KR102018536B1 - Saccharomyces cerevisiae ksd-yc which is a novel yeast - Google Patents

Abstract

The present invention relates to KSD-YC of Saccharomyces cerevisiae, which has a high fermentation rate, low acidity, excellent sensory function, and has been deposited with accession number KACC 93276P.

Description

New Saccharomyces cerevisiae KSD-YC {SACCHAROMYCES CEREVISIAE KSD-YC WHICH IS A NOVEL YEAST}

The present invention relates to Saccharomyces cerevisiae KSD-YC. This is a new yeast isolated from yeast fermented with yeast.

Recently, makgeolli using various materials has been developed and consumed (Korean Patent Publication No. 10-2017-0030672, Korean Patent Registration No. 10-1723162, etc.). However, in order to vary the material and taste, the alcohol concentration, the fermentation temperature is varied, in this case, the survival rate and fermentation efficiency of the yeast is often not as good as desired.

The inventors of the present invention, while researching yeast with a high spectrum of use in the production of makgeolli, the new yeast separated from the yeast fermented with yeast is made fermentation with low acidity, high fermentation ratio and excellent sensory and flavor, and medium and low temperature In the fermentation is possible because the fermentation heat generated during the alcohol fermentation is less necessary because the cooling water is less process cost during the fermentation, the stability of the fermentation was confirmed that the alcohol yield and productivity can be improved and completed the present invention .

It is an object of the present invention to provide novel yeasts useful in the manufacture of makgeolli.

In order to achieve the above object, the present invention provides KSD-YC to Saccharomyces cerevisiae, deposited with accession number: KACC 93276P.

Saccharomyces cerevisiae KSD-YC of the present invention is fermented with low acidity during brewing fermentation, high fermentation ratio and excellent sensory and flavor, fermentation is possible at medium and low temperatures, stability of fermentation There is a feature to be maintained.

1 is a result of comparing the standard ITS1-5.8S rRNA-ITS2 nucleotide sequence included in the same species and genus as S. cerevisiae KSD-YC.
Figure 2 shows the taxonomic phylogeny according to the ITS gene sequence of S. cerevisiae KSD-YC.
Figure 3 shows the total acid change in alcohol upon fermentation at 15 ℃.
Figure 4 shows the total acid change in the alcohol during fermentation at 20 ℃.
Figure 5 shows the total acid change in alcohol upon fermentation at 25 ℃.

The present invention,

Saccharomyces cerevisiae KSD-YC isolated from fermentation using fermented traditional yeast. Saccharomyces cerevisiae KSD-YC of the present invention has a high fermentation rate at the time of fermentation at high temperature and low temperature, high yeast survival rate at low temperature, glucose, glycerol, sorbitol, methyl-D-glucoside, cellobiose, horse A sugar selected from the group consisting of toss, saccharose and raffinose can be used and accession number: KACC 93276P.

In addition, the present invention relates to a food composition in which KSD-YC is added to Saccharomyces cerevisiae.

In addition, the present invention

Preparing a one-stage fermentation product by mixing KSD-YC with rice, water, yeast and Saccharomyces cerevisiae of the present invention; And

It relates to a method for producing makgeolli comprising the step of fermenting the two-stage fermentation by adding rice, water and yeast further to the one-stage fermentation.

Hereinafter, the present invention will be described in detail.

Saccharomyces Cerevisiae KSD - YC

Saccharomyces cerevisiae KSD-YC of the present invention was isolated from the fermentation from fermented traditional yeast, high fermentation rate at the fermentation of alcohol at medium and low temperatures, high yeast survival at low temperatures. The middle temperature means 18 ° C. or more and less than 24 ° C., and the low temperature means 12 ° C. or more and less than 18 ° C. Preferably, the Saccharomyces cerevisiae KSD-YC of the present invention has a survival rate of at least 35% and less than 95% at 15 ° C, preferably at least 40% at 15 ° C. In addition, the saccharomyces cerevisiae KSD-YC can be used a sugar selected from the group consisting of glucose, glycerol, sorbitol, methyl-D-glucoside, cellobiose, maltose, saccharose and raffinose. , Accession number: KACC 93276P.

At this time, KSD-YC in Saccharomyces cerevisiae may have a survival rate of 40% or more at 15 ° C. and a survival rate of 5% or more at 20 ° C. during fermentation using rice as a raw material. In addition, KSD-YC in Saccharomyces cerevisiae has a fermentation rate of at least 80% at 25 ℃, at least 85% at 20 ℃, 95% or more at 15 ℃, the fermentation rate is calculated according to the following formula 1.

