JPH10262652A - New yeast strain resistant to alcohol, sugar and high temperature and production of liquor with the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a new yeast strain capable of improving a raw material- utilizing rate and capable of producing conventional liquors maintained in their qualities by having resistances to alcohol, sugar and high temperature. SOLUTION: This new yeast strain belongs to Saccharomyces cerevisiae, is good in its growth in a culture medium containing alcohol in a concentration of 17.5%v/v, and has a death rate of <=10%, even when the cultured product is first extremely quickly heated and subsequently slowly heated, and has resistances against the alcohol, sugars and high temperatures. The yeast strain is deposited as FERM P-16136, and is preferably used to produce the liquor. Further, a glucoamylase preparation having a glucoamylase titer of 10000-50000 units per kg of rice is preferably added.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a novel yeast strain resistant to alcohol, sugar and high temperature, and a method for producing liquor using the same. More specifically, using a new yeast strain that is difficult to die even at the high temperature of ultra-short and gentle moromi, and using it, improving the raw material utilization rate by balancing the koji and glucoamylase preparation, the quality is not inferior to the conventional method The present invention relates to a method for producing alcoholic beverages.

[0002]

2. Description of the Related Art In the conventional sake brewing, koji rice is used in an amount of about 20% w / w and a fermentation temperature of about 15 ° C. or less based on raw rice. %
Sake is produced at a w / w lees percentage. In recent years, in order to produce sake with a high raw material utilization rate and inferior quality using coarse white rice with a milled rice ratio of 80% w / w or more, enzyme agents (gluc SB, gluc 100, etc.) ), The temperature of the mash is raised (up to 20 ° C), and the sake temperature is lowered after the middle period. (Journal of the Brewing Association, 73, 291 (1978)). In addition, in the preparation of all enzyme preparations which have been used as substitutes for koji, α-amylase and acid protease are involved in the dissolution of steamed rice. ing. However, in these production methods, it is considered that usable starch still remains in the lees, and there has been a demand for a method for producing alcoholic beverages having a higher raw material utilization rate.

[0003]

SUMMARY OF THE INVENTION In view of such circumstances, an object of the present invention is to produce alcoholic beverages with improved utilization of raw materials and maintaining the same quality as before.

[0004]

Means for Solving the Problems The present inventors have considered that the above object can be attained by allowing the temperature of the moromi to be made very short and long. Here, "Ultra-fast urgent mild" means that 3 to 7 days after charging, 5 to 10C higher than normal charging temperature.
25 ° C., hereinafter referred to as a charging method of 15 ° C. or less. However, a yeast strain that can withstand the temperature change of the moromi after a very short period of time and has grown vigorously under the conditions of high temperature in the early stage of the mash and high concentration of alcohol and sugar has not been found. Accordingly, the present inventors have intensively studied and succeeded in breeding and developing a yeast strain resistant to alcohol, sugar and high temperature. In addition to the improvement of the rate, it has been found that the use of koji is reduced, the work surface is improved by reducing the amount of lees, and furthermore, it is possible to save the cooling energy without the need for low-temperature charging, leading to the completion of the present invention. Was.

[0005] That is, one embodiment of the present invention relates to Saccharomyces cerevisiae.
, Glucose 15% w / v or more, ethanol 17.
Good growth on a medium containing 5% v / v or more, and alcohol, sugar and high temperature with 10% or less mortality in the upper tank when the temperature of the moromi is allowed to elapse shortly after It is a resistant yeast strain. Another aspect of the present invention is a method for producing alcoholic beverages, which comprises using the above-described yeast strain of the present invention.

When the yeast strain of the present invention is used, it is possible to brew in a desired ultra-short and gentle moderation, and the raw material utilization rate is improved.
In addition, while maintaining the conventional quality, there is no need to use koji, improve the work surface by reducing the amount of cake, and furthermore, there is no need for low-temperature mashing (usually cooling the mash to about 10 ° C after mashing). It is possible to provide a method for producing liquor that can save money.

