JPH06125734A - Production of protein seasoning solution - Google Patents

Abstract

PURPOSE:To provide the enzymatic agent obtained from the organic solvent immersion product of a koji, containing an exo type peptidase enzyme produced by the autodigestion of the koji, and efficiently decomposing protein to give a safe protein seasoning solution not containing a bitter taste, containing no salt, and having a body and an excellent taste. CONSTITUTION:The enzymatic agent is obtained from the organic solvent immersion product of a koji produced by the solid fermentation of microorganisms and contains an exo type peptidase enzyme produced by the autodigestion of the koji. The enzymatic agent is preferably obtained by immersing a koji such as soy koji in ethanol so as to give a final concentration of 6-15%, and subjecting the immersed koji to an autodigestion at 5-15 deg.C for 7-10 days. In order to obtain a protein seasoning solution by the use of the enzymatic agent, an animal or vegetable protein raw material is treated with a protein-solubilizing enzyme and subsequently with the enzymatic agent.

Description

Detailed Description of the Invention

[0001]

The present invention relates to an enzymatic agent containing an exo-type peptidase and a new protein seasoning solution using the same, and more specifically, amino acid or a bitter-free oligopeptide which is a proteolytic product as a taste component. The present invention relates to a method for producing a salt-free protein seasoning solution capable of significantly improving the content of glutamic acid, which is the main component of umami, and having no salt.

[0002]

2. Description of the Related Art Conventionally, a seasoning solution containing a protein hydrolyzate such as soy sauce or miso as a taste ingredient has been known, but it has been necessary to use a considerably large amount of salt. This is a cause of various restrictions on usage.
That is, soy sauce is generally produced as raw fried soy sauce by inoculating seed koji into a koji material mainly consisting of steamed soybeans to make koji, and then adding salt water to the koji to ferment the resulting moromi. , When the above-mentioned koji is charged, it prevents spoilage due to the propagation of microorganisms (miscellaneous bacteria) derived from koji, fermentation of moromi,
In order to ensure good ripening, high-concentration salt of about 23% is usually used. On the other hand, while giving a taste peculiar to soy sauce, the salt concentration of the product is set to a high concentration of 16-17%, This is a cause of restrictions on its use.

Further, conventionally, a method for producing an animal or plant extract by allowing a protein-solubilizing enzyme to act on an animal or vegetable protein raw material has been known. However, when such a protein-solubilizing enzyme is used, the obtained extract is bitter. It has a fatal drawback,
It is difficult to use it as a seasoning liquid. The bitterness can be eliminated by using exo-type protease, but the exo-type protease itself is very difficult to separate and purify, and it has not yet been commercialized. It has not been used in the production of seasoning liquids from.

On the other hand, animal and plant protein hydrolysates (HAP,
HVP), that is, a seasoning obtained by decomposing animal and plant proteins with hydrochloric acid has been conventionally known, and the demand thereof is gradually increasing as a part of natural seasonings (auxiliary seasoning) used for instant foods and the like. However, such HAP, H
Regarding VP, recently, there is one containing a mutagen, which is considered to be an organochlorine by-product formed when the above protein is hydrolyzed with hydrochloric acid, that is, a substance that damages a gene and causes a mutation in cells. It has been confirmed that its safety is becoming an issue.

Therefore, an object of the present invention is to provide the above HVP, H
An alternative to AP is to provide a new seasoning liquid which is safe, has no salt, has no bitterness, and has excellent taste.

[0006]

Means for Solving the Problems The inventors of the present invention have long conducted diligent research on a seasoning solution consisting of an enzymatic hydrolyzate of a seed animal and plant protein raw material, and as a result, as a result, the protein raw material of the edible animal protein raw material was first obtained. A method for providing a high-quality seasoning liquid containing an oligopeptide having a good taste and bitterness as a taste ingredient by allowing a solubilizing enzyme and an autodigestion solution of soy sauce koji to act was developed (Japanese Patent Publication No. 4- 20581).

Although the above seasoning liquid certainly has a good taste, it has a high salt concentration and is similar to soy sauce in this respect.

