CN106937745A - It is a kind of that there are health products of improvement function of intestinal canal and preparation method thereof - Google Patents
It is a kind of that there are health products of improvement function of intestinal canal and preparation method thereof Download PDFInfo
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- CN106937745A CN106937745A CN201610764311.4A CN201610764311A CN106937745A CN 106937745 A CN106937745 A CN 106937745A CN 201610764311 A CN201610764311 A CN 201610764311A CN 106937745 A CN106937745 A CN 106937745A
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- acetic acid
- acid bacteria
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- dried powder
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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
-
- 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
Abstract
The present invention relates to a kind of health products for having and improving function of intestinal canal and preparation method thereof.The health products are using active acetate bacterium as major ingredient, using nutrients such as milk powder, PURE WHEYs as auxiliary material, and the quantity of acetic acid bacteria is no less than 1 × 10 in preparation4Individual~1 × 1010Individual/gram, it is freeze-dried, it is fabricated to pulvis, capsule, tablets and other formulations.The health treatment of the present invention, can promote the growth of human small intestine's intestinal villus surface area, change the composition of gut flora, increase the relative amount of intestines and stomach probiotics；The relative amount of harmful bacteria in intestines and stomach is reduced, promotes the absorption of nutriment；So as to reach the good result for taking care of health and being healthy and strong.
Description
Technical field
It is specifically a kind of that there is the health care for improving function of intestinal canal the present invention relates to a kind of health products and preparation method thereof
Articles for use and preparation method thereof.
Background technology
Acetic acid bacteria belongs to aerobic Gram-negative bacteria, means：Carbohydrate, sugar can be aoxidized using oxygen as terminal electron acceptor
Alcohols and alcohols generate the general name of a class Gram-negative bacteria of corresponding sugar alcohol, ketone and organic acid.Acetic acid bacteria mainly includes vinegar
Acidfast bacilli category (Acetobacter, A.), Gluconobacter (Gluconobacter, G.) and sour zygosaccharomyces (Acidomonas,
Ac.).With the progressively in-depth recognized microorganism, some new acetic acid bacteria quasi-microorganisms are constantly found that, by 2014
Just, the acetic acid circle reported has 16 category, 84 kinds.
Acetic acid bacteria is important industrial microorganism, is mainly used in the production of vinegar and fruit vinegar beverage.With Dichlorodiphenyl Acetate bacterium
What is solved gos deep into, and the utilization for acetic acid bacteria also gradually increases.Except traditional utilization acetic acid bacteria various vinegars of fermentation manufacture and fruit
Outside vinegar beverage, due to unique metabolic pathway that acetic acid bacteria is formed in evolution, people are also turned using the biology of acetic acid bacteria
Change function, produce some special products：Such as, cocoa bean, after the microbial fermentations such as acetic acid bacteria, can possess the mouth of uniqueness
Sense, fragrance and color；And for example, using acetic acid bacteria, synthesize that good transmittance, tensile strength are high, have very strong hydrophily again and excellent
The high-performance fiber element of good biocompatibility；Acetic acid bacteria be also used for the medicines such as Tagatose, shikimic acid, Vc, Miglitol and
The production of its precursor.Meanwhile, people also gradually recognize some compositions or metabolite of acetic acid thalline, to health meeting
Beneficial effect is produced, e.g., the characteristics of using the relevant enzymes for producing acetic acid in acetic acid bacteria containing Oxidation of Alcohol, can be taken
Acetic acid bacteria reaches the effect relieved the effect of alcohol.
The content of the invention
There is document announcement：Beneficial bacteria of intestinal tract can be with fermentable carbohydrates, a variety of volatile fatty acids of generation, such as formic acid,
Acetic acid, propionic acid and butyric acid.These small molecules acid produces good effect to the health of intestine of young pigs：When prebiotic bacterial content in diet
During rise, the content of volatile fatty acid is dramatically increased in enteron aisle, while p H are significantly reduced.Because acetic acid bacteria quasi-microorganism can
To produce organic acid, while the various vitamin substances needed for being also enriched in maintenance metabolism, may have one to improving function of intestinal canal
Fixed effect；In addition, in the long history of human consumption vinegar, due to the limitation of filter method, unconsciously being taken in vinegar
A small amount of acetic acid bacteria thalline, and the vinegar does not cause damage to human body, it is seen that acetic acid bacteria is free from side effects to human body.
The zoopery that inventor is carried out by using acetic acid bacteria preparation is disclosed：Take after acetic acid bacteria preparation, can promote
Enter the increase of experimental animal small intestine intestinal villus surface area, reduce intestinal pH, change the composition of gut flora, increase beneficial bacteria of intestinal tract
Relative amount；The relative amount of harmful bacteria in enteron aisle is reduced, promotes the absorption of nutriment；So as to reach sanatory effect
Really.
It is an object of the invention to using acetic acid bacteria as primary raw material, take special process, it is made with improvement function of intestinal canal
Health products.
What the present invention was realized in：
The health products of the promotion function of intestinal canal, major ingredient is active acetate bacterium, and remaining is auxiliary material.The health-care preparation：With 1 gram
Meter, quantity contained by its acetic acid bacteria is 1 × 104Individual~1 × 1010Individual/gram；Remaining is nutrients auxiliary material.The nutrients auxiliary material is
Common dietary nutrient, such as milk powder, PURE WHEY, powdered glucose, the nutrients such as starch.
