Method for producing processed vegetable protein-containing liquid composition

Abstract

The purpose of the present invention is to provide a processing technique for improving the solubility and / or digestibility of a protein relative to a liquid composition containing a vegetable protein. By treating a liquid composition containing a vegetable protein with a protein deamidase and a cellulase, the solubility, digestibility, and / or mineral solubility of the protein in the liquid composition containing a vegetable protein can be improved.

Description

Technical Field

[0001] The present invention relates to a method for producing a processed liquid composition containing vegetable protein. More specifically, the present invention relates to a method for improving the protein solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein. Background Art

[0002] Beverages rich in nutrients such as protein have long been popular because they allow for easy intake of nutrients. Meanwhile, with the recent rise in vegetarians, allergy concerns, and religious reasons, plant-based milks, made from plant-based ingredients rich in plant protein, have become increasingly popular as an alternative to animal milk, particularly cow's milk.

[0003] Generally, plant-based milk, unlike animal milk, has a low protein content, which is cited as a technical problem. Therefore, it is desired to improve the protein solubility of a liquid composition containing plant protein in plant-based milk. For example, Patent Document 1 discloses a method of increasing the protein solubility of a plant-based milk by using amylase, β - A method for producing a protein-containing oat-based composition by treating oats with amylase and protein deamidase. Patent Document 2 discloses a method for producing an oat protein composition having a protein content of 50% or greater by treating oats with protein deamidase. Patent Document 3 discloses a method for producing a processed liquid composition containing vegetable protein by treating the liquid composition with protease and protein deamidase to increase its solubility.

[0004] Furthermore, it is known that plant proteins are less digestible than animal proteins (Non-Patent Document 1), and the discrepancy between food intake and nutrient absorption is also a problem.

[0005] Prior art literature Patent Literature Patent Document 1: International Publication No. 2014 / 123466 Patent Document 2: International Publication No. 2022 / 144452 Patent Document 3: International Publication No. 2022 / 102723 Non-patent literature Non-patent literature 1: Nutrition and Food, Vol. 20, No. 4, pp. 259-266 Summary of the Invention

[0006] Technical problem to be solved by the invention Liquid compositions containing vegetable proteins can be treated with protein deamidase to improve protein solubility. On the other hand, in view of the recent rapid expansion of plant protein foods and beverages, technologies that further improve protein solubility are desired. It should be noted that treating a liquid composition containing vegetable proteins with protein deamidase and protease can improve protein solubility, but if solubility can be improved without hydrolyzing the protein, it is believed that the properties of the protein itself (such as mouthfeel) can be easily enjoyed. Alternatively, in order to more efficiently ingest the protein and / or minerals contained in the liquid composition containing vegetable proteins, it is desired to improve food properties such as digestibility and / or mineral solubility.

[0007] Therefore, an object of the present invention is to provide a novel processing technology for improving the protein solubility, digestibility and / or mineral solubility of a liquid composition containing vegetable protein.

[0008] The present inventors conducted intensive research and discovered that treating a liquid composition containing vegetable protein with a protein deamidase and a cellulase can improve protein solubility, digestibility, and mineral solubility, compared to treating the liquid composition containing vegetable protein with the protein deamidase alone. Based on these findings, further research led to the completion of this invention.

[0009] That is, the present invention provides the following aspects.

[0010] Item 1. A method for producing a processed liquid composition containing vegetable protein, comprising the step of treating the liquid composition containing vegetable protein with a protein deamidase and a cellulase.

[0011] Item 2. The method for producing a processed liquid composition containing vegetable protein according to Item 1, wherein the vegetable protein is a cereal protein.

[0012] Item 3. The method for producing a processed liquid composition containing vegetable protein according to Item 1 or 2, wherein the vegetable protein is oat protein.

[0013] Item 4. The production method according to any one of Items 1 to 3, wherein the cellulase is a cellulase derived from the genus Aspergillus and / or a cellulase derived from the genus Trichoderma.

[0014] Item 5. The production method according to any one of Items 1 to 4, wherein the cellulase has a higher exoglucanase activity than an endoglucanase activity.

[0015] Item 6. The method for producing a processed plant protein-containing liquid composition according to any one of Items 1 to 5, wherein 2 to 5000 U of cellulase is used per 1 U of protein deamidase.

[0016] Item 7. The method for producing a processed liquid composition containing a vegetable protein according to any one of Items 1 to 6, wherein 0.01 to 200 U of the protein deamidase is used per 1 g of the vegetable protein.

[0017] Item 8. A method for improving the protein solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein, comprising the step of treating the liquid composition containing vegetable protein with a protein deamidase and a cellulase.

[0018] Item 9. An agent for improving protein solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein, comprising a protein deamidase and a cellulase.

[0019] Item 10. The agent for improving protein solubility, digestibility, and / or mineral solubility according to Item 9, comprising 2 to 5000 U of cellulase per 1 U of protein deamidase.

[0020] Item 11. An agent for improving protein solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein treated with protein deamidase, characterized in that it contains cellulase.

[0021] Item 12. A plant protein-containing food or beverage comprising a processed plant protein-containing liquid composition obtained by the production method according to any one of Items 1 to 7.

[0022] Effects of the Invention According to the present invention, there is provided a processing technology for improving the solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein. DETAILED DESCRIPTION

[0023] 1. Method for producing a processed liquid composition containing vegetable protein The method for producing a processed plant protein-containing liquid composition of the present invention is characterized by including a step of treating the plant protein-containing liquid composition with a protein deamidase and a cellulase (enzyme treatment step). The enzymatic treatment step improves the solubility, digestibility, and / or solubility of the protein in the plant protein-containing liquid composition, as well as the solubility of minerals (at least one of calcium, iron, zinc, and magnesium; the same shall apply hereinafter). This yields a processed plant protein-containing liquid composition with enhanced protein solubility, digestibility, and / or mineral solubility. The method for producing the processed plant protein-containing liquid composition of the present invention is described in detail below.

[0024] 1-1. Enzyme treatment process In the enzyme treatment step, the liquid composition containing vegetable protein is treated with protein deamidase and cellulase.

[0025] 1-1-1. Liquid composition containing plant protein The vegetable protein-containing liquid composition used in the present invention is not particularly limited as long as it is a liquid containing vegetable protein and cellulose in water. Specific examples of liquid compositions containing vegetable protein include: (i) a liquid obtained by dispersing a dry powder of at least one of a plant organ of a plant source of vegetable protein and a material obtained by increasing the protein and / or cellulose content by removing at least a portion of components other than protein and / or cellulose from the plant organ (hereinafter referred to as "plant protein material") in water, and removing impurities such as the skin of the vegetable protein material by centrifugation, filtration, a filter bag, a sieve, etc. as needed; (ii) a liquid obtained by crushing and dispersing the vegetable protein material in water, and removing impurities such as the skin of the vegetable protein material by centrifugation, filtration, a filter bag, a sieve, etc. as needed; (iii) a liquid obtained by increasing the protein and / or cellulose content by removing at least one of the components other than vegetable protein and / or cellulose from the liquid of (i) or (ii); and (iv) a liquid obtained by mixing a dry powder prepared from any of the liquids (i) to (iii) with water. Preferred examples of liquid compositions containing vegetable protein include vegetable milk.

