CN101818183A - Method for improving trehalose content of yeast cell - Google Patents
Method for improving trehalose content of yeast cell Download PDFInfo
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- CN101818183A CN101818183A CN 201010146889 CN201010146889A CN101818183A CN 101818183 A CN101818183 A CN 101818183A CN 201010146889 CN201010146889 CN 201010146889 CN 201010146889 A CN201010146889 A CN 201010146889A CN 101818183 A CN101818183 A CN 101818183A
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Abstract
The invention relates to a method for improving trehalose content of a yeast cell, which comprises the following steps of: centrifuging yeast fermentation culture solution to obtain yeast milk with cell concentration of 13 to 17 percent, adding aqueous solution of maltose, sodium chloride and calcium chloride into the yeast milk, culturing the mixture for 2 to 3 hours under the condition that the temperature is 35 DEG C, the pH value is 5.2 and the ventilation rate of 1.0 to 1.2vvm, centrifuging the cultured mixture, and collecting yeast paste, wherein the trehalose content of the yeast reaches over 16 percent. The method can improve the trehalose in the yeast cell to over 16 percent from 8 percent in shorter time; and compared with the prior art, the method has the advantages of shortening the fermentation time and saving fermentation equipment and power consumption.
Description
Technical field
The present invention relates to a kind of method that improves the yeast cell trehalose, particularly a kind of method that improves the rusk trehalose content of yeast cell belongs to fermentation engineering and microbial technology field.
Background technology
Active dry yeast is by fresh yeast or claims yeast cake to make through the granulating and drying dehydration that the activity of active dry yeast and period of storage are to estimate the important indicator of yeast quality.And the content of these indexs and yeast cell intracellular trehalose is closely related.
Trehalose (Trehalose) has another name called lentinan, deep sugar, and its molecular formula is C
12H
22O
112H
2O, and chemical name α-D-glucopyranosyl α-D-glucopyranoside (α-D-glycopyranosyl-α-D-glycopyranoside), be the irreducibility disaccharide that forms by the condensation of hemiacetal hydroxyl by two glucose molecules.Trehalose extensively is present in lower plant, algae, bacterium, filamentous fungus, yeast, the insect.
Some species is severe environment to external world; as arid; high temperature; dehydration; freezing; high osmotic pressure and toxicant etc.; the trehalose that exists in the degeneration-resistant tolerance that is showed and these species bodies is closely related; has unique biological characteristics that is different from other carbohydrate; can under above-mentioned environment, protect biological cell protein; fat; carbohydrate; components such as nucleic acid are without prejudice; can protect DNA to prevent the damage that radioactive rays cause; ectogenic trehalose also can have good nonspecific provide protection to organism and biomacromolecule; thereby this makes it that many purposes be arranged, as make the cell frostproofer in medicine and microbiology; the stablizer of diagnostic reagent and biological product; the effective ingredient of makeup; food preservatives; cultivate drought-resistant crops etc. on the agricultural.
Trehalose is as a kind of stress metabolite of yeast, can give microorganism opposing nutritive deficiencies such as yeast, high temperature, low temperature, dry, high osmotic pressure, the ability of severe environment such as toxic substance, it has the microbial film of protection organism under harsh and unforgiving environments, liposome, protein, the special efficacy of 26S Proteasome Structure and Functions such as nucleic acid, it makes and keeps moistening in the cell, prevent the loss of various nutrients in the cell, thereby allow these biologies be in the anhydrobiosis state, as when producing active dry yeast, the dry yeast of content of trehalose below 11% (in solid substance), its active preservation period is short, is difficult for storing; Content of trehalose can prolong 4-6 month between its preservation period when 12% to 14% left and right sides; And when content of trehalose surpassed 16%, its preservation period can reach more than 24 months.
Active dry yeast is to be made through the granulating and drying dehydration by fresh yeast (or claiming yeast cake), and the activity of dry yeast is represented with fermenting power usually.Generally speaking, the 3.5g fresh yeast could produce the 1g active dry yeast, but with regard to active (fermenting power), the 1g active dry yeast only is equivalent to the 2-3g fresh yeast: check is surveyed viable cell and is generally 80-90% after the rehydration.Fresh yeast is made active dry yeast, and after the storage and the reconstitution process before the use of certain hour, the certain activity loss is arranged.
