CN108865910A - A kind of application of wine waste mash, screening technique and the saccharomycete in blueberry red wine fermentation - Google Patents
A kind of application of wine waste mash, screening technique and the saccharomycete in blueberry red wine fermentation Download PDFInfo
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Abstract
The present invention relates to a kind of wine waste mash, deposit number is CGMCC NO.15931.The invention further relates to the screening technique of the saccharomycete and the applications in blueberry red wine fermentation.Best in quality by the resulting blueberry red wine of the saccharomycetes to make fermentation, clear, color is orange red, has typical red wine flavor.Blueberry Production of fruit is that red wine industry opens good development opportunity, contain the nutritional ingredients such as a large amount of amino acid, vitamin, minerals in fermented wine, and contain certain physiological activator, mainly have the function of relaxing tendons and activating collaterals, boost metabolism, is suitble to drink.
Description
Technical field
The present invention relates to microorganisms technical fields, and in particular to a kind of wine waste mash, screening technique and the ferment
Application of female bacterium in blueberry red wine fermentation.
Background technique
During red wine brewing, saccharomycete plays the role of very important.Specifically, the quality of saccharomycete is direct
Influence red wine quality, it is therefore desirable to increase the research dynamics to saccharomycete.
Carbohydrate is the highest ingredient of content in red wine, and saccharomycete has run through the whole process of red wine brewing, in fermentation
Under produce a large amount of fragrance matter and alcohol, this directly influences the quality of red wine.Therefore in order to improve the mouthfeel of red wine, need
Will form to fermentation strain and screening study, promote the sustainable development of red wine industry.But current wine brewing is used
Saccharomycete is primarily directed to grape wine, and for other plant materials, such as blueberry, and the saccharomycete not being adapted is for red
The fermentation process of wine.
In consideration of it, the present invention is specifically proposed.
Summary of the invention
The first object of the present invention is to provide a kind of wine waste mash.
The second object of the present invention is to provide the screening technique of the saccharomycete.
The third object of the present invention is to provide application of the saccharomycete in blueberry red wine fermentation.
To achieve the above object, technical scheme is as follows：
The present invention relates to a kind of wine waste mash, deposit number is CGMCC NO.15931.
The invention further relates to the screening techniques of the wine waste mash, include the following steps：
(1) blueberry juice is taken fermentation liquid to be diluted with sterile water, is diluted after 25~30 DEG C ferment 2~4 days
Liquid；
(2) dilution is coated in rose bengal medium, after 25~30 DEG C are cultivated 2~4 days, is randomly selected
Single colonie with products of typical yeast colony characteristics isolates and purifies 2~3 times, isolated yeast strain；
(3) isolated yeast strain is subjected to gas generation property analysis, fermenting speed analysis, the analysis of strain coherency and carbon source
Using analysis, filters out the wine brewing and use yeast strain.
Preferably, in step (1), the blueberry juice after cleaning, select by Blueberry, squeezing the juice by obtaining.
Preferably, sulfur dioxide is passed through in the juice-extracting process, content of the sulfur dioxide in the blueberry juice is 30
~70mg/L.
Preferably, in the step (1), the volume ratio of fermentation liquid and sterile water is 1：(3~8), preferably 1:5.
Preferably, in the step (1) and (2), fermentation and cultivation temperature are 28 DEG C, and the time is 3 days.
Preferably, in the step (3), gas generation property analysis uses Du Shi pipe fermentation method, preferably by isolated saccharomycete
Built in strain access 10ml in the YPD liquid medium of Du Shi tubule, inoculum concentration 20-40CFU, ferment 72h at 28 DEG C, measures ferment
Mother strains gas production.
Preferably, in the step (3), fermenting speed analysis includes：Isolated yeast strain is accessed in blueberry juice,
After 28 DEG C are cultivated for 24 hours, ferment 6~8d at 23 DEG C, measures the sugar consumption amount in blueberry juice, and identify in fermentation process and produce
Raw fragrance.
Preferably, in the step (3), the analysis of strain coherency includes：
(i) by isolated yeast strain access Tissue Culture Flask, 48h is placed at 28 DEG C, after distillation water washing centrifugation
Remove clear liquid, obtains wet yeast；
(ii) the Acetate Solution 12ml that the wet yeast of 1g and pH are 3~4 is placed in centrifuge tube, then centrifuge tube exists
15~20min is stood at 20 DEG C；
(iii) centrifuge tube is rocked, so that saccharomycete is suspended in centrifuge tube uniformly, suspension is then stood into 10min, is seen
It observes saccharomycete to precipitate, records ml of saccharomycete precipitating at this time.
