JP2985193B2 - Method for producing hydrolyzed gluten - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION INDUSTRIAL APPLICATION The present invention relates to an improved process for the production of gluten in the hydrous field.

[Conventional technology]

Production of hydrolyzed gluten having solubility, foaming properties, emulsifying properties, etc. by subjecting gluten, which is wheat protein, to hydrolysis treatment using a free or immobilized protease for the purpose of effective utilization of plant proteins. Has been done.

Then, the hydrolysis treatment of gluten with a protease is usually carried out by hydrolyzing the gluten derived from wheat, that is, the gluten obtained as it is from wheat, or the gluten previously treated with an acid or alkali with a protease, and then inactivating the protease. Then, the supernatant is purified after separating insoluble impurities to recover the hydrolyzed gluten. However, the hydrolysis of gluten by the protease is not sufficiently performed, and the amount of insolubles is increased, or the insolubles are often not separated and removed smoothly. It was difficult to obtain.

[Object and structure of the invention]

The present inventors have been conducting research for a long time for producing hydrolyzed gluten by subjecting wheat-derived gluten to hydrolysis and producing hydrolyzed gluten with a simple operation.

As a result, starch and fibers are present in the hydrolyzed solution after protease treatment in a swollen state, which hinders the separation and removal of insolubles from the hydrolyzed solution, It has been found that starch, fiber, and the like that are present slightly hinder the hydrolysis of gluten by protease. Further, the present inventors, when performing hydrolysis treatment of gluten derived from wheat with protease, such impurities such as starch and fiber contained in gluten are treated with hydrolyzate by amylase, hydrolysis of gluten by protease. The present inventors have found that the treatment and / or the separation and recovery thereof can be carried out smoothly, and the desired hydrolyzed gluten can be easily obtained in high yield, and the present invention has been completed.

That is, the present invention is a method for producing water-soluble hydrolyzed gluten, which comprises hydrolyzing wheat-derived gluten using a protease and an amylase.

Gluten is a mixture of proteins mainly composed of gluten and gliadin, and its composition is slightly different depending on the type of raw material and the production method. In the present invention, as gluten,
Those prepared from wheat can be used directly as they are, regardless of their composition and preparation method. At that time, the gluten may be in a raw gluten state or in a powdered form.

In the present invention, gluten obtained from wheat is subjected to a chemical treatment or a biological treatment with an enzyme or the like in advance, and the gluten obtained by reducing the molecular weight of the gluten or having a higher affinity with protease can be used. . Examples of such treated gluten include gluten, inorganic acids such as hydrochloric acid and sulfuric acid, organic acids, alkalis such as sodium hydroxide and potassium hydroxide,
Treatment under mild conditions using an enzyme (transglutaminase), etc. to form an amide bond in the side chain of gluten (such an amide bond forms many hydrogen bonds within and between gluten molecules and makes gluten insoluble. Gluten obtained from so-called deamidated gluten and wheat obtained by cutting protein) is treated with organic and inorganic reducing agents such as sodium bisulfite, 2-mercaptoethanol, dithiothreitol, L-cysteine, and reducing glutathione. There is a so-called reduced gluten in which the SS bond within and between molecules of gluten is cleaved. Therefore, the term "wheat-derived gluten" as used in the present invention has a meaning including gluten that has been previously treated as described above.

And although there is a little amount depending on the kind of gluten, all gluten contains starch and fiber, especially gluten as prepared from wheat has a large content.

In the present invention, one of the various glutenes as described above is used.
The species or two or more species are hydrolyzed with a protease and treated with an amylase.

The hydrolysis treatment with the protease and the treatment with the amylase may be performed simultaneously, the hydrolyzed gluten obtained after performing the hydrolysis treatment with the protease may be treated with the amylase, or conversely, the treatment with the amylase may be performed. After that, the treatment with the protease may be performed. Further, after amylase treatment, protease treatment or amylase treatment may be performed. However, depending on the type of protease and amylase, the working pH and temperature may be the same or different, so it is necessary to adopt a pH and temperature that is more appropriate for each enzyme actually used. When the protease and the amylase work at different pHs and temperatures, the treatment with the protease and the treatment with the amylase must be performed separately.

