JP2931022B2 - Edible peptide-containing substance and method for producing the same - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to an edible peptide-containing composition obtained by hydrolyzing a vegetable protein with an enzyme and removing aromatic amino acids, and a method for producing the same.

[Background Art] Conventionally, as a hydrolyzate of a vegetable protein, a hydrolyzate decomposed using an acid or an alkali or a hydrolyzate decomposed using an enzyme is known. These hydrolysates are used as seasonings or amino acid feedstocks.

Recently, animal proteins such as egg white and casein have been enzymatically degraded into enzymatic hydrolysates with high peptide content.
Attempts have been made to utilize this for artificial nutrition or diet therapy.

Furthermore, Japanese Patent Application Laid-Open No. 63-287462 proposes that a soybean protein is hydrolyzed by an enzyme into an oligopeptide having a peptide chain length of 2 to 10, and this oligopeptide is used as a nutritional supplement. According to the publication, this peptide is superior to amino acids in rapid absorption, and is reported to be excellent as a quick-acting nutritional supplement at the time of sports and other severe physical exhaustion.

On the other hand, various health drinks containing various amino acids obtained by chemically or enzymatically decomposing proteins have been proposed. In particular, JP-A-58-165774 proposes a beverage composition (sports drink) containing a branched-chain amino acid. According to this publication, from the results of an experiment in which a rat was ingested with a branched-chain amino acid and the amount of exercise was measured, the effect that the branched-chain amino acid improves the function of muscle exercise was recognized.

However, in the beverage composition disclosed in JP-A-58-165774, the protein is chemically or enzymatically decomposed, and a branched-chain amino acid is selectively removed therefrom and added. This necessitates the necessity and complicates the step of separating the branched-chain amino acids, resulting in an increase in product cost. In addition, since amino acids generally have a strong bitter taste, there is a problem that the flavor is impaired when the added concentration is increased.

On the other hand, the peptide disclosed in JP-A-63-287462 is superior to amino acids in the point of rapid absorption and has the advantage that it does not have the bitter taste of amino acids. It was not possible to expect a bioactive effect by selective ingestion.

The present invention has been made in view of the above-mentioned problems of the prior art, and has as its object to contain a large amount of peptide, rich in branched-chain amino acids, low in aromatic amino acids, good in flavor, and intense exercise. It is an object of the present invention to provide an edible peptide-containing composition which is suitable for nutritional supplementation when the physical strength is reduced due to the above and can be produced industrially at low cost, and a method for producing the same.

"Means for solving the problem" The present inventors have conducted various studies in order to achieve the above-mentioned object, and as a result, removed starch as an impurity from a raw material containing a vegetable protein and heat-treated with an alkali. Hydrolyzed with a high alkali protease and, if necessary, hydrolyzed with a neutral or acidic protease to remove aromatic amino acids contained in the hydrolyzate by activated carbon or adsorption resin, etc., or by isoelectric precipitation By removing by such a method,
The present inventors have found that an edible peptide-containing composition containing an unprecedented high content of branched-chain amino acids and containing almost no aromatic amino acids can be obtained, and have completed the present invention.

That is, the edible peptide-containing composition of the present invention contains a peptide having an average molecular weight of 200 to 4,000 obtained by hydrolyzing corn protein with an enzyme, and the aromatic amino acid in the amino acid composition is 3.6 mol% based on all amino acids.
It is characterized as follows.

Further, the method for producing an edible peptide-containing composition of the present invention comprises a first step of treating a raw material containing a vegetable protein with a raw starch-degrading enzyme to decompose and remove starch, and subjecting the treated substance to high alkalinity. The method is characterized by including a second step of heat treatment, a third step of hydrolysis with a high alkali protease, and a fourth step of removing aromatic amino acids.

In the above production method, after the third step of hydrolyzing with a highly alkaline protease, a fourth step of hydrolyzing with a neutral protease and / or an acid protease is performed, and then a fifth step of removing aromatic amino acids is performed. The process may be performed.

In the above, the branched-chain amino acid is leucine,
Isoleucine, means valine, aromatic amino acids,
It means phenylalanine, tyrosine, tryptophan.

Hereinafter, the present invention will be described in more detail with reference to preferred embodiments.

