CN103194400B - Low-yield acetaldehyde beer yeast and domestication method thereof - Google Patents
Low-yield acetaldehyde beer yeast and domestication method thereof Download PDFInfo
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- CN103194400B CN103194400B CN201310100330.3A CN201310100330A CN103194400B CN 103194400 B CN103194400 B CN 103194400B CN 201310100330 A CN201310100330 A CN 201310100330A CN 103194400 B CN103194400 B CN 103194400B
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Abstract
The invention discloses a low-yield acetaldehyde beer yeast D-A-14(Saccharomyces cerevisiae D-A-14) which is preserved in the China general microbiological culture collection center on November 13, 2012, wherein the preservation address is microbiology research institute of China science academy in the Chaoyang district of Beijing, and the preservation number is CGMCC No.6822. The method screens the low-yield acetaldehyde beer yeast by adopting a beer yeast as an original strain, carrying out ultraviolet mutagenesis and then coating on a disulfiram-containing synthetic culture medium. The screened optimized yeast disclosed by the invention can be enhanced in acetaldehyde metabolic capability and high in acetaldehyde metabolic speed by being domesticated for a certain period through an acetaldehyde culture medium and has important application value.
Description
Technical field
A kind of method that the present invention relates to cereuisiae fermentum and raise and train, particularly a kind of low yield acetaldehyde cereuisiae fermentum and the method for raising and train thereof.
Background technology
Acetaldehyde, the highest volatile aldehyde of content in beer, is the main source of the raw blue or green taste of beer, rotten apple taste.In the time that acetaldehyde exceedes 50mg/L, there is the excitement that cannot swallow; While exceeding 25mg/L, there are strong impulse and pungent sense and gloomy property mouthfeel; While exceeding 10mg/L, there is jejune mouthfeel.The acetaldehyde of ripe quality beer is generally at 3～8mg/L.Acetaldehyde is one of material causing " top ", and has carcinogenesis, and foreign scholar has carried out a large amount of research to its carinogenicity and toxicity.In addition, acetaldehyde also has impact to microbial growth, and the acetaldehyde of high density will suppress microorganism fermentation, promotes its fermentation when lower concentration.Therefore be, to reduce its content in beer about the research emphasis of acetaldehyde at present.
The generation approach of acetaldehyde in Process of Beer Brewing comprises biological approach and chemistry route.Wherein, the typical Biometabolic pathway of acetaldehyde is as shown below.The key enzyme that affects acetaldehyde has pyruvic oxidase, acetaldehyde dehydrogenase and ethanol dehydrogenase II etc., is bases that research yeast metabolism produces acetaldehyde to the understanding of these enzymes.The chemistry route that acetaldehyde forms can be divided into again two kinds, the first is Shi Chuike (Strecker) degraded of L-Ala, refer in browning reaction process, form dicarbonyl compound react with L-Ala, produce than the acetaldehyde of the few carbon atom of L-Ala; The second is that the melanoidin of oxidation state becomes acetaldehyde with polyphenol by oxidation of ethanol.
From the pathways metabolism of yeast, acetaldehyde dehydrogenase is the key enzyme in acetaldehyde pathways metabolism.And Tosse) is the diploid of two ethylene dithiol ammonia methyl esters, claim again disulfiram, wine fear etc., can be used as wine hypersitization medicine, carry out the treatment of abstinence from alcohol.It is one of inhibitor of acetaldehyde dehydrogenase, and certain density Tosse) can partly or entirely suppress the activity of acetaldehyde dehydrogenase, thereby makes acetaldehyde can not be oxidized to acetic acid.In the substratum taking ethanol as sole carbon source, once acetic acid can not form, the metabolism stream of yeast just cannot go on, and yeast can not be grown.The height difference of the level of the acetaldehyde dehydrogenase of different its secretions of yeast strain, to the ability difference of catalysis oxidation of acetaldehyde, finally shows the final concentration difference to some extent of acetaldehyde in tunning.The present invention utilizes Tosse) flat board, low-level acetaldehyde dehydrogenase activity is suppressed, and then suppress the growth of corresponding yeast strain, only be subject to part inhibition because enzyme is alive and produce the bacterial strain that enzyme level is relatively high, can continue growth metabolism on Tosse) flat board, seed selection obtains the bacterial strain of low yield acetaldehyde effectively rapidly thus.Afterwards, utilize acetaldehyde substratum to raise and train yeast, by add the acetaldehyde of high density in substratum, make yeast be in for a long time the acetaldehyde degradation ability of evolving out stronger in the environment of a high acetaldehyde concentration, make the further low yield of acetaldehyde, heritability is more stable.
