Schizosaccharomyces pombe with high animal and plant aroma yield and application thereof

Abstract

The invention belongs to the technical field of microorganisms, and relates to schizosaccharomyces pombe with high yield of animal and plant aroma and application of the schizosaccharomyces pombe. The schizosaccharomyces pombe with high animal and plant aroma yield is preserved in the China Center for Type Culture Collection on December 10, 2024, the preservation number is CCTCC M 20242770, the strain is separated from the natural fermentation process of blueberries in Danzhai region of Guizhou province, and the schizosaccharomyces pombe has the advantages of high yield of glycerol (10.24 g / L) and ethanol (12.5% vol), low yield of hydrogen sulfide, positive killing and remarkable capability of degrading organic acid, and can be used for preparing the high-yield schizosaccharomyces pombe with high animal and plant aroma yield. According to the present invention, the blueberry wine is fermented by using the strain, the concentration of 2, 3-butanedione (diacetyl) in the obtained wine sample is as high as 1513.1276 [mu] g / L, the OAV value is 3026.2552, the blueberry wine is endowed with the cream fragrance and the milk fat fragrance, the blueberry wine with the unique animal and plant fragrance and the rich taste can be produced, and the pursuit of consumers on the high-quality and unique-flavor beverage can be met.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms and relates to a pombe schizosaccharomyces cerevisiae with high production of animal and plant aroma and application thereof. Background Art

[0002] Blueberry wine is favored by consumers for its unique flavor and nutritional value. Traditional blueberry wine fermentation mainly relies on commercial brewing yeast, which has good fermentation ability and tolerance in blueberry juice, providing a stable quality guarantee for the production of blueberry wine. However, these yeasts often lack uniqueness and diversity in flavor. Wild non-brewing yeasts isolated from the natural fermentation of blueberries have attracted much attention in the winemaking field because they can give fruit wine a unique taste and aroma. These non-brewing yeasts are widely present in the early stage of natural fermentation of blueberry wine. They can produce a series of complex flavor compounds, thereby increasing the flavor diversity and complexity of blueberry wine. However, there are relatively few studies on non-brewing yeasts in blueberry mash, which limits the improvement of blueberry wine flavor diversity and the exploration of innovation potential.

[0003] In order to overcome this limitation, researchers in this field have explored the application of different yeasts in fruit wine fermentation in depth. For example, the patent with patent number CN117264789A discloses a strain of wine spore Hansen yeast (Hanseniasporavineae) YIMY02353, which has a strong alcohol production ability and a certain ester production ability, which can significantly improve the flavor and quality of fruit wine. Different yeast strains may differ in fermentation efficiency and flavor substance production, and further research and optimization are needed to balance the fermentation efficiency and the flavor of wine. The yeast isolated from the naturally fermented blueberry juice has a stable fermentation, a proper ratio of wine aroma and fruit aroma, moderate sourness, and a strong blueberry wine flavor. The organic acid content in blueberry wine has a significant effect on the taste, and it is necessary to regulate the organic acid content through yeast screening and fermentation condition optimization to achieve an ideal taste. For example, the patent publication number CN118755593A discloses a wild type brewer's yeast, which has not been induced by mutation or genetic modification, has high production of aromatic compounds and fruity flavor compounds, low production of sour flavor substances, and has excellent salicylic acid degradation function. This yeast can increase the content of compounds such as β-damascenone, ethyl phenylacetate, phenylethyl acetate, phenylethanol, etc. with floral, fruity and honey aroma in blueberry wine, and significantly reduce the content of salicylic acid in blueberry wine, thereby enhancing the floral, fruity and honey aroma of blueberry wine, improving the taste, reducing the sourness, reducing bitterness, and improving the sour-sweet balance.

[0004] In the flavor analysis of blueberry wine, 2,3-butanedione (diacetyl) is an important aroma component, and its content has a significant impact on the overall flavor of the wine. Studies have shown that when the content of diacetyl is 1-4 mg / L, it can significantly increase the taste and flavor complexity of wine, presenting a pleasant buttery or cheese flavor. This characteristic is particularly important for blueberry wine, as it can enhance the flavor, improve the sensory experience, and increase the complexity of the aroma, thereby improving the quality and market competitiveness of blueberry wine.

