Method for producing peptide, composition containing peptide, and food / beverage using same

Abstract

The present invention addresses the problem of providing a method for efficiently producing a specific peptide from a protein material containing therein an amino acid sequence composed of GTW. Provided is a method for producing a composition containing a GTW peptide, the production method including a step for allowing a protein material containing therein an amino acid sequence composed of GTW to interact with a protease, wherein the production method is characterized in that the pH at the start of interaction in the aforementioned step is 5.5 or higher, or in that the protease is derived from Aspergillus melleus, Streptomyces griseus, or Bacillus licheniformis, or in that a protein material containing the amino acid sequence composed of GTW in a portion thereof is a whey protein, or in that the whey protein is β-lactoglobulin.

Description

Method for producing peptides, compositions containing peptides, and foods and beverages using the same

[0001] The present invention relates to a method for producing a peptide composition by decomposing a protein material containing a GTW sequence with a protease, and in particular to a method for efficiently producing a composition containing a GTW peptide, a composition containing a GTW peptide obtained by the production method, and a food or beverage product using the same.

[0002] In recent years, the rise of lifestyle-related diseases such as vascular disease and diabetes, as well as age-related diseases, has become a major social issue in our rapidly aging society. One of the contributing factors is the increase in oxidative stress and the generation of reactive oxygen species (ROS) in the body. Humans are exposed to various oxidative stressors on a daily basis, including UV rays, air pollution, tobacco, drugs, stress, excessive exercise, and the ingestion of oxidized substances. In a healthy state, the body's natural antioxidant activity can suppress a certain level of oxidative stress. However, when excessive oxidative stress occurs or the body's antioxidant activity is weakened due to poor lifestyle habits, the generated ROS can damage cells and lead to various diseases, including cancer, cardiovascular disease, and lifestyle-related diseases. Therefore, it is important to suppress oxidative stress in the body on a daily basis, and the use of food-derived antioxidants that can be consumed daily is expected.

[0003] Known food-derived antioxidants include compounds such as polyphenols, carotenoids, vitamin C, and vitamin E. Recently, the antioxidant properties of peptides derived from milk proteins have also been reported. Among these peptides, peptides with a glycine-threonine-tryptophan (GTW) sequence (hereinafter referred to as the GTW sequence) (hereinafter referred to as the GTW peptide) have been reported to exhibit stronger antioxidant activity than glutathione (glutamic acid-cysteine-glycine), which is known to have high antioxidant activity (Non-Patent Document 1). Therefore, by developing ingredients and foods containing peptides with high antioxidant activity, such as the GTW peptide, it may be possible to suppress various inflammations and aging-related conditions in the body caused by oxidation. This is expected to contribute significantly to maintaining homeostasis, improving lifestyle-related diseases, and improving the quality of life of the general public by suppressing aging.

[0004] GTW peptides are found in whey proteins such as β-lactoglobulin, and may be produced by protein degradation in fermented dairy products such as cheese and yogurt. However, due to further degradation, the amount produced is extremely small, and no effect can be expected from the intake of ordinary dairy foods. One method involves hydrolyzing whey protein materials using commercially available general-purpose proteases, but these generally lack strict cleavage specificity and have side activities, making it difficult to produce and accumulate large amounts of GTW peptides. GTW peptides can also be produced by chemical synthesis, but this method is expensive and unsuitable for application in foods and beverages.

[0005] Furthermore, methods for concentrating specific peptides include methods using reverse osmosis membranes or ultrafiltration membranes (Patent Documents 1 and 2), and methods involving adsorption onto column resins and elution (Patent Document 3), but these methods require expensive capital investment for industrial production. Thus, no method has yet been known to effectively produce a peptide composition for food or beverage use with a high GTW peptide content at low cost without chemical synthesis or special membrane or fractionation processes.

[0006] JP-A-2003-92996 JP-A-6-7188 Patent No. 3108518

[0007] Dairy Sci. & Technol. (2015) 95:451-463Neurobiology of Aging, 2018, 72, 23-31

[0008] An objective of the present invention is to provide a method for efficiently producing a peptide composition having a high GTW peptide content, and to provide a peptide composition having a high GTW peptide content and a food or beverage product using the same.

