CN107267401A - One plant of Mucor bacteria strain and the application on fermentation green brick tea - Google Patents
One plant of Mucor bacteria strain and the application on fermentation green brick tea Download PDFInfo
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- CN107267401A CN107267401A CN201710523710.6A CN201710523710A CN107267401A CN 107267401 A CN107267401 A CN 107267401A CN 201710523710 A CN201710523710 A CN 201710523710A CN 107267401 A CN107267401 A CN 107267401A
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
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- C12R2001/00—Microorganisms ; Processes using microorganisms
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23F—COFFEE; TEA; THEIR SUBSTITUTES; MANUFACTURE, PREPARATION, OR INFUSION THEREOF
- A23F3/00—Tea; Tea substitutes; Preparations thereof
- A23F3/06—Treating tea before extraction; Preparations produced thereby
- A23F3/08—Oxidation; Fermentation
- A23F3/10—Fermentation with addition of microorganisms or enzymes
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
- C12P1/02—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using fungi
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Abstract
Application the present invention relates to mucor and on fermentation green brick tea, belongs to microbial fermentation technology application field.The strain isolation of the present invention is carrying out having the tealeaves with good hypha,hyphae hair tonic in the pile fermentation during green brick tea produces pile-fermentation from our company's workshop, through separation, identification, it is defined as mucor, is named as xingding 2, deposit number is CCTCC NO：M 2017241；The preservation time is on May 5th, 2017；Belong to mucor fungi；Its application is fermenting and producing green brick tea.Use the present invention mucor xingding 2 fermentation green brick tea method for：Liquid culture mucor xingding 2, collects the fungal spore, delivers into green brick tea fermented tea heap, and control temperature, humidity are fermented.The method of present invention fermentation green brick tea, compared with traditional green brick tea fermentation process, tea product mouthfeel is substantially similar, and fermentation period is short, greatly improves productivity ratio.
Description
Technical field
The present invention relates to mucoraceous application and its method for fermentation green brick tea, belong to microbial fermentation technology application neck
Domain.
Background technology
Green brick tea belongs to black tea, and black tea fermentation is converted using damp and hot with microorganism collective effect generation material, fermentation week
Phase is long, and qualitative characteristics are protruded.And relative to other black teas, green brick tea fermentation is more abundant, fermentation come out quality mouthfeel more alcohol and,
Chen Xiang is highlighted substantially.
Hubei green brick tea be to shine mao leafiness as raw material, by fermentation, Cheng Hua, steam pressure, drying etc. program be made.Wherein send out
Ferment process plays decisive role to the flavor formation of green brick tea.From forefathers to black teas such as Hunan Anhua black tea, yunnan puer teas
The research of series learns that the process of fermentation has very crucial effect to the quality for changing tea.
Find according to previous studies, in fermentation process, the microorganism of influence tea leaf quality flavor formation mainly has fungi, ferment
Mother, bacterium, actinomyces etc., wherein filamentous fungi are dominant strain, are played an important role during the fermentation.
The content of the invention
Present invention aims at isolating and purifying a kind of Mucor bacterial strain fermented available for green brick tea, and the bacterial strain is applied to
The method of green brick tea fermentation.
The present invention isolates and purifies one plant of bacterial strain from green brick tea fermented tea heap；It is filamentous fungi.Identified, the bacterial strain is one
Plant total shape branch Mucor bacterial strain (Mucor racemosus Fres)；It is named as mucor xingding-2；It is preserved in Chinese Typical Representative
Culture contains center；Deposit number is CCTCC M 2017241；The preservation time is on May 5th, 2017.
The mucor xingding-2 of the present invention can be used in the fermenting and producing of green brick tea.
Use the present invention mucor xingding-2 fermenting and producing green brick teas method for：Liquid culture mucor
Xingding-2, is enriched with its spore and spore suspension is made and counts.The ratio of 100,000 fungal spores is delivered according to solarization Mao Qingcha per jin
Example is inoculated with.