<Equation 1>

In addition, the Saccharomyces cerevisiae KSD-YC is 2-keto-D-gluconate, arabinose, xylose, adonitol, xylitol, galactose, inositol, N-acetyl-glucosamine, lactose, trehalose and It does not use a sugar selected from the group consisting of melezitose.

The Saccharomyces cerevisiae KSD-YC is derived from yeast fermented from yeast.

In addition, KSD-YC in the Saccharomyces cerevisiae of the present invention is capable of low acidity fermentation, so that it is possible to realize a neat quality without high-quality brewing rice, and it can have economic advantages, and yeast during fermentation at low temperature. The survival rate and fermentation rate are high, and because the spectrum of fermentation temperature is wide, it is not necessary to lower the heat of fermentation produced by alcohol fermentation with cooling water. .

Food composition

The present invention relates to a food composition in which KSD-YC is added to Saccharomyces cerevisiae. The type of the food is not limited, and may be general food, health supplement food, functional food, and the like. Preferably the food composition is a food composition in which yeast is used. More preferably, the food composition is liquor, for example, fermented liquor, distilled liquor, etc., such as medicinal liquor, takju, and sake.

Manufacturing method of rice wine

The present invention comprises the steps of preparing a single-stage fermentation by mixing KSD-YC in rice, water, yeast and Saccharomyces cerevisiae of the present invention; And adding rice, water, and yeast to the first stage fermentation to produce two stage fermentation by fermenting two stages. At this time, the manufacturing method of the makgeolli of the present invention may be performed using a general technique of preparing makgeolli using rice, water, yeast, and the like.

Advantages and features of the present invention and methods for achieving them will be apparent with reference to the embodiments described below in detail. However, the present invention is not limited to the embodiments disclosed below, but may be implemented in various different forms, only the present embodiments to make the disclosure of the present invention complete, and common knowledge in the art to which the present invention pertains. It is provided to fully inform the person having the scope of the invention, which is defined only by the scope of the claims.

<Materials and Methods>

Comparative strains were S1 in Saccharomyces cerevisiae, S2 in Saccharomyces cerevisiae and S3 in Saccharomyces cerevisiae (Table 1).

Experimental Example 1 Isolation of Strains

100 parts by weight of whole wheat flour are pulverized with a dry mill, and 40 parts by weight of water is added to make the water content 30-30% evenly, and placed on a disc with a thickness of 2.5 cm and a diameter of 20 cm. Fermentation was carried out. After that, it was dried for 7 days to make a traditional wheat yeast.

The traditional wheat malt and steamed rice and water were mixed and fermented at 15 ° C., 20 ° C. and 25 ° C. for 3 days to make drunk. Thereafter, the strains were separated by dilution with distilled water sterilized by distillation. Yeasts were selected from the isolated strains and the fermentation rate and alcohol production capacity of the yeasts were evaluated. Among them, a low total acid (total acid) and the fermentation rate and alcohol production ability, the sensory and flavor was selected to proceed to the following Experimental Example 2 (not described test results).

Experimental Example 2 Confirmation through Phylogenetic Analysis

In order to identify the species of the strain isolated from the traditional yeast, which was confirmed to have the best brewing aptitude in Experimental Example 1, the nucleotide sequence of the ITS gene was analyzed. The analysis was performed by sequencing using ITS1 and ITS4 primers from Solgent (Daejeon, Korea) (Daejeon, Korea). Specifically, the ITS gene full sequence was assembled using SeqMan software (DNASTAR) and blasted into the NCBI database. Taxonomy related ITS gene sequences were obtained from GenBank and compared with nucleotide sequences by BioEdit program.

In addition, the systematic location of the strain according to the gene sequence was constructed using the MEGA 3 program, using a neighbor-joining method having a bootstrap value based on the 1000th replication.

As a result, the nucleotide sequence of the ITS gene of the strain was as shown in SEQ ID NO: 1 (Table 2), comparing the ITS1-5.8S rRNA-ITS2 nucleotide sequence with the standard strains belonging to Saccharomyces cerevisiae species As a result, the strain was found to belong to Saccharomyces cerevisiae (Fig. 1). The taxonomy of taxonomy according to ITS gene sequence was the same as that of FIG. 2 (FIG. 2), and it was identified as a novel strain because the ITS gene sequence was significantly different from the conventional Saccharomyces cerevisiae. Therefore, it was named as KSD-YC in Saccharomyces cerevisiae and deposited with the National Academy of Agricultural Science on January 24, 2017 (Microbial Accession No .: KACC 93276P).