In the present invention, as the vegetable protein, for example, corn protein, soybean protein, wheat protein, etc., various types can be used.
Zein, a corn protein and / or a prolamin-based protein constituting corn protein, is preferably used because it has a high content of branched-chain amino acids, particularly leucine, in the constituent amino acids. As the corn protein, in the process of producing corn starch, corn protein suspension obtained from corn through wet milling (wet sulfurous acid immersion), for example, corn gluten meal suspension, 70% from corn gluten liquor and corn protein Tween eluted with hydrous alcohol or alkali is preferably used. When using these protein suspensions as raw materials, the solid content concentration is more preferably adjusted to about 5 to 15% by weight.

In the production method of the present invention, as a first step, these raw materials are previously treated with a raw starch-decomposing enzyme to decompose and remove starch. According to a preferred embodiment, an alkali such as sodium hydroxide, potassium hydroxide, or calcium hydroxide is added to the suspension of the vegetable protein to adjust the pH to about 5 to 6, and raw starch-degrading enzymes such as " Dabiase "(product name,
(Available from Daikin Industries, Ltd.) in an amount of 0.02 to 0.2 wt% based on the solid content of the raw material, and the mixture is stirred and reacted at 50 to 60 ° C. for 3 to 20 hours, followed by dehydration and filtration.

Next, as a second step, the treated product is solidified at a solid concentration of 5 to 5%.
Resuspend to 20 wt%, preferably 10-15 wt%,
An alkali such as sodium hydroxide or potassium hydroxide is added to the suspension, and the pH is preferably adjusted to 12 or more.
Heat treatment at ~ 130 ° C for 5-30 minutes. By this treatment, the vegetable protein can be denatured, and the attack by the protease can be facilitated.

Next, as a third step, the suspension is cooled to 30 to 60 ° C., more preferably to about 50 ° C., and 0.2 to 2 wt% of the high alkali protease is added to the solid content of the raw material. The reaction is preferably performed for 1 to 2 hours. By this treatment, the vegetable protein is hydrolyzed to an appropriate length.

In this case, as the highly alkaline protease, for example, "Agric. Biol. Chem. 35 (9), 1407-1414" by Horikoshi et al.
Alkalophilic bacteria (Bacillus. No.22)
1) Alkaline protease derived from Meito Sangyo Co., Ltd.
And, "Esperase 8.0L" derived from an alkaliphilic mutant,
"Savinase 8.0L" (trade name, manufactured by Novo Corporation) and the like are suitable. These highly alkaline proteases have an optimum pH for enzyme action of 10 to 12, are excellent in heat resistance, and are usually endo-type enzymes.

Next, as a fourth step, an acid such as hydrochloric acid is added to the suspension, if necessary, to obtain a neutral pH of 5.5 to 8, preferably about 7, or 3.0 to 5.5, preferably pH 3.0 to 5.5. Adjust to 3.5-4.0 acidity. When the pH is adjusted to 5.5 to 8, neutral protease is added. When the pH is adjusted to 3.5 to 4.0, an acidic protease is added in an amount of 0.2 to 2% by weight based on the solid content of the raw material. C., more preferably around 50.degree. C., for 10 minutes to 24 hours, more preferably 20 to 24 hours. In this case, the neutral or acidic protease is preferably of the exo type. By this treatment, the hydrolysis rate of the vegetable protein can be further increased, and the aromatic amino acid can be converted to the free amino acid. However, this fourth step may be omitted in order to increase the molecular weight of the peptide.

The neutral or acidic exo-type protease includes aminopeptidase, carboxypeptidase, dipeptidyl peptidase and the like, and any peptidase may be used. For example, “protease A and M”
(Trade name, manufactured by Amano Pharmaceutical Co., Ltd.), "Sumiteam AP, MP,
LP (trade name, manufactured by Shin Nippon Chemical Co., Ltd.) and the like are preferable.