[0026] It should be noted that, below, when referring to the "content of the plant protein material" in a liquid composition containing plant protein, this refers to the dry weight ratio of the components comprising the plant organs contained in the liquid composition containing plant protein. For example, if the liquid composition containing plant protein is a liquid composed solely of components derived from plant organs and water, such as the liquids (i) to (iv) above, the "content of the plant protein material" refers to the dry weight ratio of the liquid. Alternatively, if the liquid composition containing plant protein is a liquid containing the liquids (i) to (iv) above and an additive, the "content of the plant protein material" refers to the dry weight ratio of the liquid excluding the additive.

[0027] The plant-based protein materials are not particularly limited, and examples thereof include: beans such as soybeans, peas, lentils, chickpeas, black beans, broad beans, mung beans, lupines, and kidney beans; grains such as wheat, barley, oats, sorghum, rice, rye, buckwheat, millet, chestnuts, teff, corn, and potatoes; nuts such as almonds, coconuts, peanuts, cashews, hazelnuts, pecans, macadamia nuts, pistachios, walnuts, Brazil nuts, pili nuts, chestnuts, sesame seeds, and pine nuts; and seeds such as hemp seeds (industrial hemp), chia seeds, quinoa, amaranth seeds, canary seeds, and flax seeds.

[0028] In the present invention, the above-mentioned plant protein materials may be used alone or in combination. Among the above-mentioned plant protein materials, preferred are cereals, more preferred are wheat, barley, oats, sorghum, rice, rye, buckwheat, barnyard millet, chestnut, teff, corn, and potato, and even more preferred is oats.

[0029] The content of the vegetable protein in the liquid composition containing the vegetable protein is not particularly limited, and examples thereof include 0.01 to 50 wt %, 0.05 to 40 wt %, 0.1 to 20 wt %, preferably 0.25 to 10 wt %, 0.5 to 5.0 wt %, more preferably 1.0 to 2.0 wt %, or 1.2 to 1.8 wt %.

[0030] The cellulose content in the liquid composition containing vegetable protein is not particularly limited, and examples thereof include 0.005 to 5% by weight, 0.01 to 4% by weight, 0.05 to 3% by weight, preferably 0.1 to 2.5% by weight, 0.3 to 2% by weight, more preferably 0.5 to 1.5% by weight, and even more preferably 0.9 to 1.3% by weight, in terms of dietary fiber content.

[0031] In addition, the content of the vegetable protein material in the liquid composition containing vegetable protein is not particularly limited, and examples thereof include 0.05 to 50 weight %, 0.1 to 40 weight %, 0.5 to 30 weight %, preferably 1 to 25 weight %, 3 to 20 weight %, and more preferably 5 to 15 weight %, 10 to 13 weight %.

[0032] In addition, when the vegetable protein material contained in the liquid composition containing vegetable protein is a starchy material such as a cereal (preferably oats), such a liquid composition containing vegetable protein is preferably a liquid composition treated with amylase as described below in "1-2-2. Amylase Treatment Step". Examples of the starch content in the liquid composition containing vegetable protein before the amylase treatment step include 0.035 to 35% by weight, 0.07 to 28% by weight, 0.35 to 21% by weight, preferably 0.7 to 18% by weight, 2 to 14% by weight, and more preferably 4 to 10% by weight.

[0033] 1-1-2. Protein deamidase The protein deamidase used in the present invention is an enzyme that decomposes the amide group-containing side chain of a protein without cleavage of peptide bonds or protein cross-linking, and its type and origin are not particularly limited.

[0034] Examples of protein deaminases include enzymes that deamidate glutamine residues in proteins to convert them to glutamate (e.g., protein glutaminases), and enzymes that deamidate asparagine residues in proteins to convert them to aspartic acid (e.g., protein asparaginases). These protein deaminases may be used alone or in combination of two or more. Among these protein deaminases, protein glutaminases are preferred from the perspective of further improving protein solubility, digestibility, and / or mineral solubility.

[0035] More specific examples of protein deamidases include those derived from Chryseobacterium, Flavobacterium, Empedobacterium, Sphingobacterium, Aureobacterium, Myroides, Luteimicrobium, Agromyces, Microbacterium, or Leifsonia. These protein deamidases are well known, and reference can be made to, for example, JP2000-50887A, JP2001-218590A, WO2006 / 075772A1, and WO2015 / 133590.

[0036] The protein deamidating enzyme may be a natural protein deamidating enzyme or a recombinant protein deamidating enzyme.

[0037] These protein deaminases may be used alone or in combination. Among these protein deaminases, naturally occurring protein deaminases, from the perspective of further improving protein solubility, digestibility, and / or mineral solubility, are preferably protein deaminases derived from the genus Chryseobacterium, more preferably protein glutaminases derived from the genus Chryseobacterium, still more preferably protein glutaminases derived from the species Chryseobacterium proteolyticum, and still more preferably protein glutaminases derived from Chryseobacterium proteolyticum strain 9670.

[0038] Protein deamidase can be prepared from the culture fluid of a transformant that has been integrated with a gene related to a microorganism or recombinant protein deamidase that is the source of the above-mentioned protein deamidase. As a specific preparation method, a method of recovering protein deamidase from the culture fluid or bacterial cells of the above-mentioned microorganisms can be mentioned. For example, when using a protein deamidase secreting microorganism, the bacterial cells can be recovered from the culture fluid in advance by filtration, centrifugation, etc., as needed, and the enzyme can be separated and / or purified. In addition, when using a non-protein deamidase secreting microorganism, the bacterial cells can be recovered from the culture fluid in advance as needed, and then the bacterial cells can be broken by pressure treatment, ultrasonic treatment, etc. to expose the enzyme, and the enzyme can be separated and / or purified. As the enzyme separation and / or purification method, known protein separation and / or purification methods can be used without particular limitation, for example, centrifugation, UF concentration method, salting-out method, various chromatography methods using ion exchange resins, etc. can be mentioned. The enzyme obtained by separation and / or purification can be powdered by drying methods such as freeze drying and reduced pressure drying. In addition, it can also be powdered using appropriate excipients and / or drying aids in the drying method. Alternatively, the isolated and / or purified enzyme may be liquefied by adding appropriate additives and sterilizing by filtration.

