CN104357437A - Method for producing selenium-enriched yeast paste and yeast of beer - Google Patents
Method for producing selenium-enriched yeast paste and yeast of beer Download PDFInfo
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Abstract
The invention discloses a method for producing selenium-enriched yeast paste and yeast of beer, and aims to solve the problems of relatively high equipment investment, limited yeast biomass generated by fermenting and low equipment utilization due to the fact that selenium-enriched yeast is prepared by a pure yeast fermentation culture method. The invention develops another way, when beer is produced, beer and the byproduct selenium-enriched yeast can be obtained at the same time. The selenium-enriched yeast can be produced by utilizing beer brewing production equipment without arranging additional equipment, existing beer production process flow is not changed, equipment is not added, and the fermenting byproduct selenium-enriched yeast can be obtained while beer is produced. According to the method, the selenium-enriched yeast equipment investment can be effectively reduced, the production cost of the selenium-enriched yeast can be reduced, and the equipment utilization and selenium-enriched yeast yield can be greatly improved. The method has an important meaning on promoting development and application of selenium-enriched yeast.
Description
Technical field
The present invention relates to field of microbial fermentation, be specially the production method of a kind of beer yeast rich in selenium mud and yeast, the present invention utilizes cereuisiae fermentum can the feature of rich selenium, obtains byproduct yeast slurry, then yeast slurry is processed into yeast rich in selenium while production beer.
Background technology
Selenium (Selenium Se) is one of necessary trace element of human life activity, and it has enhancing body immunizing power, anti-oxidant and anti-ageing various functions of waiting for a long time.It or humans and animals body glutathion inside superoxide (glutathioneperoxidase, GSHPX) cofactor, have and remove the free radical harmful to body, prevent cytolemma to be oxidized impaired effect, be extensively present in the devices such as the liver of viable organism, kidney, the heart, lung dirty in.Lacking selenium then may cause the various diseases such as cancer, myocardial infarction to occur; the selenium enough by meal ingestion can play prophylactic effect; GSHPX utilizes gsh to make the superoxide of toxicity be reduced to harmless oxy-compound; make peroxide breakdown; scavenging capacity free radical position; Cell protection membrane structure and function, repair molecular damage.At occurring in nature, selenium exists with inorganic selenium and organoselenium two kinds of forms, the selenium of two kinds of forms all can participate in the synthesis of half Selenoperoxidase in liver, but organoselenium biological activity is higher, can effectively assimilate in vivo, and toxicity ratio inorganic selenium is little, be conducive to absorbing in vivo, specific absorption is higher.
Selenium enters in body by food chain, and therefore, the selenium in human body is inseparable with local food draft.If local Selenium in Soils content is few, local people, animal can be caused to take the photograph selenium deficiency, cause scarce selenium.Have more than 40 countries and regions to belong to scarce selenium area in the world, and China has the area of 2/3 to belong to internationally recognized scarce selenium area, has the population lives of 72% in poor selenium area, scarce selenium drastically influence the health of the mankind.Meanwhile, scarce selenium also becomes a kind of endemic illness.
In order to address this problem, the numerous countries comprising developed country adopt oral selenium medicine as the approach mending selenium.And yeast rich in selenium product is as selenium product, wherein containing organoselenium, be a kind of micro-element nutrition reinforcer and fodder additives of good functional foodstuff, its prospect that is widely used.
Biological selenium-enriched is the effective way obtaining organoselenium, and at present, biological selenium-enriched method is mainly microbe transformation method.Microorganism for rich selenium has edible fungi, yeast class and prebiotic mushroom etc., and wherein yeast is the ideal rich selenium carrier of generally acknowledging at present.Yeast rich in selenium refers to sulphur selenium being added replacement part in yeast culture medium, in yeast growth process, by yeast, selenium is absorbed, include thalline in, become a part for its protein, mainly become the result of protein with the analogue of sulfur-containing amino acid (such as selenium methionine or Gelucystine selenium).Yeast has higher selenium rich ability, and inorganic selenium higher for toxicity is converted into safe organoselenium.
