Saccharomyces cerevisiae with high yield of flower and fruit fragrance and application thereof

Abstract

The invention belongs to the technical field of microorganisms, and relates to saccharomyces cerevisiae with high yield of flower and fruit fragrance and application thereof. The strain is preserved in the China Center for Type Culture Collection, the preservation number is CCTCC M 20242771, the preservation time is December 10, 2024, the strain is separated from the natural fermentation process of blueberries in Danzhai region of Guizhou province, and the strain has excellent brewing characteristics including high ethanol (13.6% vol) and glycerol (7.48 g / L) yield, low hydrogen sulfide yield, positive killing and the like. When the strain is used for blueberry wine fermentation, the flower fragrance of an obtained blueberry wine sample is richer, the anthocyanin content exceeds that of a wine sample fermented by commercial yeast CECA, flavor analysis shows that the strain has obvious advantages in the aspect of producing key aroma compounds such as methyl benzoate, ethyl propionate, phenethyl acetate and beta-damascenone, and the strain has good application prospects. The OAV value of methyl benzoate in a wine sample fermented by the strain is 306.16, the wine can be endowed with more flower fragrance and fruit fragrance such as cherries, and richer fragrance levels are brought to a wine body.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms and relates to a high-yield floral and fruity aroma brewing yeast and application thereof. Background Art

[0002] In the process of brewing blueberry wine, the choice of yeast has a decisive influence on the flavor of the wine. At present, in the process of brewing blueberry wine, winemakers often directly use commercial wine yeast for wine brewing, and the development of wild wine yeast that can better show the unique flavor of blueberry wine is relatively lacking. At the same time, the high organic acid content and weak aroma of blueberry fruit itself make blueberry wine have certain deficiencies in taste and flavor, especially in the shaping of aroma quality, there is still a lot of room for improvement. In the aroma composition of blueberry wine, aromatic compounds such as phenylethanol bring it a lasting floral fragrance, while ester substances such as ethyl acetate and isoamyl acetate give it a variety of fruit flavors. In addition, other types of alcohol and ketone compounds also contribute to the unique flavor of blueberry wine.

[0003] In order to improve the aroma quality of blueberry wine, some technologies have been tried to solve this problem. For example, Chinese patent CN107164250A discloses a brewing method for enhancing the aroma of fruit wine, which increases the yield of β-phenylethanol in Saccharomyces cerevisiae by mutagenesis, thereby enhancing the aroma of fruit wines such as cherry wine, apple wine, and hawthorn wine. However, the limitation of this method is that it mainly focuses on the yield of phenylethanol and ignores the balance of other aroma components, which may lead to the flavor of the final wine being not harmonious enough.

[0004] As a healthy drink rich in polyphenols and anthocyanins, blueberry wine has a unique floral and fruity aroma, which is an important basis for consumers to choose. In the research on key technologies for brewing blueberry wine, the flavor substances of blueberry wine fermented by different yeasts were analyzed by GC-IMS technology, and it was found that the selection of yeast strains had a significant effect on the flavor of blueberry wine. In addition, the use of special yeast to brew high-quality blueberry wine and the comprehensive utilization of by-products also showed that specific yeast strains can improve the aroma quality of blueberry wine. In addition, the anthocyanin content in blueberry wine is also one of the important indicators to measure its quality. Anthocyanins not only give blueberry wine a deep color, but also have health benefits such as antioxidants. However, the anthocyanin content in blueberry wine samples fermented with commercial yeast is often not high, which limits the improvement of blueberry wine quality. Summary of the invention

[0005] In view of the shortcomings of the prior art, the present invention provides a high-yield floral and fruity brewing yeast and its application. The specific contents are as follows:

[0006] A high-yield floral and fruity brewing yeast, which is classified and named as Saccharomyces cerevisiae, and the laboratory strain collection number is recorded as FBKL2.9SP-L43, which is deposited in the China Center for Type Culture Collection, with a deposit number of CCTCC M 20242771 and a deposit date of December 10, 2024. The brewing yeast can produce high-yield key aroma compounds such as methyl benzoate, ethyl propionate, and phenylethyl acetate, and has the characteristics of high production of glycerol and ethanol, low production of hydrogen sulfide, and active killing. The strain was isolated by the inventor from Danzhai County, Guizhou Province.