[0039] The amount of protein deamidase used is not particularly limited, but an example of an amount of 0.01 U or more per 1 g of vegetable protein can be used. From the viewpoint of further enhancing the solubility, digestibility, and / or mineral solubility of the protein, the amount of protein deamidase used per 1 g of vegetable protein is preferably 0.1 U or more, or 0.2 U or more, more preferably 0.5 U or more, even more preferably 1 U or more, or 3 U or more, even more preferably 6 U or more, and even more preferably 8 U or more.

[0040] The upper limit of the amount of protein deamidase used per 1 g of plant protein is not particularly limited, and examples thereof include 200 U or less, 100 U or less, 50 U or less, 30 U or less, 20 U or less, 15 U or less, or 12 U or less.

[0041] Furthermore, the amount of the protein deamidase used per 1 g (dry weight conversion) of the plant protein material is, for example, 0.0013 U or more. From the viewpoint of further enhancing the solubility and digestibility of the protein and / or the effect of improving the solubility of the mineral, the amount of the protein deamidase used per 1 g (dry weight conversion) of the plant protein material is preferably 0.013 U or more or 0.026 U or more, more preferably 0.065 U or more, even more preferably 0.13 U or more or 0.4 U or more, even more preferably 0.8 U or more, and even more preferably 1 U or more.

[0042] The upper limit of the amount of protein deamidase used per 1 g of the plant protein material (dry weight conversion) is not particularly limited, and examples thereof include 26 U or less, 13 U or less, 6.5 U or less, 4 U or less, 2.6 U or less, 2 U or less, or 1.5 U or less.

[0043] Regarding the activity of protein deamidase, benzyloxycarbonyl-L-glutaminoglycine (Z-Gln-Gly) is used as substrate and 1 μ The amount of enzyme per mol of ammonia is defined as 1 unit (1U).

[0044] 1-1-3. Cellulase The cellulase used in the present invention is a hydrolyzing β The enzyme that binds the glycosidic bonds of -1,4-glucan is not particularly limited in type or origin.

[0045] As the example of cellulase, can enumerate endoglucanase and exoglucanase.These cellulases can use 1 kind separately, also can be used in combination more than 2 kinds.These cellulases all can improve protein solubility, but from the viewpoint of further improving protein solubility improving effect, preferably enumerate the cellulase that at least comprises exoglucanase (that is, the cellulase that comprises exoglucanase or the cellulase that comprises exoglucanase and endoglucanase), more preferably enumerate the cellulase that mainly comprises exoglucanase (that is, the cellulase that exoglucanase activity is higher than endoglucanase activity), further preferably enumerate exoglucanase. In addition, these cellulases can all improve the digestibility of protein, but from the viewpoint of further improving the digestibility of protein, preferably enumerate a cellulase that at least comprises an exoglucanase (that is, a cellulase that comprises an exoglucanase or a cellulase that comprises an exoglucanase and an endoglucanase), more preferably enumerate a cellulase that mainly comprises an exoglucanase (that is, a cellulase whose exoglucanase activity is higher than that of an endoglucanase), and further preferably enumerate an exoglucanase. And then, these cellulases can all improve the solubility of minerals, but from the viewpoint of further improving the solubility of zinc and magnesium, preferably enumerate a cellulase that at least comprises an exoglucanase (that is, a cellulase that comprises an exoglucanase or a cellulase that comprises an exoglucanase and an endoglucanase), more preferably enumerate a cellulase that mainly comprises an exoglucanase (that is, a cellulase whose exoglucanase activity is higher than that of an endoglucanase), and further preferably enumerate an exoglucanase.

[0046] Specific examples of cellulase include cellulase derived from microorganisms, and more specific examples include cellulase derived from the genera Aspergillus, Trichoderma, and Acremonium.

[0047] As more specific examples, cellulases derived from the genus Aspergillus include cellulases derived from Aspergillus niger, Aspergillus oryzae, Aspergillus sojae, Aspergillus saitoi, Aspergillus awamori, and Aspergillus flavus; cellulases derived from the genus Trichoderma include cellulases derived from Trichoderma reesei and Trichoderma viride; and cellulases derived from the genus Acremonium include cellulase derived from Acremonium cellulolyticus.

[0048] The cellulase may be a natural cellulase or a recombinant cellulase.

[0049] These cellulases can be used alone or in combination. Among the natural cellulases in these cellulases, from the viewpoint of further improving the solubility of protein, preferably cite a cellulase derived from Aspergillus and a cellulase derived from Trichoderma, more preferably cite a cellulase derived from Aspergillus niger and a cellulase derived from Trichoderma viride. In addition, among the natural cellulases in these cellulases, from the viewpoint of further improving the digestibility of protein, preferably cite a cellulase derived from Aspergillus and a cellulase derived from Trichoderma, more preferably cite a cellulase derived from Trichoderma, further preferably cite a cellulase derived from Trichoderma viride. Furthermore, among the natural cellulases among these cellulases, from the viewpoint of further improving the effect of improving the solubility of minerals, cellulases derived from the genus Aspergillus and cellulases derived from the genus Trichoderma are preferably exemplified; from the viewpoint of further improving the solubility of zinc and magnesium, cellulases derived from the genus Trichoderma are more preferably exemplified, and cellulases derived from Trichoderma viride are further preferably exemplified; from the viewpoint of comprehensively improving the solubility of calcium, iron, zinc and magnesium, a combination of cellulases derived from the genus Aspergillus and cellulases derived from the genus Trichoderma is more preferably exemplified, and a combination of cellulases derived from Aspergillus niger and cellulases derived from Trichoderma viride is further preferably exemplified.

[0050] Cellulase can be prepared from the culture fluid of a transformant having a gene related to a microorganism or recombinant cellulase that is the source of the above-mentioned cellulase. As a specific preparation method, a method for recovering cellulase from the culture fluid or bacterial cell of the above-mentioned microorganism can be cited. For example, when using a cellulase secreting microorganism, it is possible to recover the bacterial cell from the culture fluid in advance by filtration, centrifugation, etc. as needed, and then separate and / or purify the enzyme. In addition, when using a cellulase non-secreting microorganism, it is possible to recover the bacterial cell from the culture fluid in advance as needed, and then break the bacterial cell by pressure treatment, ultrasonic treatment, etc. to expose the enzyme, and then separate and / or purify the enzyme. As an enzyme separation and / or purification method, known protein separation and / or purification methods can be used without particular limitation, for example, centrifugation, UF concentration method, salting-out method, various chromatography methods using ion exchange resins, etc. can be cited. The enzyme obtained by separation and / or purification can be powdered by drying methods such as freeze drying and reduced pressure drying. In addition, it is also possible to use a suitable excipient and / or drying aid to powderize in the drying method. Alternatively, the isolated and / or purified enzyme may be liquefied by adding appropriate additives and sterilizing by filtration.