[0005] The selection and application of specific yeast strains have a significant impact on the flavor and quality of blueberry wine. By screening and applying yeast strains with specific characteristics, the flavor of blueberry wine can be made richer and more personalized. Summary of the invention

[0006] In view of the shortcomings of the prior art, the present invention provides a pombe schizosaccharomyces cerevisiae with high production of animal and plant aroma and its application, which are as follows:

[0007] A pombe schizosaccharomyces with high production of animal and plant aromas, whose classification name is Schizosaccharomyces pombe, the laboratory strain collection number is recorded as FBKL2.9SP-L14, and it is deposited in the China Center for Type Culture Collection with the accession number: CCTCC M20242770 and the preservation time is December 10, 2024.

[0008] Furthermore, the high-yield animal and plant aroma is high-yield 2,3-butanedione (diacetyl).

[0009] The invention discloses an application of the pombe schizosaccharomyces with high yield of animal and plant aroma in wine making.

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

[0011] Furthermore, the winemaking aspect is to enhance the animal and plant aroma of the wine.

[0012] The application of the pombe schizosaccharomyces with high production of animal and plant aroma in the degradation of organic acids, especially in the degradation of malic acid.

[0013] The invention discloses an application of the pombe schizosaccharomyces cerevisiae with high production of animal and plant aroma in producing 2,3-butanedione (diacetyl).

[0014] The application of the high-yield pombe schizosaccharomyces cerevisiae to enhance the animal and plant aroma of products. Furthermore, the product is a blueberry fermentation product.

[0015] A brewing method for enhancing the animal and plant aroma of blueberry wine adopts the above-mentioned Schizosaccharomyces pombe as a fermentation strain.

[0016] The details are as follows:

[0017] 1) preparing the raw material of concentrated blueberry juice, wherein the raw material is prepared by crushing, enzymolysis, filtering, centrifuging, concentrating and sterilizing blueberry fruits, and the sugar content of the blueberry juice before fermentation needs to be adjusted to 245 g / L using sterile distilled water; 2) inoculating the concentrated blueberry juice with high-yield animal and plant aroma Schizosaccharomyces pombe to carry out alcohol fermentation; 3) filtering and centrifuging.

[0018] The inoculation amount of the pombe schizosaccharomyces strain with high production of animal and plant aroma is 10 6 cells / mL.

[0019] The pombe schizosaccharomyces cerevisiae with high production of animal and plant aromas helps to increase the content of 2,3-butanedione (diacetyl) in the fermentation of blueberry wine. The concentration of diacetyl in the flavor data of the obtained wine sample is 1513.1276 μg / L, and its OAV value reaches 3026.2552, giving the blueberry wine a creamy and buttery aroma. The wine sample of the commercial brewing yeast CECA used in the control group has a concentration of 2,3-butanedione (diacetyl) of 0 μg / L and an OAV value of 0. It can be seen that pombe schizosaccharomyces cerevisiae has a significant advantage in improving the flavor of blueberry wine.

[0020] Beneficial effects of the present invention:

[0021] 1. The Schizosaccharomyces pombe 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 production of animal and plant aroma.

[0022] 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 12.5% ​​vol.

[0023] 3. The strain of the present invention can significantly increase the diacetyl content in blueberry wine fermented with the wine, thereby enhancing the cream flavor and milk fat flavor of the blueberry wine. The strain of the present invention is used to brew blueberry wine, and the diacetyl concentration of the obtained blueberry wine reaches 1513.1276 μg / L, and the OAV value is as high as 3026.2552, which brings a unique animal and plant aroma to the blueberry wine, significantly improves the taste and flavor complexity of the wine, and meets the consumer's demand for unique flavor fruit wine.