[0007]

DETAILED DESCRIPTION OF THE INVENTION In general, the present invention breeds and develops yeast, which is resistant to alcohol, sugar and high temperature (it is difficult to die even at high temperature) by mutation treatment, and uses the yeast to determine the ratio of koji rice as raw material. The rice is reduced to 10 to 15% w / w (conventionally about 20% w / w), and the glucoamylase agent is supplemented by 10,000 to 50,000 units per kg of raw rice, and
This is a method for producing alcoholic beverages, characterized by ultra-rapid and gentle moromi at a high temperature of 20 ° C. to 25 ° C. for 3 to 7 days after adding.

[0008] First, the necessity of the ultra-early and suddenly mild moromi will be described below. The present inventors have previously found that inactivation of α-amylase is large in mash, but inactivation of glucoamylase, acid protease, and acid carboxypeptidase is small, and the study of α-amylase supplementation has been studied. went. 40 g of total rice, 140% v / v of pumping ratio, 20% w of koji ratio
The two-stage small preparation was carried out at / w, and the yeast used was Kyokai Yeast K-701, the fermentation temperature was controlled at 15 ° C, and the upper tank was placed on the 15th day. The enzyme is a heat-stable, liquefying enzyme agent Nagase CP-40 (manufactured by Nagase Biochemicals, mainly composed of α-amylase and low in other activities) added at the time of distillation (110 mg / kg of white rice, α-amylase activity 1) (0.76 × 10,000 units / kg of white rice). Table 1 shows the results of the analysis of the upper tank liquor.

[0009]

[Table 1]

As shown in Table 1, even if α-amylase was supplemented by a small charging method at a constant temperature of 15 ° C., the amount of liquid in the upper tank could not be increased. Next, in the early stage of the mash, the conventional mashing method is generally used to reduce the temperature to 15 ° C. or lower. The method was discussed. At the same time, the addition of Nagase CP-40 was attempted. FIG. 1 shows the temperature course, and Table 2 shows the analysis results of the upper tank liquor.

[0011]

[Table 2]

As shown in Table 2, even when the temperature was increased at the beginning of the mash and Nagase CP-40 was added, the amount of the liquid in the upper tank could not be increased. Since α-amylase was considered to be sufficient in mash, next, glucoamylase supplementation was examined. Glucoamylase used was glucoamylase "Amano" (manufactured by Amano Pharmaceutical Co., Ltd.). Also,
Since there is a possibility that α-amylase may become insufficient due to supplementation with glucoamylase, a Nagase CP-40 supplementation section was also provided. Table 3 shows the analysis results of the upper tank liquor.

[0013]

[Table 3]

[0014] As shown in Table 3, the amount of liquid in the upper tank increased in the glucoamylase-added group at a constant temperature of 15 ° C and at a high temperature. However, even if α-amylase was further supplemented to the glucoamylase supplementation section, the upper tank liquid volume did not further increase. Also in this case, it is considered that α-amylase is present in the mash in a necessary and sufficient amount. Also, 1
The amount of liquid in the upper tank is larger when charged at a high temperature than when charged at a constant temperature of 5 ° C., and it is presumed that raising the temperature in the early stage of mash leads to an increase in the raw material utilization rate. However, since the result was not reflected in the dissolution rate, it was examined whether there was any index indicating the raw material utilization rate other than the upper tank liquid amount. As a result, it was found that it would be better to measure the weight of the dried sake lees. 8,000 dried sake lees
The sake lees after centrifugation at 0 rpm for 10 minutes is dried at 135 ° C. for about 5 hours. In addition, when the koji ratio was 20% w / w, the degree of amino acids increased as the temperature increased, the koji ratio could be reduced. Table 4 shows the analysis results of the upper tank liquor.

