CN1173041C - Production of alcohol by fermenting by yeast tolerant to high concentrated sugar and alcohol - Google Patents
Production of alcohol by fermenting by yeast tolerant to high concentrated sugar and alcohol Download PDFInfo
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- CN1173041C CN1173041C CNB031179312A CN03117931A CN1173041C CN 1173041 C CN1173041 C CN 1173041C CN B031179312 A CNB031179312 A CN B031179312A CN 03117931 A CN03117931 A CN 03117931A CN 1173041 C CN1173041 C CN 1173041C
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Abstract
The present invention relates to an alcohol fermentation production method by saccharomyces cerevisiae with high concentration sugar resistance and high concentration alcohol resistance, which belongs to the technical field of industrial microorganism fermentation engineering. The method of the present invention is carried out according to the existing conventional alcohol fermentation production method. In the method of the present invention, saccharomyces cerevisiae 1912 with high concentration alcohol resistance and high concentration sugar resistance is used for producing the alcohol through fermentation in two stages; when each milliliter of a culture solution has 10<8> saccharomyces cerevisiae in the first stage, the second stage of the alcohol fermentation is carried out, or strains are manufactured into immobilized carriers, and then, the two stages of alcohol fermentation production is carried out, wherein in the first stage, sterile air is introduced continuously or intermittently at every one hour for 10 to 30 minutes; in the second stage, the sugar initial concentration weight ratio is between 15 and 30%, and the sterile air is introduced at every two hours for 3 to 20 minutes. The present invention has the advantages that the implementation is convenient, the alcohol content in a fermentation solution in the alcohol fermentation process is improved, and the sugar to alcohol conversion rate and the final alcohol yield are improved, so the production cost for alcohol fermentation production is reduced.
Description
Technical field:
The present invention relates to a kind of method of utilizing the saccharomycetes to make fermentation production alcohol of resisting high-concentration sugar and alcohol, belong to industrial microorganism fermentation engineering field.
Background technology:
It is indifferent that present widely used alcohol industry fermented bacterium exists osmophilic strain power, shortcomings such as anti-high alcohol is indifferent, make the initial sugared concentration of zymamsis on the low side, assorted bacterium is growth easily therefore, the raised growth of assorted bacterium has reduced sugared utilization ratio, contained zymic growth and fermentation, the alcohol transformation efficiency is greatly reduced.Simultaneously, carrying out along with fermentation, ethanol concn constantly rises, produced the containment effect of substrate at last, cause final ethanol concn on the low side, the change between 7-10% (back together) of general volume percent, though adopt immobilization technology to be stabilized in final ethanol concn about 8-9%, the containment problem of microbiological contamination and substrate does not still solve.Its major cause is the single yeast that traditional technology of alcohol adopts, in the fermentation whole process, along with the sugar degree of fermented liquid constantly reduces, ethanol concn constantly rises, and is difficult to remain the ability of best fermentative production alcohol.
Summary of the invention:
The objective of the invention is to overcome the deficiency of existing biomass to alcohol conversion process, and a kind of method of utilizing the saccharomycetes to make fermentation production alcohol of resisting high-concentration sugar and alcohol is provided.The present invention passes through strain selection, obtain a strain alcohol in high concentration is had yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) 1912 bacterial strains (preserving number: CGMCCNo.0806 that tolerance can tolerate high concentration sugar again, the preservation on October 9 in 2002 of China Committee for Culture Collection of Microorganisms common micro-organisms center), make it under the prerequisite of utilizing existing zymamsis production unit, produce alcohol with the saccharomycetes to make fermentation of resisting high-concentration sugar and alcohol.
The present invention is undertaken by existing conventional single-stage or the multistage method of producing alcohol of continuously fermenting, wherein:
This method adopts through molecular biology method, make alcohol dehydrogenase gene disappearance or do not express and the Saccharomyces Cerevisiae in S accharomyces cerevisiae dissociant 1912 that obtains, this bacterial strain is for can tolerate high concentration sugar and alcohol in high concentration bacterial strain, its alcohol tolerance level volume percent is 20%, sugar can the tolerance level weight percent be 35%, Saccharomyces Cerevisiae in S accharomycescerevisiae1912 bacterial strain, be deposited in China Committee for Culture Collection of Microorganisms common micro-organisms center, preserving number: CGMCCNo.0806 on October 9th, 2002;
This method fed sterile air 10-30 minute in 1 hour at continuous or every interval of the seed culture stage of zymamsis, kept the gap to stir or be interrupted in the zymamsis stage and fed sterile air, promptly every interval ventilation in 2 hours 3-20 minute;
This method nutrient solution that to add sugared concentration weight percent be 15-30% in the zymamsis stage.
