JPH048280A - Production of 'mirin' - Google Patents

Abstract

PURPOSE:To obtain 'MIRIN' [sweet SAKE (Japanese rice wine)], having strong sweetness and heavy taste and improved in tastiness and flavoring properties by the presence of an alcohol-resistant glutaminase in an aging process in producing the 'MIRIN'. CONSTITUTION:Glutinous rice is washed with water and then dipped in water. Rice KOJI (yeast) and an alcohol are added to produce 'MIRIN'. At that time, an alcohol-resistance glutaminase is then added to a material in an aging process for use. Halotolerant glutamiase originating from the genus Bacilius is preferred as the alcohol-resistant glutaminase because of excellent alcohol resistance thereof. The aforementioned alcohol-resistant glutaminase may be dissolved in an alcohol in charging the KOJI and added thereto or added at an optional time in the aging period of the 'MIRIN'. The amount added thereof is preferably about 0.005-0.01 unit based on 1ml raw material alcohol solution.

Description

[Detailed Description of the Invention] Industrial Field of Application The present invention relates to a method for producing mirin, more specifically, a method for producing mirin, which has a strong sweet taste.
This invention relates to an improved method for producing mirin that has a rich flavor and improved taste.

Conventional technology Mirin is an ancient Japanese seasoning, and is generally produced by adding alcohol and rice malt to steamed glutinous rice, non-glutinous rice, etc. to create mirin moromi, which is then brewed and aged at 20 to 30 degrees Celsius. . According to the Liquor Tax Law, mirin is
It is defined as "a product made by adding shochu or alcohol to rice and rice koji and straining it", and mirin also includes "mirin and other articles specified by government ordinance", "shochu or alcohol", or "mirinkasu". In addition to water, the items specified by the above Cabinet Order include water and water.
Contains corn, glucose, starch syrup, protein decomposition products, organic acids, amino acid salts, sake lees or mirin kasu.

In recent years, as an improved method for producing mirin, various enzymes such as α-amylase, glucoamylase, α-glucosidase, debranching enzyme, acidic protease, neutral protease, alkaline protease, and peptidase have been used in the manufacturing process. Therefore, various techniques are being studied to improve the total sugar utilization rate, total nitrogen utilization rate, etc., and efficiently produce the desired product in a shorter ripening period. Furthermore, a method has also been proposed in which non-fermentable oligosaccharides are produced by adding transglucosidase to enhance the rich flavor.

However, although the technology for shortening the ripening period using the enzymes mentioned above does not reduce the flavor, which is one of the important commercial values of mirin, it does not substantially reduce the flavor, which is another important characteristic. In fact, the longer the aging period, the better the taste. Thus, an improved method that satisfies flavor and taste properties at the same time has not yet been developed.

Problems to be Solved by the Invention The purpose of the present invention is to improve the conventional mirin manufacturing method,
In particular, the object of the present invention is to develop a new manufacturing technology for mirin that simultaneously improves flavor and taste, has a strong sweetness, is rich in taste, and has improved taste.

In view of the above-mentioned current situation, the present inventors focused on the fact that the use of enzymes in the mirin manufacturing process is effective in extracting the constituent components (extracts) from the raw materials, and further developed the method by effectively utilizing the constituent components of mirin. As a result of extensive research from the perspective of improving taste, we came up with the idea of applying glutaminase, which has never been used in the mirin production process, to mirin production, and developed glutaminase derived from Escherichia coli, which is commercially available as an enzyme reagent. was added during the preparation of mirin to ripen it, but contrary to expectations, the resulting mirin did not have much of an effect on improving taste. This means that the above enzyme is /'/
As a result of the alcohol tolerance test with I'//Iρ, the activity decreased to about 10% in the presence of 10% (V/V) ethanol, and the activity was inhibited in the presence of high concentrations of alcohol, as in the mirin aging process mentioned above. This was thought to be due to its insufficient effect.

However, as a result of subsequent research, a novel salt-tolerant glutaminase, which the present inventors had previously developed and applied for a patent for, was discovered (Japanese Patent Application No.
When the enzyme No. 82444) was used in the ripening process in the production of mirin, this enzyme had high alcohol tolerance in addition to the salt tolerance, and the resulting mirin had a significant increase in the amount of glutamic acid. It has been discovered that the flavor and taste can be improved and the above objectives can be achieved, and the present invention has now been completed.