[0009] As a result of extensive research aimed at solving the above-mentioned problems, the present inventors have found that when a protein material containing a GTW sequence therein is hydrolyzed with a protease, adjusting the pH of the solution at the start of the reaction or using a specific protease can promote excision of GTW peptides from the protein material, making it possible to produce a composition with an increased GTW peptide content. Specifically, the present invention relates to a method for producing a peptide composition that can efficiently produce GTW peptides. The present invention also relates to a composition with a high GTW peptide content and foods and beverages using the same. Specifically, the present invention has the following features: <1> A method for producing a composition containing a GTW peptide, which includes a step of reacting a protein material containing a GTW sequence therein with a protease, is provided, wherein the pH at the start of the reaction in the step is set to 5.5 or higher. <2> A method for producing a composition containing a GTW peptide, comprising the step of reacting a protein material containing a GTW sequence therein with a protease, wherein the protease is derived from Aspergillus melleus, Streptomyces griseus, or Bacillus licheniformis. <3> The production method according to <1>, wherein the protease is derived from Aspergillus melleus, Streptomyces griseus, or Bacillus licheniformis. <4> The production method according to any one of <1> to <3>, wherein the protein material containing a GTW sequence therein is a whey protein. <5> The production method according to <4>, wherein the whey protein is β-lactoglobulin. <6> The composition containing a GTW peptide produced by the production method according to any one of <1> to <3>. <7> The composition containing a GTW peptide, wherein the GTW peptide content derived from a protein material containing a GTW sequence therein is 0.1% (w / w) or more. <8> A composition comprising the GTW peptide according to <7>, wherein the protein material containing the GTW sequence therein is whey protein. <9> A composition comprising the GTW peptide according to <8>, wherein the whey protein is β-lactoglobulin.<10> Provided is a food or drink comprising a composition containing a GTW peptide, wherein the content of GTW peptide derived from a protein material containing a GTW sequence therein is 0.0001% (w / w) or more in the food or drink. <11> Provided is the food or drink according to <10>, wherein the protein material containing a GTW sequence therein is whey protein. <12> Provided is the food or drink according to <11>, wherein the whey protein is β-lactoglobulin.

[0010] According to the production method of the present invention, GTW peptides can be produced selectively and efficiently by adjusting the pH when a protein material containing a GTW sequence therein is hydrolyzed with a protease or by using a specific protease.

[0011] The method for producing a composition containing a GTW peptide of the present invention is a method for hydrolyzing a protein by an enzymatic reaction (hereinafter simply referred to as a reaction) by allowing a proteolytic enzyme (hereinafter referred to as a protease) to act on a protein material containing a GTW sequence, such as whey protein, to hydrolyze the protein by adjusting the pH at the start of the reaction to a predetermined range. Specifically, the method for hydrolyzing a protein by adjusting the pH at the start of the reaction to 5.5 or higher can increase the amount of GTW peptide produced from the protein containing a GTW sequence. In this specification, the act of a protease on a substrate protein material to cause a reaction may be simply referred to as "reacting," and "start of action" may be simply referred to as "start of reaction." Furthermore, the method for producing a composition containing a GTW peptide of the present invention is a method for increasing the amount of GTW peptide produced from whey protein by using a protease derived from Aspergillus melleus, Streptomyces griseus, or Bacillus licheniformis. The present invention is described in detail below.

[0012] (Protein Material Containing a GTW Sequence) The raw material used in the method for producing a composition containing a GTW peptide of the present invention may be any protein material containing an amino acid sequence consisting of glycine-threonine-tryptophan (GTW) as part of its structure. Examples of such protein materials include whey protein, egg white lysozyme, avian collagen, and bovine fibrinogen. Of these, whey protein materials are preferred, and materials containing β-lactoglobulin are even more preferred. Protein materials containing a GTW sequence also include protein hydrolysates. Examples of materials containing β-lactoglobulin include milk, dairy products, whey, whey powder, whey protein concentrate (WPC), and whey protein isolate (WPI). The concentration of the raw material used in the hydrolysis reaction of the protein of the present invention is not particularly limited, but considering the typical production process, a protein content of approximately 0.01% to 50% (w / w) is preferred, and more preferably approximately 0.1% to 20% (w / w).