The present invention is achieved in that
1912 bacterial strains by microbial fermentation conventional liq seed culture method, cultivate with malt extract medium (get brew-house with wort 10 mother-in-law U.S. degree), when containing yeast count, every milliliter of nutrient solution are reached 1 * 10
8When individual, put into fermentor tank by the amount of fermentor cultivation liquid 1/10-1/15 and carry out the fs yeast culture, every milliliter of nutrient solution contains yeast count and reaches 10 in fermentor tank
8When individual, carry out the fermentation of subordinate phase Alcohol Production; Also this yeast bacterial strain method routinely can be fixed on the carrier, carry out two stage fermentations again and produce alcohol.Wherein: the fs is called the seed culture stage, and sugared starting weight percentage concentration is 6-14%, and all stage is continuous or be interrupted the feeding sterile air, ventilates 10-30 minute in promptly every interval in 1 hour; Subordinate phase is called fermentation stage, and sugared starting weight percentage concentration is 15-30%, and all stage keeps the gap to stir or is interrupted the feeding sterile air, ventilates 3-20 minute in promptly every interval in 2 hours.
Two stages utilizing this yeast bacterial strain to carry out fermentative production alcohol among the present invention can carry out also can carrying out in a plurality of fermentor tanks in a fermentor tank.
In the zymamsis whole process of the present invention, the pH value is controlled at 3.8-5.0, temperature, and to be controlled at 26 ℃-36 ℃, used time of fermentation be 48 hours; When its fermentative production alcohol whole process was finished, the alcohol concentration of volume percent was 11-16% in the fermented liquid.
Yeast bacterial strain 1912 of the present invention possesses following feature:
(1) basic physiological feature: cell circle, oval or foreign pyriform.In the childhood bacterium colony, cell is 4～14 * 3～7 microns, and long and wide ratio is 1: 1～2: 1; In wort, precipitate.The thecaspore circle, level and smooth.
(2) yeast bacterial strain 1912 is S. cervisiaes (Saccharomyces cerevisiae) by molecular biology method, makes the alcohol dehydrogenase gene disappearance or does not express and the dissociant that obtains, has the advantage that can tolerate high concentration sugar and alcohol in high concentration.The yeast strain alcohol dehydrogenase gene lacks or does not express and can reach by physics, chemistry or molecular biological method, also can be by physics or chemomorphosis and the acquisition of molecular biological method.
(3) yeast bacterial strain 1912 is compared with employed other yeast saccharomyces cerevisiae bacteria strain of Alcohol Production and former starting strain, can tolerate the alcohol and the sugar of higher concentration.The sugar weight percentage concentration of its tolerance is up to 35%; The alcohol concentration of volume percent is up to 20%, and has the ability of good fermentative production alcohol with this understanding.
The making method of fixation support: the first step: thalline preparation.By liquid asepsis cultured method (by " industrial microbiology laboratory manual ", Chemical Industry Press), obtaining 10 milliliters of biomasses is that every milliliter of nutrient solution contains 10
8The bacterium liquid of individual thalline (method of counting is by " microbiology experiment ", Higher Education Publishing House).Culture condition: 30 ℃, natural pH value keeps feeding sterile air.Substratum adopts malt extract medium (getting brew-house's wort 10 mother-in-law U.S. degree).Second step added 80 gram water with 10 gram polyvinyl alcohol, and heating is dissolved, and kept volume 90 milliliters (insufficient section adds water and supplies), to be cooled during to 50-55 ℃, add 8 gram diatomite, add the bacterium liquid for preparing in advance, mix, put into container, place-4 ℃ of refrigerator overnight (24 hours).The 3rd step, the fixedly thalline in the refrigerator is taken out, break away from container, dried 4 hours, and became finished product for 50 ℃.
The present invention has and implements conveniently, improves the ethanol content of fermented liquid in the molasses alcohol fermentation process, transformation efficiency that sugar is converted into alcohol and the productive rate of final alcohol, thereby reduces the production cost of fermentative production alcohol.
