CN106834196A - Lactobacillus plantarum CH126 and its application in function of polysaccharide food is prepared - Google Patents
Lactobacillus plantarum CH126 and its application in function of polysaccharide food is prepared Download PDFInfo
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- CN106834196A CN106834196A CN201710228755.0A CN201710228755A CN106834196A CN 106834196 A CN106834196 A CN 106834196A CN 201710228755 A CN201710228755 A CN 201710228755A CN 106834196 A CN106834196 A CN 106834196A
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Classifications
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- 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
-
- 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
- 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
- A61K2035/11—Medicinal preparations comprising living procariotic cells
- A61K2035/115—Probiotics
Abstract
The invention discloses Lactobacillus plantarum CH126 and its application in function of polysaccharide food is prepared.Lactobacillus plantarum (Lactobacillus plantarum) CH126 that the present invention is provided, preservation registration number is CGMCC NO.13228, abbreviation Lactobacillus plantarum CH126.The present invention also applications of the protection Lactobacillus plantarum CH126 in product is prepared.The product is food, health products or medicine.The food concretely functional food, such as functional yoghourt, feature microbial inoculum etc..The function of the product is:Hypoglycemic and/or reducing blood lipid;Treatment and/or prevention diabetes.Lactobacillus plantarum CH126 has strong reduction fasting blood-glucose, effect for reducing blood fat, can be with being widely used in food, health food and medical and health field.
Description
Technical field
The present invention relates to Lactobacillus plantarum CH126 and its application in function of polysaccharide food is prepared.
Background technology
Diabetes (Diabetes Mellitus, DM) are a kind of common chronic metabolic diseases, are often accompanied by artery athero-
The complication such as hardening, ephrosis, DPN, PVR.Diabetes B patient itself can not effectively utilize insulin or shape
Into insulin resistant, causing the glucose utilization rate of insulin-mediated reduces, and then causes the generations such as internal sugar, fat, protein
It is unrest to thank.Clinically the medicine of anti-diabetic has the agent of promotion insulin secretion, biguanides, alpha-glucosidase restrainer, thiazole
Alkane diones etc., the long-term taking serious side effect such as paropsia, gastrointestinal dysfunction, hepatic and renal function can be caused impaired.
Therefore, hypoglycemic method that is safe and reliable, having no side effect is found, more there is realistic meaning.
Probiotics is the probiotics of the work for having facilitation to host health.Probiotics be widely used to food,
The fields such as functional food, medicine.Probiotics is improving enteron aisle, enhance immunity, Reduce allergy reaction, regulation blood lipid metabolism, tune
The aspects such as section blood pressure play a significant role.Lactic acid bacteria is the superior microorganism species in some traditional fermented foods, is feature
The important sources of probiotics.
The content of the invention
It is an object of the invention to provide Lactobacillus plantarum CH126 and its application in function of polysaccharide food is prepared.
Lactobacillus plantarum (Lactobacillus plantarum) CH126 that the present invention is provided, in November 01 in 2016
Day is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, and (abbreviation CGMCC, address is:Court of Beijing
The institute 3 of positive area's North Star West Road 1, Institute of Microorganism, Academia Sinica), preservation registration number is CGMCCNO.13228.Plant breast
Bacillus (Lactobacillus plantarum) CH126CGMCC NO.13228, abbreviation Lactobacillus plantarum CH126.
The present invention also applications of the protection Lactobacillus plantarum CH126 in product is prepared.The product be food, health products or
Medicine.The food concretely functional food, such as functional yoghourt, feature microbial inoculum etc..The function of the product is
At least one in (a1) to (a31) as follows:(a1) hypoglycemic and/or reducing blood lipid;(a2) blood-sugar content in serum is reduced;
(a3) lipids contents in serum are reduced;(a4) total cholesterol level in serum is reduced;(a5) glycerine three in serum is reduced
Ester content;(a6) fasting blood sugar is reduced;(a7) oxidation resistance is increased;(a8) superoxide dismutase in serum is increased
Content and/or activity;(a9) content and/or activity of the catalase in serum are increased;(a10) the paddy Guang in serum is increased
The content and/or activity of sweet peptide peroxidase;(a11) content and/or activity of the MDA in serum are reduced;(a12) change
Kind disorders of lipid metabolism;(a13) the D-ALPHA-Hydroxypropionic acid content in serum is reduced;(a14) free fatty acid content in serum is reduced;
(a15) endotoxin content in serum is reduced;(a16) treat and/or prevention diabetes;(a17) Diabetes Mellitus are reduced
In blood-sugar content;(a18) lipids contents in Diabetes Mellitus are reduced;(a19) in reduction Diabetes Mellitus
Total cholesterol level;(a20) content of triglyceride in Diabetes Mellitus is reduced;(a21) sky of diabetic is reduced
Abdomen blood glucose value;(a22) insulin content in Diabetes Mellitus is raised;(a23) increase super in Diabetes Mellitus
The content and/or activity of superoxide dismutase;(a24) increase catalase in Diabetes Mellitus content and/or
Activity;(a25) content and/or activity of the glutathione peroxidase in Diabetes Mellitus are increased;(a26) sugar is reduced
The content and/or activity of the MDA in urine patient's serum;(a27) disorders of lipid metabolism of diabetic is improved;(a28)
Reduce the D-ALPHA-Hydroxypropionic acid content in Diabetes Mellitus;(a29) free fatty acid content in Diabetes Mellitus is reduced;
(a30) endotoxin content in Diabetes Mellitus is reduced;(a31) hyperlipidemic conditions of diabetic are improved.