The active acetate bacterium, refers to acetic acid bacteria quasi-microorganism, is：Acetobacter (Acetobacter, A.), glucose bar
Pseudomonas (Gluconobacter, G.) and the single kind or a variety of acetic acid mushrooms of sour zygosaccharomyces (Acidomonas, Ac.)
The mixture of microorganism.Acetic acid bacteria and its metabolite are conducive to strengthening function of intestinal canal in health-care preparation, promote some to plant
The growth of probiotics of class, and suppress the growth of other harmful mushrooms.
The nutrients, can be one kind in milk powder, lactalbumin, powder starch, glucose etc., two or more mixing
Thing.The nutrients can maintain the activity of acetic acid bacteria in health products, and provide certain nutrition and flavor substance for product.
The preparation method and step of the acetic acid bacteria health products be：
A. acetic acid bacteria is conventionally cultivated, thalline is collected；
B. the 2-5 times of 0.05M phosphate buffer for measuring pH=6.5 is added in collected thalline, while adding nutrients, is stirred
Mix uniform,
C. above-mentioned solution is poured into culture dish, it is 3-6mm or so to make bacterium solution thickness, -20 DEG C of pre-freeze 2h, freeze-drying is obtained
To thalline freeze-dried powder；
D. by above-mentioned thalline freeze-dried powder by conventional formulation processing technology, corresponding excipient or filler is added, is mixed and stirred
Uniformly, you can corresponding capsule, tablet or electuary is made.
The health treatment of the present invention, can promote the growth of human small intestine's intestinal villus surface area, change the group of gut flora
Into the relative amount of increase intestines and stomach probiotics；The relative amount of harmful bacteria in intestines and stomach is reduced, promotes the suction of nutriment
Receive；So as to reach the good result for taking care of health and being healthy and strong.
Brief description of the drawings
Influence of Fig. 1 acetic acid bacterias freeze-dried powders to mouse intestinal mucosa morphology
Fig. 2 16S rRNA sequence high-flux sequence dilution curve figures
The horizontal composition schematic diagram of Fig. 3 different disposal group enterobacteriaceae group's structure doors
The horizontal composition schematic diagram of Fig. 4 different disposal group enterobacteriaceae group's structure doors
Embodiment
With reference to several embodiments, the present invention will be further described, but the invention is not restricted to these embodiments.
A kind of health products for improving function of intestinal canal of embodiment 1 and its method for preparing capsule
First by Acetobacter pasteurianus subspecies (Acetobacter pasteurianus subsp. Pasteurianus,
AS1.1550) being inoculated in solid medium, (culture medium composition is glucose 100g, yeast extract 10g, calcium carbonate 20g, agar 18
~20g, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30 DEG C of cultures after cultivating 48 hours, add 2 times of amount pH=6.5
0.05M phosphate buffers, while adding skimmed milk power (0.2 gram/mL), after 5000rpm is homogenized 5 minutes in homogenizer, pour into
Culture dish, bacterium solution thickness 3mm, -20 DEG C of pre-freezes, then freeze-drying obtains thalline freeze-dried powder.Freeze-dried powder is directly encapsulated：Often
Grain weight 0.3g, every 300mg of freeze-dried powder containing acetic acid bacteria, acetic acid bacteria quantity is 1 × 104Individual~1 × 106Individual/gram.
Usage and dosage：Orally, each 3-5, twice a day, warm water is taken.
A kind of health products for improving function of intestinal canal of embodiment 2 and its method for preparing capsule
First by sour monad (Acidomonas, Ac.) be inoculated in solid medium (culture medium composition for glucose 100g,
Yeast extract 10g, calcium carbonate 20g, 18~20g of agar, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30 DEG C of cultures, training
After supporting 48 hours, 5 times of amount pH=6.5 0.05M phosphate buffers are added, while adding skimmed milk power (0.2 gram/mL), homogenate
After 5000rpm is homogenized 5 minutes in device, culture dish is poured into, bacterium solution thickness 6mm, -20 DEG C of pre-freezes, then freeze-drying obtains thalline
Freeze-dried powder.Freeze-dried powder is directly encapsulated：Every weight 0.3g, every 300mg of freeze-dried powder containing acetic acid bacteria, acetic acid bacteria quantity is 1 × 107
Individual~1 × 1010Individual/gram.
Usage and dosage：Orally, each 3-5, twice a day, warm water is taken.
Embodiment 3 it is a kind of improve function of intestinal canal health products and its prepare general-purpose glue electuary method
Gluconobacter oxydans (Gluconobacter oxydans), being inoculated in solid medium, (culture medium composition is grape
Sugared 100g, yeast extract 10g, calcium carbonate 20g, 18~20g of agar, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30 DEG C
Culture, after cultivating 72 hours, adds 3 times of amount pH=6.5 0.05M phosphate buffers, while addition PURE WHEY (0.2 gram/
ML), after 5000rpm is homogenized 5 minutes in homogenizer, culture dish is poured into, then bacterium solution thickness 6mm, -20 DEG C of pre-freezes are freeze-dried
Obtain thalline freeze-dried powder.Freeze-dried powder and Bifidobacterium freeze-dried powder are mixed according to 1: 1 ratio, and punching is directly prepared into after stirring
Agent..Freeze-dried powder directly packs sealing：Every bag of weight 0.6g, every bag of 300mg of freeze-dried powder containing acetic acid bacteria, acetic acid bacteria quantity is 1 × 107
Individual~1 × 1010Individual/gram.
Usage and dosage：Orally, each 2-3 bags, twice a day, warm water is taken.