Further, aromatic amino acids are removed from the enzymatic hydrolyzate of vegetable protein as a fourth step when the fourth step is omitted, and as a fifth step when the fourth step is performed. As one of the removing methods, for example, a method using activated carbon or various kinds of adsorption resins can be adopted. More preferably, an adsorbent obtained by activating a polystyrene-based adsorption resin, for example, “Dowex S-112” (trade name, manufactured by Dow Chemical Co., Ltd.) with hydrochloric acid, is used. Pass through the hydrolyzate and elute on the acidic side. Other adsorbent resins include, for example, JP-A-60-1365
Resins as disclosed in JP-A-43-43 can also be used.

Among the adsorbed substances, activated carbon has a property of adsorbing coloring substances and aromatic amino acids in a slightly acidic protein enzyme hydrolyzate. Further, the polystyrene-based adsorption resin has many benzene rings, and the benzene ring on the resin side and the benzene ring of the aromatic amino acid are hydrophobically bonded. Therefore,
By using these adsorbents, aromatic amino acids can be selectively adsorbed and removed.

In addition, with these adsorbents alone, aromatic amino acids, particularly tyrosine, are not sufficiently removed. Therefore, by taking advantage of the low solubility of tyrosine, the tyrosine is separated and removed by isoelectric point precipitation or the like. Group amino acids can be reduced.

The edible peptide-containing composition of the present invention can be obtained through each of the above steps.However, after commercialization, if necessary, the final step or the treatment liquid is concentrated, the pH is adjusted, and then the acid is added. It is preferable to reduce the bitterness by a treatment or an enzymatic treatment such as amidase or deaminase, desalinate with an ion exchange membrane or an ion exchange resin, etc., further treat with activated carbon, and evaporate to dryness to obtain a powder. However, depending on the application, the solution can be used as it is.

The edible peptide-containing composition of the present invention comprises a peptide obtained by hydrolyzing a vegetable protein with an enzyme as described above, and contains a peptide having a molecular weight distribution of about 200 to 4,000 and an average molecular weight of about 500 to 2,000.

The components in the edible peptide-containing composition are preferably composed of 50 to 92% by weight of a peptide, 5 to 20% by weight of a sugar, 0 to 25% by weight of an amino acid, and 3 to 5% by weight of others.

Further, the edible peptide-containing composition of the present invention, the aromatic amino acid in the amino acid composition is 3.6 amino acids to all amino acids.
Mol% or less.

In a preferred embodiment, the molar ratio of branched chain amino acid / aromatic amino acid in the amino acid composition is 10 or more, more preferably 20 or more.

In a further preferred embodiment, the branched-chain amino acid is at least 10 mol% and the aromatic amino acid is at most 2 mol% in the amino acid composition.

The edible peptide-containing composition of the present invention has the following amino acid composition as a preferred example.

Amino acid composition (mol%) Aspartic acid 3.0 to 8.7 Threonine 2.1 to 6.2 Serine 3.4 to 9.6 Glutamic acid 10.9 to 30.4 Glycine 1.9 to 4.5 Alanine 8.0 to 26.4 Valine 3.5 to 6.2 Cystine 0.0 to 0.1 Methionine 0.4 to 6.5 Isoleucine 0.9 to 6.5 Leucine 4.4 to 47.5 Tyrosine 0.1-2.0 Phenylalanine 0.0-1.6 Lysine 1.3-2.8 Histidine 1.0-2.5 Arginine 1.3-2.1 Proline 2.9-6.0 `` Action and effect '' It is mainly composed of a peptide having a molecular weight of 200 to 4,000 and has a rapid and good absorption from the digestive tract like amino acids. Therefore, it is suitable for nutritional supplementation when physical strength decreases due to intense exercise and the like.

Further, the edible peptide-containing composition of the present invention, the aromatic amino acid in the amino acid composition is 3.6 amino acids to all amino acids.
Mol% or less and rich in branched-chain amino acids derived from plant proteins, so that the effect of improving muscle exercise function as disclosed in JP-A-58-165774 can be expected. .

Furthermore, peptides have less bitterness than amino acids,
In the present invention, bitterness is further reduced by reducing the amount of aromatic amino acids as described above, so that odor and taste are improved, and the invention can be applied to a wide range of foods.

Further, according to the production method of the present invention, starch, which is an impurity, is removed from the vegetable protein, subjected to heat treatment with an alkali, and then hydrolyzed with a highly alkaline protease, and if necessary, neutral and / or acidic protease. In the hydrolysis, and further to remove the aromatic amino acids contained in the hydrolyzate, rich in branched-chain amino acids,
In addition, the edible peptide-containing composition containing almost no aromatic amino acid can efficiently produce corn protein industrially.