The content of the trehalose in the yeast has very big influence to the activity of active dry yeast and storage.Determined the activity and the long keeping ability of high activity dried yeast to a certain extent, the amount of trehalose increases, and helps the stability of cell in dry and storage process like this.Higher content of trehalose helps prolonging the prolongation of the storage period of active dry yeast.And the height of the content of phosphorus accumulates for trehalose and the activity of cell nitrogen content level and enzyme all has certain influence.The content of the trehalose of general active dry yeast reaches about 14% of dry-matter, and therefore, culturing yeast technology must be considered the accumulation degree of trehalose.The yeast trehalose synthesis need be through two stages, and the one, in a large amount of breeding stages of yeast cell, this stage yeast body intracellular trehalose content is lower, but the thalline of capacity can be provided; The 2nd, the accumulation stage of trehalose mainly is by control ambient conditions it to be accumulated.
Summary of the invention
The present invention is directed to the deficiencies in the prior art, propose a kind of method that improves trehalose content of yeast cell, can realize in the short period of time that yeast cell intracellular trehalose content reaches more than 16%.
The term explanation:
The vvm of unit of ventilation is meant the volume ratio of the amount of air capacity that per minute feeds and fermented liquid.
Cell concn: be meant in the fermented liquid or the mass concentration of yeast saccharomyces cerevisiae in the yeast-lactic.
" bread yeast " mentioned among the present invention is the product trade(brand)name of being produced by the yeast saccharomyces cerevisiae bacterial classification.
Technical scheme of the present invention is as follows:
A kind of method that improves trehalose content of yeast cell, the yeast-lactic that the yeast fermentation medium centrifugal is separated the cell concn that obtains 13-17%, in yeast-lactic, add maltose, the aqueous solution of sodium-chlor and calcium chloride, the mass volume ratio of yeast-lactic and water is 1: 1, maltose, the consumption of sodium-chlor and calcium chloride accounts for yeast-lactic total mass per-cent and is respectively 2.4-3.0% maltose, 3.2-4.0% sodium-chlor and 1.0% calcium chloride, at 35 ℃, pH5.2,1.0-1.2vvm cultivated 2-3 hour under the ventilation condition, centrifugal collection yeast slurry, measure content of trehalose 16-17% in the yeast, with the yeast cell dry weight basis.Described mass volume ratio, unit are g/mL or kg/L.Described yeast is a yeast saccharomyces cerevisiae.
The accumulation volume that can improve trehalose is cultivated in the interpolation of maltose in the method for the present invention, and this is because yeast cell contains the maltose isomerase and maltose can be converted into trehalose at short notice.
In the method for the invention described above, be starting raw material with the yeast fermentation nutrient solution, can be about the preparation of yeast fermentation nutrient solution by prior art.The invention provides the preparation method of following yeast fermentation nutrient solution, as of the present invention one of preferred:
1. substratum
(1) seed culture medium: glucose 20g, KH
2PO
41g, (NH
4)
2SO
45g, MgSO
47H
2O 0.2g, ZnSO
47H
2O 0.05g, pH5.4, distilled water 1000mL.
(2) fermention medium
1. basic medium ferments
Corn steep liquor (containing solid substance 28wt%) 20%, glucose 1%, nutrient salt solution 5%, water 74% is volume ratio.Wherein, nutrient salt solution: KH
2PO
413.0g, (NH
4)
2SO
494.16g, MgSO
47H
2O 1.4g, ZnSO
47H
2O0.28g adds water and is made into 1000mL.
2. fermenting process stream adds component
Sugar soln: glucose is made into the solution of 250g/L.
Nutrient salt solution: KH
2PO
413.0g, (NH
4)
2SO
494.16g, MgSO
47H
2O 1.4g, ZnSO
47H
2O0.28g adds water and is made into 1000mL.
The Na of alkali lye: 100g/L
2CO
3Solution.
