JPH0583222B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Application Field] The present invention relates to a method for producing natto, and more specifically, to a method for producing natto with high umami and high viscosity using Bacillus subtilis, which is isolated from nature and has high protease activity and high mucilage production ability. be. [Conventional technology] Conventionally, commercially available natto is made by spraying commercially available natto bacteria spores on soybeans that have been steamed at 1.2 to 1.8 kg/ ^cm2 for about 25 to 50 minutes.
After putting this in a suitable container, ferment it and once the temperature reaches 45-50℃, keep it at this temperature for about 2 hours, then immediately lower the temperature to below 10℃ and ferment for 1-3 days. It is produced by aging. However, there are only two to three types of natto bacteria available on the market.
All have similar mycological properties, and the quality (flavor, stickiness) of natto obtained using them is uniform and there is almost no difference. [Problems to be Solved by the Invention] As described above, natto obtained using commercially available natto bacteria lacks flavor and also has insufficient viscosity, which is characteristic of natto. Therefore, the present inventors conducted intensive studies to develop a method for producing high-tasting, highly viscous natto, and as a result, they achieved the desired goal by using Bacillus subtilis NN-1, which was isolated from nature by Mr. Kiuchi and Mr. Nagai et al. They discovered that natto can be obtained and completed the present invention. [Means for Solving the Problems] The present invention provides that when cultured in an ordinary bouillon medium,
Use Bacillus subtilis NN-1, which has two properties: secreting protease of 100 units/ml or more and producing mucilage with a relative viscosity of 25 or more (4-fold dilution) in a mucilage production medium. The present invention provides a method for producing natto with high umami and high viscosity. The quality of natto is evaluated by its taste (umami), stickiness, aroma, etc., and umami accounts for a large proportion of these. The flavor of natto comes from the decomposition of soy protein by proteolytic enzymes (proteases) secreted outside the bacterial cells during the growth of Bacillus natto on soybeans.
This is due to the release of various amino acids and peptides. In addition, glutamic acid, which is a type of amino acid produced by the action of protease, is a flavor component as well as a component of the sticky substance unique to natto. I thought I would get natto. However, since commercially available Bacillus natto does not have sufficient protease activity, it is not possible to obtain the high-tasting natto that is the objective of the present invention. Therefore, the present inventors received Bacillus subtilis NN-1, which was isolated from nature by Mr. Kiuchi and Mr. Nagai of the Food Research Institute of the Ministry of Agriculture, Forestry and Fisheries, and attempted to produce natto using this strain. This strain has been deposited with the Institute of Microbial Technology, Agency of Industrial Science and Technology, Ministry of International Trade and Industry, and its accession number is FERM P-11319. The mycological properties of this strain are shown below. A Morphological properties Gram staining Positive size 0.8x2.0-3.0μm Morphology Short bacilli Motile Spores Bacterial cells do not swell B Cultural properties Morphology of colony growing on substitute meat juice agar medium The periphery is irregular. However, the colony as a whole is circular.The color of the colony is white.The luster of the colony is dull. Bacterial cells grown in a dull substitute meat juice medium have precipitated cells in the middle, and a bacterial film with no turbidity is formed. Thick C Physiological properties Catalase reaction Positive Does not grow under anaerobic conditions. Fouges-Proskauer test Positive acid generation Positive from glucose and mannitol, negative gas generation from arabinose and xylose Negative from glucose Hydrolysis of casein Positive Hydrolysis of gelatin Positive Hydrolysis of starch Positive citric acid assimilation Positive Tyrosine degradation Negative egg yolk lecithinase reaction Negative nitrate reducing ability Positive saline growth Growth in 2, 5, and 7%. It will not grow at 10%. Growth temperature: Grows at 5, 10, 30, 40, 50℃. Does not grow at 55 or 65℃. D Growth in the presence of 0.01% lysozyme Negative When producing natto using this strain, the soybean raw material is preferably grown at low pressure for a long time, e.g., at 0.5 to 2.0 kg/cm ² for about 20 to 200 minutes. It is preferable to use it by steaming it at 0.5 to 1.1 Kg/cm ² for about 60 to 200 minutes.
Also, it is desirable to keep the temperature of the product as high as possible during fermentation, and usually when the temperature of the product is 45 to 51% after starting fermentation.
Once the temperature reaches ℃, the product is maintained in this state for 2 to 6 hours, and then the temperature of the product is lowered to about 10℃ using a conventional method. [Example] Next, the present invention will be explained in detail with reference to Examples. Reference Example The extracellular protease activity of the present strain NN-1 was measured according to the method of Mr. Kiuchi et al. below. A platinum loop of this bacterial strain 1 was added to ordinary bouillon medium (0.5% meat extract, 1.5% peptone, 0.5% sodium chloride,
5.0 ml of 0.5% potassium monohydrogen phosphate (PH7.0) was inoculated. After culturing at 37°C for 26 hours, the cells were centrifuged (10,000xG, 10 minutes, 5°C) to obtain a supernatant.
Protease activity was measured by the following method. The reagents used are as follows. 1 Substrate solution A mixed solution of 1.0% acid-precipitated soy protein (manufactured by Fuji Oil Co., Ltd., Fuji Piure SP-300) and 0.1N sodium chloride, pH adjusted to 7.3 with sodium hydroxide solution. 2 Reaction stop solution A mixture of 0.1M trichloroacetic acid, 0.22N sodium acetate and 0.33N acetic acid Add 0.1ml of the above supernatant to 1.0ml of the substrate solution,
The reaction was carried out at ℃ for 60 minutes. Next, add the reaction stop solution.
4.0 ml was added and left at room temperature for 60 minutes to allow undegraded protein to precipitate. The reaction solution was centrifuged (1600xG, 15 minutes, room temperature) to obtain a supernatant.
The absorbance of this supernatant at 275 nm was measured.
Note that the blank was prepared by adding the supernatant liquid after adding the reaction solution. One unit of enzyme is the amount of enzyme that liberates a trichloroacetic acid-soluble substance equivalent to 1 μg of tyrosine per minute. Table 1 shows the results obtained.
It was shown to. Table 1 Strain activity (units/ml) This strain 123 Commercially available Bacillus natto (Company A) 62 Commercially available Bacillus natto (Company B) 95 Commercially available Bacillus natto (Company C) 83 As is clear from the table, this strain is more active than the control strain. also has high protease activity. Example 1 1 kg of Hokkaido vine soybeans was soaked in running water at 17°C for 15 hours, drained, and then soaked at 100 ml under a pressure of 0.8 kg/ ^cm2 .
Steamed for a minute. Next, Bacillus subtilis NN-1 was added to the steamed soybeans that had reached a temperature of about 80℃.
(FERM P-11319) was sprayed and filled into a small styrofoam container at a temperature of 40°C.
(room temperature) and 95% relative humidity for 10 hours. After that, the temperature was 42℃ and the relative humidity was 70%.
After fermentation was carried out for 2 hours at a temperature of 25°C (room temperature) and a relative humidity of 60% (4 hours at a product temperature of 48 to 51°C), it was aged at 5°C for 3 days. On the other hand, natto was obtained in the same manner except that a suspension of commercially available Bacillus natto (manufactured by Company C) was used as a control. The quality of these natto is shown in Table 2.

