CN112812979B - Yeast strain QTX11 for producing aroma substances at high yield and application thereof - Google Patents
Yeast strain QTX11 for producing aroma substances at high yield and application thereof Download PDFInfo
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- CN112812979B CN112812979B CN202110108011.1A CN202110108011A CN112812979B CN 112812979 B CN112812979 B CN 112812979B CN 202110108011 A CN202110108011 A CN 202110108011A CN 112812979 B CN112812979 B CN 112812979B
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12G—WINE; PREPARATION THEREOF; ALCOHOLIC BEVERAGES; PREPARATION OF ALCOHOLIC BEVERAGES NOT PROVIDED FOR IN SUBCLASSES C12C OR C12H
- C12G1/00—Preparation of wine or sparkling wine
- C12G1/02—Preparation of must from grapes; Must treatment and fermentation
- C12G1/0203—Preparation of must from grapes; Must treatment and fermentation by microbiological or enzymatic treatment
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- C12N1/16—Yeasts; Culture media therefor
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Abstract
The invention discloses a yeast strain QTX11 for producing aroma substances with high yield and application thereof, belonging to the technical field of microorganisms. The preservation number of the strain QTX11 of the yeast strain Curvibasiumpallidicorallinum is CCTCC M2021082. The n-hexanol yield generated by fermenting grape juice with the strain is 257.93 mug/L, the 2, 3-butanedione yield is 1562 mug/L, and the ethyl acetate yield is 169.74 mug/L.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a yeast strain QTX11 for producing aroma substances at high yield and application thereof.
Background
Yeasts (Curvibasidium pallidicorallinum) have the biological characteristics of: macroscopic morphology: the culture is carried out for 3 days at 28 ℃ on a wort agar culture medium, and the bacterial colony is creamy, milky, flat, dry on the surface and incomplete on the edge. Microscopic morphology: malt extract agar culture medium, culturing at 28 deg.C for 2-3 days, and culturing in spindle or oval shape 2.5-4.0 μm × 3.5-9.0 μm.
The yeast (Curvibasidium pallidicola) is reported to be separated from sapropenia wood, the aerobic type is aerobic, but specific application and related reports about the yeast are few.
Yeast (Curvibasidium Pallidicola gallinum) capable of producing aroma substances such as hexanol, trans-2-nonenal, isovaleraldehyde and the like is never reported in the field at present.
Disclosure of Invention
Based on the blank in the field, the invention obtains a strain QTX11 of the yeast Curvibacillus pallidicola collerillum which takes grape juice as a substrate to ferment and produce aroma substances such as n-hexanol, trans-2-nonenal, isovaleraldehyde and the like at high yield by screening.
The technical scheme of the invention is as follows:
a strain of yeast Curvibacillus pallidicorallinum QTX11 with a preservation number of CCTCC M2021082.
Use of the yeast Curvivadiam pallidicorninum strain QTX11 with the collection number of CCTCC M2021082 in the production of aroma substances.
The aromatic substance is selected from the group consisting of n-heptanol, 1-octen-3-ol, linalool, damascenone, hexyl acetate, isoamyl acetate, ethyl valerate, ethyl heptanoate, ethyl hexanoate, n-hexanol, trans-2-nonenal, isovaleraldehyde, 2, 3-butanedione, 3-octanone, and ethyl acetate.
The aroma substance is selected from the group consisting of n-hexanol, trans-2-nonenal, isovaleraldehyde, 2, 3-butanedione, 3-octanone, and ethyl acetate.
Application of a yeast Curvivadiam pallidicorninum strain QTX11 with a preservation number of CCTCC M2021082 in brewing wine.
A wine brewing leaven is characterized in that fermentation active substances comprise a yeast Curvibasidium Pallidicola crallinum strain QTX11 with the preservation number of CCTCC M2021082.
The fermentation active substance also comprises saccharomyces cerevisiae.
The brewing leaven is a microbial inoculum;
preferably, the brewing leavening agent also comprises auxiliary materials acceptable for microbial inoculum;
preferably, the auxiliary material acceptable by the microbial inoculum comprises a culture medium substance of the microbial inoculum; media materials for the bacteria include, but are not limited to: starch, sucrose, peptone and water.
A method for brewing wine is characterized in that the wine is fermented and brewed by adopting a yeast Curvibasidium pallidicola rarillinum strain QTX11 with the preservation number of CCTCC M2021082.
The fermentation refers to the fermentation by using the yeast Curvibacillus pallidicola rarallinum strain QTX11 and using grape or grape juice as a substrate.