2. seed culture
Barms is available from Institute of Microorganism, Academia Sinica, title: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae), and code name: AS 2.146, AS 2.502, AS 2.503 or AS 2.504.With the yeast slant strains, insert in the 250mL triangular flask of dress 20mL seed culture medium, behind 30 ℃ of 200r/min shake-flask culture 24h, change the 500mL triangular flask that the 80mL seed culture medium is housed over to, 30 ℃ of 200r/min shake-flask culture 16h obtain seed culture fluid, and are standby.
3. fermentation culture
Carry out reality in the 50L fermentor tank of 25L fermentation basic medium and jar sterilize being equipped with, sterilising conditions is 120 ℃, 30min.When being cooled to 32 ℃ then, insert seed culture fluid 3L, regulate pH5.2,30 ℃ of control culture temperature, stream sugared soln and nutrient salt solution, the control total fermentation time is 18-21h, nutrient salt solution 3-5 before fermentation ends added in individual hour, the stream dosage of sugar soln is according to the yeast growth situation, makes that glucose content is controlled at below 0.5% always in the fermentation system.In the fermenting process, control pH5.2, mixing speed is controlled at 200r/min.Other processing condition are preferably as follows:
Culture temperature control: 30 ℃ of fermentation time 1-8h culture temperature, 32 ℃ of 9-15h culture temperature, 16h is to 35 ℃ of fermentation ends culture temperature.
Air flow: at fermentation (preceding, mid-term) time 1-15h control sterile air volume with fermentating liquid volume than per minute at 2.5vvm, in the trehalose accumulation stage of fermentation (later stage) time 16h, adjust valve to fermentation ends, air flow is dropped to 1.2vvm.
Preferably, in the above method fermentation culture step, control sugar soln dosage makes glucose content 0.30-0.50% volume ratio in the fermentation system; The pH regulator agent is 10wt%Na
2CO
3Solution.
The yeast fermentation nutrient solution that above method makes, cell concn 4.5-5.5%, wherein content of trehalose is 8-10% after measured.Further be used as the raw material of the inventive method.
Content of trehalose measuring method in the yeast
In yeast cell, contain mannosans and insoluble dextran, can react with anthrone reagent.Therefore, selecting for use suitable extraction agent to control the interference that these materials measure trehalose is the key that trehalose is measured in the yeast.Yeast extracts with the cold trichoroacetic acid(TCA) of 0.5mol/L, only has trehalose to exist in the gained solution.With trichoroacetic acid(TCA) extracting solution and 0.2% sulfuric acid anthrone solution mixed boiling water bath reaction 2min with 1: 4, and in the light absorption value of 590nm place measured reaction liquid.The content that calculates trehalose in the extracting solution is the trehalose amount in the yeast.
Adopt ply of paper to analyse the separation elution method: when analysing the quantitative analysis trehalose with ply of paper, two samples being put on the filter paper bar are same sample, and the point sample amount is controlled at 40～100 μ g, and one of them point is reference point, and another is the quantitative measurment point.With the filter paper bar behind the point sample at the developping agent propyl carbinol: acetone: water: acetic acid=10: 3: 6: launch among 2 (v/v), air-dry, and it is cut off from the centre.Half spraying colour developing of band reference point, to determine the accurate position of trehalose spot, with on second half filter paper bar therewith the part of spot correspondence cut and be cut into fragment, put into elutriant 0.1mol/LHCl room temperature wash-out 16h, supernatant liquor wherein content of trehalose of sulfuric acid-anthrone colorimetric method for determining behind the wash-out.
Method of the present invention is as follows by the technical characterstic and the excellent results of yeast culture accumulation trehalose:
1. temperature: 30 ℃ of earlier fermentation (1-8h) culture temperature, mid-term, (9-15h) culture temperature was 32 ℃, 35 ℃ of later stage (16h is to fermentation ends) culture temperature.
2. pH: in the zymic culturing process, stream adds 10%Na
2CO
3Solution is regulated pH5.2.