That is, the present invention relates to a method for producing mirin, which is characterized in that alcohol-resistant glutaminase is added during the ripening process of mirin production.

According to the method of the present invention, as mentioned above, as a result of the presence and action of alcohol-resistant glutaminase during the ripening process, the flavor is markedly improved due to the enhancement of glutamic acid, resulting in strong sweetness, rich taste, flavor and taste. You can obtain mirin products with improved quality.

The method of the present invention requires the addition and use of alcohol-resistant glutaminase during the ripening process of mirin production, and except for the use of this specific enzyme, the specific method is substantially the same as the conventional mirin production method. For example, it can be produced by washing glutinous white rice, steaming it, mixing it with alcohol and rice malt, aging it, and then separating mirin lees by press filtration.

The alcohol-resistant glutaminase used above is selected from those that have the ability to convert L-glutamine into L-glutamic acid and is alcohol-resistant, and is particularly stable in mirin produced using it. Those that can maintain and exhibit excellent enzyme activity, especially p.
High alcohol (approximately 30-40% (v/
v) Degree) It is preferable to use a substance that exhibits little deactivation and can act effectively even in the presence of the compound. The enzymes include those derived from microorganisms belonging to the genus Bacillus, Aspergillus, and the like.

Among these, salt-tolerant glutaminase (Japanese Patent Application No. 1-82444), which was previously filed by the applicant, is particularly preferred. The salt-tolerant glutaminase originates from the genus Bacillus and has excellent alcohol resistance. In this specification, alcohol resistance refers to an alcohol concentration of 10 to 30% (v/v).
, pH 5-6, temperature 20-30°C, it means showing 50% or more activity.

The above-mentioned salt-tolerant glutaminase can be obtained, for example, from a commercially available enzyme preparation by the method shown in Examples below, and has the following physical and chemical properties.

a) Action: Hydrolyzes L-glutamine to produce L-glutamic acid and ammonia.

b) Substrate specificity: Km value for L-glutamine is 37°C, p) (6,0
(acetate buffer), and the L-glutamine and D-glutamine are degraded, but D- and L-asparagine, 2-glutamine, Dnp-Pro-Gln-
Gly and Dnp-Gln-11e-Ala-Gly-
D-Arg does not decompose.

C) Optimal pH and stable pH range: The optimal pH is 6 using L-glutamine as a substrate, and the stable pH range is 5 to 8.

d) Salt tolerance: 18% (w/y) at 37°C and pH 5.5
In the presence of sodium chloride, it exhibits a relative activity of about 90% or more of that in the absence.

e) Alcohol resistance: Shows 70-80% residual activity when kept at 37°C for 20 hours in 10-15% (v/v) alcohol.

The alcohol-resistant glutaminase used in the present invention, including the salt-tolerant glutaminase related to the applicant's application, may be dissolved in an alcohol solution and added at the same time when preparing the koji, or may be added separately after the alcohol is added.
It may also be added at any time during the ripening period of mirin.

Most effectively, a method of adding at all of the above periods can be adopted.

The amount added is approximately 0.001 units or more per 1 xi of the raw alcohol solution [1 unit is la per minute at 37°C and pH 6.0! The amount of enzyme that produces l1ot of L-glutamic acid, preferably about 0.005 to 0.01
It is preferable to select from a range of units.

When the above-mentioned alcohol-resistant glutaminase is added, the pH of mirin should be approximately the same as that of normal boat-fishing mirin, that is, about 4.5 to 8.0, and particularly preferably about 5 to 6. . Further, the alcohol concentration when the enzyme is allowed to act may be any standard concentration, and is usually about 20%. This is about 35 to 45% concentration of raw alcohol to alcohol ratio (alcohol liter/total US kg)
It is prepared by charging so that the amount is about 60 to 80%. Further, as the ripening temperature in the mirin manufacturing process of the present invention, it is appropriate to use a commonly used temperature, for example, a temperature of about 10 to 30°C.

The ripening period in the method of the present invention using the above enzymes can be arbitrarily selected depending on the type of raw materials, processing conditions, preparation composition, preparation operation conditions, temperature, etc., and is generally about 20 to 20 days.
At a temperature of 30° C., it is appropriate to use a ripening period of about 20 to 60 days. This aging fully exhibits the excellent effects intended by the present invention. This is confirmed by the increase in the amount of silicogeltanic acid in the resulting mirin product.