"Example" Hereinafter, the present invention will be described in more detail with reference to examples.

Example 1 Potassium hydroxide was added to a corn protein suspension (gluten liquor) 450 obtained from the wet milling step to adjust the pH to 5.5, and "Dabiase" (trade name, manufactured by Daikin Industries, Ltd.) was used as a raw starch-degrading enzyme. ) Made 70g
And reacted at 50 ° C. for 16 hours with stirring. The suspension is subjected to solid-liquid separation using a filter press to remove the water-soluble fraction, thereby obtaining 80 kg of a corn protein wet cake. Re-suspend this wet cake in distilled water 350,
The temperature is raised to 80 ° C. with stirring. Next, the pH is adjusted to 12.0 by adding potassium hydroxide, and heat treatment is performed at 125 ° C. for 5 minutes.

Next, the suspension is cooled to 50 ° C., and 80 g of a highly alkaline protease (manufactured by Meito Sangyo Co., Ltd.) derived from an alkalophilic bacterium (Bacillus. No. 221) is added and reacted for 1.5 hours. Furthermore, hydrochloric acid was added to the suspension to adjust the pH to 7.0, and 180 g of "Denazyme AP" (trade name, manufactured by Nagase Biochemical Co., Ltd.), a neutral protease, was added, and reacted at 50 ° C for 20 hours. Let it. The reaction solution is separated into solid and liquid by a filter press,
Concentrate the water-soluble fraction to obtain Bx20,160 corn protein enzyme hydrolyzate.

A part of the hydrolyzate was adjusted to pH 4.0 with hydrochloric acid, and the polystyrene-based adsorption resin “Dowex S-112” (trade name, manufactured by Dow Chemical Co., Ltd.) was activated with hydrochloric acid. The hydrolyzate is passed through a column to separate a low fraction of aromatic amino acids. After concentrating the passing solution, desalting is performed with an ion-exchange membrane (micro-acylizer: manufactured by Asahi Kasei Corporation), treated with activated carbon, lyophilized, and dried with total nitrogen of 14.30.
%, Amino nitrogen 3.00%, sugar content 5.20%, ash content 1.0%, and a white powder having a molar ratio of branched chain amino acids to aromatic amino acids of 38.3 is obtained. The amino acid composition is shown in Table 1.

Example 2 The neutral protease "denazyme" in Example 1 above
AP "(trade name, manufactured by Nagase Seikagaku Co., Ltd.) was replaced with an acidic protease" protease Amano M "(trade name, manufactured by Amano Pharmaceutical Co., Ltd.), except that the reaction pH was changed to 4. The same operation as in Example 1 was repeated, freeze-dried,
A white powder having a composition of 13.50% of total nitrogen, 2.95% of amino nitrogen, 5.00% of sugar, and 1.2% of ash and having a molar ratio of branched-chain amino acids to aromatic amino acids of 69.1 is obtained. The amino acid composition is shown in Table 1.

Example 3 The same operation as in Example 2 was performed to pass through the adsorption resin,
Separate the low aromatic amino acid fraction and add potassium hydroxide
The pH was adjusted to 5.5 and concentrated to a solids concentration of 15% (Bx15). After the concentrate was cooled to 5 ° C., the precipitated tyrosine was removed by filtration or the like. Thereafter, desalting, activated carbon treatment, and lyophilization were performed in the same manner as in Example 2 to obtain 13.8% of total nitrogen and 25% of amino nitrogen.
A white powder having a composition of 0%, a sugar content of 4.5% and an ash content of 1.3% and having a molar ratio of the branched-chain amino acid to the aromatic amino acid of 28.5 is obtained. The amino acid composition is shown in Table 1.

Example 4 The same operation as in Example 2 was performed, and the fraction from which tyrosine was removed by filtration in Example 3 was desalted, treated with activated carbon, and treated with Bx60.
Concentrated. The obtained cloudy concentrate was separated into a precipitate section and a soluble section by centrifugation, and the precipitate section was dried under reduced pressure as it was, 13.5% of total nitrogen, 2.92% of amino nitrogen, and 4.
A white powder having a composition of 8% and an ash content of 1.1% and having a molar ratio of the branched chain amino acid to the aromatic amino acid of 46.1 is obtained. The amino acid composition is shown in Table 1.