Preferably, the yeast Curvibacillus pallidicola rarillinum strain QTX11 is added to the substrate at a concentration of 106-107CFU/ml, preferably 106CFU/ml。
The invention obtains a strain through screening, a series of aroma substances such as n-hexanol, trans-2-nonenal, isovaleraldehyde and the like can be produced at high yield through the measurement of aroma substance component identification experiments, and compared with a commercial saccharomyces cerevisiae strain F33, the strain can also produce aroma substances which cannot be produced by F33 or have lower yield, such as: the yield of the n-hexanol, the trans-2-nonenal, the isovaleraldehyde, the 2, 3-butanedione, the 3-octanone and the ethyl acetate is 257.93 mu g/L, 1562 mu g/L of the 2, 3-butanedione and 169.74 mu g/L of the ethyl acetate. The strain is identified to be a strain of the yeast Curvibacillus pallidicola rarillinum, which is named QTX11 by the applicant and is sent for preservation.
The deposited information of the yeast Curvibasidium pallidicorallinum strain QTX11 of the present invention is as follows:
naming: QTX11
And (4) classification name: yeast
The name of Latin is: curvibasidium pallidicorallinum
The preservation number is as follows: CCTCC M2021082
The preservation organization: china center for type culture Collection
The preservation date is as follows: 1 month and 15 days 2021.
Detailed Description
The following detailed description of the present invention will be made with reference to specific examples, but the scope of the present invention is not limited thereto.
Sources and documentations of biological materials
The sources and origins of the grape varieties used in experimental example 1 are shown in table 1 below:
TABLE 1
| Grape variety | Producing area | Source |
| Cabernet sauvignon | Ningxia Helan mountain east foot producing area | Collecting |
| Meile wine | Ningxia Helan mountain east foot producing area | Collecting |
| Xila | Ningxia Helan mountain east foot producing area | Collecting |
| Snake dragon bead | Ningxia Helan mountain east foot producing area | Collecting |
| Xiaowei Er duo | Ningxia Helan mountain east foot producing area | Collecting |
| Noble incense | Ningxia Helan mountain east foot producing area | Collecting |
| Weidai Er | Ningxia Helan mountain east foot producing area | Collecting |
| Rivastigmine | Ningxia Helan mountain east foot producing area | Collecting |
| Chardonnay | Ningxia Helan mountain east foot producing area | Collecting |
The grape varieties in table 1 above are all known and public grape varieties, and are also commercially available.
The grape variety used in experimental example 2 was a wital ice grape, purchased from Ningxia Bagges drunk American interline wine Co., Ltd; commercial strain Saccharomyces cerevisiae F33 was purchased from Lafford (Laffort) France.
Group 1 example, Strain QTX11 of the present invention
The embodiment of the group provides a strain of yeast Curvibasidium pallidicola rarillinum QTX11 with the preservation number of CCTCC M2021082.
Group 2 example, application of the Strain QTX11 of the invention
The group of embodiments provides the application of the yeast Curvibacillus pallidicola rarillium strain QTX11 with the preservation number of CCTCC M2021082 in the aspect of producing aromatic substances.
In some embodiments, the aroma substance is selected from the group consisting of n-heptanol, 1-octen-3-ol, linalool, damascenone, hexyl acetate, isoamyl acetate, ethyl valerate, ethyl heptanoate, ethyl hexanoate, n-hexanol, trans-2-nonenal, isovaleraldehyde, 2, 3-butanedione, 3-octanone, ethyl acetate.
In other embodiments, the aroma is selected from the group consisting of n-hexanol, trans-2-nonenal, isovaleraldehyde, 2, 3-butanedione, 3-octanone, ethyl acetate, any of which is produced in a yield of QTX11 strain that is much higher than known Saccharomyces cerevisiae F33, for specific comparisons see Table 1 below.
Group 3 example, wine brewing application of Strain QTX11 of the invention
The embodiment of the group provides application of a yeast Curvibacillus pallidicola rarillium strain QTX11 with the preservation number of CCTCC M2021082 in brewing.
Group 4 examples of the fermentation agents for brewing wine according to the invention
The embodiment of the group provides a wine brewing leavening agent. All embodiments of this group share the following common features: the fermentation active substances of the brewing leaven comprise a yeast Curvibacillus pallidicorallinum strain QTX11 with the preservation number of CCTCC M2021082.
In a further embodiment, the fermentation active further comprises saccharomyces cerevisiae. The QTX11 strain of the present invention can be used in combination with Saccharomyces cerevisiae, which is commonly used in the art, for fermenting Saccharomyces cerevisiae by one skilled in the art according to the teachings of the present invention.
In a specific embodiment, the brewing leavening agent is a microbial inoculum;
in a preferred embodiment, the brewing leavening agent further comprises auxiliary materials acceptable for microbial inoculum;
in other preferred embodiments, the microbial inoculum acceptable adjuvant comprises a medium material of the microbial inoculum; media materials for the bacteria include, but are not limited to: starch, sucrose, peptone and water.