[0051] The amount of cellulase used is not particularly limited, and an example of the amount used relative to 1g of dietary fiber is 1 U or more. From the viewpoint of further improving the effect of improving protein solubility, digestibility, and / or mineral solubility, the amount used relative to 1g of dietary fiber is preferably 3 U or more, 6 U or more, 10 U or more, 13 U or more, 17 U or more, 20 U or more, 30 U or more, 40 U or more, 50 U or more, 70 U or more, 100 U or more, 300 U or more, 400 U or more, 500 U or more, 800 U or more, 1000 U or more, or 1300 U or more.

[0052] The upper limit of the range of the amount of cellulase used relative to 1 g of dietary fiber is not particularly limited, and examples thereof include 6000U or less, 5000U or less, 4000U or less, 3000U or less, 2000U or less, 1700U or less, 1500U or less, 1200U or less, 900U or less, 600U or less, 400U or less, 200U or less, 90U or less, 80U or less, 60U or less, 50U or less, 40U or less, 30U or less, 20U or less, 15U or less, 8U or less, or 4U or less.

[0053] The amount of cellulase used per 1 g (dry weight conversion) of the vegetable protein material is, for example, 0.1 U or more. From the viewpoint of further enhancing the effects of improving protein solubility, digestibility, and / or mineral solubility, the amount of cellulase used per 1 g (dry weight conversion) of the vegetable protein material is preferably 0.3 U or more, 0.6 U or more, 1 U or more, 1.3 U or more, 1.7 U or more, 2 U or more, 3 U or more, 4 U or more, 5 U or more, 7 U or more, 10 U or more, 30 U or more, 40 U or more, 70 U or more, 100 U or more, or 130 U or more. The upper limit of the range of the amount of cellulase used per 1 g of the plant protein material (on a dry weight basis) is not particularly limited, but examples thereof include 600 U or less, 500 U or less, 400 U or less, 300 U or less, 200 U or less, 170 U or less, 150 U or less, 120 U or less, 90 U or less, 60 U or less, 40 U or less, 20 U or less, 9 U or less, 8 U or less, 6 U or less, 5 U or less, 4 U or less, 3 U or less, 2 U or less, 1.5 U or less, 0.8 U or less, or 0.4 U or less.

[0054] The ratio of protein deamidase to cellulase used is determined based on the above-described usage amounts of each enzyme. For example, the usage amount of cellulase per 1 U of protein deamidase may be 0.1 U or more. From the viewpoint of further enhancing the effects of improving protein solubility, digestibility, and / or mineral solubility, the usage amount of cellulase per 1 U of protein deamidase may preferably be 0.3 U or more, 0.6 U or more, 1 U or more, 1.3 U or more, 1.7 U or more, 2 U or more, 3 U or more, 4 U or more, 5 U or more, 7 U or more, 10 U or more, 30 U or more, 40 U or more, 70 U or more, 100 U or more, or 130 U or more. The upper limit of the range of the amount of cellulase used relative to 1 U of protein deamidase is not particularly limited, but examples thereof include 600 U or less, 500 U or less, 400 U or less, 300 U or less, 200 U or less, 170 U or less, 150 U or less, 120 U or less, 90 U or less, 60 U or less, 40 U or less, 20 U or less, 9 U or less, 8 U or less, 6 U or less, 5 U or less, 4 U or less, 3 U or less, 2 U or less, 1.5 U or less, 0.8 U or less, or 0.4 U or less.

[0055] In addition, when a combination of an Aspergillus-derived cellulase and a Trichoderma-derived cellulase (preferably a combination of an Aspergillus niger-derived cellulase and a Trichoderma-derived cellulase) is used as the cellulase, the amount of the Trichoderma-derived cellulase used relative to 1 U of the Aspergillus-derived cellulase may be, for example, 0.02 U or more. From the viewpoint of further improving the effect of improving protein solubility, digestibility, and / or mineral solubility (combined solubility of calcium, iron, zinc, and magnesium), the amount may be preferably 0.03 to 0.7 U, preferably 0.04 to 0.6 U, more preferably 0.045 to 0.3 U or 0.045 to 0.2 U, and even more preferably 0.05 to 0.1 U or 0.05 to 0.08 U.

[0056] Regarding cellulase activity, when carmellose sodium (carboxymethyl-cellulose sodium; degree of etherification 0.62 to 0.68) is used as a substrate and treated at pH 4.5 and 40°C, the amount of enzyme that produces reducing sugars equivalent to 1 mg of glucose in 1 minute is defined as 100 units (100 U).

[0057] 1-1-4. Reaction conditions, etc. In the enzyme treatment step, reactions that improve protein solubility, digestibility, and / or mineral solubility occur in the liquid composition containing vegetable protein through this treatment.

[0058] The order in which the protein deamidase and the cellulase are allowed to act is not particularly limited. The enzymes may be allowed to act sequentially in any order, or both enzymes may be allowed to act simultaneously. However, it is preferred that both enzymes be allowed to act simultaneously.

[0059] The conditions of the enzyme treatment step (temperature, pH, time, etc.) can be appropriately selected according to the properties of the enzyme used and the liquid composition containing the plant protein, and the degree of improvement in the solubility, digestibility, and / or mineral solubility of the target protein.

[0060] The temperature for performing the enzyme treatment step is not particularly limited and can be appropriately determined by those skilled in the art based on the optimal temperature for using the enzyme and / or the thermal properties of the liquid composition containing vegetable protein. For example, it can be 10°C to 70°C, preferably 25°C to 67°C, more preferably 30°C to 65°C, further preferably 45°C to 63°C, 47°C to 53°C, 55°C to 63°C, or 57°C to 63°C.

[0061] The pH of the reaction system for the enzyme treatment step is not particularly limited and can be appropriately determined by those skilled in the art based on the optimal pH of the enzyme used and / or the pH characteristics of the liquid composition containing the plant protein. Examples of the pH at 25°C include pH 1.0 to 8.0, preferably pH 3.0 to 8.0, more preferably pH 5.5 to 8.0, further preferably pH 6.5 to 8.0, and further preferably pH 7.0 to 7.5.

[0062] The time for performing the enzyme treatment step can be appropriately determined depending on the scale of the liquid composition containing vegetable protein to be subjected to the enzyme treatment step and / or the degree of improvement in the solubility, digestibility and / or mineral solubility of the target protein, and can be, for example, 5 minutes to 48 hours, preferably 30 minutes to 20 hours, and more preferably 3 hours to 5 hours.

[0063] 1-2. Other processes The method for producing a processed liquid composition containing vegetable protein of the present invention may or may not include any other steps other than the protein deamidation step and the cellulase-based enzyme treatment step described in "1-1. Enzyme Treatment Step" above, as long as the effects of the present invention are achieved. Examples of other steps include a step for preparing the liquid composition containing vegetable protein, an amylase treatment step, post-treatment steps (cooling step, enzyme inactivation step, filtration step, etc.), and a drying step. These other steps may be performed individually or in combination of two or more steps. Furthermore, the timing of performing these other steps may or may not overlap with the above-mentioned enzyme treatment step.

