism, Academia Sinica, preserving number is CGMCC No.10512.
Under light microscope, Pediococcus acidilactici JQII-5 is in MRS culture mediums in spherical.General cell is in pairs or tetrad
It is raw, not into catenation.A diameter of 0-1 μm, without gemma, cell wall structures are complete, and cytoplasmic structure is uniform, as shown in Figure 1 and Figure 2.No
Containing plasmid, metabolin does not produce the noxious materials such as D-ALPHA-Hydroxypropionic acid, indoles, nitroreductase is not expressed, does not possess generation amine yet
Ability, 9 kinds of antibiotic MIC (Gentamicin, Kanamycin, Streptomycin, Tetracycline,
Erythromycin, Clindamycin, Chloramphenicol, Amplicilin, Vancomycin) meet EFSA (Europe
Continent food and medicine surveillance authority) regulation.
Pediococcus acidilactici JQII-5 bacterial strains reduce discovery in cholesterol test in vitro, compared with the control, add lactic acid sheet
The high cholesterol solution of coccus JQII-5, after bacterium adds 24 hours, the clearance of cholesterol reaches 74.49%, shows breast
Sour piece coccus JQII-5 truly has reduction cholesterol function (Fig. 3) in vitro.
Further by sour environment that Pediococcus acidilactici JQII-5 inoculations to pH are 3.0 be inoculated into pH 6.8
Property environment in, carry out count plate after 2 hours, it is found that in pH3.0 compared with pH 6.8, viable count is maintained at an order of magnitude；
Meanwhile, Pediococcus acidilactici JQII-5 bacterial strains show in the production curve stabilization of the MRS fluid nutrient mediums containing 0.3% bile salt
Pediococcus acidilactici JQII-5 has good tolerance (Fig. 4,5) to acid and bile.
In zoopery, by Pediococcus acidilactici JQII-5 feedings high cholesterol, hyperlipidemia model mouse, feeding 28 days with
Afterwards, triglycerides and cholesterol level compared with the control group mice of feeding physiological saline, are reduced respectively in mouse blood
19.8% and more than 26.5% (Fig. 6,7), shows that Pediococcus acidilactici JQII-5 can effectively reduce the fat of high glycerine three of mouse, high
Cholesterol symptom.
Pediococcus acidilactici JQII-5 is used to prepare compound probiotic piece and acid bean milk drink, and with acid bean milk drink and height
Fat feed feeding rat, after feeding 28 days, in mouse blood triglycerides and cholesterol level reduce respectively 12% and
More than 15.5% (Fig. 8,9), shows that Pediococcus acidilactici JQII-5 can effectively reduce the fat of high glycerine three of mouse, high-cholesterol disease
Shape.
Research above shows that Pediococcus acidilactici JQII-5 bacterial strains are good to SGF and simulated intestinal fluid environmental resistance, body
Outward, cholesterol and triglycerides are can obviously reduce in vivo.Show that the Pediococcus acidilactici JQII-5 bacterial strains invented herein have been provided with
Necessary requirement as medicinal fungus, can be used to develop into and should be new drug, food, health products and food additives etc..
The present invention is described in further detail below by specific embodiment and with reference to accompanying drawing.Following examples are only right
The present invention is further detailed, and should not be construed as limiting the invention.Unreceipted particular technique or condition in embodiment
, carried out according to the technology or condition described by document in the art or according to product description.Agents useful for same or instrument
Unreceipted production firm person, be can by city available from conventional products.
Embodiment 1：The separation identification of Pediococcus acidilactici JQII-5
(1) culture medium prescription optimization
Packet optimizes culture medium experiment, finally obtains lactic acid bacteria culturing medium optimum formula：
MRS solid mediums 1L is formulated：Casein peptone 10.0g/L, beef extract 10.0g/L, yeast extract 5.0g/L, Portugal
Grape sugar 20.0g/L, dipotassium hydrogen phosphate 2.0g/L, Tween 80 1.0g/L, Triammonium citrate 2.0g/L, sodium acetate 5.0g/L, sulfuric acid
Magnesium 0.1g/L, manganese sulfate 0.05g/L, agar 17.5g；
MRS broth bouillons are formulated compared with MRS agar mediums only without agar.