[0015]

[Table 4]

The amino acid content is 15 at a koji ratio of 20% w / w.
It becomes about 1.4 times higher when the temperature is higher than the constant ℃. Koji commission 15
% W / w, about 1.1 times even at high temperature, koji rate 10% w / w
Showed similar or lower values. Therefore, it is considered that the koji ratio is preferably 10 to 15% w / w also from the viewpoint of sake quality. Next, when the temperature is raised in the early stage of the mash, the koji ratio is 20% w / w.
When comparing the temperature at 15 ° C and the high temperature over the non-enzyme-added category, the dried sake lees from 7.54g to 6.13g and the upper tank liquid volume from 58.5ml to 61.5ml. It is presumed that starch, which had become aged starch in the conventional charging method, also dissolves, leading to an increase in raw material utilization. The dissolution rate is constant at 15 ° C at a koji ratio of 20% w / w. Compared to the high temperature course, the amount of liquid in the upper tank increases, the dry sake cake decreases, and although the high temperature course clearly melts better, The dissolution rate decreases with increasing temperature. This may be caused by volatilization of alcohol due to a rise in temperature. Therefore, hereinafter, the index of the raw material utilization rate will be expressed by the weight of dried sake cake. Next, it was examined whether the high temperature progress was a temperature progress in which the amount of lees really decreased. Four additional temperature courses were added to the high temperature course at a constant temperature of 15 ° C. to carry out a small preparation. FIG. 2 shows the temperature course, and Table 5 shows the analysis results of the upper sake.

[0017]

[Table 5]

Dried sake lees tend to decrease when the integrated temperature is higher. However, even if the integrated temperature is the same 170 ° C, the dried sake lees is 4.61 g in the high temperature process, the temperature is raised in the early stage of the mash from the high temperature process, and 5.17 g in the temperature process C to raise up to 30 ° C, the higher temperature process is 5.17 g. It shows that your grades are good,
It was found that the adopted high-temperature course was a temperature course at which the amount of lees decreased. However, since the methylene blue staining rate (%) becomes slightly higher when charged with high temperature, yeast strains that are resistant to alcohol, sugar and high temperature (are difficult to die even at high temperature) are required, and breeding and development of such yeast strains are required. It was investigated.

Association yeast distributed by the Japan Brewing Association (Saccha
(B. romyces cerevisiae) K-7 strain was mutated, conditioned in a medium containing ethanol and glucose, and further increased in ethanol and glucose concentration.
Yeast strains that grew vigorously under conditions of high concentrations of alcohol and sugar were isolated. Among them, glucose 15%
Good growth on a medium containing w / v or more and ethanol of 17.5% v / v or more, and the mortality at the time of upper tank when the temperature course of mild moromi is very short and very short after is 10% or less. Has been found to be suitable for the production of the desired liquors. Such a yeast strain has not been found so far and is a novel yeast strain.

Thus, in the present invention, the ratio of koji rice is reduced to 10 to 15% w / w of the raw rice using the yeast, and the glucoamylase agent is reduced by 1 to 1 kg / kg of the raw rice.
Make up 0000 to 50,000 units, and add 3 to
Brewing of alcoholic beverages is carried out by ultra-rapid gentle moromi, which is a high temperature of 20 ° C. to 25 ° C. for 7 days. The glucoamylase agent to be used is not particularly limited and commercially available ones, for example, glucoamylase "Amano" (manufactured by Amano Pharmaceutical Co., Ltd.), Spitase M (manufactured by Nagase Seikagaku Corporation), Kokugen K (manufactured by Daiwa Kasei), Sumiteam (Nippon Kasaku Kogyo), glutase (Hankyu Bio-Industry), coclase (Sankyo) and the like can be used. In the production method of the present invention, the raw materials may be ordinary ones used for producing alcoholic beverages such as sake, and the process itself can be carried out according to a known sake brewing method. As a result, while improving the raw material utilization rate and maintaining the same quality as before, the reduction of koji consumption,
It is possible to improve the working surface by reducing the amount of cake and to eliminate the need for low-temperature charging, thereby making it possible to produce alcoholic beverages while saving cooling energy.