[0013] (Protease) In the present invention, the protease used for hydrolysis of a protein material containing a GTW sequence therein may be any protease capable of selectively cleaving GTW peptides, and commercially available general-purpose proteases may also be used. Commercially available general-purpose proteases are preferably exo-type proteases that cleave internal peptide bonds in proteins, and more preferably, proteases derived from Aspergillus melleus, Streptomyces griseus, or Bacillus licheniformis can be used, with Aspergillus melleus and Streptomyces griseus proteases being particularly preferred. Known commercial proteases derived from Aspergillus melleus include, for example, Protease P "Amano" 3SD (Amano Enzyme), Protease from Aspergillus melleus (Sigma), and Sumiteam MP (Shin-Nihon Chemical Industry). Known proteases derived from Streptomyces griseus include Protease from Streptomyces griseus (Sigma). Known proteases derived from Bacillus licheniformis include Alcalase 2.4L FG (Novozyme). The amount of protease to be added is not particularly limited, but taking into consideration an appropriate reaction time, production costs, etc., the protease concentration in the reaction solution is preferably about 0.001% (w / v) to 10% (w / v), more preferably 0.01% (w / v) to 5% (w / v).

[0014] (Protease (enzyme) reaction) Through investigations by the present inventors, it has been found that in the step of reacting a protein material containing a GTW sequence internally with a protease, GTW peptides are efficiently produced under conditions where the initial pH of the protease reaction is neutral to alkaline. The pH at the start of the reaction is preferably pH 5.5 or higher, more preferably pH 6.0 or higher, and even more preferably pH 7.0 or higher. A preferred range is pH 5.5 to 10.0, preferably pH 6.0 to 10.0, and more preferably pH 7.0 to 9.5. The pH at the start of the protease reaction can be adjusted by adding NaOH, KOH, (NH ４ ) CO ３ The reaction can be initiated by adding the protease after adjusting the pH to the above range by, for example, etc. Specifically, the pH of a solution containing a protein material containing a GTW sequence therein is adjusted to the above range in advance, and then the protease is added to the solution.

[0015] The protease reaction time may be set depending on the amount of GTW peptide of interest, and is preferably 30 minutes or more. Within this range, a time that produces a large amount of GTW peptide and does not cause degradation of the GTW peptide is desirable, for example, 30 minutes to 16 hours is preferred, and 30 minutes to 8 hours is even more preferred. Furthermore, after the GTW peptide is produced, the protease reaction time may be extended, or reaction conditions such as temperature may be changed, or other enzymes may be added to the reaction, as long as the time is within a range that does not cause significant degradation of the GTW peptide.

[0016] The reaction temperature may be about 10°C to 75°C, more preferably 30°C to 65°C, and even more preferably 35°C to 55°C. Stirring may not be required during the reaction, stirring may be performed at about 10 to 500 rpm, or the reaction may be performed while the solution is being pumped through a line (pipe). After the reaction, the protease may be inactivated by heating, or by removal through ultrafiltration membrane treatment, or by inactivation during the process of powdering the reaction solution or processing it into a food product.

[0017] (Compositions Containing GTW Peptides) The production method of the present invention enables efficient recovery of GTW peptides from raw materials containing proteins containing a GTW sequence, such as β-lactoglobulin. Specifically, it is possible to recover 10 to 95% of the GTW peptides contained in the raw material. As such, the peptide composition of the present invention is derived from proteins containing a GTW sequence, and therefore does not contain synthetic GTW peptides. It is a composition containing GTW peptides derived from protein materials containing a GTW sequence. Proteins are generally known to produce peptides and amino acids that exhibit tastes such as bitterness and umami upon protease degradation, and the more amino acids, dipeptides, and tripeptides with a molecular weight of less than 400, the stronger the perceived taste. The average molecular weight of the GTW peptide-containing composition of the present invention is preferably 400 or more, more preferably 500 or more, and most preferably 700 or more. The GTW peptide content in the GTW peptide-containing composition of the present invention is preferably 0.1% (w / w) or more. The GTW peptide content in a food or drink containing a composition comprising the GTW peptide of the present invention is preferably 0.0001% (w / w) or more. Whether the GTW peptide content of a composition comprising the GTW peptide of the present invention, or a food or drink, pharmaceutical, etc. containing the composition falls within the above range can be detected by measuring the GTW peptide content.