Preferably, in the step (3), utilization of carbon source analysis includes：By isolated yeast strain access different carbon source training
It supports in base, is cultivated at 28 DEG C and measure carbon source content afterwards for 24 hours, be determined to the carbon source utilized.The carbon source culture medium include sucrose,
Maltose, raffinose, lactose, galactolipin, cellobiose, lemon drops, inositol and soluble starch culture medium.
The invention further relates to application of the saccharomycete in blueberry red wine fermentation.
Beneficial effects of the present invention：
The present invention is studied by form to saccharomycete, physiological characteristic etc., and it is strong to filter out a kind of fermentability,
The good wine waste mash of fermenting property simultaneously.Best in quality, the clear by the resulting blueberry red wine of the saccharomycetes to make fermentation,
Color is orange red, has typical red wine flavor.Blueberry Production of fruit is that red wine industry opens good development opportunity, fermented wine
In containing the nutritional ingredients such as a large amount of amino acid, vitamin, minerals, and contain certain physiological activator, mainly have
Relaxing tendons and activating collaterals, the effect to boost metabolism, are suitble to drink.
Specific embodiment
To make the object, technical solutions and advantages of the present invention clearer, technical solution of the present invention will be carried out below
Detailed description.Obviously, described embodiments are only a part of the embodiments of the present invention, instead of all the embodiments.Base
Embodiment in the present invention, those of ordinary skill in the art are obtained all without making creative work
Other embodiment belongs to the range that the present invention is protected.
The present embodiments relate to a kind of saccharomyces cerevisiae, deposit number is CGMCC NO.15931.The saccharomyces cerevisiae
In (Saccharomyces cerevisiae) entitled triumphant edge spring No. 5, separated from the blueberry surface in Guizhou Province's Majiang area
It arrives.The strain has been saved in China Committee for Culture Collection of Microorganisms's common micro-organisms center.Preservation address is north
The institute 3 of the Chaoyang District Jing Shi North Star West Road 1, preservation time are on June 11st, 2018.
The embodiment of the present invention further relates to the screening technique of the wine waste mash, includes the following steps：
(1) blueberry juice is taken fermentation liquid to be diluted with sterile water, is diluted after 25~30 DEG C ferment 2~4 days
Liquid.
In one embodiment of the invention, blueberry juice after cleaning, select by Blueberry, squeezing the juice by obtaining.It squeezes
Sulfur dioxide is passed through during juice, content of the sulfur dioxide in blueberry juice is 30~70mg/L.The present invention selects Guizhou fiber crops
The blueberry in river area.
In one embodiment of the invention, the volume ratio of fermentation liquid and sterile water is 1：(3~8), preferably 1:5.Dilution
Effect be to keep the yeast bacterial content in culture solution moderate, conducive to the primary dcreening operation culture of next step.
(2) dilution is coated in rose bengal medium, after 25~30 DEG C are cultivated 2~4 days, randomly selecting has
The single colonie of products of typical yeast colony characteristics isolates and purifies 2~3 times, isolated yeast strain.
Rose bengal medium ingredient is：Peptone 5g, glucose 10g, potassium dihydrogen phosphate 1g, magnesium sulfate (MgSO4·
7H2O) 0.5g, agar 20g, 1/3000 rose-bengal solution 100mL, distilled water 1000mL, chloramphenicol 0.1g.Preparation method is：
Above-mentioned each ingredient is added in distilled water after dissolving, adds rose-bengal solution, 121 DEG C of sterilizing 15min after packing.
In one embodiment of the invention, in step (1) and (2), fermentation and cultivation temperature are 28 DEG C, and the time is
3 days.
Step (1) and (2) are collectively referred to as primary dcreening operation, act on to obtain the bacterium colony that different saccharomycete generate in the medium, and right
Strain in single bacterium colony successively carries out the test analysis such as gas generation property, fermenting speed, obtains having optimum performance with screening
Saccharomycete.Above-mentioned single colonie has different shape.Wherein, white strain bacterium colony is circle, locally there is lens-shaped protuberance, edge
Completely, surface is dim, opaque；Cream-colored strain bacterium colony is circle, locally has lens-shaped protuberance, edge is complete, surface light
It is bright, it is transparent；Faint yellow strain bacterium colony is spindle, and part is flat, and edge is complete, surface-brightening, transparent.Cream-colored strain bacterium
In irregular shape, edge presentation hedgehog is fallen, locally there is lens-shaped protuberance, surface dimness is opaque.