It is fast that cereuisiae fermentum has growth and breeding, and fermentation period is short, to absorption of trace elements rate high, is ideal carrier inorganic selenium being converted into organoselenium.The yeast rich in selenium production method of domestic report is generally that under certain culture condition, finally obtain yeast rich in selenium, its cell selenium content is between 300-1200mg/kg by common yeast saccharomyces cerevisiae in the fermented liquid substratum adding appropriate Sodium Selenite.
Summary of the invention
At present, mostly common yeast rich in selenium production method is pure yeast fermentation cultural method, and it by adding Sodium Selenite in substratum, and Billy makes carrier with yeast, and inorganic selenium is converted into organoselenium.The method needs to make special fermentation device---and stirred pot produces yeast, and facility investment is comparatively large, simultaneously, because carbon concentration in medium is lower, liquid amount is also less, and what carry out is low density yeast fermentation, the yeast bio amount that fermentation produces is limited, and equipment efficiency of usage is low.
Goal of the invention of the present invention is: for above-mentioned Problems existing, provides the production method of a kind of beer yeast rich in selenium mud and yeast.The present invention looks for another way, and while production beer, not only obtains beer but also can obtain byproduct yeast rich in selenium.The present invention is adopted to produce yeast rich in selenium, neither need to increase other equipment, utilize brewage production unit, do not change again original process for beer production route, do not increase equipment, while production beer, kill two birds with one stone acquisition fermentation byproduct---yeast rich in selenium.The present invention effectively can reduce the equipment investment of yeast rich in selenium, reduces the production cost of yeast rich in selenium, greatly improves the output of plant factor and yeast rich in selenium, for promoting the development of yeast rich in selenium, apply significant.
To achieve these goals, the present invention adopts following technical scheme:
A production method for beer yeast rich in selenium mud, comprises the steps:
(1) the resistance to selenium domestication of cereuisiae fermentum bacterial classification
Cereuisiae fermentum bacterial classification is carried out resistance to selenium domestication in Se solution, and in Se solution, selenium concentration gradient increase from low to high, obtains the cereuisiae fermentum of resistance to selenium bacterial classification;
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1, for the bacterial classification that sets out, adopts the process of ultraviolet radiation mutagenesis method to set out bacterial classification, filters out transgenation bacterial strain, and leavening property test is carried out to transgenation bacterial strain, obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use;
(3) yeast rich in selenium mud is produced
Divide in fermentor tank and add wheat juice four times, while adding wheat juice for the first time, add the bacterial classification two of preparation in step 2, in fermentor tank, second time, third time add wheat juice successively again, when adding wheat juice the 4th time in fermentor tank, add S-WAT, after having fermented, filter, obtain filtrate and yeast rich in selenium mud respectively;
In described step 3, divide in fermentor tank and add wheat juice four times, the timed interval at every turn adding wheat juice is 4-6 hours, and the add-on of bacterial classification two is 1%-1.5% of wheat juice total mass in fermentor tank;
The add-on of described S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 10-300mg S-WAT.
In described step 3, the add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 200-300mg S-WAT.
In described step 3, S-WAT adds from the Wort cooling Thin plate heat exchanger pipeline of fermentor tank.
In described step 3, filtrate is made as pure mellow wine, after packaging, obtains beer.
A production method for beer yeast rich in selenium, comprises the steps: that the yeast rich in selenium mud adopting abovementioned steps 3 to prepare is raw material, after being washed by yeast rich in selenium mud, successively after debitterize, centrifugal, drying, obtains yeast rich in selenium;
Or by after the drying of yeast rich in selenium mud, obtain yeast rich in selenium.
After the drying, yeast is pulverized, obtain yeast rich in selenium.
The operation steps of described debitterize is as follows: the yeast after washing is made emulsion, carries out debitterize process with the agent of tartrate debitterize.
The operation steps of described debitterize is as follows: the emulsion yeast after washing being made mass concentration 6%-15%, carry out debitterize process with the tartrate debitterize agent of yeast quality 3%-8%, the treatment time is 20-60min.
The operation steps of described debitterize is as follows: the emulsion yeast after washing being made mass concentration 8%, carry out debitterize process with the tartrate debitterize agent of yeast quality 5%, the treatment time is 30min.