PH is to 6.5 for adjustment, 121 DEG C of sterilizing 20min.Culture medium is prepared and completed, and 4 DEG C of refrigerations are standby.
(2) separation screening of Pediococcus acidilactici JQII-5 strains
The present embodiment with various traditional fermented foods as sample, such as Yoghourt, pickles, with SPSS, 10 times of ladders
To 10 after degree dilute sample-3Times, MRS solid mediums are coated, it is placed in and 24-48h is cultivated in 37 DEG C of incubators, observe bacterium
Fall morphological feature, and on picking MRS solid mediums lactic acid bacteria doubtful bacterium colony, be transferred in MRS fluid nutrient mediums to enter respectively
Row pure culture.Some plants of bacterium are isolated, every plant of bacterium is numbered.
The measure of acid and cholate tolerance and norcholesterol ability is carried out to isolating and purifying this some plants of bacterium for obtaining, in detail
Experimental procedure is shown in embodiment 2 and embodiment 3, and therefrom screening obtains acid and cholate tolerance is stronger, and norcholesterol ability is stronger
JQII-5.JQII-5 derives from Yoghourt.
(3) 16s rDNA strain idenfications
Bacteria total DNA is extracted to JQII-5,16s rDNA amplifications are carried out, primer is：
sgF:5'-AGAGTTTGATCATGGCTCAG-3'(SEQ ID NO：1)
sgR:5'-TAGGGTTACCTTGTTACGACTT-3'(SEQ ID NO：2)
Enter performing PCR amplification and agarose gel electrophoresis using above-mentioned primer, then gel extraction, sequencing.
Separate the bacterial strain 16s DNA sequencings result such as SEQ ID NO for obtaining:(1264bp) shown in 3：
5'-GCGTGAGTGAAGAAGGGTTTCGGCTCCGTAAAGCTCTGTTGTTAAAGAAGAACGTGGGTGAGAGTA
ACTGTTCACCCAGTGACGGTATTTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTG
GCAAGCGTTATCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTCTTTTAAGTCTAATGTGAAAGCCTTCGGCTC
AACCGAAGAAGTGCATTGGAAACTGGGAGACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAAT
GCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCATGG
GTAGCGAACAGGATTAGATACCCTGGTAGTCCATGCCGTAAACGATGATTACTAAGTGTTGGAGGGTTTCCGCCCTT
CAGTGCTGCAGCTAACGCATTAAGTAATCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAAGAATTGACGGG
GGCCCGCACAAGCGGTGGAGC ATGTGGTTTAATTCGAAGCTACGCGAAGAACCTTACCAGGTCTTGACATCTTCTG
CCAACCTAAGAGATTAGGCGTTCCCTTCTCTGATGGAGCAACGCCGCGTGAGTGAAGAAGGGTTTCGGCTCGTAAAG
CTCTGTTGTTAAAGAAGAACGTGGGTGAGAGTAACTGTTCACCCAGTGACGGTATTTAACCAGAAAGCCACGGCTAA
CTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTATCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCG
GTCTTTTAAGTCTAATGTGAAAGCCTTCGGCTCAACCGAAGAAGTGCATTGGAAACTGGGAGACTTGAGTGCAGAAG
AGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTG
GTCTGTAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACCCTGGTAGTCCATGCCGTAAACG
ATGATTACTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGTAATCCGCCTGGGGAGTACG
ACCGCAAGGTTGAAACTCAAAAGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCTACG
CGAAGAACCTTACCAGGTCTTGACATCTTCTGCCAACCTAAGAG-3’(SEQ ID NO：3)
Sequencing result is compared using BLAST instruments in ncbi database, separates the bacterial strain 16s rDNA alkali for obtaining
Basic sequence shows 99% homology with Pediococcus acidilactici, shows that it is Pediococcus acidilactici to separate the bacterial strain for obtaining.By the bacterial strain
Pediococcus acidilactici JQII-5 is named as, it is common that it was preserved in China Committee for Culture Collection of Microorganisms on 2 2nd, 2015
Microorganism center, preserving number is CGMCC No.10512..