3. dissolved oxygen: in the zymic culturing process, under the constant situation of air flow, improve the oxygen transfer rate that stir speed (S.S.) can effectively improve fermentor tank, thereby guarantee cell growth and the required dissolved oxygen supply of metabolism.In the inventive method, mixing speed is controlled at 200r/min, and air flow 1-15h is controlled at 2.5vvm, is the trehalose accumulation stage at fermentation time 16h to fermentation ends, and air flow is dropped to 1.2vvm.
4. after yeast culture obtains a large amount of cells, the process centrifugation obtains the yeast-lactic of the concentration of 13-17%, adds maltose in yeast-lactic, and adds sodium-chlor, calcium chloride, under 35 ℃ of ventilation conditions, cultivated 2-3 hour, to realize high concentration of trehalose accumulation in the yeast.The present invention can within a short period of time, and the trehalose in the yeast cell is brought up to more than 16% from 8%.Existing general technology has been dwindled fermentation time.
Because raw material of the present invention is a yeast-lactic, barm cell concentration is 13-17%, and zymic concentration 4.5-5.5% in the yeast fermentation broth, thereby can save fermentation equipment and power consumption.
Description of drawings
Fig. 1 the inventive method embodiment 1 yeast-lactic with salt, add trehalose accumulation under the situation of maltose;
Fig. 2 is with content of trehalose in sulfuric acid-anthrone colorimetric method for determining embodiment 1 yeast slurry: when analysing the quantitative analysis trehalose with ply of paper, two samples being put on the filter paper bar are same sample, the point sample amount is controlled at 40～100 μ g, and one of them point is reference point, and another is the quantitative measurment point.
Embodiment
Below in conjunction with embodiment, technical solution of the present invention is further elaborated, but institute of the present invention protection domain is not limited only to this.Used bacterial classification and substratum are among the embodiment:
Yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) bacterial classification is available from Institute of Microorganism, Academia Sinica, and DSMZ of Institute of Microorganism, Academia Sinica bacterial classification catalogue code name: AS 2.146; AS 2.502; AS 2.503; AS 2.504.
Seed culture medium: glucose 20g, KH
2PO
41g, (NH
4)
2SO
45g, MgSO
47H
2O 0.2g, ZnSO
47H
2O 0.05g, pH5.4, distilled water 1000mL.
Fermention medium: comprise that fermentation basic medium and fermenting process stream add component, as previously mentioned.
Barms: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) AS 2.146.With the yeast slant strains, insert in the 250mL triangular flask of dress 20mL seed culture medium, behind 30 ℃ of 200r/min shake-flask culture 24h, change the 500mL triangular flask that the 80mL seed culture medium is housed over to, 30 ℃ of 200r/min shake-flask culture 16h obtain seed culture fluid, and are standby.
Above-mentioned seed culture fluid 3L is inserted in the 25L fermentation basic medium of having sterilized (fermentor tank volume 50L), and culture condition is as follows.1. temperature: at the 1-8h of fermentation in the time, 30 ℃ of culture temperature, in the time of fermentation 9-15h, 32 ℃ of culture temperature, at fermentation 16-20h in the time, 35 ℃ of culture temperature.2. pH: in the zymic culturing process, stream adds 10%Na
2CO
3Solution is regulated pH5.2.3. dissolved oxygen: mixing speed is controlled at 200r/min, and air flow 1-16h is controlled at 2.5vvm, in the time, by valve air flow is dropped to 1.2vvm at fermentation 16-20h.Stream sugared soln and nutrient salt solution, nutrient salt solution stream adds total amount 2.5L, 16h adds at fermentation time, sugar soln then carries out stream according to ethanol feedback and zymic growing state and adds, make that glucose content is controlled at 0.40-0.45% (volume ratio) always in the fermented liquid, the 20h fermentation ends, yeast cell 4.78% (dry weight) in the gained fermented liquid, content of trehalose 8.2%.