The present invention also protects a kind of product, and its active component is Lactobacillus plantarum CH126.The present invention also protects a kind of producing
Product, its active component is the extract of the fermentate of the fermentate or Lactobacillus plantarum CH126 of Lactobacillus plantarum CH126.It is described
Product is food, health products or medicine.The food concretely functional food, such as functional yoghourt, feature microbial inoculum
Deng.The function of the product is at least one in following (a1) to (a31):(a1) hypoglycemic and/or reducing blood lipid;(a2) reduce
Blood-sugar content in serum;(a3) lipids contents in serum are reduced;(a4) total cholesterol level in serum is reduced;(a5) drop
Content of triglyceride in low serum;(a6) fasting blood sugar is reduced;(a7) oxidation resistance is increased;(a8) in increasing serum
The content and/or activity of superoxide dismutase;(a9) content and/or activity of the catalase in serum are increased;(a10)
Increase the content and/or activity of the glutathione peroxidase in serum;(a11) content of the MDA in serum is reduced
And/or activity;(a12) disorders of lipid metabolism is improved;(a13) the D-ALPHA-Hydroxypropionic acid content in serum is reduced;(a14) in reduction serum
Free fatty acid content;(a15) endotoxin content in serum is reduced;(a16) treat and/or prevention diabetes;(a17) reduce
Blood-sugar content in Diabetes Mellitus;(a18) lipids contents in Diabetes Mellitus are reduced;(a19) glycosuria is reduced
Total cholesterol level in patient's serum;(a20) content of triglyceride in Diabetes Mellitus is reduced;(a21) reduce
The fasting blood sugar of diabetic;(a22) insulin content in Diabetes Mellitus is raised;(a23) diabetes are increased
The content and/or activity of the superoxide dismutase in patients serum;(a24) hydrogen peroxide in Diabetes Mellitus is increased
The content and/or activity of enzyme;(a25) content and/or the work of glutathione peroxidase in Diabetes Mellitus are increased
Property;(a26) content and/or activity of the MDA in Diabetes Mellitus are reduced;(a27) lipid of diabetic is improved
Metabolic disorder;(a28) the D-ALPHA-Hydroxypropionic acid content in Diabetes Mellitus is reduced;(a29) trip in Diabetes Mellitus is reduced
From content of fatty acid;(a30) endotoxin content in Diabetes Mellitus is reduced;(a31) blood high of diabetic is improved
Fat symptom.
The concretely diabetes B of diabetes described in any of the above.
Lactobacillus of the invention is that the one plant of hypoglycemic work(for obtaining is screened from the lactic acid bacteria of numerous traditional fermented foods source
Can probiotics new strains, its not only have very strong reduction fasting blood-glucose, effect for reducing blood fat, also with good tolerance,
Good hydrophobicity and self-coagulation ability, there are different sensitiveness to different antibiotic, safety, and without toxicity, side effect, can use
It is widely used in food, health food and medical and health field.
Brief description of the drawings
Fig. 1 is the testing result of serum insulin
Specific embodiment
Following embodiment facilitates a better understanding of the present invention, but does not limit the present invention.Experiment in following embodiments
Method, unless otherwise specified, is conventional method.Test material used in following embodiments, unless otherwise specified, is certainly
What routine biochemistry reagent shop was commercially available.Quantitative test in following examples, is respectively provided with three repetitions and tests, and as a result makes even
Average.
Unless otherwise specified, the PBS in embodiment is the PBS of pH7,0.1M.
Solid BCP culture mediums:2.5g/L containing yeast extract, peptone 5g/L, glucose 5g/L, bromocresol purple 0.04g/L, fine jade
Fat 15g/L, balance of water;Unless otherwise specified, pH=7.0.Liquid B CP culture mediums only exist with the difference of solid BCP culture mediums
In being added without agar.
Solid MRS culture mediums:10g/L containing peptone, beef extract 10g/L, yeast extract 5g/L, KH2PO42g/L, sodium acetate
5g/L, sodium citrate 5g/L, MgSO4·7H2O 0.2g/L、MnSO4·4H2O 0.05g/L, Tween-80 1mL/L, agar
15g/L, glucose 20g/L, balance of water;Unless otherwise specified, pH=6.6.Liquid MRS culture mediums and solid MRS culture mediums
Differ only in and be added without agar.
Basal feed:This experimental animal technology Co., Ltd of Changchun hundred million.High lipid food (% represents g/100g):Base
Plinth feed 75%, lard 10%, yolk powder 10%, cholesterol (food-grade, Zhengzhou Cang Yu chemical products Co., Ltd) 5%.
C57BL/6J mouse:Changchun hundred million this experimental animal technology Co., Ltd, animal credit number:SCXK (Ji) -2011-
0004。
Using the T-CHOL in mouse T-CHOL enzyme linked immunological kit (BPE20095) the detection serum of bright of Shanghai
(Total Cholesterol, TC) content.Examined using bright of Shanghai mouse triglycerides enzyme linked immunological kit (BPE20754)
Triglycerides (Triglyceride, the TG) content surveyed in serum.Using bright of Shanghai mouse HDL-C enzyme
Linked immunoassay reagent kit (BPE20192) detection serum in HDL-C (High DensityLipoprotein,
HDL-C) content.Using bright of Shanghai mouse LDL-C enzyme linked immunological kit (BPE20100) detection serum
In LDL-C (Low Density Lipoprotein, LDL-C) content.Using bright of Shanghai mouse pancreas
Insulin (Insulin, INS) content in island element enzyme linked immunological kit (BPE20353) detection serum.Using bright of Shanghai
Mouse glutathione peroxidase immune reagent kit (BPE20748) detects serum Glutathione Peroxidase
The content of (Glutathione peroxidase, GSH-Px).Using bright of Shanghai mouse superoxide dismutase immune reagent kit
(BPE20348) content of superoxide dismutase (Superoxidedismutase, SOD) in serum is detected.Using Shang Hailang
Mouse hydrogen peroxide Enzymoimmune reagent kit (BPE20880) that pauses detects the content of catalase (Catalase, CAT) in serum.