However, in the subsequent research, when the above-mentioned koji was immersed in an organic solvent such as ethyl alcohol for self-digestion, there was no fear of spoilage even under salt-free conditions, and the exo-type which exhibited a very excellent enzyme activity. We have found a new fact that a peptidase is obtained, which is effective for the production of the salt-free seasoning liquid of the above-mentioned purpose, and by using the peptidase, the desired salt-free seasoning liquid of the desired quality can be obtained. Further, the present inventors, the self-digestion by the alcohol immersion is not limited to soy sauce koji, can be applied to solid cultures of other various microorganisms, exo that can similarly exhibit excellent enzyme activity. We have also found the fact that an autolysate containing mainly peptidase of the type can be obtained, and the desired salt-free seasoning liquid can be produced by utilizing these substances. The present invention has been completed based on this new finding.

That is, the present invention provides an enzyme preparation and an animal or plant protein raw material which are obtained from a soaked product of koji in which a microorganism is solid-cultured and which contain an exo-type peptidase enzyme by autolysis of koji. , A protein-solubilizing enzyme is allowed to act, and then the above-mentioned enzyme agent is allowed to act.

The enzyme preparation of the present invention has an excellent exo-type peptidase activity and is suitable for producing a seasoning liquid without bitterness.

Further, the seasoning liquid of the present invention has no bitterness, is salt-free and safe, and can be concentrated into an extract. It can be used in a wide variety of processed foods or food materials, for example, instant foods. It can be widely applied to various uses such as paste products, hams and sausages, pickles, dressings, sauces, beverages, etc. to improve the taste and further supplement protein nutrition. In particular, the enzyme preparation of the present invention from soy sauce koji is, as shown in Examples described later, by applying it to animal and plant protein raw materials, the content of glutamic acid, which is the main component of umami, is improved, and there is richness. It is possible to produce a salt-free seasoning liquid having a very excellent taste.

Hereinafter, the method for producing the enzyme preparation of the present invention will be described in detail. The enzyme preparation of the present invention can be obtained from an organic solvent-immersed product of koji in which microorganisms are subjected to solid culture. Here, as typical examples of koji in which the above-described microorganism is solid-cultured, various soy sauce koji and miso koji which are well known in the past, for example, thick soy sauce koji, light mouth soy koji, and soy sauce koji can be exemplified. This is a koji starting material mainly consisting of steamed soybeans. Aspergillus oryzae, that is, Aspergillus oryzae, Aspergillus soya (Aspergillus oryzae).
pergillus sojae), etc., which have been inoculated and cultured with a bacterium belonging to the genus Aspergillus, but the present invention is not limited to this, and various other well-known microorganisms that are conventionally used for food, for example, Aspergillus niger and Penicillium spp.
ium, typically Penicillium ci, which attaches to beetles
trinum, etc., Rhizopus (Rhizopus, Rizopus niveus used for tempeh, etc.), Bacillus (Bacillus, Bacillus subtilis used for paste products, etc.), etc. Available. The above Bacillus solid culture also includes natto.
The solid medium for culturing various microorganisms may be soybean, which is the same as soy sauce koji, and various nutrients that do not hinder the growth of each microorganism, such as various proteins and sugars,
Fibers and the like may be appropriately selected and contained to have characteristics derived from the raw material.

The above-mentioned organic solvent-immersed product of each koji is prepared by immersing each koji in an appropriate organic solvent solution. Here, as the organic solvent, usually ethanol can be preferably used, but the organic solvent is not particularly limited thereto and may be, for example, isopropanol. The final concentration of the organic solvent is usually about 5
It is suitable to be used in the range of about 20%, preferably about 6 to 15%.

The above-mentioned koji is dipped in the above-mentioned organic solvent in an amount of about 1 to 3 times the weight of the koji.
The desired enzyme preparation of the present invention can be obtained by subjecting it to self-digestion for about 5 to 20 days, preferably for about 7 to 10 days, preferably under a temperature condition of about 5 to 15 ° C. However, when the temperature condition is higher than 20 ° C., the self-digestion is carried out under a salt-free condition, so that the growth of spoilage microorganisms cannot be completely avoided and a desired enzyme preparation product is obtained. Can be difficult.