Summary of the invention
The technical problem to be solved in the present invention is to provide a kind of low yield acetaldehyde yeast saccharomyces cerevisiae (Saccharomyces cerevisiae), be preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on November 13rd, 2012, deposit number is CGMCC No.6822, and preservation address is Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica.
Described yeast shape approximation is in circle, and diameter is between 2.0-3.5 micron; After 48 hours, can grow the bacterium colony of size approximately 2.3 millimeter in the surface growth of YPD solid medium, bacterium colony oyster white, glossy, smooth, neat in edge, homogeneous, translucent, easily picking, has wine flavour.
The present invention also provides a kind of method that obtains low yield acetaldehyde cereuisiae fermentum of screening, and starting strain is after ultraviolet mutagenesis, and dilution spread is in Tosse) flat board, the bacterium colony that picking grows.
Tosse) is the inhibitor of acetaldehyde dehydrogenase, in the cultivation taking ethanol as sole carbon source that contains Tosse), Tosse) suppresses the growth of yeast by the activity that suppresses acetaldehyde dehydrogenase, and the relatively high bacterial strain of enzymic activity is lived and is only subject to part and suppresses can continue to grow on Tosse) flat board because of enzyme.Can produce the bacterial strain of high reactivity acetaldehyde dehydrogenase, the acetaldehyde of its metabolism can be under the effect of this enzyme be transformed by logical snperoxiaized mode rapidly, thereby acetaldehyde in end product also can be lower, can filter out thus the cereuisiae fermentum of low yield acetaldehyde.
Tosse) substratum in the present invention (ethanol is sole carbon source) component is as follows:
Ethanol 2% Tosse) 10 μ M agar 2% (NH
4)
2sO
45g/l KH
2pO
41g/l NaCl0.1g/l MgSO
47H
2o0.5g/l CaCl
20.1g/l yeast extract paste 0.1g/l
The present invention tames the bacterial strain screening, to obtaining the bacterial strain with better industrial application value, the yeast saccharomyces cerevisiae of above-mentioned acquisition is passed through to the domestication of the acetaldehyde pressure of the high density of certain hour, improve the ability that yeast self transforms acetaldehyde, the concentration of acetaldehyde in end product is further reduced.
The condition of acetaldehyde pressure domestication is:
Substratum (acetaldehyde is sole carbon source) component is as follows:
(NH
4)
2sO
45g/l KH
2pO
41g/l NaCl0.1g/l MgSO
47H
2o0.5g/l CaCl
20.1g/l yeast extract paste 0.1g/l acetaldehyde 200mg/l
Culture temperature is 11 DEG C, in case acetaldehyde volatilization.
The invention provides a kind of cereuisiae fermentum of low yield acetaldehyde, after its fermentation, acetaldehyde can lowly reach 2.86mg/L, uses acclimation method provided by the invention, and naturalized strain efficiently improves strain stability.
Embodiment
The acquisition of embodiment 1 low yield acetaldehyde cereuisiae fermentum
Respectively taking containing Tosse) concentration as 0.5,1.0,1.5,2.0,2.5,3.0, the culture medium culturing of the 3.5mg/L bacterium that sets out, only having Tosse) is that the substratum of 3.5mg/L does not have yeast to grow, so choose the resistance concentration that 3.5mg/L is screening low yield acetaldehyde yeast.