A kind of health products for improving function of intestinal canal of embodiment 4 and its method for preparing tablet
Acetobacter pasteurianus subspecies (Acetobacter pasteurianus subsp. Pasteurianus,
AS1.1550), being inoculated in solid medium, (culture medium composition is glucose 100g, yeast extract 10g, calcium carbonate 20g, agar 18
~20g, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30 DEG C of cultures after cultivating 48-72 hours, add 3 times of amount pH=
6.5 0.05M phosphate buffers, while after 5000rpm is homogenized 5 minutes in addition skimmed milk power (0.2 gram/mL), homogenizer, falling
Enter culture dish, bacterium solution thickness 6mm, -20 DEG C of pre-freezes, then freeze-drying obtains thalline freeze-dried powder.300 grams of addition dextrin of freeze-dried powder
92 grams, 8 grams of sodium carboxymethylcellulose, after fully mixing, compressing dry granulation, every 0.4 gram of weight, every freeze-dried powder containing acetic acid bacteria
300mg, acetic acid bacteria quantity is 1 × 107Individual~1 × 1010Individual/gram.
Usage and dosage：Orally, each 3-5, twice a day, warm water is taken.
A kind of health products for improving function of intestinal canal of embodiment 4 and its method for preparing general-purpose glue wafer 1000
Exemplified by preparing product general-purpose glue wafer 1000 of the present invention：Acetobacter pasteurianus subspecies (Acetobacter
Pasteurianus subsp.Pasteurianus, AS1.1550), being inoculated in solid medium, (culture medium composition is glucose
100g, yeast extract 10g, calcium carbonate 20g, 18~20g of agar, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30 DEG C of trainings
Support, after cultivating 48-72 hours, add 3 times of amount pH=6.5 0.05M phosphate buffers, while addition skimmed milk power (0.2 gram/
ML), after 5000rpm is homogenized 5 minutes in homogenizer, culture dish is poured into, then bacterium solution thickness 6mm, -20 DEG C of pre-freezes are freeze-dried
Obtain thalline freeze-dried powder.Freeze-dried powder and bacillus subtilis dry powder are mixed according to 10: 1 ratio, straight glue for connecting after stirring
Capsule, every weight 0.33g, every 300mg of freeze-dried powder containing acetic acid bacteria, acetic acid bacteria quantity is 1 × 107Individual~1 × 1010Individual/gram..
Usage and dosage：Orally, each 3-5, twice a day, warm water is taken.
Embodiment 5 it is a kind of improve function of intestinal canal health products and its prepare capsule agent method
Acetobacter aceti (Acetobacter aceti), being inoculated in solid medium, (culture medium composition is glucose
100g, yeast extract 10g, calcium carbonate 20g, 18~20g of agar, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30 DEG C of trainings
Support, after cultivating 48-72 hours, add 4 times of amount pH=6.5 0.05M phosphate buffers, while addition skimmed milk power (0.2 gram/
ML), after 5000rpm is homogenized 5 minutes in homogenizer, culture dish is poured into, then bacterium solution thickness 6mm, -20 DEG C of pre-freezes are freeze-dried
Obtain thalline freeze-dried powder.Freeze-dried powder is directly encapsulated, every weight 0.3g, every 300mg of freeze-dried powder containing acetic acid bacteria, acetic acid bacteria quantity
For 1 × 107Individual~1 × 1010Individual/gram..
Usage and dosage：Orally, each 3-5, twice a day, warm water is taken.
A kind of health products for improving function of intestinal canal of embodiment 6 and its prepare side exemplified by capsule 1000
With acetobacter aceti (Acetobacter aceti) and Acetobacter pasteurianus subspecies (Acetobacter
Pasteurianus subsp.Pasteurianus, AS1.1550) in solid medium, (culture medium composition is Portugal for combined inoculation
Grape sugar 100g, yeast extract 10g, calcium carbonate 20g, 18~20g of agar, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 30
DEG C culture, culture 48-72 hour after, add 5 times amount pH=6.5 0.05M phosphate buffers, while addition skimmed milk power
After 5000rpm is homogenized 5 minutes in (0.2 gram/mL), homogenizer, culture dish, bacterium solution thickness 6mm, -20 DEG C of pre-freezes, Ran Houleng are poured into
Jelly is dried to obtain thalline freeze-dried powder.Freeze-dried powder is directly encapsulated, every weight 0.3g, every 300mg of freeze-dried powder containing acetic acid bacteria, acetic acid
Bacterium number amount is 1 × 107Individual~1 × 1010Individual/gram..
Usage and dosage：Orally, each 3-5, twice a day, warm water is taken.
Improve the laboratory report of function of intestinal canal the following is acetic acid bacteria freeze-dried powder
First, influence of the acetic acid bacteria freeze-dried powder to function of intestinal canal and gut flora
1 experiment material and animal processing
The preparation of 1.1 acetic acid bacteria freeze-dried powders：Acetobacter pasteurianus subspecies (Acetobacter pasteurianus
Subsp.Pasteurianus, AS1.1550), being inoculated in solid medium, (culture medium composition is glucose 100g, yeast extract
10g, calcium carbonate 20g, 18~20g of agar, 20 milliliters of distilled water 1L, pH=6.8,95% ethanol), 48- is cultivated in 30 DEG C of cultures
After 72 hours, 3 times of amount pH=6.5 0.05M phosphate buffers are added, while adding skimmed milk power (0.2 gram/mL), homogenizer
After middle 5000rpm is homogenized 5 minutes, culture dish is poured into, 6mm-20 DEG C of pre-freeze of bacterium solution thickness, then freeze-drying obtains thalline and freezed
Powder..