[0018] The GTW peptide-containing composition of the present invention can be used as is, or after desalting using a dialysis membrane or ion exchange resin, or after drying by freeze-drying, spray-drying, or other methods to produce a powder. The GTW peptide-containing composition of the present invention can also be incorporated into foods and beverages. When incorporated into foods and beverages, the composition can be incorporated as is, or after desalting using a dialysis membrane or ion-exchange resin, or after drying by freeze-drying, spray-drying, or other methods to produce a powder. Examples of foods and beverages incorporating the GTW peptide-containing composition of the present invention include soft drinks, dairy drinks, fermented milk, cheese, butter, skim milk, juice, jelly, bread, ice cream, noodles, sausages, infant formula, and baby food. When using the GTW peptide-containing composition of the present invention as a pharmaceutical or functional food, it can be processed into oral formulations such as tablets, capsules, fine granules, powders, pills, lozenges, sublingual tablets, and liquids.

[0019] The present invention will be explained in more detail below by showing examples and production examples of the present invention, but the present invention is not limited to these examples at all.

[0020] Example 1: Preparation of a Composition Containing GTW Peptides from a WPC Solution 1. Test Method To investigate the proteases that can be used in the present invention, a peptide composition was prepared from WPC using a commercially available protease. Commercially available WPC was dissolved in ion-exchanged water heated to 40-50°C to a concentration of 10% (w / w), and the pH was adjusted to 8.0 with NaOH. Immediately after pH adjustment, a commercially available protease listed in Table 2 was added to the solution to a concentration of 0.1% (w / v) to initiate the reaction. Two hours after the start of the reaction, a sample was taken and heated at 90°C for 5 minutes to inactivate the protease. The sample after protease inactivation was lyophilized, and the GTW peptide content in the powder was measured. The GTW peptide content was measured using the method of Ano et al. (Non-Patent Document 2). Quantitative analysis was performed using a liquid chromatograph-tandem mass spectrometer (LC-MS / MS) according to the conditions listed in Table 1.

[0021]

[0022] 2. Test Results The measurement results are shown in Table 2. Treatment with proteases (i), (ii), and (iii) resulted in 2.1 to 4.3 mg / g of GTW peptide per material, while treatment with protease (iv) resulted in 1.1 mg / g of GTW peptide per peptide material. Treatment with proteases (v) and (vi) resulted in almost no GTW peptide. These results demonstrate that the use of proteases derived from Aspergillus melleus (i) and (ii) or proteases derived from Streptomyces griseus (iii) can prepare peptide compositions with a high GTW peptide content from protein materials containing GTW sequences, such as WPC. It was also demonstrated that proteases derived from Bacillus licheniformis (vi) can also be used to prepare peptide materials containing GTW peptides.

[0023]

[0024] Example 2 Measurement of GTW peptide content in commercially available peptide materials 1. Test method 1% aqueous solutions of commercially available peptide materials A to D shown in Table 3 were prepared and diluted, and the GTW peptide content was quantified by LC-MS / MS. The GTW peptide content in the original peptide material was calculated from the quantified value. The commercially available peptide materials were prepared by enzymatically hydrolyzing WPC.

[0025] 2. Test Results The measurement results are shown in Table 3. Although the commercially available peptide materials evaluated were prepared from WPC, they contained either no GTW peptide or only very small amounts. Even the one with the highest GTW peptide content had a GTW peptide content of 0.5 mg / g per peptide material. These results demonstrate that it is not possible to prepare a peptide material with a high GTW peptide content from WPC using conventional methods.

[0026]

[0027] Example 3: Confirmation of the effect of the initial pH of the enzymatic reaction on the production of GTW peptides from WPC 1. Test method Commercially available WPC was dissolved in ion-exchanged water heated to 40-50°C to a concentration of 10% (w / w), and the pH was adjusted to 4.0-10.0 with NaOH. Immediately after adjusting the pH, protease P "Amano" 3SD was added to a concentration of 0.1% (w / v) to initiate the reaction. Two hours after the start of the reaction, a sample was taken and heated at 90°C for 5 minutes to inactivate the protease. The inactivated sample was lyophilized, and the GTW content in the powder was measured by LC-MS / MS.