[0007] Furthermore, the high-yield floral and fruity aroma is a high-yield key aroma compound.

[0008] Furthermore, the key aroma compounds are methyl benzoate, ethyl propionate, and phenylethyl acetate.

[0009] The application of the high-yield floral and fruity brewer's yeast in brewing.

[0010] Furthermore, the winemaking aspect is the aspect of brewing blueberry wine.

[0011] Furthermore, the blueberry wine brewing aspect is to enhance the floral and fruity aromas of the blueberry wine and improve the flavor complexity of the blueberry wine.

[0012] Furthermore, the winemaking aspect is to enhance the floral and fruity aroma of the wine.

[0013] The application of the high-yield floral and fruity aroma brewer's yeast in improving the floral and fruity aroma of products.

[0014] Furthermore, the product is a blueberry fermentation product.

[0015] The high-yield floral and fruity aroma brewer's yeast strain described in the present application can enhance the floral and fruity aroma of wine by increasing the content of anthocyanin, methyl benzoate and β-damascenone during the alcohol fermentation process, and improve the flavor complexity of blueberry wine. The strain can be used in wine brewing or the preparation of blueberry-related fermented products.

[0016] A brewing method for enhancing the blueberry hop fruit aroma adopts the above-mentioned brewer's yeast as a fermentation strain.

[0017] The details are as follows:

[0018] 1) Preparation of concentrated blueberry juice raw material, the raw material is prepared by crushing, enzymolysis, filtration, centrifugation, concentration and sterilization of blueberry fruits, and the sugar content of the blueberry juice before fermentation needs to be adjusted to 245g / L using sterile distilled water; 2) Inoculating high-yield floral and fruity brewer's yeast into the concentrated blueberry juice for alcohol fermentation; 3) filtration and centrifugation.

[0019] The inoculation amount of the high-yield floral and fruity brewer's yeast strain is 10 6 cells / mL.

[0020] The high-yield floral and fruity brewing yeast can help increase the content of anthocyanins in the fermentation of blueberry wine. The anthocyanin content of the blueberry wine obtained by inoculating the strain is higher than that of the wine sample fermented by the commercial yeast CECA. According to the evaluation and analysis results, the fermented blueberry wine has a better floral aroma than that of the commercial yeast CECA. Combined with the results of flavor analysis, the OAV value of methyl benzoate in the wine sample fermented by FBKL2.9SC-L43 is 306.16, and the OAV value of β-damascenone is significantly higher than that of CECA, contributing more floral aromas and fruity aromas such as cherries. No methyl benzoate was detected in the blueberry wine fermented by the commercial yeast CECA.

[0021] Beneficial effects of the present invention:

[0022] 1. The high-yield floral and fruity aroma brewer's yeast provided by the present invention is a newly developed wild strain, which is isolated and screened from the middle and late stages of natural fermentation of blueberries and has the characteristics of high yield of floral and fruity aroma.

[0023] 2. The strain of the present invention has a strong ability to produce alcohol, and the alcohol content of the obtained blueberry wine is about 13.6% vol.

[0024] 3. The strain of the present invention has a strong ability to produce glycerol. The glycerol content in the fermented blueberry wine sample reaches 7.48 g / L, which can increase the rounded taste of the blueberry wine, making the blueberry wine more mellow and soft, thereby significantly improving its flavor characteristics and improving its overall sensory quality.

[0025] 4. The strain of the present invention has a low production of hydrogen sulfide, which can not only well preserve the natural aroma of blueberries themselves, but also effectively prevent the unpleasant odor that may be produced by hydrogen sulfide, thereby ensuring the pure aroma of the wine, significantly improving the sensory quality of the blueberry wine, making its flavor more pleasant, and contributing to the brewing of blueberry wine with better quality and better taste.