The specific method using Mucor bacteria strain Mucor racemosus Fres xingding-2 fermentation green brick teas is such as
Under：
(1) inclined-plane culture：Mucor xingding-2 is inoculated on slant medium and cultivated, and obtains inclined-plane bacterial strain；
(2) fermented bacterium expands culture：Take inclined-plane inoculation into sterilized seed culture medium, obtain seed culture
Liquid；
(3) tea leaf fermentation：Green gross tea is chosen to sprinkle water to the strain spore that water capacity is harvest in 20-30%, inoculation step (2)
Fermented after son, yeasting is dustless pollution-free constant incubator pile fermentation, it is 25 DEG C to control temperature, and humidity is 90%, hair
After ferment 3 to 4 days, when tea heap temperature is raised to 47 DEG C, turning is carried out, tea heap may proceed to grow white hypha body after turning, and
Rise with temperature, when temperature is raised to 50-55 DEG C, tealeaves color is completely converted into crineous, when giving out ferment perfume, stop；
(4) type is made in tea steaming：By the tealeaves decatize fermented, the tealeaves after decatize is put into shaped device shaping, shaping
Naturally it is cool afterwards to put, it is cool to put the time for 35-45 hours, complete the making of green brick tea loose tea.
In described step (1), mucor xingding-2 is inoculated on slant medium, 48 is cultivated at 28 DEG C small
When, obtain inclined-plane bacterial strain；Slant medium used, be per the composition contained in 1L：Glucose 20.0g, potato 200.0g, agar
15g, distilled water surplus, natural pH, 115 DEG C of sterilizing 20min.
In described step (2), take inclined-plane inoculation into sterilized seed culture medium, 175rpm, 28 DEG C
Under the conditions of isothermal vibration culture 48h, obtain seed culture fluid；Seed culture medium used, be per the composition contained in 1L：Glucose
20.0g, potato 200.0g, distilled water surplus, natural pH, 115 DEG C of sterilizing 20min.
In described step (3), it is 25% that green gross tea, which is sprinkled water to water capacity,；The strain spore harvested in inoculation step 2, is pressed
The ratio for delivering 100,000 fungal spores according to tea per jin is fermented；Yeasting is dustless pollution-free constant incubator pile fermentation, control
Temperature processed is 25 DEG C, and humidity is 90%.
In described step (4), by the tealeaves decatize fermented, steam temperature is 100 DEG C, and steaming time is 3min.
Beneficial effect：
1st, the fungal bacterial strain mucor xingding-2 that can be used in green brick tea fermenting and producing is separately cultured out first；
2nd, the present invention produces green brick tea using fungi fermentation mode, substantially reduces green brick tea fermentation period, improves production
Efficiency；
3rd, the method for the additional specific culture fungal bacterial strain of present invention application, makes fermentation process quickly form specific advantages bacterium
Group, applied to fermenting and producing green brick tea, the mode of production has new meaning.
Preservation explanation
Depositary institution：China typical culture collection center
Address：Wuhan City, Hubei Province Hongshan District Bayi Road, Wuhan University
Preservation date：On May 5th, 2017
Deposit number:CCTCC M 2017241
Classification And Nomenclature：Total shape branch Mucor Mucor racemosus Fres (xingding-2)
Brief description of the drawings
Fig. 1：Mucor xingding-2 of the present invention morphological features under the microscope.
Fig. 2：Mucor xingding-2 of the present invention positive forms on PDA plate.
Fig. 3：The form of mucor xingding-2 reverse side on PDA plate of the present invention.
Fig. 4：Add mucor xingding-2 fermentation green brick tea tea heap samples.
Embodiment
It is method commonly used in the art unless otherwise specified in following embodiments；Agents useful for same lives instrument in indicating production
Manufacturer person, being can be by the conventional products of acquisition purchased in market.