[0021]

EXAMPLES The present invention will be described in more detail with reference to the following Examples, but it should not be construed that the present invention is limited thereto. Example 1 Separation of Alcohol, Sugar and High Temperature Tolerant Yeast Strains When charged with high temperature, the concentration of alcohol and sugar is high at high temperature in the early stage of mash. Strains that vigorously grow under these conditions were isolated. B29 derived from the association yeast Saccharomyces cerevisiae strain K-7 distributed by the Japan Brewing Association
The strain was used to prepare a complete medium, YPD medium (glucose 1).
%, Peptone 1%, and 0.5% yeast extract manufactured by Difco, all in% w / v), and cultured with shaking at 30 ° C. for 1 day. This was harvested and washed, and 0.1% containing 2% w / v glucose and 3% w / v ethyl methanesulfonate.
M phosphate buffer (pH 8.0).
Minute shaking was performed by shaking for minutes. The mutated yeast was acclimated in a medium containing alcohol. Before acclimation, the concentration at which the cells vigorously grow in a medium containing alcohol and sugar was examined. Y containing 8, 10 or 12% v / v of ethanol
PD15 medium (glucose 15% w / v, peptone 1% w
/ V, yeast extract 0.5% w / v) was adjusted to 4.5 with lactic acid to prepare 3 ml each. YP in the medium
0.05 ml of the mutated B29 strain which had been statically cultured in the D medium for 2 days was inoculated at 0.05 ml each, and statically cultured at 25 ° C. for 5 days, and then the growth was examined. Table 6 shows the results.

[0022]

[Table 6]

The concentration for vigorously growing ethanol is 8% v
/ V, so that acclimation was performed with this concentration medium. Bacteria cultured in YPD medium at 30 ° C. for 2 days
5. 0.1 m in 3 ml of medium of ethanol 8% v / v, pH 4.5
and incubate at 25 ° C. for 3 to 7 days.
ml was again inoculated into the same medium, and a cycle of culturing at 25 ° C. for 3 to 7 days was repeated to separate the bacteria. This fungus was named Saccharomyces cerevisiae B29-K206-1. Subsequently, the cells were cultured on an agar medium containing higher concentrations of ethanol and glucose, and an attempt was made to isolate a resistant strain having higher resistance. Bacteria cultured on YPD agar medium at 30 ° C. for 1 day are suspended in sterilized water, and several drops are dropped on YPD 15 agar medium containing 15.8% v / v of ethanol.
After culturing for one day, bacteria having good growth were isolated. The separated bacteria were subjected to the same operation on an agar medium containing more ethanol, and bacteria that vigorously grew on a YPD15 agar medium containing 18.2% v / v of ethanol were isolated. This strain was named Saccharomyces cerevisiae B29-K206-1P. Table 7 shows the degree of growth of each bacterium.

[0024]

[Table 7]

The B29-K206-1P strain, which has been made more resistant than the B29-K206-1 strain, has glucose 15
% W / v and 18.2% v / v of ethanol, but the parent strain B29 could grow at an ethanol concentration of 15.8% v / v, but 17.5% v / v. It could not grow at a concentration higher than v. As shown in Example 2 below, normal yeast is easily killed in the course of high temperature, but B2
The 9-K206-1P strain has the same mortality as that of the control temperature course even after the high temperature course, and this resistant strain is a yeast having high temperature tolerance. The B29-K206-1P strain has been deposited on March 12, 1997 with the Institute of Biotechnology, Industrial Science and Technology under the accession number FERM P-16136.

Example 2 Using the B29-K206-1P strain, a small batch of 400 g of rice was prepared and a sensory test was conducted. The preparation method is 10% koji
In w / w, the amino acid content is somewhat low and other enzymes may be deficient. Therefore, in addition to glucoamylase "Amano", the enzyme agent Gluc 100 (manufactured by Amano Pharmaceutical) is also available. Added during distillation. FIG. 3 shows the temperature course of the control, and Table 8 shows the analysis results of the upper sake.