Group 5 example wine brewing method of the invention
The present set of embodiments provide a wine brewing method. All embodiments of this group share the following common features: fermentation brewing is carried out by adopting a yeast Curvivadiam pallidicorninum strain QTX11 with the preservation number of CCTCC M2021082.
In a specific example, the fermentation refers to fermentation using the yeast Curvibacillus pallidicola strain QTX11 with grape or grape juice as substrate.
In a preferred embodiment, the yeast Curvibacillum pallidicorallinum strain QTX11 is added to the substrate at a concentration of 106-107CFU/ml, preferably 106CFU/ml。
Experimental example 1 screening Process of the Yeast Curvibacillum pallidicorallinum Strain QTX11 of the present invention
Collecting different varieties of wine grapes (shown in table 1) at different production places of eastern foot of Ningxia Helan mountain, removing rotten, mildewed and damaged fruit grains and sundries, weighing 10.0g of uniform and complete grape fruit grains, adding the uniform and complete grape fruit grains into 90mL of sterile YPD liquid culture medium, oscillating for 10min to prepare bacterial suspension, coating the bacterial suspension with proper dilution on YPD solid culture medium added with 100mg/L chloramphenicol by adopting a gradient dilution method, culturing at 28 ℃ for 2 d-3 d, and carrying out streak purification on the YPD solid culture medium for multiple times according to colony morphology to obtain a purified strain. After the purified strain is activated in YPD culture medium for 24h, 1mL of bacterial liquid is taken to extract yeast genome according to instructions of a Biospin fungal genome DNA extraction kit, primers NL-1(5'-GCATATCAATAAGCGGAGGAAAAG-3') and NL-4(5'-GGTCCGTGTTTCAAGACGG-3') are used for PCR amplification, a gene sequence of a 26S rDNA fragment is obtained through sequencing, known standard strain information with high homology is obtained through BLAST comparison of NCBI, species identification is carried out on the strain with similarity of more than 99%, and finally strain species information is obtained.
In this experimental example, 14 strains of Curvibacillum pallidicorallinum were obtained in total. 14 strains of Curvivadiam pallidicorallinum were each introduced at an initial concentration of 106The CFU/mL is inoculated into sterilized Vidale grape juice (100 ℃, 10min), fermented for 18 days at 18 ℃ +/-2 ℃, the aroma content is measured, and finally, a Curvisadiumpallidicola strain QTX11 with the best aroma production effect is obtained and is preserved, and the preservation information is as follows:
naming: QTX11
And (4) classification name: yeast
The name of Latin is: curvibasidium pallidicorallinum
The preservation number is as follows: CCTCC M2021082
The preservation organization: china center for type culture Collection
The preservation date is as follows: 1 month and 15 days 2021.
Experimental example 2 data on aroma-producing substances of the yeast Curvibasidium pallidicola mutant QTX11 of the present invention
Grape variety: vidal iced grape
The grape juice production process comprises the following steps: pressing harvested ice grape with ice by air bag press while adding sulfur dioxide (50mg/L K)2S2O5) And 20mg/L of pectinase (more than or equal to 500U/mg), inhibits bacterial diseases and improves the juice yield.
The grape juice fermentation conditions are as follows: the steeped grape juice is respectively added at an initial concentration of 106CFU/mL inoculated F33 Saccharomyces cerevisiae and Curvibacilli pallidicoralliThe num strain QTX11, the fermentation temperature is 18 ℃ +/-2 ℃, and the fermentation is stopped when the weight loss of the grape juice is not changed for three consecutive days. All wine samples were centrifuged at 7500rpm for 8 minutes and the supernatant was stored at 4 ℃.
The detection method of each aroma substance comprises the following steps: headspace-solid phase microextraction-gas mass spectrometry (HS-SPME-GC/MS) was used. An 8mL sample of wine was accurately weighed into a headspace bottle containing 1.5g NaCl, while 394.08. mu.g/L4-methyl-1-pentanol (internal standard) was capped and sealed. Inserting CAR/DVB/PDMS extraction fiber, adsorbing at 45 deg.C for 30min, desorbing at 250 deg.C for 3min, and performing GC-MS analysis. A chromatographic column: InertCap WAX polar chromatography column (60m × 0.25mm, 0.25 μm); the temperature rising procedure is as follows: maintaining at 40 deg.C for 5min, heating to 120 deg.C at 3 deg.C/min, heating to 230 deg.C at 8 deg.C/min, and maintaining for 10 min; the flow rate of the carrier gas (He) was 0.8mL/min, and was not split. Electron bombardment ion source; electron energy 70 eV; the transmission line temperature was 275 ℃; the ion source temperature is 230 ℃; the activation voltage is 1.5V; the filament flow is 0.25 mA; the mass scanning range m/z is 33-450. Compound quantitative analysis was performed using external standard quantitation method.
