JPS60168382A - Aspergillus and preparation of malt using same - Google Patents

Abstract

PURPOSE:To produce a malt giving a soy product having characteristic flavor, with reduced carbohydrate consumption in malting, by using Aspergillus oryzae having low productivity of alpha-amylase as the malting aspergillus. CONSTITUTION:An Aspergillus oryzae having low productivity of alpha-amylase is used in the malt-preparation using an aspergillus belonging to Aspergillus oryzae. The aspergillus can be prepared by using an Aspergillus oryzae for brewing as the parent strain, subjecting the strain to a mutagenic treatment, and screening the strain having low productivity of alpha-amylase. The mutagenic treatment is e.g. the physical methods such as ultraviolet irradiation, X-ray irradiation, gamma-ray irradiation, etc., or chemical methods such as the treatment with a mutagenic agent such as N-methyl-N'-nitro-N-nitrosoguanidine, etc.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a koji mold having novel properties and a method for producing koji using the same.

Traditional seasoning foods such as soy sauce and miso are characterized by their flavor,
Each brewer has its own unique characteristics, especially when it comes to aroma. The main reason for this is said to be the koji mold used. For example, the smell of soy sauce brewing is mainly caused by Aspergillus oryzae. ) and Aspergillus sojae (
Aspergillus sojae, hereinafter referred to as "A. sojae". ) is used, but it is known that the productivity of hydrolytic enzymes and brewing characteristics are different between the two (
For example, Japan Soy Sauce Research Institute Magazine, Vol. 6, No. 8, No. 7
pp. 5-81 (1980), same magazine, Vol. 7, No. 4, No. 1
(See pages 66-172 (1981), etc.). That is, A
It has been reported that Oryzae has higher productivity of α-amylase and lower productivity of endopolygalacturonase than A and Soya, consumes more raw carbohydrates during koji making, and has a higher amount of pyrolysis. It is said that soya accounts for only about 20%, and most of the rest is A. oryzae. In order to investigate the reason why A. oryzae is used so frequently, we conducted a soy sauce brewing test under the same conditions using a large number of koji molds belonging to both species, and found that soy sauce prepared with A. oryzae and When comparing the products made by A. Oryzae in a sensory test, the products made by A. Sawyer had a strong burnt to systematic smell (most of the products were perceived as having an inferior rating).

The reason why such a difference arose even though they were prepared and aged under the same conditions is thought to be because there is a considerable difference in aroma production even at the koji stage. When A. Oryzae is used, even soy sauce koji using soybeans and wheat produces the so-called chestnut aroma that is present in rice malt, but with A. Soya, the so-called chestnut aroma is extremely weak and many produce a burnt odor.

On the other hand, during the koji making process, koji mold mainly uses carbohydrates in raw materials as an energy source to produce koji mold cells and various enzymes. At this time, a large amount of heat is generated, so operations such as ventilation and maintenance are performed to remove this heat from the system. Consumption of carbohydrates during the koji making process is essential for the formation of bacterial cells and enzyme production, but on the other hand, if the amount consumed is too large, less carbohydrates will be transferred to the strain process.
The strains produced from this will be poor in various components (sugars, alcohols, organic acids, etc.). The key point of koji making technology is how to reduce the consumption of carbohydrates and increase the efficiency (producing active ingredients such as bacterial cells and enzymes). It has been revealed that the carbohydrates consumed during koji making are mainly starch.

The present inventors investigated the relationship between the α-amylase activity of koji and the amount of carbohydrate consumed during koji production, and found a positive correlation between the two. Therefore, the present inventors took advantage of the features of A and Oryzae, which are excellent in brewing aroma production, while
We conducted research to create Aspergillus aspergillus that has the advantages of Sawyer, and finally completed the present invention. In other words, by using Aspergillus oryzae as a parent strain for brewing and applying ultraviolet irradiation to a conidial suspension of this strain, α-amylase productivity is weaker than that of the parent strain, but other properties are similar to that of the parent strain. They succeeded in obtaining mutant strains with similar characteristics.