For foregoing problems, the invention provides the production method of a kind of beer yeast rich in selenium mud and yeast.The production method of yeast rich in selenium mud comprises the domestication of resistance to selenium, strain improvement and the several step of production yeast rich in selenium mud.
Yeast is the ideal rich selenium carrier of generally acknowledging at present, and the present invention adopts cereuisiae fermentum as bacterial classification, adds appropriate Sodium Selenite when Wort cooling, while fermentative production beer, obtains yeast rich in selenium.But, if cereuisiae fermentum is directly added in wheat juice as bacterial classification, when the addition of selenium exceedes a certain amount of in wheat juice, restraining effect is produced to the rich selenium of cereuisiae fermentum, affects beer normal fermentation simultaneously.Therefore, cereuisiae fermentum direct production yeast rich in selenium mud can not simply be adopted.First applicant carries out resistance to selenium domestication to former cereuisiae fermentum bacterial classification, makes it under the prerequisite not affecting beer fermentation, progressively adapts to absorb in higher selenium concentration condition, transform inorganic selenium simultaneously, improves the selenium rich ability of barms.Prove by experiment, when the addition of selenium is in certain zone of reasonableness, less to the growth-inhibiting of yeast, and good rich selenium effect can be obtained.The present invention, under the prerequisite ensureing beer normal fermentation, takes selenium concentration gradient increase from low to high, cultivates domestication cereuisiae fermentum bacterial classification selenium rich ability under higher selenium concentration condition, and obtain the cereuisiae fermentum of resistance to selenium bacterial classification.
Meanwhile, the present invention also with the cereuisiae fermentum of resistance to selenium bacterial classification for the bacterial classification that sets out, carry out strain improvement.The present invention adopts ultraviolet (UV) to irradiate mutafacient system screening-gene mutant strain, and carries out leavening property test to mutant strain, therefrom filters out applicable produce, the better barms of selenium rich ability.By the method, the barms that selenium rich ability is strong can be obtained, and be designated as bacterial classification two.
Next, the present invention produces.The growth of yeast divide the adaptive phase, logarithmic phase, stationary phase, senescence phase four-stage, the Best Times of interpolation is logarithmic phase.Process for beer production is generally that 4 pots of wheat juice fill a fermentor tank, 4-6 hours, interval between batch, the adding technology flow process of cereuisiae fermentum is as shown in Figure 1, specific as follows: first pot of wheat juice, second pot, the 3rd pot wheat juice enter the equal uniform flow of tank and adds, and the 4th pot is not added yeast.Fermentation canful yeast growth reaches 1,600 ten thousand/ml, and yeast growth is in logarithmic phase, and the 3rd day, the 4th day yeast growth of fermentation is in peak value.Applicant considers the singularity that beer fermentation is produced, and prevents microbiological contamination of fermenting, and Sodium Selenite can not add in the canful best moment of 16-20 hours, adjustment yeast adding technology of can only trying every possible means, the logarithmic phase of yeast growth as far as possible in advance.For this reason, yeast three adding technologies are adjusted to and once add by applicant, adding total amount is 1%-1.5% of wheat juice total mass in fermentor tank, namely when first pot of Wort cooling enters tank, by yeast (i.e. bacterial classification two of the present invention) once stream add, when the 4th pot of wheat juice enters tank, the yeast added in fermentor tank substantially tides over the adaptive phase, the best entering Sodium Selenite adds opportunity---yeast growth logarithmic phase, and schema as shown in Figure 2.Sodium Selenite adds with the equal uniform flow of wheat juice before the 4th pot of (last pot) wheat juice canful.Simultaneously, the interpolation of S-WAT has material impact for the growth of yeast rich in selenium mud, applicant is found by research, as the interpolation concentration≤300mg/L of S-WAT, compared with control sample, yeast growth peak value shifts to an earlier date 1-2 days and reaches, on cereuisiae fermentum growth not impact, even also helpful to yeast growth, may be can stimulate the activity of yeast that the yeast adaptive phase is shortened after adding Sodium Selenite, illustrate that the fermentation of the present invention to beer has promoter action; When adding concentration > 300mg/L, Sodium Selenite is drawn up yeast growth, and yeast is likely poisoning, suppresses yeast activity on the contrary.