The same batch of grape juice produced by the same variety of grapes is taken as a substrate, the same amount of grape juice is fermented and brewed by a commercial strain Saccharomyces cerevisiae F33 and the yeast Curvibacilli Pallidii wine strain QTX11 under the same fermentation condition, and the content of each aroma substance (unit: mu g/L, meaning: the content of the aroma substance in each liter of grape wine) is detected by a headspace-solid phase microextraction-gas mass spectrometry method, so that the following table 2 is obtained:
TABLE 2
The aroma threshold in table 1 above refers to the lower limit of the minimum concentration at which a human can smell the substance.
SEQUENCE LISTING
<110> Ningxia agricultural product quality standard and detection technology research institute (Ningxia agricultural product quality monitoring center)
Ningxia Hui Autonomous Region State owned forest farm and tree seedling work station
<120> a strain of yeast Curvibasidium with high yield of aroma substances
Pallidicorallinum strain QTX11 and application thereof
<130> P201025/GYL
<160> 2
<170> PatentIn version 3.5
<210> 1
<211> 24
<212> DNA
<213> Artificial Sequence
<220>
<223> primer NL-1
<400> 1
gcatatcaat aagcggagga aaag 24
<210> 2
<211> 19
<212> DNA
<213> Artificial Sequence
<220>
<223> primer NL-4
<400> 2
ggtccgtgtt tcaagacgg 19
Claims (14)
1. Yeast strainCurvibasidium pallidicorallinumQTX11 with the preservation number of CCTCC NO: M2021082.
2. Yeast strain with preservation number of CCTCC NO: M2021082Curvibasidium pallidicorallinumQTX11 for use in the production of aroma.
3. Use according to claim 2, wherein the aroma substances are selected from the group consisting of n-heptanol, 1-octen-3-ol, linalool, damascenone, hexyl acetate, isoamyl acetate, ethyl valerate, ethyl heptanoate, ethyl hexanoate, n-hexanol, trans-2-nonenal, isovaleraldehyde, 2, 3-butanedione, 3-octanone, ethyl acetate.
4. Use according to claim 2, wherein the aroma substances are in the group of n-hexanol, trans-2-nonenal, isovaleraldehyde, 2, 3-butanedione, 3-octanone, ethyl acetate.
5. Yeast strain with preservation number of CCTCC NO: M2021082Curvibasidium pallidicorallinumQTX11 application in brewing wine.
6. The brewing leaven is characterized in that the fermentation active substances comprise yeast strains with the preservation number of CCTCC NO: M2021082Curvibasidium pallidicorallinum QTX11。
7. The leavening agent of claim 6, wherein the fermentation active further comprises Saccharomyces cerevisiae.
8. The wine brewing starter culture according to claim 6 or 7, wherein the wine brewing starter culture further comprises auxiliary materials acceptable for microbial inoculum.
9. The wine brewing starter culture according to claim 8, wherein the auxiliary material acceptable for the microbial inoculum comprises a culture medium material of the microbial inoculum.
10. The wine brewing starter culture according to claim 9, wherein the culture medium material of the bacteria comprises: starch, sucrose, peptone and water.
11. A method for brewing wine is characterized in that a yeast strain with a preservation number of CCTCC NO: M2021082 is adoptedCurvibasidium pallidicorallinumQTX11 fermenting and brewing.
12. A wine-making process according to claim 11, wherein said fermentation is a fermentation process usingThe yeast strainCurvibasidium pallidicorallinumQTX11 the fermentation is carried out using grape or grape juice as substrate.
13. The method of claim 12, wherein the yeast strain isCurvibasidium pallidicorallinumQTX11 was added to the substrate at a concentration of 106-107CFU/ml。
14. The method of claim 13, wherein the yeast strain isCurvibasidium pallidicorallinumQTX11 was added to the substrate at a concentration of 106CFU/ml。
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- 2021-01-27 CN CN202110108011.1A patent/CN112812979B/en active Active
Non-Patent Citations (3)
| Title |
|---|
| Yeast diversity on grapes in two German wine growing regions;Laboratory for Wine Microbiology等;《International Journal of Food Microbiology》;20150802;第214卷;全文 * |
| 新疆石河子月季花附生酵母菌多样性分析;马雪等;《广东农业科学》;20180815(第08期);全文 * |
| 黑树莓酒自然发酵过程中微生物群落演替研究;范民婷等;《中国酿造》;20200925(第09期);全文 * |
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