[0026] 5. The strain of the present invention can effectively increase the anthocyanin content in blueberry wine fermented with the strain of the present invention. This feature makes the blueberry wine brewed with the strain of the present invention surpass the wine sample fermented with the commercial yeast CECA in terms of anthocyanin level. Anthocyanin is an important antioxidant in blueberry and is also a key component that affects the color and health benefits of blueberry wine.

[0027] 6. The strain selected by the present invention significantly enhances the floral and fruity aroma characteristics of blueberry wine during the fermentation process of the wine. This characteristic has been clearly reflected and recognized in professional tasting analysis. Compared with blueberry wine fermented with the traditional commercial yeast CECA, the blueberry wine fermented with the strain of the present invention has significantly improved the richness and depth of floral and fruity aroma. This improvement is mainly due to the production and influence of the strain on specific aroma compounds during the fermentation process. In the blueberry wine fermented with the FBKL2.9SC-L43 strain, the odor activity value (OAV) of methyl benzoate reached 306.16, which is much higher than that of the wine sample fermented by CECA, in which methyl benzoate was not even detected. The presence of methyl benzoate brings obvious floral and cherry aromas to blueberry wine, and the richness of these aromas plays a key role in improving the sensory quality of blueberry wine. The OAV value of β-damascenone is significantly higher than that of the wine sample fermented by CECA. As an important aroma compound, β-damascenone further enhances the floral and fruity aroma of the wine, making the blueberry wine fermented by the strain of the present invention show more superior floral and fruity aroma characteristics in the tasting. The combination of these characteristics not only enhances the flavor complexity of blueberry wine, but also provides consumers with a richer and higher quality blueberry wine experience.

[0028] 7. The strain of the present invention has excellent comprehensive fermentation ability, high yield of floral and fruity aroma, high yield of alcohol and glycerol, low yield of hydrogen sulfide, and active killing. These characteristics make the strain of the present invention have great potential in the industrial production of blueberry wine, which is helpful to improve the economic value and production efficiency of blueberries and develop distinctive high-quality blueberry wine products. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 : Morphological characteristics of the high-yield floral and fruity brewing yeast of the present invention on WL medium;

[0030] Figure 2 : The cell morphology of the high-yield floral and fruity brewer's yeast of the present invention under a microscope;

[0031] Figure 3 : Blueberry wine fermented by using the high-yield floral and fruity brewer's yeast of the present invention, and the control group is blueberry wine fermented by using the commercial yeast CECA, and the distribution of flavor substance content and OAV value of the two wine samples.

[0032] Figure 4 : Sensory evaluation radar chart of blueberry wine fermented by using the high-yield floral and fruity brewer's yeast of the present invention and the control group of blueberry wine fermented by using the commercial yeast CECA. DETAILED DESCRIPTION

[0033] The present invention is further described in detail below in conjunction with specific embodiments, which are intended to explain the present invention rather than to limit it.

[0034] Example 1

[0035] 1. Isolation and purification of strains

[0036] Blueberry samples were collected at the Xingren planting base in Danzhai County, Guizhou Province. The sampling process was strictly carried out in accordance with the principle of random sampling. Blueberry fruits with mature and full grains, uniform color, no mold or damage were picked aseptically, crushed and fully mixed, and placed in sterile fermentation bottles. Natural fermentation was carried out at 28°C without adding exogenous microorganisms. Samples were taken during the fermentation process for dilution and coating. The fermentation process was tracked according to the amount of carbon dioxide generated. Samples were taken in the middle and late stages of fermentation and separated by culture separation technology. Classification and identification were carried out using WL medium. 3 to 5 single colonies of different morphologies on each plate were picked and purified on WL medium for 3 to 5 days. The color, morphology and other colony characteristics of single colonies on WL medium were observed and recorded. Single colonies with representative characteristics were selected and transferred to solid YPD medium using an inoculation loop under a sterile environment. The zigzag streak method was used for separation and streak to ensure that independently distributed single cells were obtained. Cultured at 28°C for 2 days. The isolated yeast was streaked on YPD medium for a second purification, and then streaked on YPD medium again and continued to be cultured. These two purification steps effectively separated and purified the target yeast, ensuring its stability and consistency in subsequent studies.