Description of drawings:
Accompanying drawing is multistage serialization zymamsis production unit schema
Embodiment:
Single-stage fermentative production alcohol
Embodiment one:
1912 bacterial strains by microbial fermentation conventional liq seed culture method, cultivate with malt extract medium (get brew-house with wort 10 mother-in-law U.S. degree), after shaking a series of cultivations such as bottle, seeding tank, when containing yeast count, every milliliter of nutrient solution are reached 1 * 10
8When individual, by the amount input 90M of fermentor cultivation liquid 1/15
3Cultivate in the fermentor tank, wherein contain 10-15M
3Sugar concentration is 10% sterile medium.Temperature is controlled at 28 ℃, and the pH value is controlled at 4.5, keeps continuing or be interrupted feeding sterile air promptly every interval ventilation in 1 hour 10 minutes.Check yeast biomass in the nutrient solution, reach every milliliter of nutrient solution and contain yeast 10
8When individual, add the nutrient solution that contains high concentration sugar, make the sugar weight percentage concentration of whole nutrient solution reach 15%, temperature is controlled at 28 ℃, and the pH value is controlled at 4.2, keeps the gap to stir or per 2 hours logical sterile airs 4 minutes, cultivated 48 hours, final alcohol concentration of volume percent reaches 11%.
Embodiment two:
Concrete operations are with embodiment one, and difference: ventilated 20 minutes in every interval of seed culture stage in 1 hour, when nutrient solution contains yeast 10
8When individual, the nutrient solution sugar weight percentage concentration of adding is 23%, keeps the gap to stir or per 2 hours logical sterile airs 12 minutes, and final alcohol concentration of volume percent reaches 13%.
Embodiment three:
Concrete operations are with embodiment one, and difference: ventilated 30 minutes in every interval of seed culture stage in 1 hour, when nutrient solution contains yeast 10
8When individual, the nutrient solution sugar weight percentage concentration of adding is 30%, keeps the gap to stir or per 2 hours logical sterile airs 20 minutes, and final alcohol concentration of volume percent reaches 14%.
Yuanjiang River sugar refinery, table 1. Yunnan Province 1912 bacterial strain single batch fermentation results
| Project | The former bacterial strain technology in Yuanjiang River sugar refinery | 1912 bacterial strains are single jar of technology |
| Average yeast number (hundred million) | 1.17 | 1.4 |
| The smart content (%) of ripe raw spirit | 7.98 | 13 |
| Ton alcohol consumption molasses amounts (ton) | 4.39 | 3.8 |
Embodiment four:
The Production Flow Chart of by specification accompanying drawing is made fixation support with 1912 bacterial strains and is put into 90M
3Cultivate in the fermentor tank (seed culture jar or first fermentor tank), wherein contain 10-15M
3The sugar weight percentage concentration is 10% nutrient solution.Temperature is controlled at 30 ℃, and the pH value is controlled at 4.5, keeps continuing or be interrupted feeding sterile air promptly every interval ventilation in 1 hour 10 minutes.Check yeast biomass in the nutrient solution, reach every milliliter of nutrient solution and contain yeast 10
8When individual, continuous adding sugar weight percentage concentration is 10% nutrient solution in seeding tank, by flowing naturally of liquid, when the liquid excessively of seeding tank was arranged in second fermentor tank, continuous adding sugar weight percentage concentration was 15% nutrient solution in second fermentor tank.Temperature is controlled at 30 ℃, and the pH value is controlled at 4.2, keeps the gap to stir or per 2 hours logical sterile airs 4 minutes, cultivates 48 hours, and finally the alcohol concentration of volume percent reaches 11%.
Wherein, seed culture tank volume (first fermentor tank) is 1 with fermentor tank (second fermentor tank) volume ratio: 4-10.
Embodiment five:
Concrete operations are with embodiment four, and difference: ventilated 20 minutes in the every interval of seeding tank in 1 hour, when nutrient solution contains yeast 10
8When individual, the nutrient solution sugar weight percentage concentration that second fermentor tank adds is 23%, keeps the gap to stir or per 2 hours logical sterile airs 12 minutes, and final alcohol concentration of volume percent reaches 13%.
