CN106434435B - One plant of acetobacter and the application in acceleration green starch separation sedimentation - Google Patents
One plant of acetobacter and the application in acceleration green starch separation sedimentation Download PDFInfo
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- CN106434435B CN106434435B CN201610814812.9A CN201610814812A CN106434435B CN 106434435 B CN106434435 B CN 106434435B CN 201610814812 A CN201610814812 A CN 201610814812A CN 106434435 B CN106434435 B CN 106434435B
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- 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
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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
- C12R2001/02—Acetobacter
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B30/00—Preparation of starch, degraded or non-chemically modified starch, amylose, or amylopectin
- C08B30/04—Extraction or purification
Abstract
One plant of acetobacterAcetobacter indonesiensis LLY11, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on July 15th, 2016, and deposit number is CGMCC No.12794.Its LLY11 pure-blood ferment wintercherry improves starch sinking speed using having the advantage that in green starch production, shortens the starch sedimentation time, improves starch yield；Acetobacter indonesiensis LLY11 ferments wintercherry for sterile fermentation, and the pollution of the pathogenic bacteria such as no miscellaneous bacteria improves the edible safety of starch product.
Description
Technical field
The present invention relates to one plant of acetobacter and accelerating the application in green starch separation sedimentation.
Technical background
Acetic acid bacteria can improve flavor, quality and the nutrition of food, extend the shelf life, and have adjusting human body intestinal canal flora flat
Weighing apparatus, the growth for inhibiting pathogen reduce cholesterol, improve the important Functions of Physiological Health Care such as immunity, anti-aging, extensive
Applied to food service industry, acetic acid bacteria fermented food is also considered as functional food.Currently, the production of green starch uses nature
Settled process or acid slurry method, settled process is not only time-consuming but also starch removal is not thorough；Acid slurry method starch separation effect is good compared with settled process,
But since wintercherry is the product of spontaneous fermentation, very big, production is influenced by the environmental factors such as raw material, climate change and operating method
Quality is very unstable, sometimes due to temperature or other factors cause yeast-bitten, wintercherry quality is deteriorated；Sometimes wintercherry vigor is not
Foot, prevent starch from protein and other impurities is timely and effective separates.The main reason for causing these problems is due to right
In wintercherry precipitation and separation starch main function bacterium understanding be not also fully aware of, therefore also never manually prepare, can
For the leavening exploitation of wintercherry production, manual fermentation can't be carried out.
Summary of the invention:
The purpose of the present invention is to the green starch precipitation and separation time is long, starch separation precipitating is not thorough, product
The problems such as unstable quality, one plant of acetobacter for promoting starch separation sedimentation capacity strong is provided, which can add
It plays a key effect during fast green starch sedimentation separation.
Another object of the present invention is to the acetobacter is accelerating the application in starch separation sedimentation, improves starch
Yield improves the stability of product.Meanwhile the wintercherry in the present invention is sterile fermentation, the pollution of the pathogenic bacteria such as no miscellaneous bacteria improves
The edible safety of starch product.
The present invention is achieved by the following technical solutions:
The present invention provides one plant of acetobacter for promoting starch separation sedimentation, which is acetobacterAcetobacter indonesiensis LLY11 is preserved in Chinese microorganism strain preservation pipe on July 15th, 2016
Reason committee common micro-organisms center, deposit number are CGMCC No. 12794.
The individual morphology feature of the acetobacter bacterial strain: size is (0.8 μm~1.2 μm) × (1.5 μm~2.5 μ
M), cell ellipse or quarter butt, Gram-negative, no gemma.Colony morphology characteristic: bacterium colony size is in 2 mm~3mm, milk yellow,
Opaque, colony edge is neat, and surface is smooth, it is glossy to moisten, and swells hemispherical.Physiological and biochemical property are as follows: oxytolerant, peroxide
Change hydrogen enzyme positive, cannot move, hydrogen sulfide, nitrate reduction, gelatin experiment are negative, and metabolizable glucose produces acid but do not produce gas, can generation
Fructose, sucrose, lactose, galactolipin, maltose, mannose, sorbierite, rhamnose, melezitose, melibiose, cellobiose is thanked to produce
Acid cannot utilize raffinose, arabinose.