[0064] 1-2-1. Preparation process of liquid composition containing vegetable protein The step of preparing a liquid composition containing vegetable protein can be performed by any method capable of preparing a liquid composition containing vegetable protein. Examples include: (I) a method of dispersing a dry powder of a vegetable protein material in water and, if necessary, removing insoluble matter such as the skin of the vegetable protein material by centrifugation, filtration, a filter bag, or a sieve; (II) a method of crushing and dispersing the vegetable protein material in water and, if necessary, removing insoluble matter such as the skin of the vegetable protein material by centrifugation, filtration, a filter bag, or a sieve; (III) a method of increasing the vegetable protein content by removing at least one component other than vegetable protein from the liquid obtained by the above method (I) or (II); and (IV) a method of mixing a dry powder prepared from a liquid obtained by any of the above methods (I) to (III) with water.

[0065] 1-2-2. Amylase treatment process When the liquid composition containing plant protein is prepared as an amylase-treated composition, the liquid obtained by any of the methods (I) to (IV) above can be treated with amylase. The amylase treatment step can be performed before or simultaneously with the enzyme treatment step. It is preferably performed before the enzyme treatment step. When the amylase treatment step is performed before the enzyme treatment step, the enzyme treatment step can be performed after inactivating the amylase or without inactivating the amylase.

[0066] Examples of amylase include α -Amylase and β -Amylase. Among these amylases, any one can be used or two can be used in combination, preferably α -Amylase.

[0067] As the α -Amylase, not particularly limited, for example, amylase derived from Aspergillus α -Amylase, derived from Bacillus α -amylase. More specifically, as derived from Aspergillus α -Amylase, such as those derived from Aspergillus oryzae and Aspergillus niger α -Amylase; as derived from Bacillus sp. α -Amylases, such as those derived from Bacillus amyloliquefaciens, Bacillus subtilis, Bacillus licheniformis, and Bacillus flexus α -Amylase. These α -Amylase can be used alone or in combination of two or more. α Among amylases, preferably those derived from Bacillus α - amylase, more preferably derived from Bacillus amyloliquefaciens α -Amylase.

[0068] about α The amount of amylase used is, for example, 0.001 U or more, preferably 0.05 U or more, more preferably 0.01 U or more, 0.05 U or more, 0.1 U or more, 0.2 U or more, or 0.5 U or more, relative to 1 g of starch contained in the liquid composition containing vegetable protein. αThe upper limit of the usage range of the amylase is not particularly limited, and examples thereof include 100 U or less, 50 U or less, 10 U or less, 5 U or less, 1 U or less, 0.6 U or less, 0.3 U or less, 0.1 U or less, 0.06 U or less, or 0.03 U or less.

[0069] about α -Amylase activity is measured as 1 unit (1U) of the enzyme that reduces the iodine color of potato starch by 10% within 1 minute.

[0070] As the β -Amylase is not particularly limited, and examples thereof include those derived from plants (wheat, soybean) and those derived from Bacillus spp. β -Amylase, preferably derived from Bacillus β -amylase, more preferably derived from Bacillus flexus β -Amylase.

[0071] about β The amount of amylase used is, for example, 0.001 to 50 U, 0.01 to 10 U, 0.02 to 5.0 U, 0.03 to 2.5 U, and preferably 0.05 to 1.5 U per 1 g of starch contained in the liquid composition containing vegetable protein.

[0072] about β -Amylase activity is defined as the amount of enzyme that increases the reducing power equivalent to 1 mg of glucose in 1 minute, and 1 unit (1U) is defined as the amount of enzyme that increases the reducing power equivalent to 1 mg of glucose in 1 minute.

[0073] The conditions for the amylase treatment (treatment pH, treatment temperature, and treatment time) can be appropriately set based on the optimal pH and temperature of the enzyme used, the scale of the liquid composition containing vegetable protein, and other factors. The treatment pH of the oat protein-containing liquid composition before amylase treatment at 25°C can be, for example, 5.5 to 8.0, preferably 6.5 to 8.0, and more preferably 7.0 to 7.5. The treatment temperature can be, for example, 35°C to 70°C, preferably 40°C to 67°C, more preferably 45°C to 65°C, 47°C to 53°C, 55°C to 63°C, or 57°C to 63°C. The treatment time can be, for example, 10 minutes to 2 hours, preferably 20 minutes to 1 hour, and more preferably 20 minutes to 40 minutes.

[0074] 1-2-3. Enzyme inactivation process In the enzyme inactivation step, the temperature and / or pH conditions for inactivating the enzymes used can be appropriately selected, preferably taking into account the temperature and / or pH characteristics of the liquid composition containing plant protein. Specific temperature conditions include, for example, 85 to 120°C, preferably 90 to 100°C, and more preferably 93 to 98°C. The treatment time includes, for example, 1 to 30 minutes, or 1 to 10 minutes, and preferably 3 to 8 minutes.

[0075] 1-2-4. Cooling process, filtration process, drying process The cooling, filtration, and / or drying steps can be performed as needed after the enzyme inactivation step. In the drying step, the processed liquid vegetable protein-containing composition can be converted into a solid vegetable protein composition with improved protein solubility, digestibility, and / or mineral solubility. The drying method is not particularly limited, and examples thereof include freeze drying, vacuum drying, and spray drying. Examples of the form of the solid vegetable protein composition include powder, granules, and pellets.

[0076] 2. Protein solubility, digestibility and / or mineral solubility of liquid compositions containing vegetable proteins Enhancer The combination of a protein deamidase and a cellulase can improve the protein solubility, digestibility, and / or mineral solubility of a liquid composition containing a vegetable protein. Therefore, the present invention also provides an agent for improving the protein solubility, digestibility, and / or mineral solubility of a liquid composition containing a vegetable protein, comprising a protein deamidase and a cellulase.

[0077] In the agent for improving protein solubility, digestibility and / or mineral solubility of a liquid composition containing plant protein, "improving protein solubility, digestibility and / or mineral solubility" means imparting at least any one of the following properties to the liquid composition containing plant protein: a property of further increasing the amount of protein dissolved in water compared to the case of treatment with protein deamidase alone; a property of further increasing the amount of free amino acids in the body environment; and a property of further increasing the amount of minerals (at least one of calcium, iron, zinc and magnesium) dissolved in water.

[0078] The protein solubility, digestibility, and / or mineral solubility enhancing agent may consist solely of protein deamidase and cellulase. Alternatively, it may further include enzymes other than protein deamidase and cellulase, and / or additives and / or bases permitted in the formulation of the enzyme agent, or it may not. Examples of such additives and bases include excipients, buffers, antioxidants, UV inhibitors, preservatives, antiseptics, pH adjusters, dispersants, emulsifiers, solubilizers, carriers, and solvents (such as water). These additives and bases may be used singly or in combination of two or more. Furthermore, the content of these additives and bases may be appropriately set based on the type of these ingredients and / or the formulation form.