The present invention relates to a novel mutant strain called A. oryzae with low α-amylase productivity. The present invention also relates to a method of using this novel mutant strain as a seed koji mold in a method for producing koji.

According to the present invention, A has excellent properties in aroma production, which is an important requirement especially in soy sauce brewing, and A has low α-amylase productivity while retaining the characteristics of A. oryzae.
For the first time, Aspergillus oryzae that also has the characteristics of Soya has been provided. By using the koji mold of the present invention in the production of noodles, the amount of carbohydrates consumed during koji production can be reduced in the same way as with A-soya, and in soy sauce brewing, sugars, alcohols, organic acids, etc. derived from carbohydrates can be reduced. It is possible to obtain a soy sauce rich in the various components described above, to reduce the weight of pasteurization, and to obtain a soy sauce product having the characteristic aroma of A. oryzae.

The koji mold of the present invention uses Aspergillus oryzae for brewing as a parent strain,
It can be obtained by subjecting it to mutation induction treatment and screening for low α-amylase producing strains.

Mutation induction treatment methods include physical methods such as ultraviolet irradiation, X-ray irradiation, r-ray irradiation, N-methyl-N'-nitro-N-nitrosoquanidine, 4-nitroquinoline-N-oxide, ethyl methanesulfonate, etc. Any chemical method by treatment with a mutagen can be employed.

For example, according to the ultraviolet irradiation method, the appearance rate is usually about 10-6.
The mutant strain of the present invention can be induced with a probability of approximately

In addition, the method for screening bacteria with low alpha-amylase virulence may be a method of selecting a strain with a small clear zone around the colony when the bacteria is grown on a flat plate medium containing corn starch.

Typical examples of such koji molds of the present invention include Aspergillus A and Oryzae-A-
287 can be mentioned. This strain was derived from Aspergillus oryzae F-1124 (Keikoken Bacteria No. 1865), which is used for soy sauce brewing, by UV irradiation treatment.
is further induced by ultraviolet irradiation treatment. This strain was transferred to the Institute of Microbial Technology, Agency of Industrial Science and Technology in February 1982.
February 6th, February 2017, February 6th, February 2018
489).

The mycological properties of A. oryzae A-287 are as follows.

1. Morphological properties (Kojijiru agar medium, cultured ao'c for 4 days at pH 6.0)
2. Physiological properties Table 3. Brewing properties 1/Into a Erlenmeyer flask, mix 1505F) watered defatted soybeans and 50g of roasted and cracked wheat, add 1200C14
After sterilizing under pressure for 0 minutes and cooling, two of each strain of Aspergillus oryzae were inoculated and cultured at 28°C for 48 hours to obtain koji. One bottle of each was used for koji analysis, and the other bottle was charged with 140 ml of 30% saline that had been sterilized under pressure for 15 minutes at 30°C.
After fermentation for 90 days, the sap and solid content were separated by filter paper filtration to obtain sap of various strains. The analysis results of the koji and the sap of various strains were as shown in Tables 8 and 4.

The enzyme activity and carbohydrate consumption were measured by the following method.

■ A method similar to the modification method of α-amylase activity Fuwa (Japanese Soy Sauce Research Institute Magazine, Vol. 6)
Vol. 8, pp. 75-81 (1980)). That is, soluble starch was used as a substrate (0.5% in the reaction solution).
The decrease in coloration of iodine starch was measured using OD7001m when reacting for 20 minutes at pH 4, 8, and 80°C. The amount of enzyme that reduces the blue iodine coloring corresponding to l tag under these conditions was defined as 1 unit.

(2) Protease activity Measured by a partially modified version of the Anson-Hagiwara method (Journal of the Japanese Society of Agricultural Chemistry, Vol. 68, p. 27 (1979)). The activity was defined as the amount of enzyme that releases 1 μq of tyrosine in 1 minute at pH 6.0 as 1 unit.

■ Glutamate colorimetric determination method using an automatic analyzer using glutaminase-activated glutamate dehydrogenase (Japanese Soy Sauce Research Institute Journal, Vol. 7,
Measured according to No. 2, pages 74-80 (1f18N).