16 S rDNA sequences of the bacterial strain totally 1411 bp, through Genbank compare withAcetobacter indonesiensisSimilitude reaches 99%；In conjunction with its individual, colony morphology feature and bio-chemical characteristics as a result, identifying the bacterium
ForAcetobacter indonesiensis, and be named as Acetobacter indonesiensis LLY11。
AcetobacterAcetobacter indonesiensis The preparation of LLY11 fermentation wintercherry: oese picking three is used
The acetobacter of ring slant preservationAcetobacter indonesiensisLLY11 is in 5 mL green bean juice culture mediums, and 30 DEG C
After 12 h of lower fermentation, this 5mL fermentation liquid is expanded and is cultivated, continued to access and ferment 12 in 100 mL green bean juice culture mediums at 30 DEG C
H is obtainedAcetobacter indonesiensisThe seed culture fluid of LLY11.By the inoculum concentration of 5%-8% by seed culture fluid
It is linked into green bean juice culture medium, ferment 12 h at 30 DEG C, can be obtained the wintercherry for settling green starch cream.
AcetobacterAcetobacter indonesiensisApplication of the LLY11 fermentation wintercherry in sedimentation starch: will
It is above-mentionedAcetobacter indonesiensis LLY11 sterile fermentation obtain wintercherry by volume 10% amount be added to it is green
In beans starch milk, 1-2min is stirred rapidly, is then allowed to stand.The bacterial strain can be specifically bound to starch particle surface, will be numerous
Starch granules is coupled to big flocculation group, since the starch granules gravity being bound up increases, accelerates starch and protein
The separation sinking speed of equal impurity, can be by sinking speed as precipitating needed for 10mL starch in 20min(100mL green starch cream
Time) foreshorten to 2min.
Acetobacter indonesiensis LLY11 pure-blood ferment wintercherry and spontaneous fermentation mung bean wintercherry are formed sediment in mung bean
It is applied to have the advantage that in powder production
(1) starch sinking speed is improved, the starch sedimentation time is shortened, improves starch yield；
⑵ Acetobacter indonesiensis LLY11 ferments wintercherry for sterile fermentation, and the pathogenic bacteria such as no miscellaneous bacteria are dirty
Dye, improves the edible safety of starch product.
Specific embodiment
Embodiment 1
Accelerate separation, screening and the identification of starch sedimentation microorganism
1 culture medium
(1) green bean juice culture medium prescription: yeast extract 5 g, K2HPO4 2g, 20 g of glucose, 2 g of lactose, 5 g of sodium acetate,
1000 mL of green bean juice.
(2) the preparation method of green bean juice: take 100 g mung beans to add distilled water to 1000 mL, 20min is boiled in heating, with 120
Purpose screen filtration is to get green bean juice.
(3) isolation medium: using glucose in soluble starch substitution green bean juice culture medium as carbon source, specific ingredient and
Its outfit is as follows: yeast extract 5 g, K2HPO4 2g, 20 g of soluble starch, 2 g of lactose, 5 g of sodium acetate, 1000 mL of green bean juice.
Adjust pH to 6.8,115 DEG C of 30 min of sterilizing.
2 detection methods
The detection method of 2.1 green starch sinking speed:
The mung bean for taking the 8-12h that is soaked adds water to be beaten in the ratio of 1:15, the green starch cream several pieces for taking 100mL to accomplish fluently,
It is separately added into spontaneous fermentation wintercherry, sterile fermentation liquid and blank green bean juice culture medium, respectively required for measurement sedimentation 10mL starch
Time.Every group is done 3 repetitions.
The detection method of 2.2 coliforms:
Referring to GB 47893-2010 " national food safety standard food microbiological examination enumeration of coliforms ".
3 accelerate starch to settle microorganism separation method
3.1 the dilution of sample
The mung bean wintercherry 5mL of spontaneous fermentation is drawn with aseptic straw, immigration fills 45mL sterile saline band bead
Triangular flask in, shake well is uniform, as 10-1Sample diluting liquid.With sterile saline to 10-1Sample diluting liquid
Gradient dilution is carried out to 10-7。
3.2 plate isolation
Take above-mentioned 10-4、10-5、10-6、10-7Each 1mL of the dilution of dilution is injected separately into sterile petri dish, each dilute
Degree of releasing does 3 repetitions.Isolation medium is injected, after culture medium solidification, is inverted in 30 DEG C of incubators and cultivates for 24 hours~48h.