Thus, the enzyme preparation containing the exo-peptidase desired in the present invention as a main component can be obtained. Of course, this can be purified by performing post-treatments and the like by various known methods, if necessary.

Next, the method for producing the seasoning liquid of the present invention will be described in detail. The seasoning liquid is prepared by allowing a protein-solubilizing enzyme to act on an animal or plant protein raw material and then acting on the enzyme preparation of the present invention obtained as described above. Can be manufactured.

The animal and plant proteins used as raw materials here include various types of foods containing proteins, which are provided for food.
For example, plant edible raw materials such as grains and corn meal, which are typified by wheat gluten, processing byproducts such as seafood, meat, and the like, such as seafood, cut meat produced during the processing of meat, dried bonito, dried sardines, etc. Examples thereof include animal food materials such as broth during production. They are used in the form of an aqueous liquid suitable for the action of the enzyme, that is, an aqueous solution, an aqueous dispersion, a suspension or the like, and particularly preferably in the form of an alkaline solution in order to facilitate the dissolution of the protein. Is desirable.

As the protein solubilizing enzyme, which is one of the enzyme agents for acting on the above-mentioned animal and vegetable protein raw materials, various known ones commonly used in this field can be used. Typical examples thereof are endo-type proteases, which include, for example, proteases of various microorganisms such as bacteria, molds, yeasts, actinomycetes, and proteases of animal origin such as rennin and pancreatin, and papain, bromelain, huaisin, etc. The plant-derived protease and the like are included. Of course, one kind thereof may be used alone, or two or more kinds may be used in combination. As the protein solubilizing enzyme,
In addition, alkaline protease, neutral protease and acidic protease can be used according to the pH condition of the raw material liquid, and the pH of the raw material liquid can be adjusted according to the type of each enzyme. Can also be adjusted to a pH at which is best exhibited. The amount of the above enzyme used can be appropriately determined depending on the type of the enzyme used and the like, and is not particularly limited, but usually about 50 per g of protein.
It may be selected from the range of 0 to 10000 units, preferably 2000 to 8000 units. In the case of an endo-type neutral protease, the above activity unit uses milk casein as a substrate, pH 7.0 and 30 μC at 1 μm / min.
It is calculated assuming that the titer of producing a decomposition product in an amount corresponding to g of tyrosine is 1 unit.

In the method of the present invention, first, the solubilizing enzyme agent is allowed to act on the animal and plant protein raw material. This is generally about 4 when the above-mentioned appropriate amount of enzyme is applied to the raw material.
0 to 65 ° C, preferably about 50 to 60 ° C, about 1 to 5
It is carried out by allowing it to act for about a time, thus hydrolyzing as much protein as possible into an oligopeptide having an average amino acid chain length of about 3 to 15 (soluble at a final trichloroacetic acid concentration of 0.17 mol). it can.

In the method of the present invention, after the completion of the above hydrolysis, the enzyme used once is inactivated, and thereafter, an enzyme agent mainly composed of exo-type peptidase by the self-digestion of koji of the present invention (koji auto-digestion liquid) To act. The pH of the solution on which the enzyme agent acts depends on the type of the raw material and the conditions under which the protein-solubilizing enzyme is acted, but generally it is acidic or neutral, which is suitable for expressing the activity of the enzyme agent. , No special adjustment is required. Of course, if desired, it is possible to adjust the pH conditions suitable for the action of the enzyme used in this step. The amount of the self-digesting solution of the above koji to be used can be appropriately determined, but it is usually selected from the range of about 0.1 to 2 g per 1 g of animal and plant protein raw material. In addition, the conditions under which the koji self-digesting solution acts are generally about 40 to 45 ° C. and about 1 to 5 hours. Thus, the seasoning liquid of the present invention, which has no bitterness and is excellent in taste, can be produced.