Using MDA in bright mouse MDA immune reagent kit (BPE20347) in Shanghai detection serum (Malondialdehyde,
MDA content).Using free fatty in mouse free fatty immune reagent kit (BPE20248) the detection serum of bright of Shanghai
The content of (Free fat acid, FFA).Using bright of Shanghai Mouse endotoxin immune reagent kit (BPE20316) detection serum
The content of middle endotoxin (Endotoxin, ET).Using bright of Shanghai mouse D-ALPHA-Hydroxypropionic acid immune reagent kit (BPE20072) detection blood
The content of D-ALPHA-Hydroxypropionic acid (D-Lactate) in clear.
Lactobacillus plantarum (Lactobacillus plantarum) Sc52, was preserved in China on 06 29th, 2015
(abbreviation CGMCC, address is Microbiological Culture Collection administration committee common micro-organisms center:BeiChen West Road, Chaoyang District, BeiJing City 1
Number institute 3, Institute of Microorganism, Academia Sinica), preservation registration number is CGMCC NO.11027.Lactobacillus plantarum
(Lactobacillus plantarum) Sc52CGMCC NO.11027, abbreviation Lactobacillus plantarum Sc52.
Separation, identification and the preservation of embodiment 1, lactobacillus plantarum strain
First, the separation of lactobacillus plantarum strain
Sample is drawn materials:Traditional fermented food is vinegar-pepper radish.
Rule repeatedly culture through BCP culture medium flat plates, the single bacterium colony that picking produces yellow circle is inoculated in solid MRS culture medium flat plates
It is upper to continue to cultivate, rule repeatedly culture purified through solid MRS culture medium flat plates, obtain many plants of Pure cultured spawns.
The strain of pure culture is inoculated into liquid MRS medium cultures, is subsequently adding 20% glycerine, -80 DEG C of Refrigerator stores.
Filtered out from the strain of pure culture Gram-positive, catalase test is negative, indole reaction is negative,
Hydrogen sulfide production test is negative, gelatin liquefaction test is negative, hydrolysis starch negative and negative some of nitrate reduction test
Bacterial strain.It is CH126 by wherein 1 plant Strain Designation.
2nd, the identification of bacterial strain
The Physiology and biochemistry qualification result of CH126:Gram-positive, catalase test is negative, benzidine test
Feminine gender, indole test is negative, and voges-Proskauer test is positive;Not hydrolysis starch, do not liquefy gelatin, does not produce hydrogen sulfide, hair
Ferment glucose produces acid not aerogenesis;No motion of bacillus;Can be grown at 15 DEG C and 45 DEG C, 37~42 DEG C of optimum growth temperature;It is suitable
Suitable pH is 5.0~7.0;Tolerance 6.5%NaCl;CH126 is in uniform turbid growth in liquid MRS culture mediums, and being long placed in thalline is in
White precipitate.
API 50CH (French Mei Liai companies) qualification result of CH126:Ribose, galactolipin, glucose, fruit can be utilized
Sugar, xylose, mannose, mannitol, sorbierite, amarogentin, ursin, N- acetyl-gucosamine, seven leaf-alcohols, salicin, fiber
Disaccharides, synanthrin, lactose, melibiose, sucrose, trehalose, sorbose, melezitose, raffinose, geraniol, gluconate;No
Adonite, arabinose, glycogen, dulcitol, maltose, inositol, Alpha-Methyl-D-Glucose glycosides, Alpha-Methyl-D- sweet dews can be utilized
Glucosides, starch, rhamnose, xylitol, D- turanoses, D-Tag, D- meters of threose, gluconate, D-R alcohol, rock
Sugar, L-arabinose alcohol, 2- keto-D-gluconates salt, 5- keto-D-gluconate salt.
16S rDNA sequence homology analysis:Genomic DNA is extracted, the primer pair (16sF constituted using 16sF and 16sR:
5 '-AGAGTTTGATCCTGGCTCAG-3 ';16sR:5 '-AGAAAGGAGGTGATCCAGC-3 ') enter performing PCR amplification, then will
Amplified production is sequenced, and 16srDNA sequences are as shown in the sequence 1 of sequence table.16s rDNA sequences are existed by blast program
Sequence analysis analysis is carried out in GenBank storehouses, with Lactobacillus plantarum strain RI-113
(GenBank:CP017406.1) homology reaches 98%.
According to above qualification result, bacterial strain CH126 belongs to Lactobacillus plantarum.
3rd, the preservation of bacterial strain
Lactobacillus plantarum (Lactobacillus plantarum) CH126, is preserved in China on November 01st, 2016
(abbreviation CGMCC, address is Microbiological Culture Collection administration committee common micro-organisms center:BeiChen West Road, Chaoyang District, BeiJing City 1
Number institute 3, Institute of Microorganism, Academia Sinica), preservation registration number is CGMCC NO.13228.Lactobacillus plantarum
(Lactobacillus plantarum) CH126CGMCC NO.13228, abbreviation Lactobacillus plantarum CH126.
The functional characteristic of embodiment 2, Lactobacillus plantarum CH126
The method of colony counting, after completing culture, doubling dilution simultaneously coats solid MRS culture medium flat plates, 37 DEG C of standings
Colony counting is carried out after culture 48h.