The Mucor bacteria strain xingding-2 of embodiment 1 Morphological Identification；
Bacterial strain xingding-2 is isolated from green brick tea fermented tea heap, and its form of micro- sem observation is as shown in Figure 1.Looking into respectively
Cultivated on the culture matrixes such as family name's agar, Cha Shi yeast agars, wort agar, 28 DEG C of cultivation temperature, incubation time 2-5d, observation
Form.
Cha Shi agar colonies are divided into two for white, mycelium, and vegetative hyphae growth is presented by mediad surrounding relatively advises
Then circular, aerial hyphae is less.Bacterium colony is that white, colonial morphology are relatively regular circle on Cha Shi yeast agars, aerial hyphae compared with
Many, compared with Cha Shi agar, growth is faster.Bacterium colony is white on wort agar, and no aerial hyphae is grown compared with Cha Shi agar
It hurry up, it is slow compared with Cha Shi yeast agars.Reference《Fungal identification handbook》(adding into bibliography) carries out shape to xingding-2 bacterial strains
State is compared, and determines that xingding-2 bacterial strains belong to bacterium for Mucor racemosus.
The bacterial strain xingding-2 Molecular Identifications of embodiment 2：
Strain gene group DNA is extracted, polymerase chain reaction is carried out to the genomic DNA using ITS1 and ITS4 universal primers
(Polymerase Chain Reaction, PCR) is answered to breed specific dna sequence, and utilizes American National Biotechnology Information
Center (National Center for Biotechnology Information, NCBI) database carries out bacterial strain comparison.
(1) 18S rDNA sequence analyses：
ITS1 rDNA primer sequences：5'-TCCGTAGGTGAACCTGCGG-3'
ITS4 rDNA primer sequences：5'-TCCTCCGCTTATTGATATGC-3'
Amplification obtains 18SrDNA sequences, sequence 617bp.By 18SrDNA sequences obtained by the amplification and GeneBank numbers
It is compared according to the gene order of the related strain in storehouse, the bacterium higher with bacterial strain xingding-2 homologys is found from NCBI
Strain is Mucor racemosus f racemosus stain CBS (EU484278), and homology reaches 99%.
(2) LSU rDNA sequence analyses：
NL1 rDNA primer sequences：5'-GCATATCAATAAGCGGAGGAAAAG-3'
NL4 rDNA primer sequences：5'-GGTCCGTGTTTCAAGACGG-3'
Amplification obtains 28SrDNA sequences, sequence 678bp.By 28SrDNA sequences obtained by the amplification and GeneBank numbers
It is compared according to the gene order of the related strain in storehouse, the bacterial strain for finding higher with bacterial strain xingding-2 homologys is
Mucor racemosus f racemosus stain CBS (HM849676.1), homology reaches 99%
Therefore can be determined that xingding-2 is total shape branch Mucor bacterial strain.Sequence has been committed to GeneBank numbers on NCBI
According to storehouse (Serial No. KY373220.1).Mucor xingding-2 was preserved in China typical culture collection center in 2017
(Wuhan University)；Deposit number is CCTCC M 2017241；This biomaterial has been survived experiment test and by the experiment.
Embodiment 3 carries out green brick tea fermenting experiment using Mucor bacterial strain：
(1) inclined-plane culture：Mucor xingding-2 is inoculated on slant medium, is cultivated 48 hours, is obtained at 28 DEG C
Inclined-plane bacterial strain.Slant medium used, be per the composition contained in 1L：Glucose 20.0g, potato 200.0g, agar 15g,
Distilled water surplus, natural pH, 115 DEG C of sterilizing 20min.
(2) fermented bacterium expands culture：Inclined-plane inoculation is taken into sterilized seed culture medium, in 175rpm, 28
Isothermal vibration culture 48h, obtains seed culture fluid under conditions of DEG C.Seed culture medium used, be per the composition contained in 1L：Portugal
Grape sugar 20.0g, potato 200.0g, distilled water surplus, natural pH, 115 DEG C of sterilizing 20min.