In the method of the present invention, the protein-solubilizing enzyme is first allowed to act on the raw material solution as described above, and then the autodigestion solution of koji is allowed to act, but in some cases, both enzymes are charged at the same time as the raw materials. Both enzymes can be allowed to act on the respective raw materials in one step to obtain a desired seasoning liquid, and such a method is also included in the present invention.

The seasoning liquid of the present invention thus obtained can be used as it is for various purposes, and if necessary, after-treatments and the like can be carried out according to various known methods to obtain products suitable for various purposes. You can also

As the above-mentioned post-processing means, for example, the following means and combinations thereof can be appropriately adopted.

(1) Solids and the like are removed by solid-liquid separation treatment such as centrifugation and filtration.

(2) Heat treatment (usually 5 at 70 to 100 ° C.)
The used enzyme is inactivated by about 30 minutes).

(3) Purification such as activated carbon treatment is performed to remove unnecessary odors and the like.

(4) Concentrate so that the solid content concentration is about 30 to 70%.

(5) A powdery seasoning is prepared by a drying means such as spray drying.

[0029]

INDUSTRIAL APPLICABILITY The self-digesting solution using the organic solvent for koji of the present invention contains, as a main component, an exo-type peptidase suitable for producing a seasoning solution without bitterness, and the activity of the enzyme is extremely high. The desired protein hydrolysis can be carried out efficiently in a very short time.

Further, according to the method of the present invention using the above-mentioned enzyme, the content of glutamic acid, which is the main component of umami, is high in a short time (usually within half a day), without bitterness, salt-free and safe,
It is possible to produce a seasoning liquid of good quality, which has a rich and very excellent taste.

[0031]

EXAMPLES Reference examples and examples will be given below to explain the present invention in more detail.

The components of the seasoning liquid in each example were analyzed by the following method.

(1) T. N (total nitrogen content, g / dl) By Kjeldahl method.

(2) F. N (formol nitrogen, g / d
l) By the formol titration method.

(3) Salt content According to the Mohr method.

[0036]

Reference Example 1 Production of Soy Sauce Koji Ethyl Alcohol Self-Digesting Solution 1 kg of concentrated soy sauce koji malt was immersed in 20% ethyl alcohol of double amount and left at 5-10 ° C. for 7 days to obtain a koji auto-digesting liquid. .

[0037]

[Test Example 1] Characteristics of auto-digestion solution of soy sauce koji with alcohol The activity expression of enzymes in koji in the alcohol soaked product of koji according to the present invention was compared with the case of normal soy sauce koji (in the presence of salt) as follows. Examined.

That is, 5 g of soy sauce koji (dark soy sauce koji) was homogenized in water to 100 ml, and the resulting solution was homogenized with a saline solution (final concentration 16%) or an ethyl alcohol solution (final concentration 15%). An enzyme solution was prepared, and each of them was allowed to act on each substrate solution containing salt or alcohol at a predetermined concentration, and each enzyme activity was examined.
Each enzyme activity was obtained and displayed by the following method.

<Protease activity> The neutral protease was as described above, and the acidic protease was titrated in the same manner as the neutral protease except that the pH was adjusted to 3.0.

<Carboxypeptidase (ACP) activity> Carbobenzoxy-glutamyl-tyrosine (Cbz)
-Glu-Tyr) as a substrate, and 1 at 60 ° C for 30 minutes at 30 ° C.
The activity of producing μg of tyrosine is defined as 1 unit.

<Leucine aminopeptidase (LAP)
Activity> Leucyl-glycyl-glycine (Leu-Gly
-Gly) as a substrate, the activity of releasing 1 μmol of amino acid (calculated as glutamic acid) in 1 minute at 30 ° C. is 1
Use as a unit.

<Glutaminase activity> Using L-glutamic acid as a substrate, the unit is the mg number of glutamic acid produced at 37 ° C. for 60 minutes.

The results obtained are shown in Table 1 below.

[0044]

[Table 1]

In Table 1, each enzyme activity is represented by a relative activity with water as a standard (100), and each enzyme activity (unit / g) in the standard is shown in parentheses below the above standard value. Is displayed.