First, acid-fast ability and bile tolerance ability
1st, acid-fast ability
The liquid MRS culture mediums that Lactobacillus plantarum CH126 or Lactobacillus plantarum Sc52 are inoculated in different pH (use 1mol/L salt
Acid adjust pH be respectively 2.0,3.0 or 6.6) in, 37 DEG C of quiescent cultures 1h or 3h, then by colony counting calculate bacteria concentration (use
Log10CFU/mL is represented).
Every kind of pH sets 3 parallel processing, and each dilution factor is once again set up 3 parallel processing, results averaged.
The results are shown in Table 1.Lactobacillus plantarum CH126:Strain growth is good under the conditions of pH6.6, extension viable bacteria over time
Number is in rising trend;Viable bacteria is not detected by after cultivating 3h under the conditions of pH2.0;Under the conditions of pH3.0, strains expressed has gone out certain resistance to
Acidity, survival rate is still 61.5% after culture 3h;Result shows that Lactobacillus plantarum CH126 has stronger sour tolerance.Plant
Lactobacillus Sc52:Under the conditions of pH3.0, survival rate is 57.0% after culture 3h.
Bacteria concentration (Logs of the Lactobacillus plantarum CH126 of table 1 in Liquid Culture system10CFU/mL)
2nd, bile tolerance ability
Lactobacillus plantarum CH126 or Lactobacillus plantarum Sc52 are inoculated in the liquid MRS culture mediums containing various concentrations bovine bile
(weight/mass percentage composition of bovine bile is respectively 0.3%, 0.5% or 1.0%), 37 DEG C of quiescent culture 4h, then by bacterium colony meter
Number calculates bacteria concentration and (uses Log10CFU/mL is represented).
Every kind of bovine bile concentration sets 3 parallel processing, and each dilution factor is once again set up 3 parallel processing, as a result makes even
Average.
The results are shown in Table 2.Lactobacillus plantarum CH126:Strain survival rate is 52% under 0.3% gallbladder salinity, in 0.5% courage
Strain survival rate is 46% under salinity, and Strain survival rate is 32% under 1% gallbladder salinity;Result shows, Lactobacillus plantarum
CH126 has stronger cholate tolerance.Lactobacillus plantarum Sc52:Strain survival rate is 49% under 0.3% gallbladder salinity,
Strain survival rate is 42% under 0.5% gallbladder salinity, and Strain survival rate is 25% under 1% gallbladder salinity.
Bacteria concentration (Logs of the Lactobacillus plantarum CH126 of table 2 in Liquid Culture system10CFU/mL)
Initial bacteria concentration | 0.3% bovine bile | 0.5% bovine bile | 1.0% bovine bile | |
Lactobacillus plantarum CH126 | 8.14±0.16 | 4.25±0.04 | 3.75±0.17 | 2.61±0.18 |
Lactobacillus plantarum Sc52 | 8.16±0.31 | 4.00±0.15 | 3.41±0.13 | 2.07±0.09 |
2nd, the hydrophobicity of Lactobacillus plantarum CH126 and self-coagulation ability are determined
Bacterium surface hydrophobic effect and self-coagulation ability be somatic cells adhere to biological and inanimate surfaces it is important because
One of element.Physical law (bacterial adhesion substance, BATS) is sticked together using bacterium and determines Lactobacillus plantarum
The hydrophobicity of CH126.Self-solidifying rate is caused by bacterial strain surface hydrophobic, at the same it is closely related with its adhesive capacity, self-solidifying rate high
Bacterial strain typically has adhesive capacity higher.
1st, hydrophobicity
By the thalline of Lactobacillus plantarum CH126 brine 2 times, 0.1mol/L KNO are then used3The aqueous solution hangs
Floating thalline, it is 1 × 10 to obtain bacteria concentration8The bacterium solution of CFU/mL, determines the OD value A of bacterium solution at 600nm wavelength0.By 3mL bacterium
Liquid is mixed with 1mL organic solvents (dimethylbenzene, chloroform or ethyl acetate), is then stored at room temperature 30min, collects water phase, 600nm ripples
Strong point determines the OD value A of water phase1.Calculate hydrophobicity (BATS).Hydrophobic evaluation criterion is as follows:BATS>50% is height
Hydrophobicity, BATS is that moderate is hydrophobic in 20%-50%, BATS<20% is non-hydrophobic.Carry out three repetitions to test, even taking of finding an excuse
Average value.
The results are shown in Table 3.Lactobacillus plantarum CH126 shows non-hydrophobic ability in dimethylbenzene, ethyl acetate and chloroform.
The hydrophobicity (%) of the Lactobacillus plantarum CH126 of table 3
Dimethylbenzene | Ethyl acetate | Chloroform |
17.0±0.90 | 5.0±0.24 | 18.7±1.52 |
2nd, self-coagulation ability
By the thalline of Lactobacillus plantarum CH126 brine 2 times, physiological saline suspension thalline is then used, obtained
Initial bacterium solution, determines the OD value A of initial bacterium solution at 600nm wavelength0It is 1.0.1mL is taken after initial bacterium solution is stored at room temperature into 1h
Upper phase, determines the OD value of bacterium solution at 600nm wavelength, be A1.Initial bacterium solution is stored at room temperature 1mL upper stratas are taken after 24h
Liquid phase, determines the OD value of bacterium solution at 600nm wavelength, be A2.Initial bacterium solution is stored at room temperature 1mL upper phases is taken after 48h,
The OD value of bacterium solution is determined at 600nm wavelength, is A3.Calculate self-solidifying rate (bacterial strain self-coagulation percentage).Carry out three repetitions
Experiment, even averaging of finding an excuse.