[0079] The properties of the agent for improving protein solubility, digestibility and / or mineral solubility are not particularly limited, and examples thereof include powdered, granular, granular dry preparations and liquid preparations.

[0080] In the protein solubility, digestibility and / or mineral solubility improving agent, the types and usage ratio (compounding ratio) of the protein deamidase and cellulase as active ingredients, as well as the method for using the solubilizing agent, are as shown in the above-mentioned "1. Method for producing a processed liquid composition containing vegetable protein".

[0081] 3. Protein soluble in liquid composition containing vegetable protein treated with protein deamidase Enhancers for enhancing digestibility, digestibility and / or mineral solubility Cellulase can improve the protein solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein treated with a protein deamidase. Therefore, the present invention also provides an agent comprising cellulase to improve the protein solubility, digestibility, and / or mineral solubility of a liquid composition containing vegetable protein treated with a protein deamidase.

[0082] It should be noted that, in the agent for improving protein solubility, digestibility, and / or mineral solubility of a liquid composition containing plant protein treated with protein deamidase, "improvement of protein solubility, digestibility, and / or mineral solubility" means imparting at least any one of the following properties to the liquid composition containing plant protein: a property of further increasing the amount of protein dissolved in water compared to the case of treatment with protein deamidase alone; a property of further increasing the amount of free amino acids in the in vivo environment; and a property of further increasing the amount of minerals (at least one of calcium, iron, zinc, and magnesium) dissolved in water.

[0083] The protein solubility, digestibility, and / or mineral solubility enhancer may consist solely of cellulase. Alternatively, it may further include enzymes other than cellulase and protein deamidase, and / or additives and / or bases permitted in the formulation of the enzyme agent, or it may not include these. Examples of such additives and bases include excipients, buffers, antioxidants, UV inhibitors, preservatives, antiseptics, pH adjusters, dispersants, emulsifiers, solubilizers, carriers, and solvents (such as water). One of these additives and bases may be used alone, or two or more may be used in combination. Furthermore, the content of these additives and bases may be appropriately set based on the type of these components and / or the formulation form.

[0084] The properties of the agent for improving protein solubility, digestibility and / or mineral solubility are not particularly limited, and examples thereof include powdered, granular, granular dry preparations and liquid preparations.

[0085] In the protein solubility, digestibility, and / or mineral solubility improving agent, the type of cellulase as an active ingredient and the method of using the protein solubility, digestibility, and / or mineral solubility improving agent are as described above in the column "1. Method for producing a processed liquid composition containing vegetable protein." Specific methods of using the protein solubility, digestibility, and / or mineral solubility improving agent include any of the following methods: a method of treating a liquid composition containing vegetable protein with the protein solubility, digestibility, and / or mineral solubility improving agent and a protein deamidase simultaneously; a method of treating a liquid composition containing vegetable protein with a protein deamidase and then treating it with the protein solubility, digestibility, and / or mineral solubility improving agent; and a method of treating a liquid composition containing vegetable protein with the protein solubility, digestibility, and / or mineral solubility improving agent and then treating it with a protein deamidase.

[0086] 4. Foods and beverages containing plant-based protein The plant protein-containing food and drink of the present invention includes a processed plant protein-containing liquid composition obtained by the above-mentioned "1. Method for Producing a Processed Plant Protein-Containing Liquid Composition." The processed plant protein-containing liquid composition is as described in the above-mentioned "1. Method for Producing a Processed Plant Protein-Containing Liquid Composition."

[0087] The specific form of the plant protein-containing food and drink of the present invention can be selected from any food and drink forms.

[0088] Specific forms of food and beverages containing plant protein include plant-based alternative milk, alternative yogurt, alternative cheese, and alternative ice.

[0089] The food and drink that contains vegetable protein can use the liquid composition that contains vegetable protein through processing, directly or through arbitrarily cooking process and obtain.In such cooking process, select the cooking method corresponding to the form of the food and drink that contains vegetable protein, specifically, can carry out seasoning, form adjustment, molding, heating cooking, fermentation, freezing etc.

[0090] Example Hereinafter, the present invention will be described in detail with reference to Examples, but the present invention is not to be construed as being limited to the following Examples.

[0091] [Enzymes and plant-based protein materials used] In the following examples, the following enzymes and plant protein materials were used.

[0092] [Table 1]

[0093] [Enzyme activity assay] (1) Protein deamidase activity assay To 1 mL of 0.2 M phosphate buffer (pH 6.5) containing 30 mM Z-Gln-Gly, add 0.1 mL of a sample solution containing protein deamidase. After incubation at 37°C for 10 minutes, add 1 mL of a 0.4 M TCA solution to stop the reaction. As a blank, add 1 mL of a 0.4 M TCA solution to 1 mL of 0.2 M phosphate buffer (pH 6.5) containing 30 mM Z-Gln-Gly. Furthermore, add 0.1 mL of a sample solution containing protein deamidase. The mixture is then incubated at 37°C for 10 minutes.

[0094] The resulting solution was then tested using the Ammonia-test Wako (Fujifilm Wako Pure Chemical Industries, Ltd.) to measure the amount of ammonia generated in the reaction solution. The ammonia concentration in the reaction solution was determined using a calibration curve plotting the relationship between ammonia concentration and absorbance (630 nm) using an ammonia standard solution (ammonium chloride).

[0095] Will generate 1 in 1 minute μ The activity of protein deamidase is calculated using the following formula: 1 unit (1 U) is defined as the amount of enzyme per mol of ammonia. Here, the reaction solution volume is 2.1, the enzyme solution volume is 0.1, and Df is the dilution factor of the enzyme solution. Furthermore, 17.03 is the molecular weight of ammonia.

[0096] [Number 1] Protein deamidase activity (U / mL) = ammonia concentration in reaction solution (mg / L) × (1 / 17.03) × (reaction solution volume / enzyme solution volume) × (1 / 10) × Df (2) Cellulase activity assay Accurately weigh approximately 1 g of sodium carmellose (sodium carboxymethylcellulose; degree of etherification 0.62-0.68), dry it at 105°C for 4 hours, and measure the weight loss. Weigh 0.500 g of the dried sodium carboxymethylcellulose and slowly (over approximately 30 minutes) add approximately 50 mL of hot water at 60-70°C to dissolve it while stirring continuously with a stirrer. Next, add 10 mL of 1 mol / L acetic acid / sodium acetate buffer (pH 4.5) and water to a total of 100 mL. This is used as the substrate solution.