Embodiment six:
Concrete operations are with embodiment four, and difference: ventilated 30 minutes in the every interval of seeding tank in 1 hour, when nutrient solution contains yeast 10
8When individual, the nutrient solution sugar weight percentage concentration that second fermentor tank adds is 30%, keeps the gap to stir or per 2 hours logical sterile airs 20 minutes, and final alcohol concentration of volume percent reaches 14%.
Yuanjiang River sugar refinery, table 2. Yunnan Province 1912 bacterial strain multiple tanks fermentation result
| Project | The former bacterial strain technology in Yuanjiang River sugar refinery | 1912 bacterial strain technologies |
| Average yeast number (hundred million) | 1.17 | 1.5 |
| The smart content (%) of ripe raw spirit | 7.98 | 11 |
| Ton alcohol consumption molasses amounts (ton) | 4.39 | 3.8 |
Brave sugar refinery forever, table 3. Yunnan Province 1912 bacterial strain multiple tanks fermentation result
| Project | The former bacterial strain technology in brave sugar refinery forever | 1912 bacterial strain technologies |
| Average yeast number (hundred million) | 1.2 | 1.4 |
| The smart content (%) of ripe raw spirit | 10.6 | 13 |
| Ton alcohol consumption molasses amounts (ton) | 4.04 | 3.8 |
Claims (1)
1. a method of utilizing the saccharomycetes to make fermentation production alcohol of resisting high-concentration sugar and alcohol is undertaken by existing conventional single-stage or the multistage method of producing alcohol of continuously fermenting, and it is characterized in that:
1.1 this method adopts through molecular biology method, make alcohol dehydrogenase gene disappearance or do not express and the Saccharomyces Cerevisiae in S accharomyces cerevisiae dissociant 1912 that obtains, this bacterial strain is for can tolerate high concentration sugar and alcohol in high concentration bacterial strain, its alcohol tolerance level volume percent is 20%, sugar can the tolerance level weight percent be 35%, Saccharomyces Cerevisiae in S accharomyces cerevisiae1912 bacterial strain, be deposited in China Committee for Culture Collection of Microorganisms common micro-organisms center, preserving number: CGMCCNo.0806 on October 9th, 2002;
1.2 this method fed sterile air 10-30 minute in 1 hour at continuous or every interval of the seed culture stage of zymamsis, kept the gap to stir or be interrupted in the zymamsis stage and fed sterile air, promptly every interval ventilation in 2 hours 3-20 minute;
1.3 this method nutrient solution that to add sugared concentration weight percent be 15-30% in the zymamsis stage.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CNB031179312A CN1173041C (en) | 2003-05-21 | 2003-05-21 | Production of alcohol by fermenting by yeast tolerant to high concentrated sugar and alcohol |
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| Application Number | Priority Date | Filing Date | Title |
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| CNB031179312A CN1173041C (en) | 2003-05-21 | 2003-05-21 | Production of alcohol by fermenting by yeast tolerant to high concentrated sugar and alcohol |
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| CN1456673A CN1456673A (en) | 2003-11-19 |
| CN1173041C true CN1173041C (en) | 2004-10-27 |
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Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100379858C (en) * | 2005-10-26 | 2008-04-09 | 云南师范大学 | Immobilized microorganism for biomass fermentation and hydrogen production and preparation method thereof |
| CN100342022C (en) * | 2005-12-20 | 2007-10-10 | 云南大学 | Method for improving alcohol yield fermented from starch material |
| CN101177695B (en) * | 2006-11-06 | 2012-05-23 | 中国科学院成都生物研究所 | High-concentration alcoholic fermentation method |
| CN1986820B (en) * | 2007-01-25 | 2010-12-01 | 黄荣胜 | Alcohol output increasing method and device |
| CN101633896B (en) * | 2009-08-27 | 2011-01-05 | 中国科学院微生物研究所 | Saccharmyces cerevisiae strain for resisting high-concentration acetic acid and application thereof |
| CN102041235B (en) * | 2009-10-21 | 2012-11-21 | 中国石油化工股份有限公司 | High-temperature resistant saccharomyces cerevisiae and application thereof |
| CN102533842A (en) * | 2012-02-14 | 2012-07-04 | 中国热带农业科学院热带生物技术研究所 | Method for improving ethanol and glucose resistance of saccharomyces cerevisiae |
| CN105200094B (en) * | 2014-05-30 | 2019-12-03 | 中粮营养健康研究院有限公司 | A method of utilizing microbial fermentation lignocellulosic material producing and ethanol |
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