[0027]

[Table 8]

From Table 8, it can be seen that the methylene blue staining rate is the same as that of the control even at a high temperature, and the B29-K206-1P strain can be used without any problem even at a high temperature. The upper tank was opened at a time when the CO ₂ reduction was about the same, but there was no problem with the general analysis values.
The amino acid content was slightly increased when Gluc 100 was also added as compared to when only glucoamylase “Amano” was used. Dried sake lees has a high koji rate of 10% w / w,
Addition of the glucoamylase "Amano" resulted in the same or slightly reduced levels as the control. Glucoamylase "Amano"-
With the addition of Gluc 100, the amount of dried sake lees decreased by about 17% w / w. The combined use of the total enzyme preparation was effective in reducing dry sake lees. Next, a sensory test was performed using this upper tank liquor. The sample was subjected to activated carbon treatment after filtration through a filter. Table 9 shows the results of the sensory test.

[0029]

[Table 9]

As a result of the sensory test, it was found that the quality of the sake brewed after the lapse of high temperature was equivalent to that of the conventional method. Although not shown here, the preparation may be repeated several times and the evaluation may be rather good. Accordingly, it was found that sake having a quality equivalent to or higher than that of the control in terms of sensory quality was obtained. Example 3 Next, the effects of differences in the amounts of glucoamylase “Amano” and Gluc 100 added on dried sake lees, acidity, amino acidity, and sensory evaluation were examined. Table 10 shows the amount of enzyme added.
The classification was as shown in

[0031]

[Table 10]

Table 11 shows the results of the analysis of the upper liquor.

[0033]

[Table 11]

Glucoamylase "Amano", Gluc 10
As more 0 was added, the amount of lees decreased, but the acidity and amino acids increased. However, glucoamylase amano 125 mg / kg of white rice, Gluc 100 375 mg / white rice
kg addition category (glucoamylase activity 6.000 × 10,0
00 units / kg of white rice) and glucoamylase "Amano"
250 mg / kg of white rice and 375 mg of gluc 100 / kg of white rice added (glucoamylase activity 9.750 × 10,000)
The dry sake lees were 2.63 g and 2.64 g, respectively, in the unit (kg / white rice), which is considered to be the condition where the lees was reduced to 55.8% w / w of the control and the rice was dissolved to the maximum.
However, it is possible to add up to 50,000 units of glucoamylase titer, judging from the functional aspect. Therefore, judging comprehensively the results so far, B29-K2 isolated as alcohol, sugar and high temperature resistant yeast
By using the yeast of the strain 06-1P, even if the temperature progressed very quickly and very slowly, the koji ratio was 10% w / w and the glucoamylase titer was up to 50,000 units. Or it turned out that it is a brewing method that can obtain a higher rating. Although not shown here, similarly, when the koji ratio is 15% w / w, the glucoamylase titer is an analysis value up to 40,000 units,
It was found that the brewing method provided a sensory evaluation equal to or higher than that of the control.

[0035]

As described above, the production of liquor according to the present invention not only improves the utilization of raw materials, but also
It is possible to improve the working surface by reducing the amount of koji used and the amount of lees, and eliminate the need for low-temperature charging, thereby saving cooling energy. Further, according to the present invention, there is no inferiority in the analysis and sensory evaluation from that of the conventional method, and it is possible to obtain sake of equal or higher quality.

[Brief description of the drawings]

FIG. 1 is a drawing showing the temperature course in the study of a preparation method for raising the temperature in the early stage of mash.

FIG. 2 is a diagram showing a temperature course in a study of a reduction in the amount of cake due to a high temperature course.

FIG. 3 is a drawing showing the temperature course of a small preparation as a control in Example 2.