[0028] 2. Test Results The GTW peptide content after the reaction is shown in Table 4. When the initial pH was 6.0 or higher, the GTW peptide content after the reaction was 1.0 mg / g or higher (0.1% (w / w) or higher). These results demonstrate that a peptide composition with a high GTW peptide content can be prepared by setting the pH at the start of the protease reaction to 6.0 or higher.

[0029]

[0030] Example 4 Effect of Holding Time After pH Adjustment on GTW Peptide Content 1. Test Method Commercially available WPC was dissolved in ion-exchanged water heated to 40-50°C to a concentration of 10% (w / w), and the pH was adjusted to 8.5 with NaOH. After pH adjustment, the solution was held for 0, 1, 3, and 5 hours, and then protease P "Amano" 3SD was added to a concentration of 0.1% (w / v) to initiate the reaction. Two hours after the start of the reaction, a sample was taken and heated at 90°C for 5 minutes to inactivate the protease. The inactivated sample was lyophilized, and the GTW peptide content in the powder was measured by LC-MS / MS.

[0031] 2. Test Results The measurement results are shown in Table 5. When the retention time after pH adjustment was 3 hours or less, the GTW content after protease treatment was 2 mg / g or more. These results demonstrate that a peptide composition with a high GTW peptide content can be prepared by setting the retention time after pH adjustment to 3 hours or less.

[0032]

[0033] Example 5: Preparation of a GTW Peptide-Containing Composition by Protease Treatment at a Constant pH 1. Test Method Commercially available WPC was dissolved in ion-exchanged water heated to 40-50°C to a concentration of 16% (w / w), and the pH was adjusted to 7.5 with NaOH. Immediately after pH adjustment, Protease P "Amano" 3SD was added to a concentration of 0.1% (w / v) to initiate the reaction. The reaction was carried out at 50°C for 5 hours using a jar fermenter (2 L total volume microbial culture system BMJ-02NC; ABLE) while adding a neutralizer (NaOH) to maintain the pH at 7.5. Samples were taken 3 and 5 hours after the start of the reaction and heated at 90°C for 5 minutes to inactivate the protease. The inactivated samples were lyophilized, and the GTW peptide content in the powder was measured by LC-MS / MS.

[0034] 2. Test Results The GTW peptide content in the reaction solution was 3.7 mg / g after 3 hours and 4.5 mg / g after 5 hours, indicating that a material with a high GTW peptide content was obtained.

[0035] Example 6: Prototype of a GTW peptide-rich composition and sensory evaluation test 1. Test method Commercially available WPC was dissolved in ion-exchanged water heated to 40-50°C to a concentration of 16% (w / w), and the pH was adjusted to 8.5 with NaOH. Immediately after pH adjustment, protease P "Amano" 3SD was added to a concentration of 0.1% (w / v), and the reaction was initiated at 50°C. Thirty minutes after the start of the reaction, a sample was collected and heated at 90°C for 20 minutes to inactivate the protease. The sample after protease inactivation was lyophilized to obtain a GTW peptide-rich composition. The resulting material was evaluated for bitterness. Ten panelists placed 20 mg of the material powder on their tongues and tasted it, and the bitterness was evaluated using a four-point scale according to the following criteria. Commercially available peptide material E was used for comparison. <Bitterness evaluation criteria> 3 points: Strong bitterness 2 points: Bitterness is felt 1 point: Slight bitterness 0 points: No bitterness felt

[0036] 2. Test Results Comparing the average scores of 10 people, the GTW peptide-rich composition of the present invention had an average score of 1.4, confirming that it had a reduced bitterness compared to commercially available peptide materials.

[0037]

[0038] Example 7 Preparation of a Composition Containing GTW Peptides from Bovine Fibrinogen 1. Test Method Commercially available fibrinogen (derived from bovine serum, Sigma-Aldrich) was used as a protein raw material containing the GTW sequence. A peptide composition was prepared by protease treatment. Fibrinogen was dissolved in 10 mM phosphate buffer (pH 8.0) to a concentration of 2% (w / w), heated to 50°C, and then a commercially available protease listed in Table 7 was added to a concentration of 0.1% (w / v) to initiate the reaction. Two hours after the start of the reaction, a sample was collected and heated at 90°C for 5 minutes to inactivate the protease. The sample was desalted using MonoSpin C18, and the GTW peptide content in the solution was measured by LC-MS / MS.