Experimental Example 3 Evaluation of Alcohol Resistance

Saccharomyces cerevisiae was evaluated for the alcohol resistance of KSD-YC. First, the incubated yeast of the log phase was inoculated in 30 mL of YPD medium containing ethanol at a concentration of 1, 5, 10, 15% (v / v), respectively, at a concentration of 1.0E + 05 cells / mL of the initial bacterial count, The cells were cultured at 30 ° C. and 150 rpm. After 24 hours of incubation, the culture solution was collected and diluted, and the absorbance was measured at 600 nm.

In general, the ethanol concentration above a certain concentration inhibits the growth of yeast and negatively affects the production of alcohol. Therefore, it is preferable as brewing yeast to have resistance to ethanol. As a result of absorbance measurement, yeast growth was generally good in the medium to which ethanol was added at a concentration of 5%. In particular, KSD-YC showed 18-42% higher resistance to Saccharomyces cerevisiae than other strains, and 15-460% higher resistance than other strains in medium containing 10% ethanol. Showed. Therefore, Saccharomyces cerevisiae KSD-YC was confirmed to have excellent alcohol resistance (Table 3).

Experimental Example 4 Evaluation of Tolerability

Generally, brewing yeast should be able to ferment alcohol while growing in high concentration of sugar. Therefore, we evaluated the glucose tolerance of KSD-YC in Saccharomyces cerevisiae. First, the incubated yeast of the log phase was inoculated at 30 mL of YPD medium containing glucose at concentrations of 2, 10, 20, 30 and 40% (v / v), respectively, at a concentration of 1.0E + 05 cells / mL of the initial bacterial count. Then incubated at 30 ℃, 150 rpm conditions. After 24 hours of incubation, the culture solution was collected, diluted appropriately, and the absorbance was measured at 600 nm.

As a result, the strains generally showed good growth in medium added with 20% glucose. In particular, Saccharomyces cerevisiae KSD-YC of the present invention showed a higher sugar resistance of 19 to 44% compared to other strains (Table 4).

Experimental Example 5 Brewing Characteristics

900 g of rice, 1350 mL of water, 8.6 g of yeast and yeast culture were mixed. At this time, the mixture was mixed so as to have a concentration of 6.5E + 07 cells / mL per mL of the lower liquor, and immersion was performed in one step. Then, the fermentation was carried out for 1 day at 15 ° C., 20 ° C. and 25 ° C. to carry out single stage fermentation.

After one-stage fermentation, 2,100 g of rice, 3,150 mL of water, and 20.1 g of yeast were additionally immersed in two stages, and then fermented in two stages under 15 ° C, 20 ° C, and 25 ° C, respectively. At this time, the temperature of the first and second stage fermentation was the same, each experiment was carried out using four types of yeast by type (S. cerevisiae KSD-YC, S. cerevisiae S1, S. S2 in cerevisiae, and S. S3 in cerevisiae).

The fermentation products, ie total acid content, alcohol concentration and fermentation ratio of the fermentation products were evaluated. At this time, the fermentation ratio was calculated according to the following formula (1).

On the other hand, the amino acid degree is neutralized with 0.1N NaOH to 10mL of the sample of the fermentation finished with filter paper, and then mixed with 5mL of neutral formalin solution and titrated with 0.1N NaOH solution to pH 8.3 It was evaluated by expressing it in the amount of 0.1N NaOH.

In addition, the survival rate of yeast during fermentation was calculated according to the following Equation 2.

<Equation 1>

<Equation 2>

As a result, KSD-YC in S. cerevisiae was lower than the other yeasts, it was judged that the total acidity of the liquor, which is a fermentation product, could be given a soft and light liquor (Tables 5 to 8, FIGS. 3 to 5). In addition, alcohol was produced at a high concentration, and the fermentation rate was high. In particular, it was confirmed that the alcohol concentration is higher than the other yeast produced at low temperature conditions, and the yeast survival rate is also high (Table 10, Table 11).

