CN104388477B - Utilize the method for one plant of safe strain fermentation vinasse production 2,3- butanediol - Google Patents
Utilize the method for one plant of safe strain fermentation vinasse production 2,3- butanediol Download PDFInfo
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- CN104388477B CN104388477B CN201410748634.5A CN201410748634A CN104388477B CN 104388477 B CN104388477 B CN 104388477B CN 201410748634 A CN201410748634 A CN 201410748634A CN 104388477 B CN104388477 B CN 104388477B
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- C—CHEMISTRY; METALLURGY
- 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
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/18—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic polyhydric
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Abstract
Present invention firstly discloses one plant of Environmental security bacterial strain Bacillus amyloliquefaciens CCTCC M2012349 is utilized, using vinasse as unique carbon nitrogen source, the method for 2,3 butanediol of fermenting and producing belongs to fermentation technical field in bioengineering.This method mainly includes seed culture and fermented and cultured.Specifically, seed liquor is inoculated in by 4% inoculum concentration in fermentation medium, at 37 DEG C, air mass flow 120m3/h·m3Culture medium, speed of agitator carry out fermented and cultured under the conditions of being 350r/min, and ferment 60h, stop fermentation at once, and zymotic fluid is to contain 2,3 butanediol liquid.Liquor production cost, raising resource utilization, extension industry chain can be effectively reduced, is the important measures for establishing efficient, economic biomass energy comprehensive utilization industry, the overall technology level and circulating benefit of liquor industry will be greatly promoted.
Description
Technical field
Using one plant of safe strains B. amyloliquefaciens fermentation vinasse production 2,3-butanediol, belong in bioengineering and send out
Ferment technical field.
Background technology
With the worsening shortages of energy resources and the getting worse of environmental problem, reproducible biomass resource is developed as original
Material is got over by the biorefinery technology of chemistry, biology and its integrated approach production various chemicals, functional material and energy substance
To get over by great attention both domestic and external.In recent years, white wine annual output in China's was about 8,660,000 tons, and the annual output of vinasse is about
98660000 tons.Solid spirit produces vinasse as the Main By product during brewed spirit, while is also likely to be that it is main dirty
Contaminate object.Fully and effectively vinasse are comprehensively utilized, can not only mitigate environmental pollution, but also can save food, reduce production
Cost.On the one hand due to being used as primary raw material using sorghum, wheat, glutinous rice etc. so that vinasse containing protein, cellulose, fat,
Vitamin and various trace elements etc. create favorable conditions for its comprehensive utilization；Another aspect is special due to brewage process
Property, cause to lose poor acidity height and easily go mouldy, it is necessary to which otherwise timely processing would seriously pollute the environment, how to carry out depth to vinasse
Exploitation, it has also become the key of liquor industry sustainable development and guarantee, while be also the critical issue that can not avoid.It if can
Using the starchy material exploitation high value added product (such as 1,3-PD, ethylene glycol, 2,3-butanediol) in vinasse, with regard to energy
It is effectively reduced liquor production cost, improves resource utilization, extension industry chain, be the biomass energy established efficiently, economic
The important measures of comprehensive utilization industry will greatly promote the overall technology level and circulating benefit of liquor industry.
2,3-butanediol is important industrial chemicals and liquid fuel, is widely used in chemical industry, food, fuel and aviation boat
The fields such as it.2,3-butanediol dehydrogenation generates biacetyl, is a kind of high value flavour of food products agent, has certain fungistatic effect；
Under the action of dehydrogenase, the 3- hydroxy-2-butanones (3-hydroxy-2-butanone) of generation are a kind of widely used natural flavourings；Dehydration
The methyl ethyl ketone of generation can also be important low boiling point solvent as a kind of liquid fuel additive of high value, applied to coating,
The industries such as binding agent, lubricant, fuel, ink；The 1,3- butadiene of dehydration generation can be used for synthetic rubber, ABS resin and SBS
Elastomer etc.；Esterification products are the precursors of synthesizing polyester and polyurethane.And 2,3- butanediols are a kind of valuable liquid
Fuel, fuel value are suitable with methanol (22.1kJ/g), ethyl alcohol (29.0kJ/g).Due to the high-octane rating of 2,3-butanediol, it
Can as gasoline octane number elevator or aviation fuel in addition, 2,3-butanediol and its derivative also can be widely used for medicinal load
The production of body, plasticizer, softening agent etc..