The results are shown in Table 4.Increase over time, Lactobacillus plantarum CH126 self-solidifyings rate is raised, upper phase is in clear after 48h
Clear state, self-solidifying rate reaches 84.2%.
The self-solidifying rate (%) of the Lactobacillus plantarum CH126 of table 4
1h | 24h | 48h |
2.5±0.4 | 75.3±0.57 | 84.2±1.00 |
3rd, antibiotic susceptibility test
1st, antibiotic is configured to the mother liquor that concentration is 5120 μ g/mL, with 0.22 μm of filter membrane mistake as solvent with distilled water
Bacterium is filtered, then stepwise dilution is carried out with sterilized water, following various antibiotic solutions are obtained:
Concentration is 0.016 μ g/mL, μ g/mL0.032 μ g/mL, 0.064 μ g/mL, 0.125 μ g/mL, 0.25 μ g/mL, 0.5 μ
The erythromycin solution of g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL or 8 μ g/mL;
Concentration be 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/mL, 32 μ g/mL, 64 μ g/mL,
The streptomycin sulfate solution of 128 μ g/mL or 256 μ g/mL;
Concentration be 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/mL, 32 μ g/mL, 64 μ g/mL,
The gentamycin solution of 128 μ g/mL or 256 μ g/mL;
Concentration be 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/mL, 32 μ g/mL, 64 μ g/mL, 128 μ g/mL, 256 μ g/mL,
The kanamycin sulfate solution of 512 μ g/mL or 1024 μ g/mL;
Concentration is 0.125 μ g/mL, 0.25 μ g/mL, 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/
The quadracycline solution of mL, 32 μ g/mL or 64 μ g/mL;
Concentration be 0.25 μ g/mL, 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/mL, 32 μ g/mL,
The vancomycin solution of 64 μ g/mL or 128 μ g/mL;
Concentration is 0.125 μ g/mL, 0.25 μ g/mL, 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/
The chloromycetin solution of mL, 32 μ g/mL, 64 μ g/mL or 128 μ g/mL;
Concentration is 0.032 μ g/mL, 0.063 μ g/mL, 0.125 μ g/mL, 0.25 μ g/mL, 0.5 μ g/mL, 1 μ g/mL, 2 μ g/
The ampicillin sodium solution of mL, 4 μ g/mL, 8 μ g/mL or 16 μ g/mL;
Concentration is 0.125 μ g/mL, 0.25 μ g/mL, 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/
The rifampin solution of mL, 32 μ g/mL or 64 μ g/mL;
Concentration is 0.125 μ g/mL, 0.25 μ g/mL, 0.5 μ g/mL, 1 μ g/mL, 2 μ g/mL, 4 μ g/mL, 8 μ g/mL, 16 μ g/
The trimethoprim solution of mL, 32 μ g/mL, 64 μ g/mL or 128 μ g/mL.
2nd, Lactobacillus plantarum CH126 dilutes 1000 times with liquid MRS culture mediums, and it is 1 × 10 to obtain bacteria concentration8CFU/mL's
Bacterium solution.
3rd, 96 orifice plates are taken, test hole adds antibiotic prepared by the bacterium solution and 70 μ L steps 1 of the preparation of 70 μ L steps 2 molten per hole
Liquid, Positive control wells add the bacterium solution and 70 μ L sterilized waters of 70 μ L steps 2 preparation per hole, and negative control hole adds 70 μ L liquid per hole
Body MRS culture mediums and 70 μ L sterilized waters, 37 DEG C of quiescent culture 16h, then observe the growing state of bacterial strain in 96 orifice plates, hence it is evident that nothing
Antibiotic concentration corresponding to the hole of strain growth is MIC of the antibiotic to bacterial strain.Each of every kind of antibiotic
Dilution factor sets 3 parallel processing.
The results are shown in Table 5.Lactobacillus plantarum CH126 is more sensitive to chloramphenicol, rifampin, penicillin, to vancomycin, card
That mycin shows strong drug resistance.
Minimum inhibitory concentrations (μ g/mL) of the Lactobacillus plantarum CH126 of table 5 to different antibiotic
4th, alpha-glucosaccharase enzyme inhibition is tested
1st, Lactobacillus plantarum CH126 thalline are taken, is suspended with PBS (pH6.8,0.1mol/L), it is 1 to obtain bacteria concentration
×109The bacterium solution of CFU/mL.
2nd, the bacterium solution of 50 μ L steps 1 preparation is taken, 100 μ L PBSs (pH6.8,0.1mol/L) are added, 100 are added
P-nitrophenyl-α-D- the glucopyranosides aqueous solution (PNPG solution) of μ L 20mmol/L, 37 DEG C of hatching 10min after mixing,
60 μ L 20U/mL alpha-glucosaccharase enzyme aqueous solutions are subsequently adding, then 37 DEG C of hatching 20min, are subsequently adding 100 μ L 1mol/L
Na2CO3Aqueous solution terminating reaction, the densitometric value at 405nm.
In formula:A contains bacterium solution and alpha-glucosaccharase enzyme aqueous solution for sample sets;B for sample sets contain bacterium solution without α-
Glucoside enzyme aqueous solution (is replaced) with isometric PBS;C is (slow with isometric PBS for blank control group does not contain bacterium solution
Fliud flushing replacement) enzyme aqueous solution containing alpha-glucosaccharase;D does not contain bacterium solution (being replaced with isometric PBS) for blank control group
Alpha-glucosaccharase enzyme aqueous solution (being replaced with isometric PBS) is not contained yet.
Lactobacillus plantarum CH126 is shown in Table 6 to alpha-glucosaccharase enzyme inhibition rate.Lactobacillus plantarum CH126 is to alpha-glucosaccharase
Enzyme shows certain inhibitory action, can improve the metabolism of glucose.