3.3 plating cultures
With the single petite of oese picking, in isolation medium plate, 30 DEG C of cultures are for 24 hours, repeatedly pure for streak inoculation
Changed for 3 generations, obtain single colonie and be inoculated on green bean juice slant medium, 30 DEG C of cultures are for 24 hours.
3.4 bacterial strain screening
In the green bean juice culture solution that access after the pure bacterial strain activation in the inclined-plane being separated to is substituted to glucose with soluble starch,
30 DEG C of cultures for 24 hours, using the acceleration starch separation sinking speed of fermentation liquid as index, filter out one plant of rush starch separation sinking speed
High bacterial strain LLY11.
3.5 strain idenfication
3.5.1 the morphological feature observation of bacterial strain LLY11
Bacterial strain LLY11 is cultivated for 24 hours at 30 DEG C in green bean juice agar medium, then carries out Gram's staining,
Volume morphing feature is shown in Table 1.3.5 strain idenfication
3.5.1 the morphological feature observation of bacterial strain LLY11
Bacterial strain LLY11 is cultivated for 24 hours at 30 DEG C in green bean juice agar medium, then carries out Gram's staining,
Volume morphing feature is shown in Table 1.
The individual morphology feature of 1 LLY11 bacterial strain of table
| Bacterial strain | Thallus size | Cellular morphology | Arrangement mode after cell division | Gram's staining | Gemma | Motility |
| LLY11 | 0.8 μ~1.2 μm) × (1.5 μm~2.5 μm) | Cell ellipse or quarter butt | Chain | Gram-negative | Nothing | It does not move |
Bacterial strain LLY11 cultivates 48h, observation bacterium colony size, form and color, knot at green bean juice agar medium, 30 DEG C
Fruit is shown in Table 2.
2 colony morphology characteristic of table
3.5.2 the physiological and biochemical property of LLY11
It according to customary physiological biochemical identification, the results are shown in Table 3, referring to " Berger bacterial identification manual ", tentatively judge bacterial strain
LLY11 is acetobacter.
3 bacterial strain physio-biochemical characteristics of table
| Test item | As a result | Test item | As a result |
| Catalase | + | Lactose | + |
| Mobility test | - | Galactolipin | + |
| H2S test | - | Maltose | + |
| Nitrate reduction | - | Mannitol | + |
| Gelatin liquefaction | - | Sorbierite | + |
| 15 DEG C of growths | + | Gossypose | - |
| 45 DEG C of growths | - | Mannose | + |
| Gluconate | + | Rhamnose | + |
| Glucose produces gas | - | Melezitose | + |
| Glucose produces acid | + | Melibiose | + |
| Xylose | - | Seven leaves sugar | + |
| Fructose | + | Arabinose | - |
| Sucrose | + | Cellobiose | + |
3.5.3 the 16S rDNA the sequencing results of acetobacter LLY11
The 16 S total 1411bp of rDNA sequence of the bacterial strain, through Genbank compare withAcetobacter indonesiensisSimilitude reaches 99%, comprehensive its colony morphology observation, individual morphology observation, physiological and biochemical test as a result,
Determine that the bacterial strain isAcetobacter indonesiensisAnd it is named asAcetobacter indonesiensis
LLY11 has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center (referred to as on July 15th, 2016
CGMCC, address are as follows: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, institute of microbiology, the Chinese Academy of Sciences, postcode 100101), preservation is compiled
Number be CGMCC No. 12794.
Embodiment 2
Acetobacter indonesiensisApplication of the LLY11 fermentation wintercherry in starch processing
1 experimental material
The bacterial strain present invention of test separatesAcetobacter indonesiensisLLY11, Starch Production are former
Material selection mung bean.