(3) colony characteristicses
JQII-5 cell Gram's staining is positive, and form is spherical.General cell is into opposite, and single survivor is rare, not into
Catenation, does not move, and does not produce gemma, and without pod membrane, catalase experiment is negative, is the bacterium of amphimicrobian.In MRS solids
37 DEG C of culture 18h on culture medium, colony diameter is 0.8~1.4mm, and milky is glossy, opaque, and neat in edge is flat.
(see Fig. 1,2)
(4) biochemical identification
API-50 biochemical identifications are carried out to JQII-5, authentication method is with reference to saying that manufacturer in API-50 identification kits is equipped with
Bright book (API 50CHL, French biology Mei Liai).
Purposes and principle：API50CH is used for carrying out the metabolism of the carbohydrate of microorganism using 50 biochemical tests
Research.API50CH strips are made up of 50 micro holes, are used to study the fermentation of carbohydrate and its derivative substrates.With
API50CHL culture medium inoculated strips, make substrate hydrolysis, then detect fermentation test.It is small in strip during strip is incubated
Acid is produced in pipe under anaerobic environment, pH changes and makes indicator color change in culture medium, and fermentation thus can be observed.
Any active component is free of in strip in first hole, in this, as negative control.The results are shown in Table 1 (+represent positive, it is possible to use
This fermenting substrate；- represent negative).
Table 1：API-50 biochemical identifications result (+represent positive, it is suspicious to utilize this fermenting substrate；- represent negative)
| Carbon source | JQII-5 | Carbon source | JQII-5 |
| Blank | ﹣ | Aesculin | ﹢ |
| Glycerine | ﹣ | Salicin | ± |
| Red tinea alcohol | ﹣ | Cellobiose | ﹢ |
| D-R | ﹣ | Maltose | ﹣ |
| L-arabinose | ﹢ | Lactose | ﹣ |
| Ribose | ﹢ | Melibiose | ﹣ |
| D- xyloses | ﹢ | Sucrose | ﹣ |
| L- xyloses | ﹣ | Trehalose | ﹢ |
| Adonite | ﹣ | Synanthrin | ﹣ |
| Beta-methyl-D- xylosides | ﹣ | The sugar of pine three | ﹣ |
| Galactolipin | ﹢ | Gossypose | ﹣ |
| Glucose | ﹢ | Starch | ﹣ |
| Fructose | ﹢ | Glycogen | ﹣ |
| Mannose | ﹢ | Xylitol | ﹣ |
| Sorbose | ﹣ | Gentiobiose | ± |
| Rhamnose | ± | D- turanoses | ﹣ |
| Melampyrin | ﹣ | D- lyxoses | ﹣ |
| Inositol | ﹣ | D-Tag | ± |
| Mannitol | ﹣ | D- rocks sugar | ﹣ |
| Sorbierite | ﹣ | L- rocks sugar | ﹣ |
| Alpha-Methyl-D-MANNOSE glycosides | ﹣ | D-R alcohol | ﹣ |
| Alpha-Methyl-D-Glucose glycosides | ﹣ | L-arabinose alcohol | ﹣ |
| N- acetyl-aminoglucose | ± | Gluconate | ﹣ |
| Amarogentin | ± | 2- ketone group gluconates | ﹣ |
| Ursin | ﹣ | 5- ketone group gluconates | ﹣ |
The result input API50CHL databases of table 1 are shown that JQII-5 is Pediococcus acidilactici.
(5) bacterium security
A, antibiotics sensitivity test
JQII-5 antibiotic resistances are detected using micro-dilution method.Will test antibacterials (Gentamicin,
Kanamycin,Streptomycin,Tetracycline,Erythromycin,Clindamycin,Chloramphenicol,
Amplicilin, initial concentration is respectively 256ug/ml, 1024ug/ml, 256ug/ml, 64ug/ml, 8ug/ml, 16ug/ml,
64ug/ml, 16ug/ml) make a series of 2 doubling dilutions and totally 12 times and be separately added into 96 orifice plate corresponding apertures, then corresponding aperture is added
The experiment bacterium solution of 2ml dilutions, observes, with antibacterials minimal inhibitory concentration (MIC) hole, as test strain after being cultivated through 48h
Susceptibility.Experimental result (being shown in Table 2) shows that antibiotic MIC is in safe range.