Gluten protease treatment and amylase treatment are usually preferably performed in a state in which glutenase is dispersed or dissolved in a liquid such as water, from the viewpoints of easiness of operation, purity and yield of the target substance, and the like.

Gluten is hydrolyzed and cleaved in the peptide bond of the main chain by protease treatment to reduce the molecular weight and increase the solubility in water and the like.

In addition, the amylase treatment hydrolyzes starch and fiber contained as impurities in gluten to reduce the molecular weight, which hinders the protease treatment of gluten, and the recovery of the produced hydrolyzed gluten. Interference is prevented.

During the protease hydrolysis treatment and the amylase hydrolysis treatment, the pH of the gluten-containing solution, the gluten concentration,
This is performed by appropriately adjusting the temperature and the like.

The protease used in the present invention is not particularly limited as long as it can hydrolyze and cleave a peptide bond in gluten. For example, pepsin, trypsin, chymotrypsin, acid protease derived from Hitake mushroom, acidic protease derived from Aspergillus, papain, bromelain, and many others can be used. As long as the proteases do not adversely affect each other, a plurality of proteases may be used in combination. For example, pepsin and another acidic protease may be used in combination.

Further, as the amylase, α-amylase, β-amylase, glucoamylase, isoamylase, oligosaccharide-forming amylase, and the like can be used, and a plurality of amylase may be used in combination as long as they do not adversely affect each other. Although α-amylase can be used, α-amylase is preferred in view of the purity and yield of hydrolyzed gluten.

The conditions of the protease hydrolysis treatment and the amylase hydrolysis treatment are different depending on the situation (for example, the type of protease or amylase, the difference in the treatment method such as whether the protease treatment and the amylase treatment are performed simultaneously or separately,
Optimal conditions such as pH and temperature may be selected according to the use form of each enzyme. It does not prevent the removal of impurities in the middle of both processing steps.

For example, after performing protease treatment using pepsin, an acidic protease derived from Hilotake mushroom, and an acidic protease derived from Aspergillus, α-produced from Bacillus
When the treatment is performed using amylase, the pH is about 1.5 to 4.
5.After performing protease treatment at a temperature of about 30-50 ° C, pH
Is adjusted to about 4.5 to 7.0, and then treated with amylase.

The protease and / or amylase may be added to the treatment solution in a free state, or may be used after immobilization. When used in a free state, it is usually preferable to use about 0.1 to 0.5 g of protease and about 0.1 to 5.0 g of amylase per 100 g of dried gluten. When used by immobilization, any method such as a carrier binding method and a cross-linking method can be employed regardless of the immobilization method. It is practical to immobilize it on a carrier such as a bead, a membrane or a net. When using immobilized protease, the amount of gluten in the treatment solution should be about 20-60 g /
When the treated liquid is passed through a bed on which about 10 to 50 mg of protease is immobilized per 1 g of the carrier at a rate of about 1.0 to 6.0 hr ^-1 (residence time is about 10 to 60 minutes), the yield and the treatment time,
Good results are obtained in terms of the foaming properties of the product. When the amylase is immobilized and used, the amount of gluten in the treatment solution is set to about 20 to 60 g /, and the treatment solution is applied to a bed having immobilized about 20 to 150 mg of amylase per 1 g of the carrier in an amount of about 1.0 to 1.0 g. When the solution is passed at a rate of up to 6.0 hr ^-1 (residence time about 10 to 60 minutes), good results are obtained in terms of yield, treatment time, and the like. When the protease and / or amylase is immobilized and used, the yield decreases when the concentration of gluten in the treatment liquid is higher than the above range, and the protease is inactivated when the treatment temperature is 60 ° C. or higher.

If the protease hydrolysis treatment and the amylase hydrolysis treatment are performed separately, the enzyme used first must be removed or completely inactivated if the enzyme used first does not interfere with the subsequent enzyme treatment. There is no.