1.2 experimental animals are grouped and handled：60 male and female half and half of Kunming mouse, body weight (20 ± 2) g, are purchased from the 4th army
Experimental Animal Center, quality certification number are learned by medical university：SCXK (army) 20013-007, raises in standardization Animal House, adapts to environment 4d,
Free water acquisition, feeding, room ventilation condition are good, normal day-night change, relative humidity 40%~70%, 18~22 DEG C of room temperature.
Animal is randomly divided into 5 groups after being layered by body weight, every group of 12 mouse, male and female half and half, acetic acid bacteria freeze-dried powder is divided into height
In low three dosage groups (being denoted as AH, AM, AL respectively), respectively according to 0.2,0.4,0.8g/kgd dosage gavage gives acetic acid
Bacterium freeze-dried powder, positive drug group (PCK) gives probiotics preparation (Medilac-Vita, Beijing S. Korea and the USA medicine according to 0.2g/kgd dosage
Co., Ltd), blank control group (NCK) gives the solvent of respective volume, successive administration 2 weeks, after last dose, animal fasting
12h, during animal fasting, proper amount of fresh mouse excrement sample is taken with sterilizing tweezers or disinfecting cotton swab, is extracted for DNA.Mouse is weighed
Afterwards, anaesthetize, eye socket takes blood (being used to determine blood glucose, blood fat) to put to death animal afterwards, and separating mouse small intestine of cutting open the belly collects intestinal contents
Afterwards, take close to ileum portion about 1cm intestinal tubes, clean intestinal contents with physiological saline, put in 10% formaldehyde and fix stand-by.
2 detection methods and result
2.1 blood sugar detections
According to glucose (Glu) testing cassete, (glucose determination reagent box (the limited public affairs of Changchun remittance power biotechnology) illustrate behaviour
Make, measure mouse blood sugar, it is as a result as follows：
Influence (n=12) of the acetic acid bacteria freeze-dried powder to mouse blood sugar
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL freezes for acetic acid bacteria
Dry powder low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
Examined through t-, each group blood glucose value data difference nonsignificance, illustrate that acetic acid bacteria freeze-dried powder influences on mouse blood sugar
It is smaller.
2.2 changes of weight
Acetic acid bacteria freeze-dried powder influence (n=12) increased on Mouse Weight
| Group | NCK | PCK | AL | AM | AH |
| Original body mass (g) | 20.45±1.30 | 20.21±1.18 | 20.82±1.36 | 20.51±1.15 | 20.80±1.22 |
| Whole body weight (g) | 22.55±1.54 | 23.03±1.47 | 24.08±1.40★ | 24.52±1.07※ | 24.86±1.32※ |
★Represent compared with Normal group, t- is examined, p ＜ 0.05,※Represent compared with Normal group, t- inspections
Test, p ＜ 0.01.
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL freezes for acetic acid bacteria
Dry powder low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
From the results, it was seen that giving after acetic acid bacteria freeze-dried powder, Mouse Weight increase is very fast, compared with control group, number
According to difference significance.Positive drug group Mouse Weight also has increase trend, but after statistical disposition, data difference is without notable meaning
Justice.
2.3 Analysis of blood lipid
T-CHOL (TC) determines kit, triglycerides (TG) and determines kit, HDL-C (HDL)
Determine kit, LDL-C (LDL) measure kit to be purchased from Nanjing and build up Bioengineering Research Institute, according to examination
Agent box specification is operated, as a result as follows.
Influence (n=12) of the acetic acid bacteria freeze-dried powder to lipid of mice
| Test index | NCK | PCK | AL | AM | AH |
| TG mmol/L | 1.34±0.30 | 1.58±0.40 | 1.59±0.61 | 1.43±0.40 | 1.38±0.18 |
| TC mmol/L | 3.32±0.21 | 3.21±0.20 | 3.43±0.10 | 3.56±0.29 | 3.54±0.31 |
| LDL mmol/L | 2.12±0.34 | 2.04±0.12 | 2.05±0.21 | 2.30±0.30 | 2.29±0.27 |
| HDL mmol/L | 0.82±0.03 | 0.87±0.14 | 0.90±0.07 | 1.21±0.06※ | 1.03±0.04※ |
※Represent compared with Normal group, t- is examined, p ＜ 0.01.
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL freezes for acetic acid bacteria
Dry powder low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
As a result show, give after acetic acid bacteria freeze-dried powder, mice serum T-CHOL, total triglycerides, low-density lipoprotein
Have no significant change, the hdl concentration of the middle high dose group of acetic acid bacteria freeze-dried powder rises, and data difference has notable meaning
Justice, acetic acid preparation is taken in prompting can increase mice serum hdl concentration.
2.4 intestinal pHs change
The dilution of 0.2g intestinal contents redistilled water is taken to cause 100mL, with acidometer (S220 SevenCompactTM
PH pH) is determined, it is as a result as follows：
Influence (n=12) of the acetic acid bacteria freeze-dried powder to mouse intestinal pH
| Group | NCK | PCK | AL | AM | AH |
| pH | 7.33±0.20 | 6.95±0.13※ | 7.23±0.11 | 7.21±0.15 | 7.18±0.11★ |
★Represent compared with Normal group, t- is examined, p ＜ 0.05,※Represent that t- is examined compared with Normal group,
P ＜ 0.01.