And bacterial strain is stripped plasmid, do not it is found that JQII-5 carries any plasmid, show that it can without Horizontal transfer
Can property.
Table 2：9 kinds of determination of antibiotic susceptibility results
B, metabolin toxicity detection
Lactic acid optical activity is detected：Use the D-/L- lactate acid detection kits of Irish Megazyme companies.Experimental result is not
Produce D-ALPHA-Hydroxypropionic acid.
Nitrate reductase activity is detected：Aseptically, the bacterial strain that will have been activated is linked into nitre by 3% bacterium amount that connects
In hydrochlorate culture medium after 37 DEG C of culture 5d, liquor kalii iodide and each 10 drop of starch solution, observation experiment result is added dropwise.Do sky simultaneously
White experiment.Experimental result shows the unchanged indigo plant of all bacterium solutions, is negative reaction.
Indoles is tested：Aseptically, the bacterial strain that will have been activated is linked into peptone water medium by 3% bacterium amount that connects
In.37 DEG C of culture 72h, add indole reagent 8~10 to drip, observation experiment result.Do blank assay simultaneously.Experimental result does not occur
Red annulus, shows its metabolism edwardsiella hoshinae.
Amino decarboxylase is detected：Bacterium with amino acid decarboxylases, can decompose amino acid makes its decarboxylation generate amine
(lysine → cadaverine, ornithine → putrescine, arginine → spermine) and carbon dioxide, make culture medium become alkalescence, and indicator is added dropwise
(bromocresol purple), is feminine gender in yellow, is the positive in purple.It is in yellow that experimental result display determines pipe, is negative.
The measure of c, cell adherence ability
Cell monolayer culture in the 6 orifice plates full DMEM (without serum and dual anti-) that cannots be used up is changed into 37 DEG C of constant temperature trainings of liquid
Support 1h；Nutrient solution is suctioned out, is washed with aseptic PBS 3 times, cleaning solution is blotted；Take the experimental strain of 500 μ L 108CFU/mL with
After the incomplete DMEM of 500 μ L are mixed, it is added in 6 well culture plates, while the blank of acellular only bacterium solution is set, 37 DEG C,
Incubated 1.5h；Culture plate is taken out, nutrient solution is blotted, washed with aseptic PBS 5 times, blot cleaning solution；10% is added per hole
Formaldehyde, room temperature fixes 2h；Fixer is blotted, is washed with aseptic PBS 3 times, blot cleaning solution, drying at room temperature；After Gram's staining
Microscopy, randomly selects 20 visuals field and is counted, and calculate adhesion index：
Adhesion index=bacterium/cell number (the bacterium number of i.e. average each cell adherence)
Bacterial strain randomly selects 20 visuals field, and the total and cell number of the bacterium to being adhered on the cell in the visual field is counted
And determining adhesion rate, experimental result JQII-5 is 5.9 to CCL188 HT-29 adhesion indexes, and compare the newborn bar of cheese
The bacterial strain of bacterium 334 is suitable, with certain cell adherence ability.
Embodiment 2：The external function test of Pediococcus acidilactici JQII-5 norcholesterols
The external removal rate of cholesterol of bacterial strain for obtaining is separated according to following steps detection embodiment 1, it is specific as follows：
(1) activation of bacterial strain：Strains tested is transferred in MRS fluid nutrient mediums, 24h are cultivated in 37 DEG C, connect with 1%
After the amount of kind passes on 2 times, 15h is cultivated in 37 DEG C, it is stand-by；
(2) MRS culture mediums are prepared, is sterilized stand-by；
(3) MRS culture mediums (nutrient solution MRSO-CHOL) containing cholesterol is prepared：By MRS fluid nutrient mediums, sulfydryl second
Sour sodium, cholate and cholesterol composition, wherein, concentration of the sodium thioglycolate in nutrient solution MRSO-CHOL is 2g/L, and cholate is in training
Concentration in nutrient solution MRSO-CHOL is 0.3%, the final concentration of 120ug/ml of cholesterol.
(4) measure of norcholesterol ability
Experimental group：Activation bacterium solution is inoculated in 10ml nutrient solutions MRSO-CHOL by 1% inoculum concentration.
Control group：10ml does not have the nutrient solution MRSO-CHOL of inoculating strain.