In recovering the hydrolyzed gluten from the treatment solution, the pH and temperature of the hydrolyzed gluten-containing solution are adjusted at the time when both the protease treatment and the amylase treatment have been completed to inactivate the enzyme. For example, as a protease, for example, pepsin, an acidic protease of Hilotake origin, using an acidic protease of Aspergillus origin,
When α-amylase is used as the amylase, the enzyme is inactivated at a pH of about 4.5 to 7.0 and a temperature of about 60 to 100 ° C. Next, the inactivated enzyme, undegraded gluten,
It is preferable to employ a method in which insoluble impurities such as starch substances are separated and removed by an appropriate method, and the hydrolyzed gluten contained in the residual liquid after the removal of the insoluble matter is recovered by drying or the like.

Hydrolyzed gluten produced by the method of the present invention,
It is soluble in water and usually has an average molecular weight of about 5,000 to 20,000. The hydrolyzed gluten is generally white and has no unpleasant taste and off-flavor.

Furthermore, the hydrolyzed gluten produced according to the present invention is:
It is also excellent in foaming properties such as foaming power and foam powder stability, and has good dispersibility and processability. In addition, since it is derived from grains such as wheat and is highly safe, it is extremely effective as an additive for food processing, and especially produces confectionery such as cakes, cookies and icing and kneaded products such as bread, kamaboko and hampon. It is suitable as a foaming agent. The hydrolyzed gluten produced according to the present invention can be stored and used in any form of powder, paste and solution.

〔The invention's effect〕

In the method of the present invention, high-purity hydrolyzed gluten can be obtained in a high yield by a simple operation, compared with the conventional technique in which only protease treatment is performed without treatment with amylase.

〔Example〕

Hereinafter, the present invention will be described by way of examples, but the present invention is not limited to those examples.

Example 1 134 g of powdery gluten (100 g as protein) prepared from wheat flour, 30 ml of 1N hydrochloric acid and 834 ml of distilled water were mixed and dispersed by stirring with a stirrer to prepare a dispersion having a pH of 3.0. After adding 0.5 g of pepsin derived from swine stomach (manufactured by Amano Pharmaceutical Co., Ltd.) to the dispersion and reacting at 45 ° C. for 3 hours, the liquid was divided into five equal parts.

In the first section, pepsin was inactivated by directly heating at 80 ° C. for 30 minutes, then cooled to room temperature, and centrifuged at 6500 G for 10 minutes to remove undissolved substances to obtain a supernatant.

In the second category, the pH of the solution was adjusted to 4.N with 5N sodium hydroxide.
After adjusting to 7, 0.1 g (0.5% based on the weight of gluten) of α-amylase (manufactured by Ueda Chemical Industries, Ltd.) was added, and the mixture was treated at 80 ° C. for 0.5 hours.
For 30 minutes to inactivate the α-amylase. Next, the solution was cooled to room temperature, and centrifuged at 6500 G for 10 minutes to remove an undissolved substance to obtain a supernatant.

0.2 g of α-amylase (1.0 g based on the weight of gluten)
%) Was applied to the third section in the same manner as for the second section, to obtain a supernatant.

0.4 g of α-amylase (2.0 g based on the weight of gluten)
%) Was processed in the same manner as for the second section, except that%) was added to obtain a supernatant.

0.8 g α-amylase (4.0 g based on the weight of gluten)
%) Was processed in the same manner as for the second section except that%) was added to obtain a supernatant.

While measuring the amount of the supernatant obtained in each of the above with a measuring cylinder, using a spectrophotometer wavelength 280n
The absorbance (hereinafter, referred to as “Abs ₂₈₀ ”) of each supernatant at m was measured and fitted to a calibration curve previously determined by the following method to measure the protein concentration in the supernatant.

[How to obtain calibration curve]

Hydrolyzed gluten whose protein content was measured in advance by the Kjeldahl method was dissolved in water to a concentration of 1 mg / ml. This was diluted to give hydrolyzed gluten solutions of 0.2, 0.4, 0.6 and 1.0 mg / ml, respectively. When Abs _{280 of} each solution was measured, the following results were obtained. Hydrolyzed gluten concentration (mg / ml) Abs ₂₈₀ 00 0.2 0.112 0.4 0.230 0.6 0.334 1.0 0.584 From the above results, the concentration of hydrolyzed gluten in the supernatant was
The relationship with Abs ₂₈₀ , that is, the calibration curve is represented by the following equation.