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL freezes for acetic acid bacteria
Dry powder low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
As a result show, take after acetic acid bacteria freeze-dried powder, mouse intestinal pH has downward trend, after statistical disposition, acetic acid bacteria is high
The pH difference significances of dosage group, positive drug also has obvious reduction to act on to intestinal pH.
2.5 intestinal mucosa morphologies change
The intestinal tube fixed is taken, is cut into slices after H-E dyeing, observation Small intestinal mucosa morphology change is soft with Image-pro plus
Part measures small intestinal villous height, and Crypt depth calculates height of naps/Crypt depth ratio.-
Influence (n=10) of the acetic acid bacteria freeze-dried powder to mouse intestinal mucosa morphology
★Represent compared with Normal group, t- is examined, p ＜ 0.05,※Represent that t- is examined compared with Normal group,
P ＜ 0.01.
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL freezes for acetic acid bacteria
Dry powder low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
Fig. 1 is the photo of histotomy, and H-E dyeing, wherein AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria
Freeze-dried powder middle dose group, AL is acetic acid bacteria freeze-dried powder low dose group, and PCK is probiotics preparation Medilac-Vita group, NCK for positive drug group
For blank control group.
As a result show, compared with control group, the intestinal villus length of positive drug group and acetic acid bacteria freeze-dried powder group has increase trend,
Wherein, positive drug group, the intestinal villus length of acetic acid bacteria freeze-dried powder high dose group is compared with control group, and data difference has notable meaning
Justice；The crypts of acetic acid bacteria freeze-dried powder group highly has downward trend, wherein the data difference significance of middle high dose group；Fine hair
In terms of highly/Crypt depth ratio, positive drug group and acetic acid bacteria freeze-dried powder group are raised, data difference significance.Prompting
Intestinal absorption area can be increased by taking acetic acid bacteria freeze-dried powder, promote the absorption of nutriment；Higher height of naps/crypts is deep
Degree ratio reflects higher intestinal villus cell growth and renewal speed faster
The change detection of 2.6 gut floras
Fresh mouse excrement sample (every group parallel take 5 samples), glues mill, DNA (TIANamp is extracted according to kit explanation
Stool DNA Kit, excrement genome DNA extracting reagent kit (centrifugation column type).Complete after genome DNA extraction, utilize 1% fine jade
The genomic DNA of sepharose electrophoresis detection extracting., primer, 16S rRNA primers are designed using bacterial 16 S rRNAV3-V4 areas
For bacterial 16 S rRNA 338F 5 '-ACTCCTACGGGAGGCAGCA-3 ' ,-GGACTACHVGGGTWTCTAAT- of 806R 5 '
3 ' amplification target genes, pcr amplification product, after purification.Purpose fragment is carried out using Illumina Miseq microarray datasets high
Flux is sequenced, and the OTU under 97% similar level carries out biological information statistical analysis.With Usearch (vsesion
7.1http：//drive5.com/uparse/) software platform progress
The statistical analysis of OTU cluster analyses, Taxonomic analysis and structure of community.
2.6.1OUT distribution statisticses
652 kinds (Operational Taxonomic Units, OTU) are detected altogether, according to 97% similitude to non-heavy
Complex sequences (being free of simple sequence) carries out OTU clusters.
2.6.2 dilution curve is analyzed
Fig. 2 is shown, with increasing for sequencing sequence number, and curve tends to flat, illustrates that sequencing data amount rationally, can be covered
Most strains.
In Fig. 2, ordinate rarefaction measure represent the OTU quantity that can be built based on specific sequencing bar number,
I.e. according to 97% the identifiable grouping sheet digit of similarity；Abscissa Number of reads sampled digital table
Show the sequencing bar number extracted immediately in each sample；What the letter such as AH1C, AM1C, NCK2 was represented is the specific sample extracted immediately
This, the sample that such as AH1C is represented is acetic acid bacteria freeze-dried powder high dose No. 1 sample of group, and the sample that AM1C is represented is acetic acid bacteria freeze-dried powder
No. 1 sample of middle dose group.
2.6.3 gut flora diversity compares
Shannon (the Shannon index) index is for estimating one of biodiversity index in sample.It
It is usually used in reflecting alpha diversity indices with Simpson diversity indices.Under the conditions of different disposal it can be seen from table
The equal very little of Shannon, Simpson index variation, illustrates that the diversity of gut flora is basically identical.
The multifarious change (n=5) of mouse intestinal flora under the conditions of different disposal
| Test index | NCK | PCK | AL | AM | AH |
| shannon | 4.78 | 4.82 | 4.87 | 4.78 | 4.75 |
| simpson_hci | 0.019 | 0.015 | 0.016 | 0.019 | 0.019 |
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL freezes for acetic acid bacteria
Dry powder low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
2.6.4 comparison in difference figure of the intestinal microflora in door level
Fig. 3 shows, composition of the different disposal lower intestinal tract flora in door level
In Fig. 3：Ordinate Relative abundence represent relative abundance, and abscissa is different treatment groups, wherein
AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL is acetic acid bacteria freeze-dried powder low dose group,
PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
Different disposal group is consistent in the species composition of door level, by Firmicutes (Firmicutes), Bacteroidetes
(Bacteroidetes), Proteobacteria (Proteobacteria), candidate_division_tm7, deferrization bacillus door
(Deferribacteres), Tenericutes (Tenericutes), actinomyces door (Actinobacteria), conveyor screw door
(Spirochaetae) composition, wherein Firmicutes (Firmicutes), Bacteroidetes (Bacteroidetes), mycetozoan such as
Door (Proteobacteria) in each group relative amount more than 98%, be main dominant bacteria；In door level, not fellow disciple
Relative amount slightly have difference, compared with control group, positive group and the Firmicutes/ of each treatment group of acetic acid bacteria
Bacteroidetes ratio has the Firmicutes/ Bacteroidetes of reduction trend, wherein acetic acid bacteria low dose group
Compared with control group, data difference significance.