(3) tea leaf fermentation：It is 25% to choose the green gross tea of three-level and sprinkle water to water capacity.The strain spore harvested in inoculation step 2
Son, the ratio for delivering 100,000 fungal spores according to tea per jin is fermented.Yeasting is that dustless pollution-free constant incubator is wet
Heap, it is 25 DEG C to control temperature, and humidity is 90%.After fermentation 3 to 4 days, fermented fungal grows in tea heap surface colonization, tealeaves leaf
Visible white mycelium on piece, simultaneous tea heap temperature rises, when tea heap temperature is raised to 47 DEG C, carries out turning, turning
Tea heap may proceed to grow white hypha body afterwards, and rise with temperature, treat that temperature is raised to 53 DEG C or so, tealeaves color is totally converted
Into crineous, when giving out ferment perfume, stop.
(4) type is made in tea steaming：By the tealeaves decatize fermented, steam temperature is 100 DEG C, and steaming time is 3min, is obtained
Tealeaves after to decatize is put into shaped device shaping, naturally cool after shaping to put, cool to put the time for 40 hours, completes green brick tea loose tea
Making.
The fermentation green brick tea of embodiment 4 is evaluated：
Reference is evaluated to fermented tea《Tealeaves organoleptic evaluation method》, to this in terms of profile, soup look, fragrance, flavour
The green brick tea that the tealeaves of experiment fermenting and producing is produced with traditional mode of production mode is contrasted, as a result such as table 1.As a result show with this
Too big difference is had no in the tea of method fermenting and producing and the green brick tea quality of produced in conventional processes, illustrates that this method can be used for green grass or young crops
The production of brick tea, and this method used time is shorter, production efficiency is higher.
The present invention of table 1 production green brick tea evaluates contrast with traditional green brick tea
Claims (8)
1. one plant of Mucor bacteria strainMucor racemosusFresXingding-2, Chinese allusion quotation is preserved on May 5th, 2017
Type culture collection, deposit number is CCTCC NO：M 2017241, preservation address is Hubei, Wuhan, Wuhan University.
2. the mucor strain isolation described in claim 1 is in pile-fermentation mid-term from green brick tea production pile fermentation technique
Pile fermentation, the tealeaves for having hypha,hyphae hair tonic.
3. the Mucor bacteria strain described in claim 1 or 2Mucor racemosusFresXingding-2 is on fermentation green brick tea
Application.
4. the application described in claim 3, it is characterised in that comprise the following steps：
（1）Inclined-plane culture：Mucor xingding-2 is inoculated on slant medium and cultivated, and obtains inclined-plane bacterial strain；
（2）Fermented bacterium expands culture：Take inclined-plane inoculation into sterilized seed culture medium, obtain seed culture fluid；
（3）Tea leaf fermentation：It is 20-30%, inoculation step to choose green gross tea and sprinkle water to water capacity（2）After the strain spore of middle harvest
Fermented, yeasting is dustless pollution-free constant incubator pile fermentation, it is 25 DEG C to control temperature, and humidity is 90%, fermentation 3 to 4
After it, when tea heap temperature is raised to 47 DEG C, turning is carried out, tea heap may proceed to grow white hypha body after turning, and with temperature
Degree rises, and when temperature is raised to 50-55 DEG C, tealeaves color is completely converted into crineous, when giving out ferment perfume, stops；
（4）Type is made in tea steaming：By the tealeaves decatize fermented, the tealeaves after decatize is put into shaped device shaping, after shaping from
It is so cool to put, it is cool to put the time for 35-45 hours, complete the making of green brick tea loose tea.