Taking cereuisiae fermentum (deposit number is as CGMCC NO.2448) as starting strain, by ultraviolet mutagenesis (15W ultraviolet lamp, at 30cm place, the yeast liquid of breaing up is carried out the uv irradiating 27s of different time), dilution spread is in Tosse) flat board, the bacterium colony that picking grows, number consecutively is 1,2,3,4,5,6,7,8,9 etc.
In table 1 Tosse) resistant panel, choose the fermentation situation of bacterium colony
Note: partial data only in table, because of length finite part data unlisted.
Bacterial strain in table 1 is that the yeast that sets out is longer than the bacterial strain on Tosse) substratum after ultraviolet mutagenesis, experimental results show that through the yeast of Tosse) plate screening are all bacterial strains that acetaldehyde reduces, just to have height to have low for the reduction amplitude of acetaldehyde, need to further tame.
The domestication of embodiment 2 bacterial strains
The bacterial strain obtaining in embodiment 1 is placed in to the certain density liquid nutrient medium taking acetaldehyde as sole carbon source, and every 48-72h switching once, to ensure the concentration of acetaldehyde, is cultivated 14 days for 11 DEG C.After raising and train, carry out triangular flask fermentation, investigate its acetaldehyde.
The yeast fermentation situation of table 2 after acetaldehyde substratum is raised and train
Note: partial data only in table, because of length finite part data unlisted.
As can be seen from Table 2, the yeast acetaldehyde rate of descent of raising and train through acetaldehyde substratum is 89.5%, and yeast acetaldehyde declines and more than 50% account for 68.4%, thereby proves that acetaldehyde substratum raises and train yeast and can effectively reduce acetaldehyde amount.
After raising and train, obtain the extremely excellent yeast saccharomyces cerevisiae D-A-14 of a strain, its acetaldehyde output is 2.86mg/L, and preservation is to China Committee for Culture Collection of Microorganisms, preserving number is: CGMCC No.6822, preservation address is Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica.
Claims (2)
- A low yield acetaldehyde yeast saccharomyces cerevisiae ( saccharomyces cerevisiae) D-A-14, being preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on November 13rd, 2012, preservation address is Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, deposit number is CGMCC No.6822.
- 2. yeast saccharomyces cerevisiae D-A-14 according to claim 1, is characterized in that described yeast saccharomyces cerevisiae D-A-14 shape approximation is in circle, and diameter is between 2.0-3.5 micron; After 48 hours, can grow the bacterium colony of big or small 2-3 millimeter in the surface growth of YPD solid medium, bacterium colony oyster white, glossy, smooth, neat in edge, homogeneous, translucent, easily picking, has wine flavour.
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| CN105861206B (en) * | 2016-05-23 | 2019-11-26 | 江南大学 | A kind of fingered citron beer and its brew method |
| CN106148211B (en) * | 2016-09-08 | 2019-07-02 | 江南大学 | The brewer's yeast of one plant of low-acetaldehyde and its application |
| CN109337896A (en) * | 2018-09-18 | 2019-02-15 | 皖西学院 | A kind of selection of low-acetaldehyde glug brewer's yeast |
| CN111662835B (en) * | 2020-06-28 | 2022-03-01 | 青岛啤酒股份有限公司 | Beer yeast capable of producing beer with low acetaldehyde content and domestication method thereof |
| CN112608917A (en) * | 2021-01-08 | 2021-04-06 | 青岛啤酒股份有限公司 | High-throughput screening method for low-acetaldehyde-yield beer industrial yeast |
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| CN102220253A (en) * | 2011-06-16 | 2011-10-19 | 天津大学 | Saccharomyces cerevisiae strain with vacuum tolerability |
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Non-Patent Citations (6)
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| Development of industrial brewing yeast with low acetaldehyde production and improved flavor stability;Jinjing Wang et al.;《Appl Biochem Biotechnol》;20130109;第169卷(第3期);第1016-1025页 * |
| Jinjing Wang et al..Development of industrial brewing yeast with low acetaldehyde production and improved flavor stability.《Appl Biochem Biotechnol》.2013,第169卷(第3期),第1016-1025页. |
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