Different disposal group enterobacteriaceae structure of community belongs to level composition change (n=5)
| Taxon | AH | AL | AM | NCK | PCK |
| Bacteroidetes | 47.06 | 43.5 | 51.88 | 39.98 | 43.6 |
| Firmicutes | 42.76 | 46.59 | 38.3 | 47.64 | 45.55 |
| Proteobacteria | 8.97 | 7.63 | 8.28 | 10.94 | 8.87 |
| Candidate_division_TM7 | 0.46 | 1.15 | 0.62 | 0.35 | 0.26 |
| Deferribacteres | 0.34 | 0.45 | 0.18 | 0.72 | 0.78 |
| Tenericutes | 0.19 | 0.27 | 0.21 | 0.15 | 0.21 |
| Actinobacteria | 0.12 | 0.36 | 0.32 | 0.2 | 0.16 |
| Spirochaetae | 0.05 | 0 | 0.03 | 0.01 | 0.05 |
| Verrucomicrobia | 0.05 | 0.02 | 0.15 | 0.01 | 0.41 |
| Cyanobacteria | 0.01 | 0.01 | 0.03 | 0.01 | 0.1 |
| Firmicutes/Bacteroidetes | 0.91 | 1.07 | 0.74 | 1.19 | 1.04 |
2.6.5 intestinal microflora is belonging to the comparison in difference figure of level
In Fig. 4, ordinate Relative abundence represent relative abundance, and abscissa is different treatment groups, wherein
AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL is acetic acid bacteria freeze-dried powder low dose group,
PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
Detect the enterobacteriaceae species of 108 category altogether, relative abundance no significant differences of most of category between each group, under
Table lists the relatively large category of some relative abundance differences and its statistical disposition result (is arrived with each microorganism detection in sample
Sequence number represents the relative abundance of each microorganism)
Different disposal group enterobacteriaceae structure of community belongs to level composition change (n=5)
| Taxon | NCK | PCK | AL | AM | AH |
| S24-_norank | 3037±984 | 3660±1442 | 3765±699 | 5174±852※ | 5232±848※ |
| Ruminococcaceae_uncultured | 621±345 | 585±173 | 752±571 | 875±317 | 1542±332※ |
| Allobaculum | 253±150 | 249±132 | 1239±379※ | 841±264※ | 751±135※ |
| Parasutterella | 256±104 | 228±109 | 489±107 | 539±118※ | 481±233 |
| Coprococcus | 42±3 | 110±83 | 78±20※ | 51±29 | 64±21 |
| Akkermansia | 3±1 | 67±25※ | 3.6±3.5 | 25±8※ | 8±6.5 |
| Prevotella | 396±202 | 279±238 | 374±157 | 690±346 | 732±424 |
| Helicobacter | 1029±373 | 679±334 | 313±149※ | 178±34※ | 259±144※ |
| Prevotellaceae_uncultured | 556±150 | 562±371 | 164±70※ | 488±293 | 192±120※ |
| Roseburia | 397±154 | 296±81 | 333±125 | 76±56※ | 108±68※ |
| Rikenella | 120±37 | 91±26 | 36±27※ | 44±23※ | 101±50 |
| Anaerotruncus | 273±152 | 215±131 | 163±42 | 138±36 | 225±48 |
| Odoribacter | 110±42 | 284±75※ | 82±80 | 122±79 | 144±19 |
| RC9_gut_group | 198±178 | 142±86 | 53±43 | 106±47 | 71±32 |
| Lachnospiraceae_uncultured | 3844±1500 | 4647±2324 | 2919±481 | 2333±1120 | 3349±692 |
| Bacteroides | 1611±380 | 978.±282 | 1498±822 | 1287±920 | 1257±668 |
※Represent that t- is examined compared with Normal group, p ＜ 0.05.
In table：AH represents acetic acid bacteria freeze-dried powder high dose group, and AM is acetic acid bacteria freeze-dried powder middle dose group, and AL is bacterium freeze-dried powder
Low dose group, PCK is probiotics preparation Medilac-Vita group for positive drug group, and NCK is blank control group.
The relative abundances for having 9 category it can be seen from table have downward trend, through statistical check, Helicobacter,
The data difference of Prevotellaceae_uncultured, Roseburia, Rikenella in the relative abundance for the dosage group having
There is significant, the relative abundance of 7 category is on the rise, statistical analysis, S24-7_norank,
Ruminococcaceae_uncultured, Allobaculum, Parasutterella, Coprococcus, Akkermansia
There is significant in the data difference of the relative abundance for the dosage group having
3. conclusion
It can be seen from the results that gavage is given after mouse acetic acid bacteria freeze-dried powder, Mouse Weight increase is fast compared with control group, explanation
Acetic acid bacteria freeze-dried powder can promote mouse growth, and the research to mouse intestinal mucosa morphology shows, acetic acid bacteria freeze-dried powder can increase
Mouse intestinal height of naps, improves the absorption area and small intestine of the ratio of height of naps/Crypt depth, the two indexs and small intestine
Function is closely related【1】, point out acetic acid bacteria freeze-dried powder to promote the absorption of nutriment.The small size decline of pH in digestive tract can be with
Regulate and control animal intestinal tract microbial balance, propagation beneficial bacterium, suppress harmful bacteria, improve the effect such as digestive tract enzyme activity【2,3】, this experiment
As a result, acetic acid bacteria freeze-dried powder is taken, mouse intestinal pH can reduce 0.1-0.15 units, points out acetic acid bacteria freeze-dried powder that there is regulation
The effect of mouse intestinal function, it is consistent with the result of Mouse Weight increase, intestinal villus metamorphosis.