With the yeast rich in selenium mud obtained for raw material, by yeast rich in selenium mud wash after, successively through debitterize, centrifugal, dry, pulverizing after, obtain medicinal yeast rich in selenium; Or with the yeast rich in selenium mud obtained for raw material, after the drying of yeast rich in selenium mud, pulverizing, obtain feed yeast rich in selenium, it can act on animal feedstuff additive, for growth of animal provides sufficient selenium.
Yeast rich in selenium product of the present invention is as the basic material of selenium product development, have broad application prospects, the method is produced simple, do not change original process for beer production, only need add appropriate Sodium Selenite before Wort cooling, cereuisiae fermentum is utilized to have stronger selenium rich ability, inorganic selenium can be converted into organoselenium, its more than Transformation efficiency is up to 90 %, the selenium content of yeast can reach more than 300mg/kg, and the waste yeast mud of collection can obtain yeast rich in selenium powder by general yeast working method.
The present invention is fermented by yeast S, while preparing beer, produces yeast rich in selenium mud and yeast rich in selenium.The present invention, without the need to additionally adding yeast rich in selenium production unit, effectively can reduce equipment investment, and meanwhile, the present invention can shorten the production cycle of beer, improves the economic benefit of beer; Further, the present invention can also by-product yeast rich in selenium mud and yeast rich in selenium, and has higher output, achieves the effect of killing two birds with one stone.
Accompanying drawing explanation
Examples of the present invention will be described by way of reference to the accompanying drawings, wherein:
Fig. 1 is the adding technology schema of existing cereuisiae fermentum.
Fig. 2 is the adding technology schema of bacterial classification two and S-WAT in the present invention.
Embodiment
All features disclosed in this specification sheets, or the step in disclosed all methods or process, except mutually exclusive feature and/or step, all can combine by any way.
Arbitrary feature disclosed in this specification sheets, unless specifically stated otherwise, all can be replaced by other equivalences or the alternative features with similar object.That is, unless specifically stated otherwise, each feature is an example in a series of equivalence or similar characteristics.
1 measuring method
Embodiments of the invention are adopted and are measured with the following method.
The mensuration of 1.1 biomasss
By centrifugal for fermented liquid 5000rpm/min 10min, collect thalline, distilled water wash twice, weigh after collecting the lyophilize of yeast thalline, obtain yeast bio amount, represent with g/L.
The mensuration of 1.2 yeast cell selenium and the mensuration of total selenium
The mensuration of selenium content: Se+4 can generate azo ion by catalytic chlorination acid potassium oxidation phenylhydrazine hydrochloride, then becomes red azo dyestuff with chromotropic acid coupling.The light absorption value of the red azo dyestuff generated is directly proportional to the selenium of finite concentration scope, utilizes ultraviolet-visible spectrophotometer to survey its absorbancy under 520nm wavelength, with calibration curve method, can try to achieve the content of selenium.
The selenium content of the dry mycelium obtained after measuring said determination biomass, is the selenium in yeast cell.Because the selenium in yeast cell mainly exists with the form of organoselenium, wherein organoselenium can account for more than 90% of total selenium in its cell, and therefore, using the selenium content in yeast cell as the how many index of the organoselenium weighing yeast enrichment, corresponding calculation formula is as follows.
Selenium content (ug/L) in selenium total content (ug/L)=yeast cell × biomass (ug/L)
The concentration (ug/L) × 100% of transformation efficiency (%)=selenium total content (the ug/L)/interpolation selenium of selenium
Embodiment 1
(1) the resistance to selenium resistance domestication of cereuisiae fermentum bacterial classification
By cereuisiae fermentum bacterial classification in liquid malt extract medium, add the Sodium Selenite of the 10wt% of different concns, tame yeast by the method for gradient concentration.
Solid medium: 10 ° of BX wort+1% agar.