[0024] 4. The strain of the present invention also has the ability to degrade organic acids, mainly manifested in the degradation of malic acid and tartaric acid, which are reduced to 7.12 g / L and 4.63 g / L respectively, which helps to balance the acidity of blueberry wine, reduce excessive sourness and astringency, make the wine more harmonious and the taste more mellow.

[0025] 5. The strain of the present invention has a strong ability to produce glycerol. The glycerol content in the fermented blueberry wine sample reaches 10.38 g / L, which can improve the roundness of the taste of blueberry wine and significantly improve the sensory quality of wine.

[0026] 6. The strain of the present invention has low hydrogen sulfide production. The fermented blueberry wine sample maintains the original aroma of blueberries while avoiding the unpleasant odor caused by hydrogen sulfide, thereby improving the overall sensory quality of the wine. This feature is of great significance for the production of blueberry wine.

[0027] 7. The strain of the present invention has excellent comprehensive fermentation ability, high alcohol and glycerol production, low hydrogen sulfide production, active killing, and good ability to degrade organic acid (malic acid). It is expected to play an important role in the industrial production of blueberry wine, improve the economic value of blueberries, enhance the production efficiency of blueberry wine, and has great potential for developing distinctive high-quality blueberry wine. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 : Morphological characteristics of the Schizosaccharomyces pombe with high production of animal and plant aroma on WL medium;

[0029] Figure 2 : The cell morphology of the Schizosaccharomyces pombe with high production of animal and plant aroma under a microscope;

[0030] Figure 3 : Blueberry wine fermented with the pombe schizosaccharomyces cerevisiae of the present invention with high production of animal and plant aroma, and the control group with the blueberry wine fermented with the commercial yeast CECA, the distribution of flavor substance content and OAV value of the two wine samples.

[0031] Figure 4 : Sensory evaluation radar chart of blueberry wine samples fermented with the pombe schizosaccharomyces cerevisiae of the present invention and the control group fermented with the commercial yeast CECA. DETAILED DESCRIPTION

[0032] 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.

[0033] Example 1

[0034] 1. Isolation and purification of Schizosaccharomyces pombe strains

[0035] 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.

[0036] 2. Identification of Schizosaccharomyces pombe strains

[0037] 2.1 The morphological characteristics of the Schizosaccharomyces pombe strain FBKL2.9SP-L14 are that the colony edge is light green, the central outer circle is light green, the inner circle is dark green, and the shape is a spherical protrusion with a tray-like bottom.

[0038] 3. Study on the fermentation characteristics of Schizosaccharomyces pombe strain FBKL2.9SP-L14

[0039] 3.1 Detection of the fermentation capacity of S. pombe strain FBKL2.9SP-L14

[0040] 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, the sample is diluted to within 0-1g / L, 200μL of the diluted test solution is taken, 150μL of DNS reagent is added, mixed and placed in a water bath for 5min, cooled in an ice bath, 400μL of distilled water is added, mixed and allowed to stand for 20min, and the absorbance is detected at a wavelength of 540nm. Distilled water is used as a blank control.

[0041] 3.2 Determination of glycerol production capacity of Schizosaccharomyces pombe strain FBKL2.9SP-L14

[0042] The glycerol content was determined using the glycerol enzymatic method (kit from: Beijing Pulilai Gene Technology Co., Ltd.).

[0043] 3.3 Determination of H2S production capacity of Schizosaccharomyces pombe strain FBKL2.9SP-L14

[0044] 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.

[0045] 3.4 Killing test of Schizosaccharomyces pombe strain FBKL2.9SP-L14

[0046] The yeast was inoculated into YPD-MB medium to test the killing activity and sensitivity of yeast.

[0047] 3.5 Detection of alcohol production ability of Schizosaccharomyces pombe strain FBKL2.9SP-L14

[0048] The alcohol production capacity of the strain was tested by the alcoholometer method (refer to GB / T15038-2006).