The measurement result of blood glucose and blood fat shows, it is smaller to blood glucose, lipid to take acetic acid bacteria freeze-dried powder, to serum
HDL-C content has a rise effect, and other lipid contents are without clearly changing.Epidemiological study is disclosed, HDL-C content
Occur with cardiovascular and cerebrovascular disease negatively correlated【4】, this experimental result prompting acetic acid bacteria freeze-dried powder there is potential health value.
The change of gut flora and health are closely related, it has already been proven that many disease such as colon cancer, diabetes, cardiovascular and cerebrovasculars
The generation of disease etc. is related to the change of gut flora, and the TMAO (TMAO) that such as enteric microorganism metabolism is produced can be led
Atherogenicity【5】, enterobacter cloacae B29, melaninogenicus is related to fat generation【6-7】, autoimmune disease,
Diabetes, chronic kidney disease, anaphylactia, self-closing disease etc. are related to the change of gut flora【8】.But it is due to gut flora
Composition is complicated, and human intestine possesses 100,000,000,000,000 nearly 1 000-11500 kind bacteriums, and most enterobacteriaceaes can not still succeed in vitro
Culture, the classification position of some strains is also indefinite, so currently known probiotics species is less, such as Bifidobacterium, newborn bar
Bacterium, hay bacillus etc., and the physiological function of most bacterium is unclear.This result of study is shown, is given acetic acid bacteria and is freezed
After powder, mouse intestinal flora there occurs that some change, and in door level, Firmicutes/Bacteroidetes ratio has
Reduction trend, document shows that the ratio, which reduces, may indicate in preferable health status, compared with common old man, long-lived
The gut flora Firmicutes/Bacteroidetes ratios of old man are relatively low【9】.In category level, the 13 of relative abundance change
In individual category, it is considered that Allobaculum, Coprococcus category bacterium participate in the synthesis of short chain fatty acids, and participate in maintaining intestines
The stability of road flora【10-11】.Endocannabinoids can influence the change of gut flora, such as Akkermansia【12】, diabetes
Rat is given after Or Metformin In Treating, and Akkermansia relative abundance also rises, and simple oral Akkermansia can also
Improve diabetic symptom【13】, this research is shown, is given after acetic acid bacteria freeze-dried powder, Akkermansia relative abundance is carried in enteron aisle
Height, points out acetic acid bacteria freeze-dried powder to have potential value in terms of diabetes generation is prevented.Prevotella belongs in long lived elder intestines
The relative abundance in road is higher than common old man【9】, the Prevotella relative amounts in obesity patient's body are low, give arabinose
After sill glycan, Prevotella relative amounts rise, and the metabolism state of obesity patient is relative with Roseburia spp. to be contained
Amount is negatively correlated, negatively correlated with Bacteroides/Prevotella levels【14】, this result of study display, acetic acid bacteria freeze
Prevotella relative amounts rise after powder processing, and Roseburia relative amounts decline.Helicobacter be chronic gastritis,
The Etiological of peptic ulcer, stomach cancer and gastric mucosa-associated lymphoid tissue lymphoma, gives after acetic acid bacteria,
Helicobacter relative amount is substantially reduced.
To sum up state, acetic acid bacteria freeze-dried powder can increase intestinal villus surface area, promote the absorption of nutriment, thus it is possible to vary intestines
The composition of road flora, with potential sanatory function.
Bibliography
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[J] HEILONGJIANG ANIMAL SCIENCE AND VETERINARY MEDICINEs, 2014,05：91-92
[3] Xiao Qin, Shen Yiru, Zhang Shan, Chen Jie, apply Shou Rong compound acidulants to broiler chicken pH in digestive tract and digestive enzyme activity
Influence [J] Animal nutrition journals, 2015,08：2527-2533.
[4] the Chinese artery sclerosis magazines of present Research [J] of Zhao Shuiping HDLs, 2005,06：673-675.
5th, Koeth RA, Wang Z, Levison BS, et al.Intestinal microbiota metabolism
Of l-carnitine, a nutrient in red meat, promotes atherosclerosis [J] .Nature
Medicine, 2013,19 (5)：576-585
[6] Zhang H, DiBaise JK, Zuccolo A, et al.Human gut microbiota in obesity
and after gastric bypass[J].Proceedings of the National Academy of Sciences
Of the United States of America, 2009,106 (7)：2365-2370
[7] Zhang C, Zhang M, Pang X, et al.Structural resilience of the gut
microbiota in adult mice under high-fat dietary perturbations[J].ISME
Journal, 2012,6 (10)：1848-1857
[8] Guo Huiling, Shao Yuyu, Meng He, wait progress [J] the microbiologies of gut floras and disease relationship to lead to
Report, 2015,42 (2)：400-410.