From Table 1 above, it is clear that the alcohol-soaked soy sauce koji according to the present invention significantly improves the activity of any of the enzymes as compared with the case of using salt. In particular, the glutaminase activity was only 5% in the case of utilizing salt, whereas it was actually 95% in the case of utilizing alcohol according to the present invention, which shows substantially no activity inhibition or reduction. Therefore, it can be confirmed that this is very useful and useful for producing a seasoning liquid having excellent taste.

[0047]

[Reference Example 2] Production of isopropyl alcohol auto-digestion solution of Rhizopus koji Rhizopus koji was prepared as follows. That is, 50% corn protein grains, 42% wheat malt and 8% cornstarch (however, cornstarch was mixed with a portion of water by enzyme liquefaction)
A mixture of 20 kg of raw material containing 20 kg of water was used as a koji raw material, which was sterilized under pressure steam, cooled, and inoculated with Rhizopouniveus as an inoculum, and inoculated at 30 to 35 ° C.
It took 2 days to carry out koji culture.

Then, 30 liters of a 20% isopropyl alcohol solution was added to the koji, and the mixture was allowed to stand at 5 to 10 ° C. for 10 days to obtain a self-digesting solution of koji of the genus Rhizopus.

[0049]

[Example 1] Production of seasoning liquid of the present invention from wheat gluten 10 kg of wheat raw gluten (protein content of about 25%), caustic soda (granule) 32 g, protein solubilizing enzyme agent (Orientase 22B, Ueda Chemical Industry Co., Ltd.) (Manufactured by Bacterial Alkaline Protease) (68 g) and water (10 liters) are mixed to pH 1
A raw material-enzyme mixture solution of 0.5 was prepared, and this was mixed at 60 ° C. for 3 hours.
After leaving it to stand for a period of time to allow the enzyme to act on the raw wheat gluten,
The enzyme was inactivated by heating at 0 ° C for 15 minutes.

Next, 2.5 kg of the enzyme preparation (soy sauce koji ethyl alcohol self-digesting solution) obtained in Reference Example 1 was added to the obtained solution.
Was added and reacted at 45 ° C for 4 hours, then at 90 ° C for 1 hour.
The mixture was heated for 0 minutes to obtain a seasoning liquid (stock solution).

2 kg of activated carbon and 1 kg of perlite as a filter aid were added to the obtained stock solution, and the mixture was filtered using a small filter press to obtain 14.5 liters of filtrate.
T. N was 2.8%.

Then, the above filtrate was concentrated in vacuum to obtain 4.8 liters of concentrated seasoning liquid. This one is N8.4
%, F.I. The N content was 2.7%, the pH was 6.7, and the salt content was 0.08%.

From the above, it is clear that the use of soy sauce koji self-digested with alcohol gives a seasoning liquid of excellent quality.

[0054]

[Example 2] Production of seasoning liquid of the present invention from corn meal A mixture of 60 kg corn meal stock solution (protein content of about 15%), 300 ml of caustic soda solution (40%) and 250 g of a protein solubilizing enzyme agent (Orientase 22B) was mixed. A raw material-enzyme mixture solution (pH 9.5) is prepared by leaving it at 60 ° C for 3 hours to allow the enzyme to act on corn meal, and then at 45 ° C.
10.8 of the self-digesting solution of isopropyl alcohol of the knot of Rhizopus obtained in Reference Example 2
kg, and react at 45 ° C for 3 hours, then 90 ° C
And heated for 15 minutes to obtain a seasoning liquid (stock solution).

6 kg of activated carbon and 3 kg of perlite as a filter aid were added to the resulting seasoning liquid stock solution and filtered using a small filter press to obtain 43 liters of filtrate. T. N was 3.0%.

Then, the above filtrate was concentrated in vacuum to obtain 14.5 liters of concentrated seasoning liquid. This one is N8.8
%, F.I. The N content was 2.9%, the pH was 6.8, and the salt content was 0.06%.

Further, the obtained seasoning liquid had a high amount of glutamic acid, which is a main component of the taste component, and had an excellent taste property.