With above-mentioned fermented liquid, use the large vol whizzer, at 3500r/min, under the condition of time 10min, centrifugal collection yeast-lactic.Abandon the centrifuge tube supernatant liquor, take out yeast-lactic.In 1000mL distilled water, add sodium-chlor 40g, calcium chloride 10g, maltose 30g, dissolving fully, 100 ℃ of sterilizations with water-bath temperature control to 30 ℃, are added in the 1000g yeast-lactic, place the 10L fermentor tank.35 ℃ of control culture temperature are used 10wt%Na
2CO
3Solution is regulated pH5.2; 1.20vvm under the ventilation condition, incubation time 3 hours, centrifugal collection yeast slurry, content of trehalose is 16.88% in the mensuration yeast cell dry weight.
Barms: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) AS 2.502.The seed culture fluid preparation is with embodiment 1.
With the yeast slant strains, insert in the 250mL triangular flask of dress 20mL seed culture medium, behind 30 ℃ of 200r/min shake-flask culture 24h, change the 500mL triangular flask that the 80mL seed culture medium is housed over to, 30 ℃ of 200r/min shake-flask culture 16h obtain seed culture fluid, and are standby.
Above-mentioned seed culture fluid 3L is inserted in the 25L fermentation basic medium of having sterilized (fermentor tank volume 50L), and culture condition is as follows.1. temperature: at the 1-8h of fermentation in the time, 30 ℃ of culture temperature, in the time of fermentation 9-15h, 32 ℃ of culture temperature, at fermentation 16-19h in the time, 35 ℃ of culture temperature.2. pH: in the zymic culturing process, stream adds 10%Na
2CO
3Solution is regulated pH5.2.3. dissolved oxygen: mixing speed is controlled at 200r/min, and air flow 1-16h is controlled at 2.5vvm, in the time, by valve air flow is dropped to 1.2vvm at fermentation 16-20h.Stream sugared soln and nutrient salt solution, nutrient salt solution stream adds total amount 2.5L, 16h adds at fermentation time, sugar soln then carries out stream according to ethanol feedback and zymic growing state and adds, make that glucose content is controlled at 0.40-0.48% (volume ratio) always in the fermented liquid, the 19h fermentation ends, yeast cell 4.56% (dry weight) in the gained fermented liquid, content of trehalose 9.0%.
With above-mentioned fermented liquid,, collect yeast-lactic in the centrifugal 10min of 3500r/min.Abandon the centrifuge tube supernatant liquor, take out yeast-lactic.In 1000mL distilled water, add sodium-chlor 32g, calcium chloride 10g, maltose 24g, dissolving fully, 100 ℃ of sterilizations with water-bath temperature control to 30 ℃, are added in the 1000g yeast-lactic, place the 10L fermentor tank.35 ℃ of control culture temperature are used 10%Na
2CO
3Solution is regulated pH5.2; 1.20vvm under the ventilation condition, incubation time 3 hours, centrifugal collection yeast slurry, content of trehalose is 16.00% in the mensuration yeast cell dry weight.
Embodiment 3
Barms: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) AS 2.503.The seed culture fluid preparation is with embodiment 1.
With the yeast slant strains, insert in the 250mL triangular flask of dress 20mL seed culture medium, behind 30 ℃ of 200r/min shake-flask culture 24h, change the 500mL triangular flask that the 80mL seed culture medium is housed over to, 30 ℃ of 200r/min shake-flask culture 16h obtain seed culture fluid, and are standby.
Above-mentioned seed culture fluid 3L is inserted in the 25L fermentation basic medium of having sterilized (fermentor tank volume 50L), and culture condition is as follows.1. temperature: at the 1-8h of fermentation in the time, 30 ℃ of culture temperature, in the time of fermentation 9-15h, 32 ℃ of culture temperature, at fermentation 16-20h in the time, 35 ℃ of culture temperature.2. pH: in the zymic culturing process, stream adds 10%Na
2CO
3Solution is regulated pH5.2.3. dissolved oxygen: mixing speed is controlled at 200r/min, and air flow 1-16h is controlled at 2.5vvm, in the time, by valve air flow is dropped to 1.2vvm at fermentation 16-20h.Stream sugared soln and nutrient salt solution, nutrient salt solution stream adds total amount 2.5L, 16h adds at fermentation time, sugar soln then carries out stream according to ethanol feedback and zymic growing state and adds, make that glucose content is controlled at 0.40-0.45% (volume ratio) always in the fermented liquid, the 20h fermentation ends, yeast cell 5.72% (dry weight) in the gained fermented liquid, content of trehalose 7.72%.