After this two groups of samples are positioned over into 37 DEG C of constant incubator culture 24h, taking-up shakes up, 4000r/m centrifugation 15min,
Supernatant is taken, cholesterol level in supernatant is determined with OPA developer, and calculate the clearance of cholesterol, wherein, courage
Difference/control group the cholesterol level of sterol clearance=control group cholesterol and experimental group cholesterol.
Experimental result is shown in Fig. 3, and Pediococcus acidilactici JQII-5 norcholesterols reduced rate is 74.49%, shows Pediococcus acidilactici
JQII-5 has relatively strong external reduction cholesterol ability.
Embodiment 3：The external acid of Pediococcus acidilactici JQII-5 and the experiment of cholate tolerance
SGF SGF (2.0g NaCl+3.2g pepsins (pepsin)+appropriate concentrated hydrochloric acid adjusts pH value 3.0) is prepared,
With stand-by after 0.2 μm of filtering with microporous membrane.In MRS fluid nutrient mediums, 37 DEG C are cultivated 16~18h to picking slant strains.By bacterium
Suspension 4000r/min is centrifuged 15min, removes supernatant, weighs thalline weight in wet base, and suspension thalline is arranged again in life according to the ratio of 0.1g/ml
In reason salt solution, according to 1：10 ratio row are added in SGF liquid, are placed in 37 DEG C of constant incubators after fully mixing and are cultivated 2h,
Carry out cell count.Experimental result is shown in Fig. 4, and viable count is maintained at an order of magnitude, shows that Pediococcus acidilactici JQII-5 has in vitro
Stronger acid-fast ability.
The bacterium solution that will be activated twice is pressed in the 10mL MRS fluid nutrient mediums containing 0.3% cholate that 1% amount accesses sterilizing
Cultivate and carry out growth curve measure, experimental result is shown in Fig. 5, show that Pediococcus acidilactici JQII-5 has stronger anti-bile energy in vitro
Power.
Embodiment 4：The internal study on the efficiency (zoopery) of Pediococcus acidilactici JQII-5
1st, the preparation of experimental strain：
The Pediococcus acidilactici JQII-5 for activating twice is inoculated in MRS fluid nutrient mediums, 18h is cultivated in 37 DEG C,
6000r/m is centrifuged 10min, collects thalline after being washed with sterile saline.Then, 0.85% physiological saline is added, bacterium is adjusted
Number about 1.0 × 109CFU/ml, then by viable bacteria by daily usage amount be sub-packed in 15ml centrifuge tubes (taking dose be daily 2ml/
Only, every group 10, totally 4 groups, gavage is needed 28 days.
2nd, experimental animal packet and feeding manner：
The week old male rat of Sprague-Dawley systems 5, freely feeds to the 28th day by 40, is equally divided into 4 groups, every group
10：
JQII-5 groups：Gavage viable bacteria suspension, feeds high lipid food；
Model group：The physiological saline of gavage 0.85%, feeds high lipid food；
Normal group：The physiological saline of gavage 0.85%, feeding standard feed；
Effects of Xuezhikang group：Gavage standard dose Effects of Xuezhikang, feeds high lipid food.
Daily every 2ml gavage, continues 28 days, then stops gavage, and each group feed, water are constant to continue 28 days.
3rd, sample collection is tested with analysis：
Before formal test, the 28th day and time period taken a blood sample.After blood-sampling method is for a night of going on a hunger strike, in rat femoral vein
Blood sampling, 4000r/m centrifugations 10min separates serum after blood coagulation, using kit (kit title：T-CHOL determines kit,
Company：Zhejiang Dong'ou Diagnosis Producuts Co., Ltd.) and chromotropic acid development process, mark instrument (company：Molecular Devices, type
Number：Spectra MAX190) difference cholesterol detection and triglycerides.
4th, experimental result
Experimental result is shown in Fig. 6,7.Femoral vein after fat mouse high 28 days is fed with the bacteria suspension of bacterial strain containing JQII-5 to draw blood inspection
Survey, it is found that triglycerides (TG) and cholesterol (TC) decline 19.8% and 26.5%, table respectively in mouse blood compared with the control
Bright Pediococcus acidilactici JQII-5 truly has reduction cholesterol and triglycerides effect in Mice Body.