(3) isolated yeast strain is subjected to gas generation property analysis, fermenting speed analysis, the analysis of strain coherency and carbon source
Using analysis, filters out wine brewing and use yeast strain.
In one embodiment of the invention, gas generation property analysis uses Du Shi pipe fermentation method in step (3), will preferably divide
From yeast strain access 10ml built in Du Shi tubule YPD liquid medium in, inoculum concentration 20-40CFU is issued at 28 DEG C
Ferment 72h measures yeast strain gas production.
Wherein, YPD culture medium is yeast extract peptone glucose agar medium, the culture for saccharomycete.Formula is：1%
Yeast Extract (yeast extract), 2%Peptone (peptone), 2%Dextrose (glucose) (glucose), if system is solid
Body culture medium adds 2% agar powder.
Learnt according to professional experiences, if Du Shi pipe in 72h there are no by saccharomycete and its producing gas and being full of, the degree of fermentation is logical
Often be unable to satisfy the basic demand of blueberry red wine brewing, therefore corresponding bacterial strain can be given up, and will ferment preferable bacterial strain into
Row comparison screening again.It finds that the gas generation property of triumphant No. 5 strains of edge spring is most strong in this course, illustrates its fermentation efficiency and hair
Ferment degree is high.
In one embodiment of the invention, fermenting speed analysis includes in step (3)：Isolated yeast strain is accessed
In blueberry juice, after 28 DEG C are cultivated for 24 hours, ferment 6~8d at 23 DEG C, measures the sugar consumption amount in blueberry juice, and identify
The fragrance generated in fermentation process.This process can filter out excellent, healthy and strong bacterial strain, while find the production perfume (or spice) ability of the bacterial strain
By force, suitable for the fermentation process of blueberry red wine.
In one embodiment of the invention, the analysis of strain coherency includes in step (3)：
(i) by isolated yeast strain access Tissue Culture Flask, 48h is placed at 28 DEG C, distilled water progress can be used
Centrifuge washing more than twice, then removes clear liquid, obtains wet yeast；
(ii) Acetate Solution (such as aqueous solution of sodium acetate or potassium acetate) for being 3~4 by the wet yeast of 1g and pH value
12ml is placed in in graduated centrifuge tube, then centrifuge tube is stood to 15~20min at 20 DEG C；
(iii) centrifuge tube is rocked, so that saccharomycete is suspended in centrifuge tube uniformly, suspension is then stood into 10min, is seen
It observes saccharomycete to precipitate, records ml of saccharomycete precipitating at this time, number is bigger, and its dispersibility is better.
In one embodiment of the invention, in step (3), utilization of carbon source analysis includes：Isolated yeast strain is connect
Enter in different carbon source culture medium, is cultivated at 28 DEG C and measure carbon source content afterwards for 24 hours, to be determined to the carbon source utilized.Carbon source culture
Base includes sucrose, maltose, raffinose, lactose, galactolipin, cellobiose, lemon drops, inositol and soluble starch culture medium.
Wherein, carbon source liquid medium includes 0.2 gram of magnesium sulfate, 1 gram of ammonium dihydrogen phosphate, 1 gram of dipotassium hydrogen phosphate, 5 grams of carbon source, above-mentioned object
Matter is dissolved in 1000ml sterile water.Wherein, magnesium ion is the confactor in all kinds of metabolism, and ammonium dihydrogen phosphate is nitrogen source, phosphoric acid
Hydrogen dipotassium is buffer, and above-mentioned carbohydrate is carbon source.When yeast strain is inoculated in the liquid medium, saccharomycete can utilize phosphoric acid
Ammonium dihydrogen grows in liquid medium as sole carbon source as only nitrogen source, certain carbohydrate and produces gas.
In experimental identification, it is found by the applicant that the triumphant edge spring, No. 5 strains were in the carbon sources culture medium such as sucrose, maltose, raffinose
In become muddy, illustrate that No. 5 strains of triumphant edge spring can be very good to utilize these carbon sources.No. 5 strains of triumphant edge spring are in lactose, half simultaneously
Will not be muddy in the carbon sources culture medium such as lactose, cellobiose, lemon drops, inositol, soluble starch, illustrate that the bacterial strain can not utilize
These carbon sources.In sugared fermenting experiment qualification process, the triumphant edge spring, No. 5 strains had in the culture mediums such as glucose sugar, sucrose, maltose
Bubble generates, this is enough to illustrate that No. 5 strains of triumphant edge spring can use these types of carbon source.Link, application are identified in other physiology
People has found that No. 5 strains of triumphant edge spring can be grown in the glucose that concentration is 50%, and has very strong glucose resisting high-concentration
Performance.Color is constant in starch compound, and this demonstrate No. 5 strains of triumphant edge spring can not be healthy and strong in the environment of being deficient in vitamin
Growth, this also illustrates vitamin is not the growth factor of triumphant No. 5 strains of edge spring.