[9] Park S H, Kim K A, Ahn Y T, et al.Comparative analysis of gut
microbiota in elderly people of urbanized towns and longevity villages[J].Bmc
Microbiology, 2015,15 (1)：1-9.
[10] Tap J, Furet J P, Bensaada M, et al.Gut microbiota richness promotes
its stability upon increased dietary fibre intake in healthy adults[J]
.Environmental microbiology, 2015,17 (12)：4954-4964.
[11] Ferrario C, Taverniti V, Milani C, et al.Modulation of fecal
Clostridiales bacteria and butyrate by probiotic intervention with
Lactobacillus paracasei DG varies among healthy adults.[J].Journal of
Nutrition, 2014,144 (11)：1787-96.
[12] Cani P D, Geurts L, Matamoros S, et al.Glucose metabolism：focus on
Gut microbiota, the endocannabinoid system and beyond. [J] .Diabetes＆Metabolism,
2014,40 (4)：246-257.
[13] Shin N R, Lee J C, Lee H Y, et al.An increase in the Akkermansia
spp.population induced by metformin treatment improves glucose homeostasis in
Diet-induced obese mice. [J] .Molecular Medicine Reports, 2014,63 (5)：727-35.
[14] Neyrinck A M, Possemiers S, Druart C, et al.Prebiotic effects of
Wheat arabinoxylan related to the increase in bifidobacteria, Roseburia and
Bacteroides/Prevotella in diet-induced obese mice. [J] .Plos One, 2011,6 (6)：230-
234.
[15] Shi Leqin, Zheng Rongliang, king grasp auspicious helicobacter pyloris [J] microbiologies immunology progress, 2007,03：51-
68.
2nd, the studies on acute toxicity of acetic acid bacteria freeze-dried powder
1st, test medicine
Acetic acid bacteria freeze-dried powder, ditto, before use with distillation water dissolves, preparing turns into Cmax 1.5g/mL..
2nd, experimental animal
ICR mouse are provided by Xi'an Jiaotong University Medical College's medical experiment animal center, 20~22g of body weight, mouse age 6~7
Week.
3rd, experimental method
Through trial test, acetic acid bacteria freeze-dried powder LD50 can not be measured, so determining the maximal tolerance dose of acetic acid bacteria freeze-dried powder.Take
Healthy ICR mouse 20,20~22g of body weight, male and female half and half, fasting 12 hours before experiment, by Cmax, maximum volume
(40ml/kg) gastric infusion, Continuous Observation 7 days after administration record animal dead number.
4th, experimental result：Mouse situation is without exception in seven days after perfusion, and none is only dead.
5 conclusions：When acetic acid bacteria freeze-dried powder gives mouse with administration by gavage, maximal tolerance dose is more than 30g/kg, changes with the medicine
Maximum dose level 0.8g/kg used by kind function of intestinal canal is compared, more than 37 times, illustrates that the product uses safety.
Claims (3)
1. a kind of have the health products for improving function of intestinal canal, it is characterised in that：Its major ingredient is active acetate bacterium, and remaining is nutrients
Auxiliary material；
The active acetate bacterium, refers mainly to acetic acid bacteria quasi-microorganism, can be acetobacter, Gluconobacter and sour monad
The mixture of the single kind of category or a variety of acetic acid bacteria quasi-microorganisms；The quantity of acetic acid bacteria is no less than 1 × 10 in preparation4Individual~
1×1010Individual/gram.
2. there are the health products for improving function of intestinal canal according to claim 1, it is characterised in that：The nutrients auxiliary material, be
Refer to containing protein, amino acid, carbohydrate, lipid, vitamin material, can be a kind of, two or more nutrients therein
The mixture of matter.
3. a kind of method for production with the health products for improving function of intestinal canal, it is characterised in that carried out according to following method and steps：
A. the acetic acid bacteria conventionally cultivated, collects thalline；
B. the 2-5 times of 0.05M phosphate buffer for measuring pH=6.5 is added in collected thalline, while adding nutrients, stirring is equal
It is even,
C. above-mentioned solution is poured into culture dish, it is 3-6mm or so to make bacterium solution thickness, -20 DEG C of pre-freeze 2h, freeze-drying obtains bacterium
Body freeze-dried powder；
D. corresponding excipient or filler are added by above-mentioned thalline freeze-dried powder by conventional formulation processing technology, uniform mixing,
It can be made into corresponding capsule, tablet or electuary.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103820368A (en) * | 2014-02-28 | 2014-05-28 | 天津替代医学科技有限公司 | Preparation method and applications of freeze-dried powder of acetic acid bacteria |
| US20140193383A1 (en) * | 2005-02-11 | 2014-07-10 | Technologies Biolactis Inc. | Use of lactobacillus kefiranofaciens as a probiotic and a synbiotic |
-
2016
- 2016-08-31 CN CN201610764311.4A patent/CN106937745A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20140193383A1 (en) * | 2005-02-11 | 2014-07-10 | Technologies Biolactis Inc. | Use of lactobacillus kefiranofaciens as a probiotic and a synbiotic |
| CN103820368A (en) * | 2014-02-28 | 2014-05-28 | 天津替代医学科技有限公司 | Preparation method and applications of freeze-dried powder of acetic acid bacteria |
Non-Patent Citations (1)
| Title |
|---|
| BABAK HAGHSHENAS，等: "Anticancer impacts of potentially probiotic acetic acid bacteria isolated from traditional dairy microbiota", 《LWT-FOOD SCIENCE AND TECHNOLOGY》 * |
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