Example 5 The same operation as in Example 3 was performed, and the fraction from which tyrosine had been removed by filtration was passed through a cation exchange resin and an anion exchange resin. The elution fraction was separated, and the pH was adjusted to 4.0 with potassium hydroxide. Thereafter, desalting, activated carbon treatment, and lyophilization were performed in the same manner as in Example 1 to obtain a total nitrogen of 14.0%, amino nitrogen of 0.90%, and sugar content.
A white powder having a composition of 5.0% and an ash content of 1.0% and having a molar ratio of branched-chain amino acids to aromatic amino acids of 25.6 is obtained. The amino acid composition is shown in Table 1.

Example 6 The same operation was performed up to the enzymatic reaction with the highly alkaline protease in Example 1, and hydrochloric acid was added to the reaction solution.
The pH was adjusted to 5.5. The reaction solution is subjected to solid-liquid separation using a filter press, and the water-soluble fraction is concentrated to Bx15 to obtain corn protein enzyme hydrolyzate 230.

Next, the fraction passed through the adsorption resin was passed through a cation exchange resin and an anion exchange resin as in Example 1, the elution fraction was separated, the pH was adjusted to 5.5 with potassium hydroxide, and the salt was desalted. Activated carbon treatment, freeze-dried, total nitrogen 14.1%, amino nitrogen 0.55%, sugar 4.8%, ash 0.9% composition,
The molar ratio of branched-chain amino acids to aromatic amino acids is 21.1
To obtain a white powder. The amino acid composition is shown in Table 1.

Table 1 does not include data on tryptophan, which is a kind of aromatic amino acid, because tryptophan was scarcely contained.

From the results in Table 1, in the enzymatic hydrolyzates obtained in Examples 1 to 6, the ratio of the branched-chain amino acids (leucine, isoleucine, valine) to the aromatic amino acids (phenylalanine, tyrosine) was significantly increased in the amino acid composition. You can see that it is. Further, in Examples 3 to 6, it can be seen that the branched-chain amino acids are contained in a ratio equal to or higher than the starting material. In addition, since the total nitrogen is 14 and the protein content is high, and the amino nitrogen is 3.0 or less, it can be understood that the peptide is rich in peptides. From the above, it can be seen that the composition containing the edible peptide can be sufficiently used for various oral nutritional supplements and the like.

Example 7 The peptide compositions obtained in Examples 1 and 3 were analyzed by high performance liquid chromatography (HPLC).
The molecular weight distribution of each of the peptide compositions was determined.

Using "OH Pak KB-802.5" (trade name, manufactured by Showa Denko KK) as a separation column, 0-1% trifluoroacetic acid-
Using a 40% acetonitrile eluent, flow rate 0.5 ml / min
The analysis was performed with the detector set to UV.210 nm. In addition, tyrochrome C (MW ≒ 10,000) as a standard sample,
Gly-Gly-Gly which is an oligopeptide (MW ≒ 189.2), Gl
y-Gly (MW ≒ 132) and free amino acids tyrosine (MW ≒ 181.2) and phenylalanine (MW ≒ 165.2) were used.

FIG. 1 shows the results of the analysis of the peptide composition of Example 1, and FIG. 2 shows the results of the analysis of the peptide composition of Example 3. In the figure, the elution positions of the standard samples are indicated by arrows.

In FIGS. 1 and 2, complete separation and identification of the free amino acid and the peptide are not performed, but taking into consideration that the minimum molecular weight of the peptide is usually around 200,
And the molecular weight of the peptide contained in the peptide composition of 3.
It turns out that it is in the range of 200-4,000.

[Brief description of the drawings]

FIG. 1 is a chart showing an elution curve when the peptide composition obtained in Example 1 was subjected to high performance liquid chromatography. FIG. 2 is a table showing the peptide composition obtained in Example 3 subjected to high performance liquid chromatography. 4 is a chart showing an elution curve at the time.