With above-mentioned fermented liquid at the centrifugal 10min of 3500r/min,, centrifugal collection yeast-lactic.Abandon the centrifuge tube supernatant liquor, take out yeast-lactic.In 1000mL distilled water, add sodium-chlor 36g, calcium chloride 10g, maltose 26g, dissolving fully, 100 ℃ of sterilizations with water-bath temperature control to 30 ℃, are added in the 1000g yeast-lactic, place the 10L fermentor tank.35 ℃ of control culture temperature are used 10%Na
2CO
3Solution is regulated pH5.2; 1.20vvm under the ventilation condition, incubation time 3 hours, centrifugal collection yeast slurry, content of trehalose is 16.01% in the mensuration yeast cell dry weight.
Embodiment 4
Barms: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) AS 2.504.The seed culture fluid preparation is with embodiment 1.
With the yeast slant strains, insert in the 250mL triangular flask of dress 20mL seed culture medium, behind 30 ℃ of 200r/min shake-flask culture 24h, change the 500mL triangular flask that the 80mL seed culture medium is housed over to, 30 ℃ of 200r/min shake-flask culture 16h obtain seed culture fluid, and are standby.
Above-mentioned seed culture fluid 3L is inserted in the 25L fermentation basic medium of having sterilized (fermentor tank volume 50L) culture condition such as embodiment 1,20h fermentation ends, yeast cell 5.11% (dry weight) in the gained fermented liquid, content of trehalose 6.2%.
Above-mentioned fermented liquid at the centrifugal 10min of 3500r/min, is collected yeast-lactic.Abandon the centrifuge tube supernatant liquor, take out yeast-lactic.In 1000mL distilled water, add sodium-chlor 38g, calcium chloride 10g, maltose 28g, dissolving fully, 100 ℃ of sterilizations with water-bath temperature control to 30 ℃, are added in the 1000g yeast-lactic, place the 10L fermentor tank.35 ℃ of control culture temperature are used 10%Na
2CO
3Solution is regulated pH5.2; 1.20vvm under the ventilation condition, incubation time 3 hours, centrifugal collection yeast slurry, content of trehalose is 16.22% in the mensuration yeast cell dry weight.
Claims (4)
1. method that improves trehalose content of yeast cell, the yeast-lactic that the yeast fermentation medium centrifugal is separated the cell concn that obtains 13-17%, in yeast-lactic, add maltose, the aqueous solution of sodium-chlor and calcium chloride, the mass volume ratio of yeast-lactic and water is 1: 1, maltose, the consumption of sodium-chlor and calcium chloride accounts for yeast-lactic total mass per-cent and is respectively 2.4-3.0% maltose, 3.2-4.0% sodium-chlor and 1.0% calcium chloride, at 35 ℃, pH5.2,1.0-1.2vvm cultivated 2-3 hour under the ventilation condition, centrifugal collection yeast slurry, measure content of trehalose 16-17% in the yeast, with the yeast cell dry weight basis;
Described yeast is a yeast saccharomyces cerevisiae;
Described mass volume ratio, unit are g/mL or kg/L.