[0097] Weigh 4 mL of substrate solution (pH 4.5) into a 50 mL Nessler colorimetric tube. After incubating at 40°C for 10-15 minutes, add 1 mL of a sample solution containing cellulase and immediately shake to mix. After incubating at 40°C for exactly 30 minutes, add 2 mL of Somogyi test solution 1 and shake to mix. Stopper the tube and heat in a boiling water bath for exactly 20 minutes before immediately cooling. After cooling, add 1 mL of Nelson's reagent and shake thoroughly until the red precipitate of cuprous oxide completely dissolves. Incubate at room temperature for approximately 20 minutes, add 17 mL of water to a total volume of 25 mL, and shake thoroughly to mix. The absorbance (A30) of the resulting solution is measured at a wavelength of 500 nm using water as a control. Separately, add 2 mL of Somogyi test solution 1 to 4 mL of substrate solution (pH 4.5) and shake to mix. Then, add 1 mL of a sample solution containing cellulase and repeat the same procedure to measure the absorbance (A0) of the resulting solution.

[0098] The amount of enzyme that generates reducing sugars equivalent to 1 mg of glucose in 1 minute was defined as 100 units (100 U), and the cellulase activity was calculated according to the following formula.

[0099] [Number 2] Cellulase activity relative to 1g or 1mL of sample (U / g, U / mL) = G×1 / 30×100×n G: The amount of glucose generated (mg) calculated from the (A30-A0) value using the glucose standard curve 1 / 30: Converted into a factor of 1 minute 100: coefficient for conversion to 100 units n: dilution factor relative to 1g or 1mL of sample a: Slope of the glucose standard curve b: intercept of the glucose standard curve (3) α -Amylase activity assay After heating 10 mL of 1% potato starch substrate solution (0.1 mol / L acetic acid (pH 5.0)) at 37°C for 10 minutes, add α Prepare 1 mL of a 1-amylase sample solution and immediately shake to mix. After incubating the solution at 37°C for 10 minutes, add 1 mL of this solution to 10 mL of a 0.1 mol / L hydrochloric acid solution and immediately shake to mix. Next, weigh 0.5 mL of this solution and add 10 mL of a 0.0002 mol / L iodine solution (Japanese Pharmacopoeia). After shaking to mix, measure the absorbance (AT) at a wavelength of 660 nm using water as a control. Also, add 1 mL of water in place of the sample solution and repeat the same procedure to measure the absorbance (AB). The amount of enzyme that reduces the color development caused by iodine in potato starch by 10% within 1 minute is defined as 1 unit (1 U).

[0100] [Number 3] α -Amylase activity (U / g, U / mL) = (AB-AT) / AB×1 / W AT: absorbance of the reaction solution AB: absorbance of blank solution W: The amount of sample in 1 mL of sample solution (g or mL) (4) Protease activity assay After heating 5 mL of a 0.6% (w / v) casein solution (0.05 mol / L sodium hydrogen phosphate, pH 8.0) at 37°C for 10 minutes, 1 mL of a sample solution containing a protease was added and immediately shaken to mix. After incubating the solution at 37°C for 10 minutes, 5 mL of a trichloroacetic acid test solution (containing 1.8% (w / v) trichloroacetic acid, 1.8% (w / v) sodium acetate, and 0.33 mol / L acetic acid) was added and shaken to mix. The solution was again incubated at 37°C for 30 minutes and filtered. The initial 3 mL of filtrate was removed, and the remaining 2 mL of the filtrate was weighed. 5 mL of a 0.55 mol / L sodium carbonate test solution and 1 mL of a Folin test solution (1→3) were added, the solution was thoroughly shaken to mix, and the solution was incubated at 37°C for 30 minutes. The absorbance (AT) of this solution (enzyme reaction solution) was measured at a wavelength of 660 nm, using water as a control.

[0101] Separately, 1 mL of a sample solution containing protease was weighed, 5 mL of a trichloroacetic acid test solution (containing 1.8% (w / v) trichloroacetic acid, 1.8% (w / v) sodium acetate, and 0.33 mol / L acetic acid) was added, and the mixture was shaken. 5 mL of a 0.6% (w / v) casein solution (0.05 mol / L sodium hydrogen phosphate, pH 8.0) was then added, the mixture was immediately shaken, and the mixture was allowed to stand at 37°C for 30 minutes. In addition, the absorbance AB of a liquid (blank) obtained by the same procedure as the above enzyme reaction solution was measured.

[0102] Will generate the equivalent of 1 in 1 minute μ The amount of enzyme that increases the color development substance of Folin test solution with g tyrosine is regarded as 1 unit (1U).

[0103] Weigh 1mL, 2mL, 3mL, and 4mL of 1mg / mL tyrosine standard stock solution (0.2mol / L hydrochloric acid), and add 0.2mol / L hydrochloric acid test solution to make 100mL respectively. Weigh 2mL of each liquid, add 5mL of 0.55mol / L sodium carbonate test solution and 1mL of Folin test solution (1→3), immediately shake and mix, and place at 37℃ for 30 minutes. For these liquids, weigh 2mL of 0.2mol / L hydrochloric acid test solution and use the same operation as above as a control to measure the absorbance A1, A2, A3, and A4 at a wavelength of 660nm. Take the absorbance A1, A2, A3, and A4 on the vertical axis, and the amount of tyrosine in 2mL of each liquid on the horizontal axis ( μ g), prepare a standard curve, and calculate the amount of tyrosine corresponding to an absorbance difference of 1 ( μ g).

[0104] [Number 4] Protease activity (U / g, U / mL) = (AT-AB) × F × 11 / 2 × 1 / 10 × 1 / M AT: absorbance of enzyme reaction solution AB: absorbance of blank solution F: The amount of tyrosine when the absorbance difference is 1, obtained from the tyrosine standard curve ( μ g) 11 / 2: Conversion factor of the total liquid volume after the reaction stops 1 / 10: Conversion factor per 1 minute of reaction time M: The amount of sample in 1 mL of sample solution (g or mL) [Test Example 1] (1) Manufacturing of processed oat milk 18.1 g of oat flour was mixed with 150 g of water and suspended in a 300 mL conical flask. 0.7 U / g-starch was added to the suspension (oat flour content 10.8 wt%, protein content 1.4 wt%, dietary fiber content 1.1 wt%, starch content 7.3 wt%). α oat milk (pH 7.0-7.5 at 25°C) was prepared by adding amylase and reacting at 60°C for 0.5 hours. Subsequently, 10 U / g protein of protein deamidase and 1500 U / g dietary fiber of cellulase 1 were added and reacted at 60°C for 2 or 4 hours. After the reaction, the enzymes were inactivated by heating at 95°C for 5 minutes. This produced processed oat milk (Examples 1 and 2). Comparative processed oat milk (Comparative Examples 1 and 2) was prepared by the same procedure except that cellulase 1 was not added. The resulting processed oat milk was used in the following experiments.

[0105] (2) Experiment on processing oat milk (2-1) Protein solubility determination Preparation method of sample solution for determination of processed oat milk The processed oat milk was centrifuged at 9000 rpm for 10 minutes, and the supernatant was recovered and further diluted 50-fold to prepare a sample solution for measurement.