In recent years preferable results of laboratory, but bioanalysis are achieved with saccharine material fermenting and producing 2,3-butanediol
Production 2,3- butanediols not yet realize industrialization.However application prospect, production technology maturity, knowledge production from 2,3- butanediols
Power situation and future developing trend are seen, are developed using cheap non-grain saccharic as raw material (such as vinasse, biodiesel) fermenting and producing
2,3- butanediols have good development prospect.The 2,3- butanediols production bacterial strain reported at present is mostly Klebsiella
(Klebsiella) Enterobacter (Enterobacter) and Serratia (Serratia) etc., but these bacterium are mostly with sugar
Matter fermenting raw materials produce 2,3-butanediol, and produce 2,3-butanediol for raw material using vinasse at home and have not been reported；In addition,
These production strains have it is potential pathogenic, do not meet meet industrialization safety in production requirement.Therefore, strain profit safe to use
There is good industrial prospect with glycerol fermentation 2,3- butanediols.The present invention utilizes one for the first time using vinasse as unique carbon nitrogen source
The safe bacterial strain Bacillus amyloliquefaciens CCTCC M2012349 fermenting and producing 2,3-butanediols of strain, for wine brewing
Industry waste material carries out industrialization deep development and provides the foundation.
Invention content
The object of the present invention is to provide one kind using vinasse as unique carbon nitrogen source, one plant of safe bacterial strain Bacillus is utilized
The method of amyloliquefaciens CCTCC M2012349 fermentation vinasse production 2,3- butanediols.
Realization process of the present invention is as follows:
(bacterial strain is for Bacillus amyloliquefaciens CCTCC M2012349 for strain used herein
Disclosed in the patent applied early period in applicant, related application number：201310342415.2；It has been preserved in Chinese Typical Representative training
Object collection is supported, deposit number is：CCTCC M2012349).Recently, we found that B.amyloliquefaciens CCTCC
M2012349 can efficiently use vinasse and efficiently synthesize 2,3-butanediol for unique carbon nitrogen source, and specific production technology is as follows：
(1) the B.amyloliquefaciens CCTCC M2012349 single bacterium colonies of preservation are inoculated in seed on picking tablet
It is cultivated in culture medium, preferred incubation time is 12h, and cultivation temperature is 37 DEG C, and seed culture medium composition is as follows：
Starch 10g/L, yeast extract 5g/L, tryptone 10g/L, NaCl10g/L；
Seed culture fluid uses 250mL shaking flask cultures, and seed liquor is cultivated 12h or so, cultivation temperature 37 on shaking table
DEG C, shaking speed 160r/min.
(2) B.amyloliquefaciens CCTCC M2012349 fermented and cultureds
Fermentation medium composition is as follows：
Vinasse 300g/L；Above-mentioned fermentation medium is adjusted into its pH to 6.5 with the NaOH of 5mol/L, the high temperature at 121 DEG C
Sterilize 30min.Installation for fermenting be 5L fermentation tanks, initial liquid amount be 2.5L, inoculum concentration 4%.Fermentation condition is：Air mass flow
For 120m3/h·m3Culture medium, speed of agitator 350r/min, fermentation time 60h.After fermentation, zymotic fluid is through centrifugation etc.
After processing, 2,3-butanediol content in the i.e. detectable zymotic fluid of the fid detector of gas-chromatography is utilized.
Beneficial effects of the present invention：
The present invention utilizes Xie Dian shallow lakes bacillus B.amyloliquefaciens CCTCC M2012349 fermented wines for the first time
Grain efficiently synthesizes biology base platform chemicals 2,3-butanediol, and 2,3-butanediol yield reaches 50.6g/L, is prior art institute
It is inaccessiable, achieve unexpected technique effect；In addition, its fermentation medium is sole component only with vinasse, send out
In ferment culture medium without additionally add yeast extract, peptone, corn pulp, the nitrogen sources such as dregs of beans and phosphorus source, amino acid, vitamin and
Other nutriments such as trace element, so as to reduce production cost.So there are higher industrial applications to be worth for the invention.
In addition, the 2,3-butanediol production bacterial strain reported at present is mostly Klebsiella (Klebsiella) Enterobacter
(Enterobacter) and Serratia (Serratia) etc., these strains are mostly with saccharine material fermenting and producing 2,3- fourths
Glycol；In addition, these strains have it is potential pathogenic, do not meet meet industrialization safety in production requirement, solve starch gemma bar
Pseudomonas disclosure satisfy that requirement of the industrialization to safety in production in Environmental security bacterial strain.In addition, the present invention utilizes the pair of brewed spirit
Product vinasse waste material develops high value added product 2,3-butanediol, with regard to that can be effectively reduced liquor production cost, improve resource profit
With rate, extension industry chain, it is the important measures for establishing efficient, economic biomass energy comprehensive utilization industry, will greatly promotes
The overall technology level and circulating benefit of liquor industry.