The Lactobacillus plantarum CH126 of table 6 is to alpha-glucosaccharase enzyme inhibition rate
Bacterial strain | Inhibiting rate |
CH126 | 29.35 ± 2.16% |
The adjustment effect of embodiment 3, Lactobacillus plantarum CH126 to blood sugar
First, packet transaction mode
80 male C 57 BL/6 J mouses, are randomly divided into 4 groups, and every group 20, packet transaction is as follows:
Blank control group:Fed using normal diet from on-test to end;9th week, mouse was on an empty stomach after 12 hours
Disposable celiac injecting normal saline 30mg/kg;It is administered within 10th week to the 13rd week, dosage is " every per kg body weight
It is secondary to give 12mL sterile PBS buffers ";
Model group:Fed with high lipid food within 1st week to 8 weeks, fed with normal diet within the 9th week to off-test;9th week,
Mouse empty stomach disposable intraperitoneal injection Streptozotocin 30mg/kg after 12 hours;It is administered within 10th week to the 13rd week;To medicament
Measure is " giving 12mL sterile PBS buffers every time per kg body weight ";
Positive controls:Fed with high lipid food within 1st week to 8 weeks, fed with normal diet within the 9th week to off-test;9th
Week, mouse empty stomach disposable intraperitoneal injection Streptozotocin 30mg/kg after 12 hours;It is administered within 10th week to the 13rd week;Give
Pharmaceutical quantities are for " it is (double containing 120mg diformazans in 12mL melbine solution that every kg body weight gives 12mL melbine solution every time
Guanidine;The solvent of melbine solution is PBS) ";
CH126 groups:Fed with high lipid food within 1st week to 8 weeks, fed with normal diet within the 9th week to off-test;9th week,
Mouse empty stomach disposable intraperitoneal injection Streptozotocin 30mg/kg after 12 hours;It is administered within 10th week to the 13rd week;To medicament
Measure " to give the 12mL bacterium solutions (preparation method of bacterium solution every time per kg body weight:With PBS suspension plant lactobacillus
CH126, it is 1.0 × 10 to obtain Lactobacillus plantarum CH126 concentration9The bacterium solution of CFU/mL) ".
Administering mode is:Gastric infusion, once a day.
Test within 1st week to 8 weeks the 1-56 days, be within the 10th week to 13 weeks experiment the 64-91 days, by that analogy.Note in abdominal cavity
Penetrate Streptozotocin or physiological saline is carried out on the 57th day in experiment.
2nd, judge to set up diabetes model standard
9th week (experiment the 63rd day), tail vein blood determines mouse fasting blood sugar (FBG).Meet the small of following standard
Mouse is diabetes B model mice:Double measure FBG >=11.1mmol/L.Result shows, tests the 57th day, model
Group, positive controls and CH126 groups each mouse are diabetes B model mice, and blank control group each mouse is sky
The orthoglycemic mouse of abdomen.
3rd, blood sugar test
Respectively at the 10th week, the 11st week, the 12nd week, the 13rd week and the 14th week, tail vein blood determined mouse fasting blood-glucose
Value (detection, calculates the average value in this week daily).
Fasting blood sugar the results are shown in Table 7.Compared with model group, CH126 group mouse fasting blood sugars are on a declining curve.
CH126 group mouse fasting blood sugars were remarkably decreased the 12nd week, the 13rd week, the 14th week, and blood glucose value is dropped at the 14th week
12.35mmol/L, there is pole significant difference (P < 0.01) compared with model group mouse.Result shows, Lactobacillus plantarum CH126
There is therapeutic action to the hyperglycemia of diabetes B, fasting blood sugar can be significantly reduced.
Influences of the Lactobacillus plantarum CH126 of table 7 to diabetes B mouse fasting blood-glucose
Note:Compare with model group, * has significant difference (P < 0.05), * * have pole significant difference (P < 0.01).
4th, in serum index of correlation detection
During off-test (experiment the 100th day), each group mouse is taken, pluck eyeball and take blood, 3 000r/ after room temperature solidification 60min
Min is centrifuged 15min, collects supernatant (serum).Detection serum in T-CHOL, triglycerides, HDL-C,
LDL-C, insulin, glutathione peroxidase, superoxide dismutase, catalase, the third two
Aldehyde, free fatty, endotoxin, the content of D-ALPHA-Hydroxypropionic acid.
The testing result of blood fat correlation factor is shown in Table 8 in serum.TC, TG, LDL-C content are bright in model group mice serum
Aobvious is higher than blank control group, and HDL-C contents are significantly lower than blank control group, and this is because diabetes cause Mice Body intracellular metabolite disorderly
Disorderly.Compared with model group, TC, TG, LDL-C content decline in CH126 group mice serums.Result shows, Lactobacillus plantarum CH126
The hyperlipidemic conditions of diabetes B can be obviously improved.
Influences of the Lactobacillus plantarum CH126 of table 8 to diabetes B lipid of mice
Note:Compare with model group, * has significant difference (P < 0.05), * * have pole significant difference (P < 0.01).
The testing result of serum insulin is shown in Fig. 1.Sugar environment high suppresses glucose stimulated insulin secreting function, causes
Serum insulin content declines, and gavage Lactobacillus plantarum CH126 can effectively alleviate internal metabolic disorder, controls blood sugar, improves pancreas
Island function, improves serum insulin content (P < 0.05).