Embodiment 5：The preparation of medicine and food
(1) compound probiotic piece：
Dispensing：Compound probiotic (streptococcus thermophilus, Lactobacillus rhamnosus, Lactobacillus casei, lactobacillus acidophilus, plant breast
Bacillus, Pediococcus pentosaceus, Pediococcus acidilactici), inulin, dietary fiber (resistant dextrin), D-sorbite, microcrystalline cellulose, tristearin
Sour magnesium.After being mixed uniformly, compressing tablet is made tablet.
(2) acid bean milk drink (original flavor)
Bean powder, inulin, white granulated sugar, water is by weight 18:3:6:73 carry out batch mixing, homogeneous, with 121 DEG C of UHTSs
300s is sterilized, is cooled to 42 DEG C, the leavening that access has been activated:Lactobacillus bulgaricus and lactobacillus thermophilus bacterium powder totally 0.4%.
Ferment 10h at 42 DEG C.
Product is allocated after fermentation, is cooled to 37 DEG C of pentoses for adding 0.1% Pediococcus acidilactici JQII-5 and 0.1%
Piece coccus JQI-7, two kinds of bacterium are 108CFU/ml.Curdled milk, it is filling.
(3) acid bean milk drink (Sugarless type)
Bean powder, inulin, antierythrite, water is by weight 18:3:6:73 carry out batch mixing, homogeneous, with 121 DEG C of UHTSs
300s is sterilized, is cooled to 42 DEG C, the leavening that access has been activated:Lactobacillus bulgaricus and lactobacillus thermophilus bacterium powder totally 0.4%.
Ferment 10h at 42 DEG C.
Product is allocated after fermentation, is cooled to 37 DEG C of pentoses for adding 0.1% Pediococcus acidilactici JQII-5 and 0.1%
Piece coccus JQI-7, two kinds of bacterium are 108CFU/ml.Curdled milk, it is filling.
(4) test of pesticide effectiveness
The week old male rat of Sprague-Dawley systems 5, is divided into 3 groups, every group 10 by 30：
Soy yogurt group：Daily feeding 50ml soy yogurts+high lipid food；
Model group：Daily feeding 50ml ordinary milks+feeding high lipid food；
Normal group：Daily feeding 50ml ordinary milks+feeding standard feed；
Continue 28 days, before formal test, the 28th day and time period taken a blood sample.After blood-sampling method is for a night of going on a hunger strike, in
Rat femoral vein blood, 4000r/m centrifugations 10min separates serum after blood coagulation, using kit (kit title：T-CHOL
Determine kit, company：Zhejiang Dong'ou Diagnosis Producuts Co., Ltd.) and chromotropic acid development process, mark instrument (company：Molecular
Devices, model：Spectra MAX190) difference cholesterol detection and triglycerides.
Experimental result is shown in Fig. 8 and Fig. 9, and femoral vein is drawn blood detection after the High fat diet rats 28 days fed with Sugarless type soy yogurt,
It was found that compared with the control, triglycerides and cholesterol decline 12% and 15.5% respectively in blood, and this product soy yogurt is in Mice Body
Inside truly have reduction cholesterol and triglycerides effect.
In the description of this specification, reference term " one embodiment ", " some embodiments ", " example ", " specifically show
The description of example " or " some examples " etc. means to combine specific features, structure, material or spy that the embodiment or example are described
Point is contained at least one embodiment of the invention or example.In this manual, to the schematic representation of above-mentioned term not
Necessarily refer to identical embodiment or example.And, the specific features of description, structure, material or feature can be any
One or more embodiments or example in combine in an appropriate manner.
Although an embodiment of the present invention has been shown and described, it will be understood by those skilled in the art that：Not
Can these embodiments be carried out with various changes, modification, replacement and modification in the case of departing from principle of the invention and objective, this
The scope of invention is limited by claim and its equivalent.
Claims (10)
1. a kind of Pediococcus acidilactici JQII-5, its preserving number is CGMCC No.10512.
2. purposes of the Pediococcus acidilactici JQII-5 described in claim 1 in food or medicine is prepared.
3. the purposes described in claim 2, it is characterised in that the food or medicine are used for reduction blood fat；Specifically, courage is reduced
Sterol or triglycerides.