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[Procedure amendment]

[Submission date] April 10, 1997

[Procedure amendment 1]

[Document name to be amended] Statement

[Correction target item name] 0020

[Correction method] Change

[Correction contents]

Thus, in the present invention, the ratio of koji rice is reduced to 10 to 15% w / w of the raw rice using the yeast, and the glucoamylase agent is reduced by 1 to 1 kg / kg of the raw rice.
Make up 0000 to 50,000 units, and add 3 to
Brewing of alcoholic beverages is carried out by ultra-rapid gentle moromi, which is a high temperature of 20 ° C. to 25 ° C. for 7 days. The glucoamylase agent to be used is not particularly limited and commercially available ones, for example, glucoamylase "Amano" (manufactured by Amano Pharmaceutical Co., Ltd.), Spitase M (manufactured by Nagase Seikagaku Corporation), Kokugen K (manufactured by Daiwa Kasei), Sumiteam (Manufactured by Nippon Chemical Industries), glutase (manufactured by Hankyu Bio-Industry), coclase (manufactured by Sankyo) and the like can be used. In the production method of the present invention, the raw materials may be ordinary ones used for producing alcoholic beverages such as sake, and the process itself can be carried out according to a known sake brewing method. As a result, while improving the raw material utilization rate and maintaining the same quality as before, the reduction of koji consumption,
It is possible to improve the working surface by reducing the amount of cake and to eliminate the need for low-temperature charging, thereby making it possible to produce alcoholic beverages while saving cooling energy.

[Procedure amendment 2]

[Document name to be amended] Statement

[Correction target item name] 0027

[Correction method] Change

[Correction contents]

[0027]

[Table 8]

[Procedure amendment 3]

[Document name to be amended] Statement

[Correction target item name] 0034

[Correction method] Change

[Correction contents]

Glucoamylase "Amano", Gluc 10
As more 0 was added, the amount of lees decreased, but the acidity and amino acid content increased. However, glucoamylase amano 125 mg / kg of white rice, Gluc 100 375 mg / white rice
kg addition category (glucoamylase activity 6.000 × 10,0
00 units / kg of white rice) and glucoamylase "Amano"
250 mg / kg of white rice and 375 mg of gluc 100 / kg of white rice added (glucoamylase activity 9.750 × 10,000)
The dry sake lees were 2.63 g and 2.64 g, respectively, in the unit (kg / white rice), which is considered to be the condition where the lees was reduced to 55.8% w / w of the control and the rice was dissolved to the maximum.
However, it is possible to add up to 50,000 units of glucoamylase titer, judging from the functional aspect. Therefore, judging comprehensively the results so far, B29-K2 isolated as alcohol, sugar and high temperature resistant yeast
By using yeast of the strain 06-1P, the koji ratio is 10% w / w and the glucoamylase titer up to 50,000 units is equivalent to the control value in sensory evaluation and sensory evaluation even if the temperature progresses very quickly and quickly. Or it turned out that it is a brewing method that can obtain a higher rating. Although not shown here, similarly, when the koji ratio is 15% w / w, the glucoamylase titer is an analysis value up to 40,000 units,
It was found that the brewing method provided a sensory evaluation equal to or higher than that of the control.

 ──────────────────────────────────────────────────続 き Continuing from the front page (72) Inventor Masaaki Hamachi 4-9, Imazu, Iizazu-cho, Nishinomiya-shi, Hyogo Inside the Ozeki Research Institute (72) Inventor Chieko Kumagai 4-9, Imazu, Iizazu-cho, Nishinomiya-shi, Hyogo No. Ozeki Research Institute Co., Ltd.

Claims (6)

[Claims] 1. Saccharomyces cerevisiae (Saccha
romyces cerevisiae), good growth on a medium containing glucose 15% w / v or more and ethanol 17.5% v / v or more. Alcohol with a mortality of 10% or less in the tank,
A yeast strain that is resistant to sugars and high temperatures. 2. The yeast strain according to claim 1, which is FERM P-16136. 3. A method for producing liquor, comprising using the yeast strain according to claim 1. 4. The method for producing alcoholic beverages according to claim 3, wherein the ratio of koji rice to the raw rice used for brewing is 10 to 15% w / w. 5. The method for producing liquor according to claim 4, wherein a glucoamylase preparation having a glucoamylase titer of 10,000 to 50,000 units per kg of raw rice is supplemented. 6. 20 to 25 ° C. for 3 to 7 days after charging.
6. The method for producing alcoholic beverages according to claim 5, wherein the temperature of the liquor is high and the temperature of the liquor is set to 15 ° C. or less.