The testing result of anti-oxidant correlation factor is shown in Table 9 in serum.The peroxide injury that diabetes B causes causes model
Anti-oxidant vigor declines in group mice serum, SODs of the mouse gavaging Lactobacillus plantarum CH126 after 4 weeks in mice serum, CAT,
GSH-Px vigor is significantly improved, and SOD, CAT are respectively increased 56.48U/mL, 23.89U/mL in CH126 group serum, with model
Group is compared to difference extremely significantly (P<0.01);GSH-Px levels bring up to 91.24nmol/mL in CH126 groups serum at the end of experiment
(P<0.05);MDA contents are significantly reduced in serum.During result shows that Lactobacillus plantarum CH126 can significantly improve mice serum
Superoxide dismutase, catalase and activity of glutathione peroxidase, suppress MDA vigor, show obvious
Oxidation resistance.
Influences of the Lactobacillus plantarum CH126 of table 9 to Antioxidant Indexes in diabetes B mice serum
Note:Compare with model group, * has significant difference (P < 0.05), * * have pole significant difference (P < 0.01).
D-ALPHA-Hydroxypropionic acid, free fatty, endotoxic testing result are shown in Table 10 in serum.The body Portugal that diabetes B causes
Grape sugar, protein and disorders of lipid metabolism, cause substantial amounts of D-ALPHA-Hydroxypropionic acid, free fatty, endotoxin to enter blood, and influence is normal
Physiological metabolism.The intake of Lactobacillus plantarum CH126 can be different degrees of improvement serum in D-ALPHA-Hydroxypropionic acid, free fatty, interior
Content of toxins, experiment terminates D-ALPHA-Hydroxypropionic acid in rear CH126 groups mice serum, free fatty acid content and obviously drops to 2.27 μm of ol/
ML, 689.27 μ g/L, endotoxin content are reduced to 71.34EU/L (P<0.05).Result shows that Lactobacillus plantarum CH126 may
By adjusting diabetes B mouse body lipid metabolic disorder, suppress D-ALPHA-Hydroxypropionic acid, free fatty, endotoxin and largely enter blood
Liquid.
The Lactobacillus plantarum CH126 of table 10 to diabetes B D-ALPHA-Hydroxypropionic acid, free fatty, endotoxin content influence
Group | D-Lactate(μmol/mL) | FFA(μg/L) | ET(EU/L) |
Blank control group | 1.96±0.23 | 567.14±6.48 | 59.47±4.92 |
Model group | 3.05±0.47 | 722.48±5.34 | 82.16±5.84 |
Melbine group | 2.11±0.26* | 665.38±4.67 | 64.55±7.69** |
CH126 groups | 2.27±0.38 | 689.27±4.59 | 71.34±6.60* |
SEQUENCE LISTING
<110>Jilin Academy of Agricultural Science
<120>Lactobacillus plantarum CH126 and its application in function of polysaccharide food is prepared
<130> GNCYX170653
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1504
<212> DNA
<213>Lactobacillus plantarum(Lactobacillus plantarum)
<400> 1
cgatctcgct cttgttacga cttcacccta atcatctgtc ccaccttagg cggctggttc 60
ctaaaaggtt accccaccga ctttgggtgt tacaaactct catggtgtga cgggcggtgt 120
gtacaaggcc cgggaacgta ttcaccgcgg catgctgatc cgcgattact agcgattccg 180
acttcatgta ggcgagttgc agcctacaat ccgaactgag aatggcttta agagattagc 240
ttactctcgc gagttcgcaa ctcgttgtac catccattgt agcacgtgtg tagcccaggt 300
cataaggggc atgatgattt gacgtcatcc ccaccttcct ccggtttgtc accggcagtc 360
tcaccagagt gcccaactta atgctggcaa ctgataataa gggttgcgct cgttgcggca 420
cttaacccca cactctcacg acacgagctg acgacaacca tgcaccccct gtatccatgt 480
ccccgaaggg aacgtctaat ctcttagatt tgcatagtat gtcaagacct ggtaaggttc 540
ttcgcgtagc ttcgaattaa accacatgct ccaccgcttg tgcgggcccc cgtcaattcc 600
tttgagtttc agccttgcgg ccgtactccc caggcggaat gcttaatgcg ttagctgcag 660
cactgaaggg cggaaaccct ccaacactta gcattcatcg tttacggtat ggactaccag 720
ggtatctaat cctgtttgct acccatactt tcgagcctca gcgtcagtta cagaccagac 780
agccgccttc gccactggtg ttcttccata tatctacgca tttcaccgct acacatggag 840
ttccactgtc ctcttctgca ctcaagtttc ccagtttccg atgcacttct tcggttgagc 900
cgaaggcttt cacatcagac ttaaaaaacc gcctgcgctc gctttacgcc caataaatcc 960
ggacaacgct tgccacctac gtattaccgc ggctgctggc acgtagttag ccgtggcttt 1020
ctggttaaat accgtcaata cctgaacagt tactcctcag atatgttctt ctttaacaac 1080
agagttttac ggagccgaaa cccttcttca ctcacgcggc gttgctccat cagactttcg 1140
tccattgtgg aagattccct actgctgcct cccgtaggag tttgggccgt gtctcagtcc 1200
caatgtggcc gattaccctc tcaggtcggc tacgtatcat tgccatggtg agccgttacc 1260
ccaccatcta gctaatacgc cgcgggacca tccaaaagtg atagccgaag ccatctttca 1320
agctcggacc atgcggtcca agttgttatg cggtattagc atctgtttcc aggtgttatc 1380
ccccgcttct gggcaggttt cccacgtgtt actcaccagt tcgccactca ctcaaatgta 1440
aatcatgatg caagcaccaa tcaataccag agttcgttcg acttgcatgt attaggcagg 1500
gccg 1504
Claims (10)
1. Lactobacillus plantarum (Lactobacillus plantarum) CH126, its deposit number is CGMCC NO.13228.
2. application of the Lactobacillus plantarum described in claim 1 in product is prepared.