In addition, according to the method of the present invention, it is also possible to shorten this ripening period by using the above-mentioned specific enzyme.

Thus, according to the method of the present invention, mirin with significantly improved flavor and taste can be obtained. The resulting mirin product has a particularly rich and enhanced sweetness, and this sweetness is mild and has a pleasant aftertaste. Furthermore,
The mirin obtained by the present invention has the effect of removing the fishy odor inherent in mirin, alleviating the taste, preventing the elution of ingredients such as meat, increasing the texture and luster, preventing the cooking from collapsing, and enhancing the umami (hidden taste). Improvements have also been observed in this method, and the unique and excellent cooking effects can be achieved.

In addition, in the method of the present invention, appropriate amounts of various other conventionally known enzymes, such as protease, peptidase, amylase, etc., can be added in addition to the alcohol-resistant glutaminase, if necessary. Ba,
It may be possible to further improve quality such as taste.

Effects of the Invention According to the method of the present invention, by utilizing alcohol-resistant glutaminase, it is possible to provide mirin products of good quality, which are strong in sweetness, rich in flavor, and have improved flavor and taste properties.

EXAMPLES Hereinafter, in order to explain the present invention in more detail, examples of preparing enzymes used in the present invention will be given as reference examples, and then examples of producing mirin of the present invention will be given as examples.

The glutaminase activity on each side was measured by the following method.

<Glutaminase activity measurement method> 1% (w/v%) L-glutamine 0.5yA', 1M acetate buffer (pH 6,0) 0. 1xL test enzyme solution 0.1
yA' and water 0.311! After reacting the solution consisting of the above at 37°C for a predetermined time, the reaction was stopped by heat treatment at 100°C for 5 minutes, and the reaction solution was cooled in ice water. It is quantified using L-glutamic acid (manufactured by Boehringer Mannheim Yamanouchi).

One unit (U) of this enzyme is defined as the amount of enzyme that produces 1 Mmol of glutamic acid per minute at 37°C.

Reference Example 1 Preparation of glutaminase (1) Preparation of salt-tolerant glutaminase derived from the genus Bacillus (referred to as "enzyme A") Salt-tolerant glutaminase derived from the genus Bacillus was prepared by the following method.

That is, a commercially available enzyme preparation [Protin M3XJ (manufactured by Daiwa Kasei Co., Ltd.) 1300 g at 20 mM] was prepared using Lis-HCl buffer (pH 7).
, 5), and after thorough dialysis against the same buffer, insoluble materials were removed by centrifugation (12,000 rpm x 15 minutes).
DEAE-cellulose column (H7,5) equilibrated with
5.6 x 27cm.

Wako Pure Chemical Industries, Ltd.) and washed with 20mM) Lis-HCl buffer (pH 7.5, containing O, LM NaCl).
Glutaminase was eluted with NaCJ). Then the obtained 0.3M NaC containing glutaminase
Add ammonium sulfate to 1 elution fraction at a concentration of 0.8 saturation and incubate overnight for 4
After being allowed to stand at ℃, the mixture was centrifuged (12,00 rpm x 15 minutes) to collect a glutaminase active fraction.

The ammonium sulfate precipitate obtained was again diluted with 20mM'J acid buffer (p
Hydroxyl apatite (2,8
X20cm, Nacalai tesque
adsorbed on a 100-350 mesh (manufactured by que),
After washing with the same buffer, 20mM phosphate buffer (pH 7,4)
) to 400mM phosphate buffer (pH 7,4) while continuously increasing the buffer concentration, and the buffer concentration was 350mM.
The glutaminase active fraction eluted in M was then diluted with 10mM
DEAE-Toyopearl (1) was dialyzed against Tris-HCl buffer (pH 7.5) and the dialyzed solution was equilibrated with the same buffer.
, 6 x 7.5 cm, manufactured by Tosoh Corporation), and after washing with the same buffer, 10
Sephacryl S was eluted stepwise with Lis-HCl buffer (pH 7,5) and the eluted fraction of 0.15M NaCl was further equilibrated with 20mM Tris-HCl buffer (pH 7,5, containing 0,2M NaCl). -200 columns (2,
8x41cm, manufactured by Pharmacia), 1211/
2 zl portions were collected at a rate of 1 hour.