Specific embodiment
Embodiment 1
(1) B.amyloliquefaciens CCTCC M2012349 seed cultures
It is inoculated in seed culture medium from picking single bacterium colony on activation tablet and carries out seed culture, 37 DEG C of cultivation temperature is shaken
Bed rotating speed 160r/min, incubation time are 12h or so, and seed culture medium forms：Starch 10g/L, yeast extract 5g/L, pancreas egg
White peptone 10g/L, NaCl10g/L；
(2) B.amyloliquefaciens CCTCC M2012349 fermented and cultureds
Preliminary fermentation volume of culture is 2.5L, and the fermentation medium components of use are as follows：
Vinasse 300g/L；Above-mentioned fermentation medium is adjusted into its pH to 6.5 with the NaOH of 5mol/L, the high temperature at 121 DEG C
Sterilize 30min.
Above-mentioned cultured seed liquor is inoculated in by 4% inoculum concentration in fermentation medium and carries out fermented and cultured, fermentation temperature
37 DEG C of degree, air mass flow 120m3/h·m3Culture medium, speed of agitator 350r/min.Timing sampling measures cell concentration, bottom
Object residual quantity and 2,3-butanediol output, fermented and cultured 60h stop fermentation at once.After fermentation, 2,3-butanediol yield
Reach 50.6g/L.
Claims (1)
1. utilize the method for one plant of safe strain fermentation vinasse production 2,3-butanediol, it is characterised in that using vinasse as sole substrate
And nutrient source, utilize one plant of safe strains B. amyloliquefaciens Bacillus amyloliquefaciens CCTCC
M2012349 ferments, and it produces 2,3- butanediols；Wherein fermentation medium is made of following component：Vinasse 300g/L, surplus are
Water, pH 6.5；
Specific production method is as follows：
(1) B.amyloliquefaciens CCTCC M2012349 seed cultures
The B.amyloliquefaciens CCTCC M2012349 single bacterium colonies of preservation on picking tablet are inoculated in seed culture medium
Middle culture 12h, obtains seed culture fluid, and seed culture medium composition is as follows：
Starch 10g/L, yeast extract 5g/L, tryptone 10g/L, NaCl 10g/L, surplus is water；
(2) B.amyloliquefaciens CCTCC M2012349 fermented and cultureds
The fermentation medium adjusts its pH to 6.5 with the NaOH of 5mol/L, the high-temperature sterilization 30min at 121 DEG C；It will be above-mentioned
Seed culture fluid is inoculated in by 4% inoculum concentration in fermentation tank, 37 DEG C of fermentation temperature, air mass flow 120m3/(h·m3) culture
Base, speed of agitator 350r/min, fermented and cultured 60h stop fermentation at once；Obtain the fermented and cultured containing 2,3- butanediols
Liquid.
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| CN106636238A (en) * | 2016-11-17 | 2017-05-10 | 江南大学 | Method for producing gamma-amino butyric acid through distillers' grains fermentation by microorganisms |
| CN111154812B (en) * | 2020-02-26 | 2022-08-26 | 北京联合大学 | Method for producing 2, 3-butanediol by fermenting kitchen waste serving as substrate |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103314101A (en) * | 2010-11-03 | 2013-09-18 | 加利福尼亚大学董事会 | Biofuel and chemical production by recombinant microorganisms via fermentation of proteinacious biomass |
| CN103525877A (en) * | 2013-08-08 | 2014-01-22 | 江南大学 | Method for selectively producing 3-hydroxyl-2-butanone and 2, 3-butanediol through microbial fermentation |
| CN103602624A (en) * | 2013-08-08 | 2014-02-26 | 江南大学 | Method for increasing 2,3-butylene glycol output of Bacillus amyloliquefaciens by strengthening coenzyme cyclic regeneration speed |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103314101A (en) * | 2010-11-03 | 2013-09-18 | 加利福尼亚大学董事会 | Biofuel and chemical production by recombinant microorganisms via fermentation of proteinacious biomass |
| CN103525877A (en) * | 2013-08-08 | 2014-01-22 | 江南大学 | Method for selectively producing 3-hydroxyl-2-butanone and 2, 3-butanediol through microbial fermentation |
| CN103602624A (en) * | 2013-08-08 | 2014-02-26 | 江南大学 | Method for increasing 2,3-butylene glycol output of Bacillus amyloliquefaciens by strengthening coenzyme cyclic regeneration speed |
Non-Patent Citations (2)
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| pH 与溶氧控制对解淀粉芽胞杆菌发酵粗甘油生产 2,3-丁二醇的影响;杨套伟;《生物工程学报》;20131231;第29卷(第13期);1860-1864 * |
| Screening of bacterial strains capable of converting biodiesel-derived raw glycerol into 1,3-propanedio l,2,3-butanediolandethanol;Metsoviti,M.et al.;《Eng.life sci.》;20120229;第12卷(第1期);全文 * |
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