3. application as claimed in claim 2, it is characterised in that:The product is food, health products or medicine.
4. application as claimed in claim 2 or claim 3, it is characterised in that:The function of the product is in following (a1) to (a31)
At least one:
(a1) hypoglycemic and/or reducing blood lipid;
(a2) blood-sugar content in serum is reduced;
(a3) lipids contents in serum are reduced;
(a4) total cholesterol level in serum is reduced;
(a5) content of triglyceride in serum is reduced;
(a6) fasting blood sugar is reduced;
(a7) oxidation resistance is increased;
(a8) content and/or activity of the superoxide dismutase in serum are increased;
(a9) content and/or activity of the catalase in serum are increased;
(a10) content and/or activity of the glutathione peroxidase in serum are increased;
(a11) content and/or activity of the MDA in serum are reduced;
(a12) disorders of lipid metabolism is improved;
(a13) the D-ALPHA-Hydroxypropionic acid content in serum is reduced;
(a14) free fatty acid content in serum is reduced;
(a15) endotoxin content in serum is reduced;
(a16) treat and/or prevention diabetes;
(a17) blood-sugar content in Diabetes Mellitus is reduced;
(a18) lipids contents in Diabetes Mellitus are reduced;
(a19) total cholesterol level in Diabetes Mellitus is reduced;
(a20) content of triglyceride in Diabetes Mellitus is reduced;
(a21) fasting blood sugar of diabetic is reduced;
(a22) insulin content in Diabetes Mellitus is raised;
(a23) content and/or activity of the superoxide dismutase in Diabetes Mellitus are increased;
(a24) content and/or activity of the catalase in Diabetes Mellitus are increased;
(a25) content and/or activity of the glutathione peroxidase in Diabetes Mellitus are increased;
(a26) content and/or activity of the MDA in Diabetes Mellitus are reduced;
(a27) disorders of lipid metabolism of diabetic is improved;
(a28) the D-ALPHA-Hydroxypropionic acid content in Diabetes Mellitus is reduced;
(a29) free fatty acid content in Diabetes Mellitus is reduced;
(a30) endotoxin content in Diabetes Mellitus is reduced;
(a31) hyperlipidemic conditions of diabetic are improved.
5. application as claimed in claim 4, it is characterised in that:The diabetes are diabetes B.
6. a kind of product, its active component is Lactobacillus plantarum described in claim 1.
7. a kind of product, its active component plant described in the fermentate or claim 1 of Lactobacillus plantarum described in claim 1
The extract of the fermentate of lactobacillus.
8. product as claimed in claims 6 or 7, it is characterised in that:The product is food, health products or medicine.
9. the product as described in claim 6 or 7 or 8, it is characterised in that:The function of the product is following (a1) to (a31)
In at least one:
(a1) hypoglycemic and/or reducing blood lipid;
(a2) blood-sugar content in serum is reduced;
(a3) lipids contents in serum are reduced;
(a4) total cholesterol level in serum is reduced;
(a5) content of triglyceride in serum is reduced;
(a6) fasting blood sugar is reduced;
(a7) oxidation resistance is increased;
(a8) content and/or activity of the superoxide dismutase in serum are increased;
(a9) content and/or activity of the catalase in serum are increased;
(a10) content and/or activity of the glutathione peroxidase in serum are increased;
(a11) content and/or activity of the MDA in serum are reduced;
(a12) disorders of lipid metabolism is improved;
(a13) the D-ALPHA-Hydroxypropionic acid content in serum is reduced;
(a14) free fatty acid content in serum is reduced;
(a15) endotoxin content in serum is reduced;
(a16) treat and/or prevention diabetes;
(a17) blood-sugar content in Diabetes Mellitus is reduced;
(a18) lipids contents in Diabetes Mellitus are reduced;
(a19) total cholesterol level in Diabetes Mellitus is reduced;
(a20) content of triglyceride in Diabetes Mellitus is reduced;
(a21) fasting blood sugar of diabetic is reduced;
(a22) insulin content in Diabetes Mellitus is raised;
(a23) content and/or activity of the superoxide dismutase in Diabetes Mellitus are increased;
(a24) content and/or activity of the catalase in Diabetes Mellitus are increased;
(a25) content and/or activity of the glutathione peroxidase in Diabetes Mellitus are increased;
(a26) content and/or activity of the MDA in Diabetes Mellitus are reduced;
(a27) disorders of lipid metabolism of diabetic is improved;
(a28) the D-ALPHA-Hydroxypropionic acid content in Diabetes Mellitus is reduced;
(a29) free fatty acid content in Diabetes Mellitus is reduced;
(a30) endotoxin content in Diabetes Mellitus is reduced;
(a31) hyperlipidemic conditions of diabetic are improved.
10. product as claimed in claim 9, it is characterised in that:The diabetes are diabetes B.
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CN108085285A (en) * | 2018-01-25 | 2018-05-29 | 吉林省命之元生物科技有限公司 | One DM-50 plants of lactobacillus plantarum and its application |
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CN115135749A (en) * | 2019-12-31 | 2022-09-30 | Gi生物群系公司 | Lactobacillus plantarum strain and composition for preventing or treating metabolic diseases containing the same |
CN113308421A (en) * | 2021-07-30 | 2021-08-27 | 北京量化健康科技有限公司 | Lactobacillus plantarum BUFX and application thereof in metabolic syndrome |
CN113308421B (en) * | 2021-07-30 | 2022-01-07 | 北京量化健康科技有限公司 | Lactobacillus plantarum BUFX and application thereof in metabolic syndrome |
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