[0058]

Example 3 Production of seasoning liquid of the present invention from skipjack extract 20 kg of skipjack extract (protein content about 40%), 400 ml of caustic soda aqueous solution (40%) and protein solubilizing enzyme agent (Orientase 90N, Ueda Chemical Industry Co., Ltd. ), Bacterial neutral protease) (50 g) is mixed to prepare a raw material-enzyme mixture having a pH of 8.5, and the mixture is left at 60 ° C. for 3 hours to allow the enzyme to act on the skipjack extract and then cooled to 45 ° C. Then
6 kg of the enzyme preparation (soy sauce koji ethyl alcohol self-digesting solution) obtained in Reference Example 1 was added to the obtained solution, reacted at 45 ° C for 3 hours, and then heated at 90 ° C for 15 minutes to prepare a seasoning solution (stock solution). ) Got.

To the obtained stock solution, 1.5 kg of activated carbon and 1.5 kg of perlite as a filter aid were added and filtered using a small filter press to obtain 38 liters of filtrate. T. N was 3.6%.

Then, the above filtrate was concentrated in vacuum to obtain 18.5 liters of concentrated seasoning liquid. This one is N7.4
%, F.I. N3.2%, pH 6.5, salt content 2.02
%, And the taste was excellent.

The amino acid analysis of the seasoning liquid of the present invention obtained above was carried out by an amino acid analyzer (manufactured by Hitachi Ltd.,
835 high-speed amino acid analyzer).

The obtained results are shown in T.W. The amount of amylic acid per N1.0 (mg / dl) is shown in Table 2 below.

[0063]

[Table 2]

From Table 2, the seasoning liquid of the present invention has an amino acid composition corresponding to the constituent amino acid composition of the skipjack extract,
In particular, it can be seen that the content of histidine, taurine, glutamic acid, etc. is high, and that it has a good taste.

[0065]

Example 4 Production of Seasoning Solution of the Present Invention from Salmon Fish Approximately 3.2 kg of salmon fish was disassembled and only the meat portion was crushed with a mixer to obtain a crushed product having a protein content of about 20%).

2 kg of the crushed salmon fish body, 600 ml of an aqueous solution of caustic soda (5N), 400 ml of water and 11 g of a protein-solubilizing enzyme agent (Orientase 22B) were mixed to pH.
A raw material-enzyme mixture of 7.7 was prepared, and this was mixed at 60 ° C. for 2 hours.
After allowing the salmon fish body to react with an enzyme for a period of time, it was cooled to 45 ° C, and 400 g of the enzyme preparation (soy sauce koji ethyl alcohol self-digesting solution) obtained in Reference Example 1 was added to the resulting liquid, and the mixture was heated to 45 ° C.
For 3 hours and then heated at 90 ° C. for 15 minutes to obtain a seasoning liquid (stock solution).

To the obtained stock solution, 200 g of activated carbon and 200 g of perlite as a filter aid were added and filtered using Nutze to obtain 2.1 liters of filtrate. T. N was 1.9%.

Then, the above filtrate was concentrated in vacuum to obtain 0.6 liter of concentrated seasoning liquid. This one is N6.5
%, F.I. N2.7%, pH 6.8, salt content 0.04
%, And the taste was excellent.

With respect to the seasoning liquid of the present invention obtained in each of the above-mentioned examples, the taste of each of them was evaluated by eight panelists in three levels according to the following criteria, and each sensory test was conducted.

Rating 1 ... No taste is felt Rating 2 ... A little taste is felt Rating 3 ... Umami is strongly felt The results (shown by the total score of 8 persons) are shown in Table 3 below, and any seasoning liquid is delicious. It is clear that the

[0071]

[Table 3]

Claims (2)

[Claims] 1. An enzyme preparation, which is obtained from a soaked product of koji in which microorganisms are solid-cultured in an organic solvent, and which contains an exo-type peptidase enzyme by autolysis of the koji. 2. A method for producing a protein seasoning solution, which comprises allowing a protein-solubilizing enzyme to act on an animal or vegetable protein raw material, and then causing the enzyme agent according to claim 1 to act on the animal or vegetable protein raw material.