Applicant has also carried out isolated yeast strain the analysis of sensory evaluation.Two bacterial strains are specially selected, it is right
Analysis is made in its alcoholic strength, pol, sensory evaluation etc., it is found that this two plants of strains grow fine, fermenting property is fabulous.It is commented in sense organ
Mainly to the liquor-producing ability of bacterial strain, alcoholic strength, the evaluation for producing the science of making such as fragrant ability during valence.It is found in this process：
The fermentability of No. 2 strains of triumphant edge spring and triumphant No. 5 strains of edge spring and the fragrant ability of production are stronger, produce fragrant ability and liquor-producing ability all has
There is apparent advantage, therefore they have good utility value.
The invention further relates to application of the saccharomycete in blueberry red wine fermentation.
Alcohol content changes in fermentation process：During the fermentation, the total reducing sugar in blueberry juice, ethyl alcohol, total acid content be all
Present different variations.Further, total sugar content is 10.9% in blueberry juice Normal juice, is unable to satisfy the finished product of red wine
It is required that.According to the rules, alcoholic strength content should be 11.2%~12.0% in finished product.Hair after saccharomycete is added to blueberry juice
Ferment the 4th day, 9.3% sugar is added to it, finds that the activity of saccharomycete is higher and higher, and liquor-producing ability increases at fermentation initial stage therewith
By force.With the decline of total sugar content, such as ferment the 10th day in fermentation whole process, total reducing sugar is only 2.9g/L, this explanation
Fermentation process has been completed.The content for needing to calculate total acid in yeast in actual production in time, in ferment middle, total acid content
It is gradually increasing, while producing the products such as acid, pyruvic acid, acetic acid cream and having arrived the fermentation later period, polyacid becomes monoacid, total acid
Content decline, in this link, pH value also declines therewith, but variation is little on the whole.After fermentation, blueberry wine base essence body
Fraction has had reached 12.1%, and wherein pH value is 2.98, and total acid also reaches 7.95g/L, and it is real that these indexs have reached fermentation
Test the main technical requirements of screening and blueberry red wine production.
Tannin content changes in fermentation process：
In Blueberry, anthocyanin and tannin are main nutritional ingredients.But during the fermentation, anthocyanin contains
Amount has dropped, and development trend presents the form of slip, and this variation shows unobvious in premise.Until fermentation
The downward trend of mid-term, anthocyanin content is gradually steady, and the storage rate that ferments at this time is 65%.Anthocyanin and fresh fruit there are environment
Somewhat like, since the oxygen demand of saccharomycete is larger, while fermenting step oxygen content is low, therefore in the increased situation of acidity,
The speed of growth of saccharomycete is restricted, and produces a large amount of alcohol, seldom to the consume of oxygen content, so that oxidation is gradually
Enhancing.
Therefore, the present invention is using blueberry red wine as research object, and after the fruit of blueberry is squeezed the juice, being placed on interior sends out it naturally
Ferment, while respectively shaking sooner or later daily primary.The yeast of different numbers is subjected to sugar degree regulation respectively, is sent out in microscopy after separating three times
Existing, these bacterium colonies are purebred.Form and physiological characteristic to yeast carry out scientific and effective analysis and identification, triumphant edge spring No. 5
The gas generation property of strain is most strong, and fermentation efficiency is high, and the degree of fermentation is high.During the fermentation, total reducing sugar, ethyl alcohol, total acid etc. all show
Different variations, barms sensory evaluations analysis, ferment middle, total acid content is gradually increasing, and produces acid, acetone
The products such as acid, acetic acid cream, the bacterial strain are produced suitable for blueberry red wine.
Embodiment 1 produces blueberry red wine using No. 5 strains of triumphant edge spring, and comparative example 1 produces blueberry red wine using commercial yeast.
In the identical situation of other fermentation conditions, 1 is shown in Table at the component content in wine.
Table 1
| Wine sample number | Residual sugar (g/l) | Alcoholic strength (%) | Total acid (g/l) | Volatile acid (g/l) |
| Embodiment 1 | 2.1 | 12.1 | 7.95 | 0.38 |
| Comparative example 1 | 2.9 | 10.5 | 5.77 | 0.45 |
As known from Table 1, the saccharomycete that present invention screening obtains is used for the fermentation process of blueberry red wine, it is residual after fermentation
Sugared content is lower, and alcoholic strength is higher.