Hydrolyzed gluten concentration (mg / ml) = 1.72 x Abs ₂₈₀ The amount of the supernatant obtained above and the hydrolyzed gluten concentration are shown in Table 1 below.

The yield of the finally obtained hydrolyzed gluten was determined by the following equation.

After diluting the supernatant obtained in each case 5-fold,
The transmittance at 0 nm was measured and expressed as a ratio (%) to the transmittance of distilled water at 610 nm of 100.

The yield of the hydrolyzed gluten obtained above and the transmittance of the supernatant are shown in Table 1 below.

From the results in Table 1 above, it can be seen that, in the present invention in which the amylase treatment was performed during the hydrolysis treatment of gluten with the protease, the yield of hydrolyzed gluten was increased as compared with the case where the amylase treatment was not performed, and the supernatant was used. Liquid permeability is also high.

Example 2 134 g of powdered gluten prepared from wheat flour, 1N citric acid
10 ml and 856 ml of distilled water were mixed and stirred and dispersed by a stirrer to prepare a dispersion having a pH of 4.5. An acid protease of Aspergillus origin (Protease M manufactured by Amano Pharmaceutical)
After adding 1 g and 2 g of Bacillus-derived α-amylase (manufactured by Ueda Chemical Industries, Liquefaction Enzyme T), the mixture was reacted at 45 ° C. for 5 hours, and then heated to 80 ° C. and maintained for 60 minutes. Thereafter, the temperature of the solution was maintained at 90 ° C. for 10 minutes to inactivate the acidic protease and α-amylase. Then, after cooling the liquid to room temperature, 6500
The supernatant was obtained by removing the undissolved material by centrifugation at G for 10 minutes.

The supernatant volume was 785 ml, the protein concentration was 85.0 mg / ml, and the transmittance was 86.5%.

Example 3 Gluten dispersion prepared in the same manner as in Example 2
Add 1 g of Bacillus-derived α-amylase (manufactured by Yakult Yakuhin Kogyo: Uniase), heat to 90 ° C, and bring to 60 ° C.
Hold for 30 minutes. After cooling to 50 ° C., an acidic protease originating from Hilo mushroom (rapidase, manufactured by Takeda Pharmaceutical Co., Ltd.) was added and reacted for 7 hours. Then, it was kept at 80 ° C. for 30 minutes to inactivate the acidic protease.

Next, the solution was cooled to room temperature, and centrifuged at 6500 G for 10 minutes to remove an undissolved substance to obtain a supernatant.

The supernatant volume was 780 ml, the protein concentration was 85.6 mg / ml, and the transmittance was 87.2%.

Continued on the front page (72) Inventor Shinichi Fukudome 3-4-8 Suehirocho, Kawagoe-shi, Saitama Prefecture (72) Inventor Takaki Yamagata 672 Minamitakada-cho, Nagahama-shi, Shiga Prefecture Corpora Takata 105 (72) Inventor Toshio Tanaka Shiga Prefecture 10-52 Jinmae-cho, Nagahama-shi Excellent 105 Shinmae 105 (72) Inventor Jun Nakamura 8-5-5 Okusawa, Setagaya-ku, Tokyo (56) References JP-A-54-95760 (JP, A) JP-A-54 -122762 (JP, A) JP-A-1-47353 (JP, A) Mitsubishi Heavy Industries Technical Report Vol. 26, No. 5 (1989. Sep.) p. 425-429

Claims (2)

(57) [Claims] 1. Gluten derived from wheat is hydrolyzed using a protease and an amylase, and then the enzyme is inactivated at a pH of 4.5 to 7.0 and a temperature of 60 to 100 ° C., and insoluble impurities are separated and removed. A method for producing water-soluble hydrolyzed gluten having an average molecular weight. 2. Hydrolysis by a protease and hydrolysis by an amylase are carried out simultaneously, hydrolysis by a protease followed by hydrolysis by an amylase, or hydrolysis by a protease followed by hydrolysis by a protease. 2. A method for producing the hydrolyzed gluten according to 1.