──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. ⁶ Identification code FI C12P 21/06 C12P 21/06 (72) Inventor Son-ichi Yamaguchi 2954 Imaizumi, Fuji City, Shizuoka Prefecture Nisshinkinomiya Shrine 2-307 (72) Inventor Koichi Ogawa 2954 Imaizumi, Fuji City, Shizuoka Prefecture Nisshinkinomiya Dormitory (56) References JP-A-57-7490 (JP, A) JP-A-64-14274 (JP, A) (58) Survey ⁶ (Int. Cl. ⁶ , DB name) C07K 14/415, 1/12, 1/14 A23J 3/34, 3/14 C12P 21/06 CA (STN) BIOSIS (DIALOG)

Claims (12)

(57) [Claims] 1. A corn protein containing a peptide having an average molecular weight of 200 to 4,000 obtained by hydrolyzing a corn protein with an enzyme, wherein an aromatic amino acid in the amino acid composition is 3.6 mol% or less based on all amino acids. An edible peptide-containing composition. 2. The edible peptide-containing composition according to claim 1, wherein the molar ratio of branched chain amino acid / aromatic amino acid in the amino acid composition is 10 or more. 3. An amino acid composition comprising 10 branched-chain amino acids.
The edible peptide-containing composition according to claim 1 or 2, wherein the content of the aromatic amino acid is 2 mol% or less in an amount of 2 mol% or more. 4. The edible peptide-containing composition according to claim 1, which has the following amino acid composition. Amino acid composition (mol%) Aspartic acid 3.0 to 8.7 Threonine 2.1 to 6.2 Serine 3.4 to 9.6 Glutamic acid 10.9 to 30.4 Glycine 1.9 to 4.5 Alanine 8.0 to 26.4 Valine 3.5 to 6.2 Cystine 0.0 to 0.1 Methionine 0.4 to 6.5 Isoleucine 0.9 to 6.5 Leucine 4.4 to 47.5 Tyrosine 0.1 to 2.0 Phenylalanine 0.0 to 1.6 Lysine 1.3 to 2.8 Histidine 1.0 to 2.5 Arginine 1.3 to 2.1 Proline 2.9 to 6.0 5. The solid content of 50 to 92% by weight of peptide and 5 to 5% of sugar
20% by weight, 0-25% by weight of amino acids, 3-5% by weight of others
The edible peptide-containing composition according to any one of claims 1 to 4, which contains the edible peptide-containing composition. 6. A first step of treating a raw material containing a vegetable protein with raw amylolytic enzyme to degrade and remove starch.
A second step of heat-treating the treated product under a high alkali;
A method for producing an edible peptide-containing composition, comprising a third step of hydrolyzing with a highly alkaline protease and a fourth step of removing aromatic amino acids. 7. A first step of treating a raw material containing a vegetable protein with raw amylolytic enzyme to degrade and remove starch.
A second step of heat-treating the treated product under a high alkali;
A third step of hydrolyzing with a highly alkaline protease, a fourth step of hydrolyzing with a neutral protease and / or an acidic protease, and a fifth step of removing aromatic amino acids.
7. The method for producing an edible peptide-containing composition according to claim 6, comprising the steps of: 8. The method for producing an edible peptide-containing composition according to claim 6, wherein a raw material containing corn protein is used. 9. The edible product according to claim 6, wherein the second step is performed in a highly alkaline aqueous solution having a pH of 12 or more at 100 to 130 ° C. for 5 to 30 minutes. A method for producing a peptide-containing composition. 10. The method for producing an edible peptide-containing composition according to any one of claims 7 to 9, wherein an endo-type enzyme derived from an alkalophilic bacterium is used as the highly alkaline protease in the third step. . 11. The method according to claim 7, wherein the step of hydrolyzing with a high alkaline protease, a neutral protease and / or an acidic protease is carried out at 30 to 60 ° C. for 10 minutes to 24 hours. 5. A method for producing an edible peptide-containing composition according to any one of the above. 12. The method for removing aromatic amino acids by a method using an adsorbent obtained by activating a polystyrene-based adsorption resin with hydrochloric acid, a method using activated carbon, and / or an isoelectric point precipitation method. Item 12. The method for producing the edible peptide-containing composition according to any one of Items 7 to 11.