The above description is merely a specific embodiment, but scope of protection of the present invention is not limited thereto, any
Those familiar with the art in the technical scope disclosed by the present invention, can easily think of the change or the replacement, and should all contain
Lid is within protection scope of the present invention.Therefore, protection scope of the present invention should be based on the protection scope of the described claims.
Claims (10)
1. a kind of wine waste mash, which is characterized in that deposit number is CGMCC NO.15931.
2. the screening technique according to claim 1 for making waste mash, which is characterized in that include the following steps：
(1) blueberry juice is taken fermentation liquid to be diluted with sterile water, obtains dilution after 25~30 DEG C ferment 2~4 days；
(2) dilution is coated in rose bengal medium, after 25~30 DEG C are cultivated 2~4 days, randomly selecting has
The single colonie of products of typical yeast colony characteristics isolates and purifies 2~3 times, isolated yeast strain；
(3) isolated yeast strain is subjected to gas generation property analysis, fermenting speed analysis, the analysis of strain coherency and utilization of carbon source
Analysis filters out the wine brewing and uses yeast strain.
3. according to the method described in claim 2, it is characterized in that, in step (1), the blueberry juice is by by Blueberry
Clean, select, squeeze the juice after obtain, be passed through sulfur dioxide in the juice-extracting process, sulfur dioxide containing in the blueberry juice
Amount is 30~70mg/L.
4. according to the method described in claim 2, it is characterized in that, in the step (1), the volume ratio of fermentation liquid and sterile water
It is 1：(3~8), preferably 1:5.
5. according to the method described in claim 2, it is characterized in that, fermentation and cultivation temperature are equal in the step (1) and (2)
It is 28 DEG C, the time is 3 days.
6. according to the method described in claim 2, it is characterized in that, gas generation property analysis is managed using Du Shi in the step (3)
Fermentation method preferably accesses isolated yeast strain built in 10ml in the YPD liquid medium of Du Shi tubule, inoculum concentration 20-
40CFU, ferment 72h at 28 DEG C, measures yeast strain gas production.
7. according to the method described in claim 2, it is characterized in that, in the step (3), fermenting speed analysis includes：It will divide
From yeast strain access blueberry juice in, 28 DEG C cultivate for 24 hours after, at 23 DEG C ferment 6~8d, measure blueberry juice in
Sugar consumption amount, and identify the fragrance generated in fermentation process.
8. according to the method described in claim 2, it is characterized in that, in the step (3), the analysis of strain coherency includes：
(i) by isolated yeast strain access Tissue Culture Flask, 48h is placed at 28 DEG C, is removed after distillation water washing centrifugation
Clear liquid obtains wet yeast；
(ii) the Acetate Solution 12ml that the wet yeast of 1g and pH are 3~4 is placed in centrifuge tube, then by centrifuge tube at 20 DEG C
15~20min of lower standing；
(iii) centrifuge tube is rocked, so that saccharomycete is suspended in centrifuge tube uniformly, suspension is then stood into 10min, observes ferment
Female bacterium is precipitated, and records ml of saccharomycete precipitating at this time.
9. according to the method described in claim 2, it is characterized in that, in the step (3), utilization of carbon source analysis includes：It will divide
From yeast strain access different carbon source culture medium in, 28 DEG C cultivate for 24 hours afterwards measure carbon source content, be determined to the carbon utilized
Source.The carbon source culture medium includes sucrose, maltose, raffinose, lactose, galactolipin, cellobiose, lemon drops, inositol and can
Soluble starch culture medium.
10. application of the saccharomycete as described in claim 1 in blueberry red wine fermentation.
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| CN109929768A (en) * | 2019-01-04 | 2019-06-25 | 浙江省农业科学院 | A kind of Wine brewing yeast strain and its application in the low alcohol essence wine pyrus nivalis fruit wine of production |
| CN113717868A (en) * | 2021-09-15 | 2021-11-30 | 黑龙江忠芝科远科技有限公司 | Method for optimizing extraction, separation, cultivation and utilization of saccharomyces cerevisiae in wild persimmons |
| CN114958629A (en) * | 2022-03-14 | 2022-08-30 | 常州大学 | Non-saccharomyces cerevisiae RM12 and application thereof in blueberry wine |
| CN114958629B (en) * | 2022-03-14 | 2023-08-29 | 常州大学 | Non-saccharomyces cerevisiae RM12 and application thereof in blueberry wine |
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