[0039] 2. Test Results The measurement results are shown in Table 7. It was found that GTW peptides were produced by treatment with proteases (i) and (ii), and that a composition containing GTW peptides can also be prepared from fibrinogen using this technology.

[0040]

[0041] Example 8 Preparation of GTW peptide-containing capsules 10 g of the GTW peptide-rich composition prepared in Example 5, 33.5 g of lactose, 56.0 g of soluble starch, and 0.5 g of magnesium stearate were mixed, granulated by a conventional method, and filled into capsules to produce GTW peptide-containing capsules of the present invention.

[0042] Example 9 Preparation of GTW Peptide-Containing Beverage 10 g of the GTW peptide-rich composition prepared in Example 5 was dissolved in 690 g of deionized water, heated to 40°C, and then stirred and mixed for 20 minutes at 9,500 rpm using an Ultra Disperser (ULTRA-TURRAX T-25; manufactured by IKA Japan). 100 g of maltitol, 2 g of acidulant, 20 g of reduced starch syrup, 2 g of flavoring, and 176 g of deionized water were added, and the mixture was filled into 100 ml glass bottles, sterilized at 95°C for 15 seconds, and then tightly sealed to prepare 10 bottles (100 ml) of the GTW peptide-containing beverage of the present invention.

[0043] Example 10 Preparation of GTW Peptide-Containing Milk Drink 100 g of the GTW peptide-rich composition prepared in Example 5 was mixed with 9.9 kg of milk, heated to 40°C, and then stirred and mixed for 10 minutes at 6,000 rpm in a TK homomixer (TK ROBO MICS; manufactured by Tokushu Kika Kogyo Co., Ltd.). The mixture was heat-sterilized at 130°C for 2 seconds and then cooled to 10°C or below to produce 10 kg of the GTW peptide-containing milk drink of the present invention.

[0044] Example 11 Preparation of GTW Peptide-Containing Fermented Milk 100 g of the GTW peptide-rich composition prepared in Example 5, 1600 g of skim milk powder, 300 g of glucose, and 7700 g of deionized water were mixed and sterilized by heating at 95°C for 2 hours. This mixture was cooled to 37°C, and 300 g of a lactic acid bacteria starter (Lb. casei) was inoculated. After stirring and mixing, the mixture was fermented in an incubator maintained at 37°C until the pH reached 4.0. After the pH reached 4.0, the mixture was cooled to 10°C or below to produce 10 kg of the GTW peptide-containing fermented milk of the present invention.

[0045] Example 12 Preparation of GTW peptide-containing cheese 9.5 kg of Gouda cheese, 9.5 kg of Cheddar cheese, 100 g of the GTW peptide-rich composition prepared in Example 5, 200 g of sodium citrate, and 700 g of deionized water were mixed and emulsified at 85° C. After emulsification, the cheese was packed into a carton and cooled at 5° C. for two days and nights to produce 20 kg of the GTW peptide-containing cheese of the present invention.

[0046] Example 13: Preparation of GTW peptide-containing cream 4.5 kg of hardened rapeseed oil, 40 g of lecithin, 10 g of monoglycerin fatty acid ester, and 10 g of sorbitan fatty acid ester were mixed to prepare an oil phase. Next, 100 g of the GTW peptide-rich composition prepared in Example 5, 308 g of skim milk powder, 10 g of sodium caseinate, 20 g of sugar ester, 10 g of phosphate, 5 g of xanthan gum, and 4.987 kg of deionized water were mixed to prepare an aqueous phase. The aqueous phase was heated to 65°C, and the oil phase heated to 70°C was added in small portions with stirring. The mixture was stirred and mixed for 10 minutes at 6,000 rpm in a TK homomixer (TK ROBO MICS; manufactured by Tokushu Kika Kogyo Co., Ltd.). This mixture was homogenized in a homogenizer to produce 10 kg of the GTW peptide-containing cream of the present invention.