4. a kind of composition, it is characterised in that including the Pediococcus acidilactici JQII-5 described in claim 1.
5. the purposes of composition according to claim 4, it is characterised in that the composition is used for reduction blood fat；Specifically,
Reduce cholesterol or triglycerides.
6. a kind of medicine, it is characterised in that the medicine is included：
Pediococcus acidilactici described in claim 1；And
Pharmaceutical non-toxic carrier.
7. the medicine described in claim 6, it is characterised in that the formulation of the medicine be selected from tablet, granule, powder, capsule,
Suspending agent, emulsion, at least one of lyophilized formulations；Optionally, the medicine is compound probiotic piece.
8. medicine according to claim 6, it is characterised in that the medicine is used for reduction blood fat；Specifically, courage is reduced to consolidate
Alcohol or triglycerides.
9. a kind of food, it is characterised in that the food is included：
Pediococcus acidilactici described in claim 1, and
Acceptable additive in bromatology；
Optionally, the food is acid bean milk drink.
10. food according to claim 9, it is characterised in that the food is used for reducing blood lipid；Specifically, courage is reduced to consolidate
Alcohol or triglycerides.
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| CN108504601A (en) * | 2018-04-10 | 2018-09-07 | 北京好实沃生物技术有限公司 | One plant of Pediococcus acidilactici HEW-AP27 and its application |
| CN109182225A (en) * | 2018-10-30 | 2019-01-11 | 吉林省农业科学院 | One plant of Pediococcus acidilactici and its application in antiatherosclerosis |
| CN110973635A (en) * | 2019-12-26 | 2020-04-10 | 汤臣倍健股份有限公司 | Auxiliary material composition and probiotic tablet |
| CN111035658A (en) * | 2018-10-15 | 2020-04-21 | 深圳华大生命科学研究院 | Use of pediococcus pentosaceus |
| CN111269850A (en) * | 2020-01-20 | 2020-06-12 | 吉林农业大学 | A strain of Pediococcus pentosaceus PP04 with high adhesion ability and hypolipidemic effect |
| CN113278552A (en) * | 2021-05-21 | 2021-08-20 | 深圳市华大农业应用研究院 | Pediococcus acidilactici HG-7 strain and application thereof |
| CN114369546A (en) * | 2021-12-29 | 2022-04-19 | 中粮营养健康研究院有限公司 | Pediococcus acidilactici, fermentation microbial inoculum and application thereof, and preparation method of coarse cereal fermented food |
| CN116814480A (en) * | 2023-06-08 | 2023-09-29 | 山东凤凰生物科技股份有限公司 | A strain of Pediococcus acidilactici MP1001, its bacterial powder and its application |
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| CN108504601A (en) * | 2018-04-10 | 2018-09-07 | 北京好实沃生物技术有限公司 | One plant of Pediococcus acidilactici HEW-AP27 and its application |
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| CN111269850B (en) * | 2020-01-20 | 2022-07-22 | 吉林农业大学 | A strain of Pediococcus pentosaceus PP04 with high adhesion ability and hypolipidemic effect |
| CN111269850A (en) * | 2020-01-20 | 2020-06-12 | 吉林农业大学 | A strain of Pediococcus pentosaceus PP04 with high adhesion ability and hypolipidemic effect |
| CN113278552A (en) * | 2021-05-21 | 2021-08-20 | 深圳市华大农业应用研究院 | Pediococcus acidilactici HG-7 strain and application thereof |
| CN114369546A (en) * | 2021-12-29 | 2022-04-19 | 中粮营养健康研究院有限公司 | Pediococcus acidilactici, fermentation microbial inoculum and application thereof, and preparation method of coarse cereal fermented food |
| CN114369546B (en) * | 2021-12-29 | 2023-12-22 | 中粮营养健康研究院有限公司 | Pediococcus acidilactici, fermentation inoculant, application of Pediococcus acidilactici and fermentation inoculant, and preparation method of coarse cereal fermented food |
| CN116814480A (en) * | 2023-06-08 | 2023-09-29 | 山东凤凰生物科技股份有限公司 | A strain of Pediococcus acidilactici MP1001, its bacterial powder and its application |
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