2 Acetobacter indonesiensisThe preparation of LLY11 fermentation wintercherry
(1) green bean juice culture medium prescription: yeast extract 5g, K2HPO42g, glucose 20g, lactose 2g, sodium acetate 5g, green bean juice
1000mL。
(2) the preparation method of green bean juice: take 100g mung bean to add distilled water to 1000mL, 20min is boiled in heating, with 120 mesh
Screen filtration to get green bean juice.
(3) acetobacter LLY11 seed culture liquid and preparation method thereof: with the acetobacter of oese picking tricyclic slant preservation
LLY11 ferments in 5mL green bean juice culture medium at 30 DEG C after 12h, this 5mL fermentation liquid is expanded and is cultivated, and it is green to continue access 100mL
Ferment at 30 DEG C 12h in fermented bean drink culture medium, obtains the seed culture fluid of acetobacter LLY11.
(4) acetobacter LLY11 seed culture fluid is linked into green bean juice culture medium by the inoculum concentration of 5%-8%, 30 DEG C
Lower fermentation 12h can be obtained the fermentation wintercherry for settling green starch cream.
3 are applied to that starch is accelerated to separate sedimentation with impurity such as protein
It willAcetobacter indonesiensisLLY11 fermentation obtain wintercherry by volume 10% amount be added to
In green starch cream, 1-2min is stirred rapidly, is then allowed to stand.
Addition is taken respectivelyAcetobacter indonesiensis Green starch cream before and after LLY11 fermentation liquid, aobvious
The distribution of micro- microscopic observation starch granules, is shown in Fig. 1, Fig. 2, as seen from the figure,Acetobacter indonesiensis
LLY11 can be specifically bound to starch particle surface, and numerous starch granules are coupled to big flocculation group.Due to being bound up on one
Rise starch granules gravity increase, accelerate the separation sinking speed of the impurity such as starch and protein, can by sinking speed by
In 20min(100mL green starch cream the time required to precipitating 10mL starch) foreshorten to 2min.
Detailed description of the invention
Fig. 1 is that the green starch distribution of particles figure before fermentation liquid is added in observation under 100 power microscopes；
Fig. 2 is to be added 10%Acetobacter indonesiensis LLY11 ferments after wintercherry, green starch particle connection
Form big flocculation group.
Effect of settling is relatively shown in Table 4；
The amount detection of 4 coliforms:
It is right respectivelyAcetobacter indonesiensisLarge intestine in LLY11 fermentation wintercherry and spontaneous fermentation mung bean wintercherry
The quantity of flora is detected, as a result such as the following table 5:
The present invention filters out one plant from spontaneous fermentation mung bean wintercherry can accelerate starch to settleAcetobacter indonesiensis LLY11, and be applied in mung bean fermented wintercherry, fermentation liquid can accelerate green starch separation heavy
Drop.The invention not only shortens the time of starch separation sedimentation, improves starch yield, improves the stability of product；And
Wintercherry in invention is sterile fermentation, and the pollution of the pathogenic bacteria such as no miscellaneous bacteria improves the edible safety of starch product.
SEQUENCE LISTING
<110>Jinzhou medical university
<120>one plants promote the acetic acid bacteria that starch separation settles
<130>accelerating the application in green starch separation sedimentation
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 1411
<212> DNA
<213> Acetobacter indonesiensis
<400> 1
gtggtcggct gcgccccttg cgggttcgct caccggctta aggtcaaacc aactcccatg 60
gtgtgacggg cggtgtgtac aaggcccggg aacgtattca ccgcggcatg ctgatccgcg 120
attactagcg attccacctt catgcactcg agttgcagag tgcaatccga actgagacgg 180
cttttagaga tcagcacgat gtcgccatct agcttcccac tgtcaccgcc attgtagcac 240
gtgtgtagcc caggacataa gggccatgag gacttgacgt catccccacc ttcctccggc 300