[0037] 2. Identification of Saccharomyces cerevisiae strains

[0038] 2.1 Observation of morphological characteristics of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0039] The colony characteristics of the Saccharomyces cerevisiae strain are that it is milky white in color, spherical with dome-shaped protrusions, and has a rough but shiny surface.

[0040] 3. Study on the fermentation characteristics of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0041] 3.1 Detection of fermentation capacity of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0042] This study preliminarily determines the strength of yeast fermentation ability by testing the ability of yeast to consume glucose. The fermentation ability is reflected by monitoring reducing sugar during the fermentation process, and the reducing sugar detection is performed using the DNS method. Every 24 hours during the fermentation process, samples are diluted to within 0-1g / L. Referring to the method of Wang Chunxiao et al. (2012), 200μL of the diluted test solution is taken, 150μL of DNS reagent is added, mixed, and then bathed in water for 5min, cooled in an ice bath, 400μL of distilled water is added, mixed and allowed to stand for 20min, and then the absorbance is detected at a wavelength of 540nm. Distilled water is used as a blank control.

[0043] 3.2 Determination of glycerol production capacity of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0044] The glycerol content was determined using the glycerol enzymatic method (kit from: Beijing Pulilai Gene Technology Co., Ltd.).

[0045] 3.3 Determination of H2S production capacity of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0046] First, the activated strain was mixed with 5 mL of liquid YPD medium and cultured for 2 days. Then, 2 μL of the cultured bacterial solution was taken out with a pipette and spotted on BIGGY medium for three times. After the bacterial solution on the sample dried, it was placed at 28°C for 4 days to observe the color characteristics of the colonies to determine whether the strain produced H2S.

[0047] 3.4 Test for the killer toxicity of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0048] The yeast was inoculated into YPD-MB medium to test the killing activity and sensitivity of yeast.

[0049] 3.5 Detection of ethanol production ability of Saccharomyces cerevisiae strain FBKL2.9SC-L43

[0050] The alcohol production capacity of the strain was tested by the alcoholometer method (refer to GB / T15038-2006).

[0051] Example 2

[0052] The invention discloses an application of a high-yield floral and fruity brewing yeast strain in the development of blueberry wine; the high-yield floral and fruity brewing yeast is classified and named as Saccharomyces cerevisiae, and the laboratory strain collection number is recorded as FBKL2.9SP-L43, which is preserved in the China Center for Type Culture Collection with a preservation number of CCTCC M 20242771 and a preservation date of December 10, 2024.

[0053] The blueberry wine brewing method comprises the following steps:

[0054] 1) Preparation of concentrated blueberry juice raw material, the raw material is prepared by crushing, enzymolysis, filtration, centrifugation, concentration and sterilization of blueberry fruit, and the blueberry juice before fermentation needs to be adjusted to 245g / L with sterile distilled water; 2) 6cells / mL, inoculated with high-yield floral and fruity aroma brewer's yeast in concentrated blueberry juice for alcohol fermentation, the fermentation temperature was controlled at 28°C, and the fermentation process was monitored by CO2 production; 3) after the fermentation was completed, filtered with sterile gauze and centrifuged; 4) after removing the precipitate, the supernatant was taken and properly stored at 4°C.

[0055] The inoculation amount of the high-yield floral and fruity brewer's yeast strain is 10 6 cells / mL.