[0049] Example 2

[0050] Application of a pombe schizosaccharomyces strain with high animal and plant aroma in the development of blueberry wine; the pombe schizosaccharomyces strain with high animal and plant aroma is classified and named as Schizosaccharomyces pombe, the laboratory strain collection number is recorded as FBKL2.9SP-L14, and it is deposited in the China Center for Type Culture Collection with a deposit number of CCTCC M20242770 and a deposit date of December 10, 2024. The diacetyl content of the wine sample obtained by fermentation with the strain was measured to be 1513.1276 μg / L, and the OAV value reached 3026.2552, which gave the blueberry wine a unique animal and plant aroma, such as cream aroma and milk fat aroma, and had a significant advantage in improving the flavor of the blueberry wine.

[0051] The blueberry wine brewing method comprises the following steps:

[0052] 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) 6 cells / mL, inoculated with pombe Saccharomyces pombe, which has high production of animal and plant aroma, into concentrated blueberry juice for alcohol fermentation. The temperature of the fermentation process 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 collected and properly stored at 4°C.

[0053] The inoculation amount of the pombe schizosaccharomyces strain with high production of animal and plant aroma is 10 6 cells / mL.

[0054] The method for determining flavor substances in blueberry wine comprises the following steps:

[0055] 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.

[0056] contrast:

[0057] The pombe Saccharomyces cerevisiae and the commercial Saccharomyces cerevisiae CECA isolated from Example 1 were used to brew blueberry wine in the form of single-strain inoculation. The flavor data of the obtained blueberry wine are as follows: Figure 3 According to the results of flavor analysis, the blueberry wine fermented by FBKL2.9SP-L14 has more flavor substances than CECA, and its aroma is more complex. Both wines are mainly composed of alcohols and esters. The substances with the highest content in the wine samples fermented by FBKL2.9SP-L14 are ethyl acetate and ethyl propionate. These ester compounds are usually related to fruity and floral aromas, and have a positive effect on enhancing the aroma of blueberry wine; β-damascenone and α-terpineol are more abundant in the blueberry wine fermented by FBKL2.9SP-L14. The higher content in the wine helps to provide richer floral and fruity aromas; the strain of the invention has significant advantages in producing diacetyl, and the diacetyl concentration of the brewed blueberry wine reaches 1513.1276μg / L, and the OAV value is as high as 3026.2552, which contributes to the creamy aroma of the wine. The strain significantly enhances the animal and plant aroma of the blueberry wine, increases the richness of the taste and the complexity of the flavor of the wine, meets the needs of consumers for unique flavor fruit wine, and shows great application prospects in the production of blueberry wine.

[0058] 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 pombe schizosaccharomyces cerevisiae with high yield of animal and plant aroma, characterized in that: It is deposited in the China Center for Type Culture Collection with the accession number: CCTCC M 20242770 and the preservation date: December 10, 2024.

2. The Schizosaccharomyces pombe having high yield of animal and plant aroma according to claim 1, characterized in that The high-yield animal and plant aroma is high-yield 2,3-butanedione (diacetyl).

3. The Schizosaccharomyces pombe having high production of animal and plant aroma according to claim 1, characterized in that At the same time, it also has excellent brewing characteristics of high glycerol and ethanol production, low hydrogen sulfide production, active killing, and degradation of organic acids.

4. Use of the pombe schizosaccharomyces cerevisiae with high production of animal and plant aromas according to claim 1 in wine making.

5. The use according to claim 3, characterized in that: The winemaking aspect is the brewing of blueberry wine.

6. Use of the Schizosaccharomyces pombe having high production of animal and plant aroma according to claim 1 in degrading organic acids.

7. Use of the Schizosaccharomyces pombe with high production of animal and plant aromas according to claim 1 in producing 2,3-butanedione (diacetyl).

8. Use of the Schizosaccharomyces pombe with high production of animal and plant aroma according to claim 1 in improving the animal and plant aroma of products.

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

10. A method for brewing blueberry wine to enhance the animal and plant aroma, characterized in that: The Schizosaccharomyces pombe described in claim 1 is used as a fermentation strain.

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