[Table] * mg%
Example 2 1 kg of small soybeans was immersed in running water at 17° C. for 14 hours, drained, and then steamed for 35 minutes under a pressure of 1.8 kg/cm ² . Next, Bacillus subtilis NN-1 (FERM P-
11319) was sprayed, filled into a small styrofoam container, and fermented for 10 hours at a temperature of 39°C (room temperature) and a relative temperature of 90%. after that,
After fermenting for 8 hours at a temperature of 37℃ (room temperature) and 70% relative humidity (approximately 2 hours at a product temperature of 45 to 48℃), and 2 hours at a temperature of 25℃ and 60% relative humidity, 5℃ It was aged for 2 days. On the other hand, natto was obtained in the same manner except that a suspension of commercially available Bacillus natto (manufactured by Company A) was used as a control.
The quality of these natto is shown in Table 3, and the evaluation results of the sensory test by 20 panelists are shown in Table 4.

[Table] * mg%

〔Effect of the invention〕

According to the method of the present invention, natto with high flavor and high viscosity can be efficiently produced.

Claims (1)

[Claims] 1. A bacillus that has two properties: secretes protease of 100 units/ml or more when cultured in ordinary bouillon medium, and produces mucilage with a relative viscosity of 25 or more (4-fold dilution) in mucilage production medium. - A method for producing high-umami, high-viscosity natto characterized by using Subtilis NN-1.