Liquid seed culture medium: 10 ° of BX worts, concrete grammar is as follows:
Slant strains (cereuisiae fermentum bacterial classification)-→ shake-flask culture (in substratum, selenium content is 5ug/ml)-→ gradient dilution be coated with dull and stereotyped (in substratum, selenium content is 10ug/ml)-→ choose single bacterium colony in inclined-plane-→ shake-flask culture (in substratum, selenium content is 10ug/ml)-→... ...-→ repeat aforesaid operations process, until the content of selenium is 25ug/ml in substratum.With this understanding, measure barms to the resistance of selenous acid, the single bacterium colony selecting the strong selenium rich ability of growth strong is starting strain, is the cereuisiae fermentum of resistance to selenium bacterial classification.
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1, for the bacterial classification that sets out, adopts the process of ultraviolet radiation mutagenesis method to set out bacterial classification, filters out transgenation bacterial strain, and leavening property test is carried out to transgenation bacterial strain, obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use.
Wherein, ultraviolet (UV) mutafacient system is as follows.
First will be in the bacteria suspension dilution of logarithmic phase with malt extract medium, and make the concentration of bacterial classification reach 1 × 10
6individual/ml.Open 20W ultraviolet lamp preheat 30min, treat that its power reaches stable.Get 7 9cm culture dish and draw the bacteria suspension of 4ml dilution respectively in culture dish, liquid layer thickness is about 2mm, and 7 culture dish are placed in ultraviolet lamp 20cm place, takes a culture dish away at interval of 30s, then gets a culture dish and oppose in the same old way without uv irradiating.Mutagenesis liquid after 7 ware mutagenesis and the non-mutagenesis of 1 ware are carried out 10 times of dilution method dilutions and are coated with dull and stereotyped, and Duplicate Samples 3, cultivates 48h for 28 DEG C, calculate mutation rate and lethality rate.
Select mortality ratio be about 85% UV irradiation time be test the mutagenic exposure time, being in logarithmic phase yeast irradiates after mutagenesis through UV, coat wheat juice solid medium and cultivate 48h, number with inoculating needle random choose 7 bacterium colonies, that 7 mutant strains and starting strain are received respectively corresponding numbering is equipped with in the test tube of identical wheat juice, be placed in 28 DEG C of constant temperature 130r/min, after shaking culture 56h, make seed for subsequent use.Draw 15ml seed liquor respectively, be inoculated in the triangular flask that 300ml wheat juice is housed, 26 DEG C, 130r/min oscillation and fermentation 6d, weigh once every day, adopts CO
2weight-loss method measures the leavening property of different strains.
Through verification experimental verification: fermentation inoculation 14h bacterium liquid is in logarithmic phase, get bacterium liquid and carry out ultraviolet mutagenesis, time of ultraviolet irradiation is 90s, 28 DEG C of dull and stereotyped constant temperature culture 48h of solid-state malt extract medium, choose 7 yeast colonies grown fine, 1-No. 7 numberings, No. 8 is control sample, carries out leavening property test, and the final selected good bacterial strain of 1 strain leavening property is for produce bacterial strain greatly.Above liquid state or solid malt extract medium are the malt extract medium of selenium content 25ug/ml.
(3) yeast rich in selenium mud is produced
Divide in fermentor tank and add wheat juice four times, while adding wheat juice for the first time, add the bacterial classification two of preparation in step 2, in fermentor tank, second time, third time add wheat juice successively again, when adding wheat juice the 4th time in fermentor tank, add S-WAT, after having fermented, filter, obtain filtrate and yeast rich in selenium mud respectively.Wherein, the timed interval at every turn adding wheat juice is 4-6 hours.
In the present embodiment, former three adding technologies are adjusted to and once add, meet to greatest extent and add Sodium Selenite in yeast growth logarithmic phase, impact is dropped to minimum.The add-on of bacterial classification two is 1.5% of wheat juice total mass in fermentor tank; The add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 300mg S-WAT.The amount of dividing equally three times is once added when first pot of wheat juice enters tank, relative first pot of wheat juice amount is equivalent to increase yeast-inoculated amount, enter rapidly logarithmic phase to the yeast growth shortening adaptive phase and provide Time and place, second pot, the 3rd pot, the 4th pot wheat juice enters tank timed interval 4h-6h, relative first pot of wort fermentation, can be considered and mend sugar operation.The adjustment of yeast adding technology, can meet the processing requirement that yeast growth logarithmic phase adds Sodium Selenite to greatest extent, after the adjustment of yeast addition means, does not substantially affect beer fermentation and final beer quality.