5. the application described in claim 4, it is characterised in that step（1）In, mucor xingding-2 is inoculated in inclined-plane culture
On base, cultivated 48 hours at 28 DEG C, obtain inclined-plane bacterial strain；Slant medium used, be per the composition contained in 1L：Glucose 20
.0g, potato 200.0g, agar 15g, distilled water surplus, natural pH, 115 DEG C of 20 min of sterilizing.
6. the application described in claim 4, it is characterised in that step（2）In, take inclined-plane inoculation to be trained to sterilized seed
Support in base, isothermal vibration culture 48h, obtains seed culture fluid under conditions of 175 rpm, 28 DEG C；Seed culture medium used, often
The composition contained in 1L is：The .0g of glucose 20, potato 200.0g, distilled water surplus, natural pH, 115 DEG C of sterilizing 20min.
7. the application described in claim 4, it is characterised in that step（3）In, it is 25% that green gross tea, which is sprinkled water to water capacity,；Inoculation step
The strain spore harvested in rapid 2, the ratio for delivering 100,000 fungal spores according to tea per jin is fermented；Yeasting is dustless nothing
Constant incubator pile fermentation is polluted, it is 25 DEG C to control temperature, and humidity is 90%.
8. the application described in claim 4, it is characterised in that step（4）In, by the tealeaves decatize fermented, steam temperature is
100 DEG C, steaming time is 3min.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107647010A (en) * | 2017-10-23 | 2018-02-02 | 鑫鼎生物科技有限公司 | A kind of method of mixed fermentation with various bacterium green brick tea |
| CN108272909A (en) * | 2018-01-31 | 2018-07-13 | 三峡大学 | A kind of preparation method and application of tea leaf fermentation product |
| CN109055273A (en) * | 2018-09-10 | 2018-12-21 | 安徽农业大学 | A kind of green brick tea pile-fermentation strain composition and application |
| CN109136100A (en) * | 2018-08-21 | 2019-01-04 | 鑫鼎生物科技有限公司 | One plant of Tabin aspergillus bacterial strain and the application on fermentation green brick tea |
| CN109287784A (en) * | 2018-09-10 | 2019-02-01 | 安徽农业大学 | A kind of method of the quick pile-fermentation of green brick tea |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105875884A (en) * | 2016-04-28 | 2016-08-24 | 王婧 | Novel green brick tea manufacturing process |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105875884A (en) * | 2016-04-28 | 2016-08-24 | 王婧 | Novel green brick tea manufacturing process |
Non-Patent Citations (2)
| Title |
|---|
| GENEBANK: "Mucor racemosus isolate LC-2 internal transcribed spacer 1 , partial sequence", 《GENEBANK》 * |
| 胥伟: "四川康砖茶渥堆过程中真菌种群的鉴定", 《中国优秀硕士学位论文全文数据库 工程科技Ⅰ辑》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107647010A (en) * | 2017-10-23 | 2018-02-02 | 鑫鼎生物科技有限公司 | A kind of method of mixed fermentation with various bacterium green brick tea |
| CN108272909A (en) * | 2018-01-31 | 2018-07-13 | 三峡大学 | A kind of preparation method and application of tea leaf fermentation product |
| CN109136100A (en) * | 2018-08-21 | 2019-01-04 | 鑫鼎生物科技有限公司 | One plant of Tabin aspergillus bacterial strain and the application on fermentation green brick tea |
| CN109136100B (en) * | 2018-08-21 | 2021-11-09 | 鑫鼎生物科技有限公司 | Aspergillus tubingensis strain and application thereof in fermenting green brick tea |
| CN109055273A (en) * | 2018-09-10 | 2018-12-21 | 安徽农业大学 | A kind of green brick tea pile-fermentation strain composition and application |
| CN109287784A (en) * | 2018-09-10 | 2019-02-01 | 安徽农业大学 | A kind of method of the quick pile-fermentation of green brick tea |
| CN109055273B (en) * | 2018-09-10 | 2022-03-04 | 安徽农业大学 | Green brick tea pile fermentation strain composition and application thereof |
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