ttgtcaccgg cagtctctct agagtgccca cccaaacatg ctggcaacta aagatagggg 360
ttgcgctcgt tgcgggactt aacccaacat ctcacgacac gagctgacga cagccatgca 420
gcacctgtgc ggtaggtccc ttgcgggaaa tatccatctc tgaatacagc ctacccatac 480
aagccctggt aaggttctgc gcgttgcttc gaattaaacc acatgctcca ccgcttgtgc 540
gggcccccgt caattccttt gagtttcaac cttgcggccg tactccccag gcggtgtgct 600
tatcgcgtta gctacgacac tgaataacta agttacccaa catccagcac acatcgttta 660
cagcgtggac taccagggta tctaatcctg tttgctcccc acgctttcgc gcctcagcgt 720
cagtaatgag ccaggttgcc gccttcgcca ccggtgttct tcccaatatc tacgaatttc 780
acctctacac tgggaattcc acaaccctct ctcacactct agtctgcacg tctcaaatgc 840
agctcccagg ttaagcccgg ggatttcaca tctgactgta caaaccgcct acacgccctt 900
tacgcccagt cattccgagc aacgctagcc cccttcgtat taccgcggct gctggcacga 960
agttagccgg ggcttcttct gcgggtaccg tcatcatcgt ccccgccgaa agtgctttac 1020
aatccgaaaa ccttcttcac acacgcggca ttgctggatc agggttgccc ccattgtcca 1080
atattcccca ctgctgcctc ccgtaggagt ctgggccgtg tctcagtccc agtgtggctg 1140
atcatcctct cagaccagct attgatcatc gccttggtag gccattaccc caccaacaag 1200
ctaatcaaac gcaggctcct ccgcaggcga cttgcgcctt tgaccctcag gtgtcatgcg 1260
gtattagctc cagtttccca gagttatccc ccacccacgg atagattcct acgcgttact 1320
cacccgtccg ccactaaggc cgaaaccttc gtgcgacttg catgtgttaa gcatgccgcc 1380
agcgttcgct ctgagccagg attcaaacac t 1411
Claims (3)
1. one plant of acetic acid bacteria for accelerating starch separation to settle, it is characterised in that the bacterium is acetobacterAcetobacter indonesiensisIt is commonly micro- to be preserved in China Committee for Culture Collection of Microorganisms on July 15th, 2016 by LLY11
Bio-Centers, deposit number are CGMCC No. 12794.
2. the one plant according to claim 1 acetic acid bacteria for accelerating starch separation to settle, which is characterized in that acetobacterAcetobacter indonesiensisThe fermentation wintercherry of LLY11 is applied in green starch separation.
3. the one plant according to claim 2 acetic acid bacteria for accelerating starch separation to settle, characterized in that separated in green starch
In, acetobacter usedAcetobacter indonesiensisLLY11 ferments wintercherry using the preparation of following method:
(1) green bean juice culture medium prescription: yeast extract 5g, K2HPO4 2g, glucose 20g, lactose 2g, sodium acetate 5g, green bean juice
1000 mL；
(2) the preparation method of green bean juice: take 100g mung bean to add distilled water to 1000 mL, 20 min are boiled in heating, with 120 purposes
Screen filtration is to get green bean juice；
(3) acetobacter LLY11 seed culture liquid and preparation method thereof: with the acetobacter of oese picking tricyclic slant preservation
LLY11 ferments in 5 mL green bean juice culture mediums at 30 DEG C after 12 h, this 5mL fermentation liquid is expanded and is cultivated, access 100 is continued
Ferment at 30 DEG C 12 h in mL green bean juice culture medium, obtains the seed culture fluid of acetobacter LLY11；
(4) acetobacter LLY11 seed culture fluid is linked into green bean juice culture medium by the inoculum concentration of 5%-8%, 30 DEG C issue
12 h of ferment, can be obtained the wintercherry for settling green starch cream.
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| CN103876005A (en) * | 2014-03-19 | 2014-06-25 | 湖北工业大学 | Compound microbial agent for promoting settlement of sweet potato starch slurry |
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| CN103194409A (en) * | 2013-04-03 | 2013-07-10 | 天地壹号饮料股份有限公司 | Bacillus aceticus and application thereof in preparing apple vinegar |
| CN103876005A (en) * | 2014-03-19 | 2014-06-25 | 湖北工业大学 | Compound microbial agent for promoting settlement of sweet potato starch slurry |
| CN104041726A (en) * | 2014-05-21 | 2014-09-17 | 湖北工业大学 | Complex microbial inoculant capable of promoting settlement of starch in lotus root starch slurry |
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