Food grade Sodium Selenite non-contaminant brew water is configured to the solution of 10wt% concentration by applicant, the addition that concentration calculates Sodium Selenite is added by 300ug/ml, add place: Wort cooling Thin plate heat exchanger preceding pipeline adds (utilize the heat of 90-95 DEG C of hot wheat juice self, carry out sterilization to sodium selenite solution); Addition manner: add with the mobile equal uniform flow of pump that adds during canful last pot of Wort cooling.
After beer Primary Fermentation terminates, main ferment temperature is cooled to yeast recovered temperature 0-2 DEG C (not needing the waste yeast of reserving seed for planting) by 10-12 DEG C, collects yeast slurry.Yeast itself is owing to adsorbing the impurity such as some hop resins, protein chip, wheat skin, and also have very large bitter taste due to hop resin, so yeast slurry could must obtain for rich selenium series foods, pharmaceutical production base-material yeast rich in selenium through operations such as washing, debitterize, centrifugal, dry, pulverizing.If produce feed yeast rich in selenium, more simple, only need by yeast slurry drying, pulverize.
Wherein, to prepare the operation steps of yeast rich in selenium as follows for yeast slurry.
Washing: the pure water agitator treating yeast adding yeast slurry volume several times, according to processing requirement, repetitive scrubbing 2-3 time.
Removal of impurities: removal of impurities of sieving when at every turn washing with 80 object sieves, the impurity such as removing wheat skin, protein chip.
Debitterize: the yeast emulsion being prepared into 8% concentration after washing is sieved, adds the tartrate debitterize agent debitterize process 30min of 5%.
Centrifugal: the yeast-lactic liquid precipitate after debitterize is centrifugal, the centrifugal 10min of 2000rpm obtains clean wet yeast.
Dry: oven drying put into by the wet yeast after centrifugal.
Pulverize: dried yeast can obtain finished product yeast rich in selenium powder through pulverizing.
After measured, in yeast rich in selenium prepared by the present embodiment, the transformation efficiency of selenium is 95%, and the selenium content of yeast can reach 350mg/kg.
Embodiment 2
(1) the resistance to selenium resistance domestication of cereuisiae fermentum bacterial classification
Cultivate with the cereuisiae fermentum bacterial classification of Mianyang, Sichuan brew-house, other operations are identical with embodiment 1.
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1, for the bacterial classification that sets out, adopts the process of ultraviolet radiation mutagenesis method to set out bacterial classification, filters out transgenation bacterial strain, and leavening property test is carried out to transgenation bacterial strain, obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use.
Wherein, ultraviolet (UV) mutafacient system is as follows.
First will be in the bacteria suspension dilution of logarithmic phase with malt extract medium, and make the concentration of bacterial classification reach 1 × 10
6individual/ml.Open 20W ultraviolet lamp preheat 30min, treat that its power reaches stable.Get 12 9cm culture dish and draw the bacteria suspension of 4ml dilution respectively in culture dish, liquid layer thickness is about 2mm, and 12 culture dish are placed in ultraviolet lamp 20cm place, takes a culture dish away at interval of 30s, then gets a culture dish and oppose in the same old way without uv irradiating.Mutagenesis liquid after 12 ware mutagenesis and the non-mutagenesis of 1 ware are carried out 10 times of dilution method dilutions and are coated with dull and stereotyped, and Duplicate Samples 3, cultivates 48h for 28 DEG C, calculate mutation rate and lethality rate.
Select mortality ratio be about 85% UV irradiation time be test the mutagenic exposure time, being in logarithmic phase yeast irradiates after mutagenesis through UV, coat wheat juice solid medium and cultivate 48h, number with inoculating needle random choose 10 bacterium colonies, that 10 mutant strains and starting strain are received respectively corresponding numbering is equipped with in the test tube of identical wheat juice, be placed in 28 DEG C of constant temperature 130r/min, after shaking culture 56h, make seed for subsequent use.Draw 15ml seed liquor respectively, be inoculated in the triangular flask that 300ml wheat juice is housed, 26 DEG C, 130r/min oscillation and fermentation 6d, weigh once every day, adopts CO
2weight-loss method measures the leavening property of different strains.