[0047] Example 14 Preparation of GTW Peptide-Containing Pudding 2000 g of honey, 100 g of the GTW peptide-rich composition prepared in Example 5, 770 g of skim milk powder, 300 g of mascarpone, 700 g of liquid starch syrup, 500 g of granulated sugar, 250 g of fresh cream, 200 g of butter, 400 g of sweetened egg yolk, 40 g of gelatin, 15 g of agar, 120 g of locust bean gum, and 4605 g of deionized water were mixed to prepare a pudding mix. This pudding mix was stirred and mixed at 6,000 rpm for 10 minutes in a TK homomixer (TK ROBO MICS; manufactured by Tokushu Kika Kogyo Co., Ltd.), heated to 60°C to dissolve, and then packed into containers in 100 g portions and cooled to produce 100 GTW peptide-containing puddings of the present invention.

[0048] Example 15 Preparation of GTW Peptide-Containing Powdered Milk 1 kg of the GTW peptide-rich composition prepared in Example 5, 8.3 kg of skim milk powder, and 90 kg of deionized water were mixed and heated to 40°C, followed by stirring and mixing for 10 minutes at 6,000 rpm in a TK homomixer (TK ROBO MICS; manufactured by Tokushu Kika Kogyo Co., Ltd.). The solution was spray-dried to produce approximately 10 kg of GTW peptide-containing powdered milk of the present invention.

[0049] Example 16: Preparation of a GTW Peptide-Containing Soft Drink A GTW peptide-containing soft drink was prepared by enzyme treatment during the soft drink manufacturing process. 16 g of commercially available WPC was dissolved in 84 g of distilled water, heated to 50°C, and the pH was adjusted to 8.5 with NaOH. Protease P "Amano" 3SD was added to a concentration of 0.1% (w / v), and the enzyme reaction was carried out at 50°C for 30 minutes. In a separate container, 10 g of skim milk powder, 100 g of sugar, 5 g of citric acid, 2.5 g of trisodium citrate, and 5 g of pectin were dissolved in 778 ml of distilled water. This was then mixed with the enzyme-treated WPC solution, sterilized by heating at 90°C for 20 minutes, and filled into a 100 ml plastic bottle.

[0050] According to the production method of the present invention, when a protein material containing a GTW sequence, such as whey protein, is hydrolyzed with a protease, the GTW peptide content can be increased by adjusting the pH or by using a specific protease, thereby allowing for selective concentration of the GTW peptide. Furthermore, since no expensive equipment is required, the method can be easily incorporated into conventional production processes. Furthermore, since the composition containing the GTW peptide of the present invention has a high GTW peptide content, it can be added in reduced amounts to foods and beverages, minimizing the impact on flavor, making it highly useful.

Claims

1. A method for producing a composition containing a GTW peptide, comprising the step of reacting a protein material containing a GTW sequence therein with a protease, characterized in that the pH at the start of the reaction in said step is 5.5 or higher.

2. A method for producing a composition containing a GTW peptide, comprising a step of reacting a protein material containing a GTW sequence therein with a protease, wherein the protease is derived from Aspergillus melleus, Streptomyces griseus or Bacillus licheniformis.

3. The method according to claim 1, wherein the protease is derived from Aspergillus melleus, Streptomyces griseus or Bacillus licheniformis.

4. The method according to any one of claims 1 to 3, wherein the protein material containing the GTW sequence therein is whey protein.

5. The method according to claim 4, wherein the whey protein is β-lactoglobulin.

6. A composition comprising a GTW peptide produced by the method according to any one of claims 1 to 3.

7. A composition comprising a GTW peptide, the GTW peptide content being 0.1% (w / w) or more from a proteinaceous material containing a GTW sequence therein.

8. A composition comprising the GTW peptide of claim 7, wherein the protein material containing the GTW sequence therein is whey protein.

9. A composition comprising the GTW peptide according to claim 8, wherein the whey protein is β-lactoglobulin.

10. A food or beverage comprising a composition containing a GTW peptide, wherein the GTW content derived from a protein material containing a GTW sequence therein is 0.0001% (w / w) or more.

11. The food or drink according to claim 10, wherein the protein material containing the GTW sequence therein is whey protein.

12. The food or drink according to claim 11, wherein the whey protein is β-lactoglobulin.

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