Finally, the active fractions were collected and dialyzed against 10mM phosphate buffer (pH 6,0) containing 25% ammonium sulfate, and then dialyzed against phenyl-Sepha 0-S C pre-equilibrated with the same buffer.
After adsorbing onto L-4B (1.6 x 5 cm, manufactured by Pharmacia) and washing with the same buffer, the ammonium sulfate concentration was reduced to 25.
% to 15% (W/V) while increasing ethylene glycol from 0 to 20% (V/V) to elute the glutaminase and obtain the desired salt-tolerant glutaminase (Enzyme A
) was obtained.

As a result of disk electrophoresis using a pH 8,017% gel, a single pore was revealed, and its specific activity was 17 units/■.

(2) Preparation of glutaminase derived from the genus Bacillus (derived from Proleather) (referred to as "enzyme B").
DEAE prepared by equilibrating a commercially available ProLeather (manufactured by Ohno Pharmaceutical Co., Ltd.) solution with the same buffer and thoroughly dialyzing it against 7,2).
) Passed through Yopar (manufactured by Tosoh Corporation) to adsorb glutaminase, followed by washing and removing impure proteins with the same buffer, eluting the glutaminase with the same buffer containing 0.3 M NaC, and separating the active fraction with 0.8 Salting out and recovery with saturated ammonium sulfate was then carried out in 10mM phosphate buffer (pH 7, 2
A gel was obtained using a Sephacryl S-200 column (manufactured by Pharmacia) equilibrated with ). According to the above procedure, a sample that does not contain protease and has a specific activity increased by 274 times can be obtained.

Example 1 After washing commercially available glutinous rice, it was soaked in water for 4 hours, drained for 30 minutes, and then steamed at normal pressure for 40 minutes.

For 200g of steamed glutinous rice obtained above, 5 pieces of commercially available rice koji
0 g 1 white liquor (35 degrees) 174111 0.01 unit/yl of various glutaminase was added, and after thorough stirring and mixing, the mixture was sealed tightly and aged at 20° C. for 30 days. still,
During the ripening period, stirring was performed once a week.

For comparison, commercially available glutaminase derived from Escherichia coli (manufactured by Sigma) was used.

Hereinafter, this will be referred to as a comparative enzyme C-added sample.

After the completion of the above-mentioned aging, the mash was centrifuged to obtain a mirin product sample as a supernatant.

The amount of monoglutamic acid contained in the obtained sample was measured using a Yamasa Shi-glutamic acid measurement kit (manufactured by Yamasa Soy Sauce Co., Ltd.).

The results obtained are shown in Table 1 below. For comparison, Table 1 also shows the same measurement results for mirin prepared in the same manner as above, except that glutaminase was not used (no addition).

Table 1 From the above table, it can be seen that in the sample to which enzyme A was added according to the method of the present invention, the amount of monoglutamic acid was increased by 225 μg/yl compared to sample G) without additive.

At the same time, a taste test was conducted on the three types of samples by a panel of 11 people as follows. That is, each panelist tasted two of each sample and asked them to answer which one they thought was tastier.

In addition, the amount of food eaten at one time was 0.5xA'.

As a result, 9 out of 11 people answered that the sample with the enzyme A of the present invention was more delicious when comparing the sample with the enzyme A of the present invention and the control sample without the enzyme C. In comparison with the sample, 9 out of 11 people used the enzyme A of the present invention.
They answered that the added sample tasted better. In addition, when comparing the comparative enzyme C-added sample and the control enzyme-free sample, 6 out of 11 people answered that the control enzyme-free sample was more delicious, and 5 people said that the comparative enzyme C-added sample was more delicious. .

From this, it can be seen that the taste of mirin obtained by the method of the present invention is clearly improved by the addition of a specific glutaminase (enzyme A) and is of excellent quality. There was no significant difference in taste between the enzyme C-added mirin and the control enzyme-free mirin, and no quality improvement effect was observed.

Furthermore, a taste test was conducted by 11 panelists similar to the above for the sample containing enzyme A of the present invention and commercially available hon mirin. As a result, 7 out of 11 people answered that the sample containing enzyme A of the present invention was more delicious, and 2 people answered that the commercially available mirin was delicious.

From the results of the above taste test, it is clear that the mirin obtained by the method of the present invention has a strong sweetness and a rich taste, and has been significantly improved in flavor, taste, and quality.

(that's all) (=Sa.

Agent: Patent Attorney Eiji Mitsue [“

Claims (1)

[Claims] (1) A method for producing mirin, which comprises adding and using alcohol-resistant glutaminase during the ripening process.