Through verification experimental verification: fermentation inoculation 14h bacterium liquid is in logarithmic phase, get bacterium liquid and carry out ultraviolet mutagenesis, time of ultraviolet irradiation is 90s, 28 DEG C of dull and stereotyped constant temperature culture 48h of solid-state malt extract medium, choose 12 yeast colonies grown fine, 1-No. 12 numberings, No. 13 is control sample, carries out leavening property test, and the final selected good bacterial strain of 6 strain leavening property is for produce bacterial strain greatly.Above liquid state or solid malt extract medium are the malt extract medium of selenium content 25ug/ml.
(3) yeast rich in selenium mud is produced
The add-on of bacterial classification two is 1.2% of wheat juice total mass in fermentor tank; The add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 100mg S-WAT.Other are identical with embodiment 1
After measured, in yeast rich in selenium prepared by the present embodiment, the transformation efficiency of selenium is 91.3%, and the selenium content of yeast can reach 308mg/kg.
Embodiment 3
(1) the resistance to selenium resistance domestication of cereuisiae fermentum bacterial classification
Cultivate with the cereuisiae fermentum bacterial classification of Sichuan brew-house, other operations are identical with embodiment 1.
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1 is for the bacterial classification that sets out, and other operations are identical with embodiment 1.Obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use.
(3) yeast rich in selenium mud is produced
The add-on of bacterial classification two is 1.5% of wheat juice total mass in fermentor tank; The add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 220mg S-WAT.Other are identical with embodiment 1
After measured, in yeast rich in selenium prepared by the present embodiment, the transformation efficiency of selenium is 93.6%, and the selenium content of yeast can reach 323mg/kg.
Embodiment 4
(1) the resistance to selenium resistance domestication of cereuisiae fermentum bacterial classification
The cereuisiae fermentum bacterial classification getting Sichuan brew-house is cultivated, and other operations are identical with embodiment 1.
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1 is for the bacterial classification that sets out, and other operations are identical with embodiment 1.Obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use.
(3) yeast rich in selenium mud is produced
The add-on of bacterial classification two is 1.3% of wheat juice total mass in fermentor tank; The add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 300mg S-WAT.Other are identical with embodiment 1
After measured, in yeast rich in selenium prepared by the present embodiment, the transformation efficiency of selenium is 92.9%, and the selenium content of yeast can reach 317mg/kg.
Embodiment 5
(1) the resistance to selenium resistance domestication of cereuisiae fermentum bacterial classification
By the cereuisiae fermentum bacterial classification of somewhere, Sichuan brew-house in liquid malt extract medium, add the Sodium Selenite of the 8wt% of different concns, tame yeast by the method for gradient concentration.
Solid medium: 10 ° of BX wort+1% agar.
Liquid seed culture medium: 10 ° of BX worts, concrete grammar is as follows:
Slant strains (cereuisiae fermentum bacterial classification)-→ shake-flask culture (in substratum, selenium content is 5ug/ml)-→ gradient dilution be coated with dull and stereotyped (in substratum, selenium content is 10ug/ml)-→ choose single bacterium colony in inclined-plane-→ shake-flask culture (in substratum, selenium content is 10ug/ml)-→... ...-→ repeat aforesaid operations process, until the content of selenium is 25ug/ml in substratum.With this understanding, measure barms to the resistance of selenous acid, the single bacterium colony selecting the strong selenium rich ability of growth strong is starting strain, is the cereuisiae fermentum of resistance to selenium bacterial classification.
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1, for the bacterial classification that sets out, adopts the process of ultraviolet radiation mutagenesis method to set out bacterial classification, filters out transgenation bacterial strain, and leavening property test is carried out to transgenation bacterial strain, obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use.Other operations are identical with embodiment 1.
(3) yeast rich in selenium mud is produced
Divide in fermentor tank and add wheat juice four times, while adding wheat juice for the first time, add the bacterial classification two of preparation in step 2, in fermentor tank, second time, third time add wheat juice successively again, when adding wheat juice the 4th time in fermentor tank, add S-WAT, after having fermented, filter, obtain filtrate and yeast rich in selenium mud respectively.Wherein, the timed interval at every turn adding wheat juice is 6 hours.