2. the method for raising trehalose content of yeast cell as claimed in claim 1 is characterized in that the preparation of described yeast fermentation nutrient solution, and step is as follows:
(1) substratum
Seed culture medium: glucose 20g, KH
2PO
41g, (NH
4)
2SO
45g, MgSO
47H
2O 0.2g, ZnSO
47H
2O 0.05g, pH5.4, distilled water 1000mL;
Fermention medium:
1. basic medium ferments
Corn steep liquor (containing solid substance 28wt%) 20%, glucose 1%, nutrient salt solution 5%, water 74% is volume ratio; Wherein, the nutrient salt solution prescription is KH
2PO
413.0g, (NH
4)
2SO
494.16g, MgSO
47H
2O 1.4g, ZnSO
47H
2O0.28g adds water and is made into 1000mL;
2. fermenting process stream adds component
Sugar soln: glucose is made into the solution of 250g/L; Nutrient salt solution: KH
2PO
413.0g, (NH
4)
2SO
494.16g, MgSO
47H
2O 1.4g, ZnSO
47H
2O 0.28g adds water and is made into 1000mL; The Na of alkali lye: 100g/L
2CO
3Solution;
(2) seed culture
Barms is yeast saccharomyces cerevisiae (Saccharomyces cerevisiae), code name: AS 2.146, and AS 2.502, AS 2.503 or AS 2.504; With the yeast slant strains, insert in the 250mL triangular flask of dress 20mL seed culture medium, 30 ℃ of 200r/min shake-flask culture 24h change the 500mL triangular flask that the 80mL seed culture medium is housed over to, and 30 ℃ of 200r/min shake-flask culture 16h obtain seed culture fluid, and are standby;
(3) fermentation culture
Carry out reality in the 50L fermentor tank of 25L fermentation basic medium and jar sterilize being equipped with, sterilising conditions is 120 ℃, 30min; When being cooled to 32 ℃ then, insert seed culture fluid 3L, regulate pH5.2,30 ℃ of control culture temperature, stream sugared soln and nutrient salt solution, the control total fermentation time is 18-21h, nutrient salt solution 3-5 before fermentation ends added in individual hour, the stream dosage of sugar soln is according to the yeast growth situation, makes that glucose content is controlled at below 0.5% always in the fermentation system; In the fermenting process, control pH5.2, mixing speed is controlled at 200r/min.
3. the method for raising trehalose content of yeast cell as claimed in claim 2, it is characterized in that the control of culture temperature described in step (3) fermentation culture is as follows: 30 ℃ of fermentation time 1-8h culture temperature, 32 ℃ of 9-15h culture temperature, 16h is to 35 ℃ of fermentation ends culture temperature.
4. the method for raising trehalose content of yeast cell as claimed in claim 2, it is characterized in that air flow in step (3) fermentation culture: at fermentation time 1-15h control sterile air volume with fermentating liquid volume than per minute at 2.5vvm, in the trehalose accumulation stage of fermentation time 16h to fermentation ends, adjust valve, air flow is dropped to 1.2vvm.
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| CN105861344A (en) * | 2016-05-30 | 2016-08-17 | 湖北工业大学 | Synchronous culture method for improving yeast biomass and intracellular trehalose content |
| CN106399423A (en) * | 2016-09-14 | 2017-02-15 | 天津替代医学科技股份有限公司 | Method for preparing trehalose by using waste beer yeast under adverse stress conditions |
| CN109868229A (en) * | 2019-03-11 | 2019-06-11 | 江南大学 | A method of keeping yeast activity |
| CN110656080A (en) * | 2019-11-22 | 2020-01-07 | 顾霆 | Directional culture method of yeast cells |
| CN113151367A (en) * | 2021-05-10 | 2021-07-23 | 湖北工业大学 | Fermentation method for synthesizing xylitol de novo by self-protected zygosaccharomyces rouxii |
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| CN105861344A (en) * | 2016-05-30 | 2016-08-17 | 湖北工业大学 | Synchronous culture method for improving yeast biomass and intracellular trehalose content |
| CN106399423A (en) * | 2016-09-14 | 2017-02-15 | 天津替代医学科技股份有限公司 | Method for preparing trehalose by using waste beer yeast under adverse stress conditions |
| CN106399423B (en) * | 2016-09-14 | 2019-11-29 | 中美食品有限公司 | A method of trehalose is prepared under the conditions of environment stress using beer waste yeast |
| CN109868229A (en) * | 2019-03-11 | 2019-06-11 | 江南大学 | A method of keeping yeast activity |
| CN110656080A (en) * | 2019-11-22 | 2020-01-07 | 顾霆 | Directional culture method of yeast cells |
| CN113151367A (en) * | 2021-05-10 | 2021-07-23 | 湖北工业大学 | Fermentation method for synthesizing xylitol de novo by self-protected zygosaccharomyces rouxii |
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