■ Carbohydrate consumption After drying the sample in vacuum, crush it and collect a certain amount.
Hydrolysis was carried out with 6-hydrochloric acid in a boiling water bath for 3 hours, reducing sugars were determined by the Lane method, and carbohydrate consumption (%) was determined by the following formula.

8th table! As shown in Table 4, the strain of the present invention is almost similar to the parent strain, except for its lower productivity of α-amylase compared to the parent strain. The v1 quality is converted to V-f.The strain of the present invention having such properties can be used for making koji.

The degree of α-amylase productivity in the koji mold of the present invention is A
The standard is a level equivalent to Sawyer's α-amylase productivity. That is, the ability to produce alpha-amylase in amounts sufficient to degrade starch in the strains during brewing, such as during the maturation of soy sauce, is necessary. In the above α-amylase activity measurement method, approximately 0.5 to 5 x 10'
It is sufficient if the α-amylase productivity is within the range of 0.07 g.

The koji mold of the present invention can be used for producing koji for soy sauce and miso. The method for making koji at that time may be according to the usual method, and no special methods are required.

For example, in the production of soy sauce 7I11 koji, the seed koji of the koji mold of the present invention is inoculated into a mixture of ordinary koji raw materials, such as soybean raw materials that have been sprinkled with water and steamed, and wheat raw materials that have been roasted, steamed, and crushed;
A method of culturing at 5 to 85°C for 12 to 4 days may be used.

In preparing the final product from the koji thus obtained, conventional methods can be applied to the fermentation and maturation steps. In other words, in the case of soy sauce, mix soy sauce koji with salt water at an appropriate concentration according to the usual mixing ratio, stir as appropriate according to the usual method, and add 8 to 6
The final product can be obtained by fermentation and maturing for several months, followed by compression filtration and firing and clarification.

Hereinafter, an example in which the koji mold of the present invention is used for making koji in soy sauce brewing will be given as an example. However, the present invention is not limited to this in any way.

Example 1 15096 water was sprinkled on 5 kti of defatted soybeans, 2 kti/d
After pressure steaming for 18 minutes, 4.8 kQ of fried, steamed and crushed wheat was mixed with the mixture, which was cooled to 40°C to obtain powdered raw materials. (Microtechnical Research Institute No. 7489
The seed koji of No. 1) was inoculated and mixed, and koji was made in a small ventilation koji making device at a blowing temperature of 28°C for 24 hours, then at 26°C for 20 hours to obtain soy sauce koji.

Kono koji 10 kti lc 24.596 saline solution 15.
81 was added to one small container, and the mixture was fermented and aged at 15°C for 1 month and then in ao'c for 5 months. In addition, in order to promote lactic acid fermentation and alcohol fermentation, 14 hours after preparation.
On day 1, add 2 x 105/l of lactic acid bacteria isolated from various soy sauce strains.
85 days after preparation, yeast isolated from various soy sauce strains was added to 2x1
Added on 05/9.

``These strains were pressed using a small press to obtain raw soy sauce.

This raw soy sauce was heat-treated to reach a temperature of 80°C, and then kept at 50°C for 2 days to perform pasteurization.

Soy sauce was produced in the same manner using A. Oryzae 1065 as a control.

The analysis results and fired weights of these raw soy sauces are shown in Table 5. As is clear from Table 5, the raw soy sauce prepared using the strain of the present invention contained a significant amount of components derived from carbohydrates, and had a significantly low cooked weight.

Further, when the obtained heated soy sauce was tasted, it was found that the soy sauce produced using the bacterial cells of the present invention was sensually very unpleasant. The taste test method was conducted by a panel of 15 people using a 2-point preference test method, and the evaluation was performed using a rating of 1 or 2 in order of preference among the two points.

Table 6

Claims (1)

[Claims] 1) Aspergillus oryzae with low α-amylase production. 2) A method for producing koji, which comprises using Aspergillus oryzae with low α-amylase productivity as the koji mold.