Wherein, the add-on of bacterial classification two is 1.5% of wheat juice total mass in fermentor tank; The add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 200mg S-WAT.
After measured, in yeast rich in selenium prepared by the present embodiment, the transformation efficiency of selenium is 94.1%, and the selenium content of yeast can reach 335mg/kg.
The present invention is not limited to aforesaid embodiment.The present invention expands to any new feature of disclosing in this manual or any combination newly, and the step of the arbitrary new method disclosed or process or any combination newly.
Claims (9)
1. a production method for beer yeast rich in selenium mud, is characterized in that, comprises the steps:
(1) the resistance to selenium domestication of cereuisiae fermentum bacterial classification
Cereuisiae fermentum bacterial classification is carried out resistance to selenium domestication in Se solution, and in Se solution, selenium concentration gradient increase from low to high, obtains the cereuisiae fermentum of resistance to selenium bacterial classification;
(2) strain improvement
The cereuisiae fermentum of the resistance to selenium bacterial classification prepared with step 1, for the bacterial classification that sets out, adopts the process of ultraviolet radiation mutagenesis method to set out bacterial classification, filters out transgenation bacterial strain, and leavening property test is carried out to transgenation bacterial strain, obtain the barms that selenium rich ability is strong, be designated as bacterial classification two, for subsequent use;
(3) yeast rich in selenium mud is produced
Divide in fermentor tank and add wheat juice four times, while adding wheat juice for the first time, add the bacterial classification two of preparation in step 2, in fermentor tank, second time, third time add wheat juice successively again, when adding wheat juice the 4th time in fermentor tank, add S-WAT, after having fermented, filter, obtain filtrate and yeast rich in selenium mud respectively;
In described step 3, divide in fermentor tank and add wheat juice four times, the timed interval at every turn adding wheat juice is 4-6 hours, and the add-on of bacterial classification two is 1%-1.5% of wheat juice total mass in fermentor tank;
The add-on of described S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 10-300mg S-WAT.
2. the production method of beer yeast rich in selenium mud according to claim 1, it is characterized in that, in described step 3, the add-on of S-WAT is with the entire volume of wheat juice in fermentor tank, and 1L wheat juice adds 200-300mg S-WAT.
3. the production method of beer yeast rich in selenium mud according to claim 1 or 2, it is characterized in that, in described step 3, S-WAT adds from the Wort cooling Thin plate heat exchanger pipeline of fermentor tank.
4. the production method of beer yeast rich in selenium mud according to claim 1 or 2 or 3, is characterized in that, in described step 3, filtrate is made as pure mellow wine, after packaging, obtains beer.
5. the production method of a beer yeast rich in selenium, it is characterized in that, comprise the steps: to adopt the yeast rich in selenium mud that described in any one of claim 1-4 prepared by step 3 to be raw material, after yeast rich in selenium mud is washed, successively after debitterize, centrifugal, drying, obtain yeast rich in selenium;
Or by after the drying of yeast rich in selenium mud, obtain yeast rich in selenium.
6. according to the production method of claim 5 beer yeast rich in selenium, it is characterized in that, after the drying, yeast is pulverized, obtain yeast rich in selenium.
7. according to the production method of claim 5 or 6 beer yeast rich in selenium, it is characterized in that, the operation steps of described debitterize is as follows: the yeast after washing is made emulsion, carries out debitterize process with the agent of tartrate debitterize.
8. the production method of beer yeast rich in selenium according to Claim 8, it is characterized in that, the operation steps of described debitterize is as follows: the emulsion yeast after washing being made mass concentration 6%-15%, carry out debitterize process with the tartrate debitterize agent of yeast quality 3%-8%, the treatment time is 20-60min.
9. according to the production method of claim 9 beer yeast rich in selenium, it is characterized in that, the operation steps of described debitterize is as follows: the emulsion yeast after washing being made mass concentration 8%, carry out debitterize process with the tartrate debitterize agent of yeast quality 5%, the treatment time is 30min.
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