Total acid content of fermentation product in liquor fermentation according to fermentation temperature (% (w / v))

Total Acid Variation with Fermentation Time During Alcohol Fermentation at 15 ℃

Total Acid Changes with Fermentation Time at 20 ° C

Total Acid Changes During Alcohol Fermentation at 25 ° C

Amino Acid Degree in Alcohol Fermentation by Fermentation Temperature and Yeast Type

Survival Rate of Yeast by Fermentation Temperature and Yeast Types

Final alcohol concentration and fermentation rate according to fermentation temperature and yeast type

Experimental Example 6 Sensory Characteristics

In Experimental Example 5, the sensory test was carried out for the final fermentation by brewing yeast fermented and manufactured at 20 ° C. First, 15 trained inspectors were selected and the taste, aroma, and overall preferences were determined using a 9-point scale (1- Not Good, 3- Not Good, 5- Normal, 7- Good, 9- Good). Was investigated.

As a result, S. cerevisiae KSD-YC showed similar or high preference in all the items of flavor, taste and overall (Table 12). Therefore, it was judged that S. cerevisiae could give sensory excellent quality such as taste and aroma when brewed using KSD-YC.

Example 6, sensory test results for the final fermentation 20 ℃ ℃ by brewing yeast

Experimental Example 7 Evaluation of Sugar Utilization

The sugar utilization characteristics of KSD-YC in Saccharomyces cerevisiae were analyzed using API 20C AUX kit (Biomerieux, France). In this case, S1 was used as Saccharomyces cerevisiae and S2 was used as Saccharomyces cerevisiae.

As a result, KSD-YC in the Saccharomyces cerevisiae of the present invention was able to use glucose, glycerol, sorbitol, methyl-D-glucoside, cellobiose, maltose, saccharose and raffinose. However, Saccharomyces cerevisiae of the present invention contains 2-keto-D-gluconate, arabinose, xylose, adonitol, xylitol, galactose, inositol, N-acetyl-glucosamine, lactose, trehalose and melezitose. There was no availability (Table 13).

Depositary Name: Institute of Agricultural Biotechnology

Accession number: KACC93276P

Deposit Date: 2017124

<110> KOOKSOONDANG <120> SACCHAROMYCES CEREVISIAE KSD-YC WHICH IS A NOVEL YEAST <130> KNP170020 <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 789 <212> DNA <213> Saccharomyces cerevisiae <400> 1 attaaagaaa tttaataatt ttgaaaatgg atttttttgt tttggcaaga gcatgagagc 60 ttttactggg caagaagaca agagatggag agtccagccg ggcctgcgct taagtgcgcg 120 gtcttgctag gcttgtaagt ttctttcttg ctattccaaa cggtgagaga tttctgtgct 180 tttgttatag gacaattaaa accgtttcaa tacaacacac tgtggagttt tcatatcttt 240 gcaacttttt ctttgggcat tcgagcaatc ggggcccaga ggtaacaaac acaaacaatt 300 ttatttattc attaaatttt tgtcaaaaac aagaattttc gtaactggaa attttaaaat 360 attaaaaact ttcaacaacg gatctcttgg ttctcgcatc gatgaagaac gcagcgaaat 420 gcgatacgta atgtgaattg cagaattccg tgaatcatcg aatctttgaa cgcacattgc 480 gccccttggt attccagggg gcatgcctgt ttgagcgtca tttccttctc aaacattctg 540 tttggtagtg agtgatactc tttggagtta acttgaaatt gctggccttt tcattggatg 600 tttttttttc caaagagagg tttctctgcg tgcttgaggt ataatgcaag tacggtcgtt 660 ttaggtttta ccaactgcgg ctaatctttt ttatactgag cgtattggaa cgttatcgat 720 aagaagagag cgtctaggcg aacaatgttc ttaaagtttg acctcaaatc aggtaggagt 780 acccgctga 789

Claims (5)

Glucose, glycerol, sorbitol, methyl-D-glucoside, cellobiose, maltose, saccharose and raffinose can be used,
It has less than 0.168% total acid in brewing fermentation with rice as raw material at 15 ~ 20 ℃
Accession number: Saccharomyces cerevisiae KSD-YC, deposited under KACC 93276P.
The method of claim 1,
Saccharomyces cerevisiae KSD-YC. Has a survival rate of 40% or more during brewing fermentation using rice as a raw material at 15 ℃, the survival rate was calculated according to the following formula (2).

<Equation 2>

delete The method of claim 1,
Saccharomyces cerevisiae KSD-YC. It has a fermentation rate of 84% or more during fermentation with rice as a raw material at 15 ~ 25 ℃, the fermentation rate was calculated according to the following formula 1.


<Equation 1>

The food composition to which KSD-YC is added to Saccharomyces cerevisiae of Claim 1.

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