[0056] The method for determining flavor substances in blueberry wine comprises the following steps:

[0057] 5 mL of wine sample was taken for volatile substance analysis. The sample was processed by solid phase microextraction, and 20 μL of 2-octanol with a concentration of 20 mg / L was added as an internal standard. The aged extraction head was used for extraction head adsorption at 50°C for 30 min, and then desorbed at 250°C for 3 min. At the same time, the instrument was started to collect data. 26 The retention index RI of each substance was calculated after the normal alkanes were analyzed under the same chromatographic conditions as the samples.

[0058] contrast:

[0059] The high-yield floral and fruity brewing yeast isolated from Example 1 and the commercial brewing yeast CECA were used in the form of single-plant inoculation to brew blueberry wine. The flavor data of the obtained blueberry wine are as follows: Figure 3 As shown. According to the flavor analysis results,

[0060] Based on the provided flavor substance content distribution map and odor activity value (OAV) distribution map, we conducted the following analysis on the flavor characteristics and advantages of blueberry wine fermented by FBKL2.9SC-L43 strain:

[0061] The aroma components of blueberry wine brewed by FBKL2.9SC-L43 and CECA are similar, with ethyl acetate, ethyl hexanoate, isoamyl alcohol and phenylethyl alcohol as the main substances, which make blueberry wine have fruity, winey, creamy and rose aromas. The aroma components of FBKL2.9SC-L43 are more complex, forming substances such as ethyl propionate, methyl benzoate, phenylethyl acetate, n-octanoic acid, and 4-ethylguaiacol, which means that the FBKL2.9SC-L43 strain has obvious advantages in producing these key aroma compounds, which can give the wine a rich fruity aroma and full ester aroma, bringing a richer aroma layer to the wine. In the blueberry wine fermented by FBKL2.9SC-L43, the content of methyl benzoate was 159.2013μg / L, and its OAV value was 306.16. However, no methyl benzoate was detected in the blueberry wine fermented by commercial yeast CECA. Methyl benzoate brings obvious floral and cherry aromas to blueberry wine. The richness of these aromas plays a key role in improving the sensory quality of blueberry wine. The FBKL2.9SC-L43 strain has advantages in producing a variety of aroma compounds, which play an important role in improving the aroma richness of blueberry wine. It is better in the balance of aroma compounds, especially in producing floral and fruity aromas, which helps to improve the flavor characteristics and sensory quality of blueberry wine.

[0062] Finally, it should be pointed out that the above embodiments are only representative examples of the present invention. Obviously, the technical solution of the present invention is not limited to the above embodiments, and there are many variations. All variations that can be directly derived or associated with the content disclosed by ordinary technicians in this field should be considered as the protection scope of the present invention.

Claims

1. A high-yield floral and fruity brewing yeast, characterized in that: It is deposited in the China Center for Type Culture Collection with the accession number: CCTCC M 20242771 and the preservation date: December 10, 2024.

2. The high-yield floral and fruity brewing yeast according to claim 1, characterized in that: The high-yield floral and fruity aroma is a high-yield key aroma compound.

3. The high-yield floral and fruity brewing yeast according to claim 2, characterized in that: The key aroma compounds are methyl benzoate, ethyl propionate, phenylethyl acetate, and β-damascenone.

4. The high-yield floral and fruity brewing yeast according to claim 1, characterized in that: The brewer's yeast also has excellent brewing characteristics, such as high production of ethanol and glycerol, low production of hydrogen sulfide, and active killing.

5. Use of the high-yield floral and fruity brewing yeast of claim 1 in brewing.

6. The use according to claim 5, characterized in that: The winemaking aspect is the brewing of blueberry wine.

7. The use according to claim 5, characterized in that: The winemaking aspect is to enhance the floral and fruity aroma of the wine.

8. Use of the high-yield floral and fruity aroma brewer's yeast according to claim 1 in improving the floral and fruity aroma of products.

9. The use according to claim 8, characterized in that: The product is a blueberry fermentation product.

10. A brewing method for enhancing the fruity aroma of blueberry hops, characterized in that: The brewer's yeast according to claim 1 is used as the fermentation strain.

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