[0106] Preparation method of BCA solution Thoroughly mix 50 mL of BCA Protein Assay Reagent (Pierce Reagent A) and 1 mL of BCA Protein Assay Reagent (Pierce Reagent B) to prepare a BCA solution.

[0107] Protein solubility determination method Weigh 4 mL of BCA solution and incubate at 37 ± 0.5°C for 10 minutes. Then, add 0.2 mL of the sample solution for measurement and immediately shake to mix. Incubate this solution at 37 ± 0.5°C for 30 minutes, then cool it in running water (20-25°C). Measure the absorbance (AT) of this solution at a wavelength of 562 nm using water as a reference. Separately, incubate 4 mL of BCA solution at 37 ± 0.5°C for 10 minutes, then add 0.2 mL of water and shake to mix. Repeat the same procedure to measure the absorbance (AB).

[0108] The albumin concentration Tmg in the sample dilution was calculated using the absorbance difference (AT-AB) and an albumin standard curve. The protein solubility (mg / g) in the processed oat milk was calculated based on the obtained albumin concentration Tmg. The relative solubility (%) of the protein in the processed oat milks of Examples 1 and 2 was calculated, assuming the protein solubility in the processed oat milks of Comparative Examples 1 and 2 was 100%, respectively. The results are shown in Table 2.

[0109] (2-2) Results [Table 2]

[0110] As shown in Table 2, the protein solubility was improved in the processed oat milks (Examples 1 and 2) obtained by the combined treatment with protein glutaminase (PG) and cellulase 1, compared with the processed oat milks (Comparative Examples 1 and 2) obtained by the treatment with protein glutaminase (PG) alone.

[0111] In the processed oat milks (Examples 3 and 4) produced in the same manner as in Examples 1 and 2 except that Cellulase 2 (cellulase derived from Trichoderma viride) was used instead of Cellulase 1 (cellulase derived from Aspergillus niger), improvement in protein solubility was also confirmed as in Examples 1 and 2.

[0112] Furthermore, in the processed oat milk produced by changing the amount of cellulase 1 added to 74.6 U / g-dietary fiber in Examples 1 and 2 (Examples 1a and 2a), and in the processed oat milk produced by changing the amount of cellulase 2 added to 14 U / g-dietary fiber in Examples 3 and 4 (Examples 3a and 4a), it was confirmed that the solubility of protein in the produced processed oat milk was improved, similar to Examples 1 and 2 and Examples 3 and 4.

[0113] [Test Example 2] (1) Manufacturing of processed oat milk In "(1) Production of Processed Oat Milk" of Test Example 1, the enzymes added to the oat milk were set as shown in Table 3, and the enzyme reaction time was set to 4 hours, thereby preparing processed oat milk (Comparative Examples 2, 4, and Example 2). The same procedures were followed except that no enzyme was added, and this is referred to as Comparative Example 3.

[0114] (2) Experiment on processing oat milk (2-1) Protein solubility determination The processed oat milk was centrifuged at 9000 rpm for 5 minutes, and the supernatant and residue were recovered. The residue was dried at 80°C for 6 hours. The supernatant and dried residue were used as samples for measurement.

[0115] The protein content of each of the measurement samples (supernatant and dry residue) was measured as nitrogen content by the Kjeldahl method. The results are shown in Table 3.

[0116] (2-2) Results [Table 3]

[0117] As shown in Table 3, when the protein content of the untreated oat milk (Comparative Example 3) was compared with the protein content of the enzyme-treated oat milk (Comparative Examples 2, 4, and Example 2), it was confirmed that the protein solubility of the processed oat milk (Example 2) obtained by the combined treatment with PG and cellulase 1 was improved, as compared with the processed oat milk obtained by the treatment with protein glutaminase (PG) alone (Comparative Example 2) and the processed oat milk obtained by the treatment with cellulase 1 alone (Comparative Example 4).

[0118] The same procedure was also performed on processed oat milks (Comparative Example 4a and Example 2a), produced in Comparative Example 4 and Example 2, by changing the amount of Cellulase 1 added to 74.6 U / g-dietary fiber. The results, comparing the protein content of the untreated oat milk (Comparative Example 3) with the protein content of the enzyme-treated oat milk (Comparative Examples 2, 4a, and Example 2a), showed, as shown in Table 3, that the processed oat milk treated with protein glutaminase (PG) alone (Comparative Example 2) and the processed oat milk treated with Cellulase 1 alone (Comparative Example 4a), showed a decrease in protein in the residue and a synergistic increase in protein in the supernatant, confirming improved protein solubility.

[0119] [Test Example 3] (1) Manufacturing of processed oat milk In the "(1) Production of Processed Oat Milk" of Test Example 1, the oat milk used in the production of oat milk was α -Amylase addition was set at 0.01U / g-starch, based on α The amylase treatment temperature was set at 60°C, the enzymes added to the resulting oat milk were as shown in Tables 4-6, and the enzyme treatment temperature was set at 50°C to prepare processed oat milk (Comparative Examples 6-7, 9-13, and Examples 5-9). The same procedures were followed except that no enzyme was added, and these are referred to as Comparative Examples 5 and 8.

[0120] (2) Experiment on processing oat milk (2-1) Protein solubility determination The measurement was performed in the same manner as in "(2-1) Protein solubility measurement" of Test Example 2. The results are shown in Tables 4 to 6.

[0121] (2-2) Results [Table 4]

[0122] [Table 5]

[0123] [Table 6]

[0124] As shown in Table 4, when the protein content of the oat milk without enzyme treatment (Comparative Examples 5 and 8) is compared with the protein content of the oat milk treated with enzyme (Comparative Examples 6 and 7, Example 5, Comparative Examples 9 and 10, Example 6), it can be confirmed that the protein solubility is improved in the processed oat milk obtained by the combined use of PG and cellulase 2 (Examples 5 and 6), as compared with the processed oat milk obtained by the treatment with protein glutaminase (PG) alone (Comparative Examples 6 and 9) and the processed oat milk obtained by the treatment with cellulase 2 alone (Comparative Examples 7 and 10).

[0125] As shown in Table 5, when the protein content of the untreated oat milk (Comparative Example 8) was compared with the protein content of the enzyme-treated oat milk (Comparative Examples 9 to 11, Examples 6 to 8), the protein content in the supernatant of the processed oat milk obtained by treating with protein glutaminase (PG) alone (Comparative Example 9) and the processed oat milk obtained by treating with cellulases 1 and 2 alone (Comparative Examples 11 and 10) was synergistically increased. Furthermore, the protein content in the supernatant of the processed oat milk obtained by treating with PG together with cellulases 1 and 2, respectively (Examples 7 and 6) was further synergistically increased. Therefore, it was confirmed that the solubility of the protein was improved.

[0126] As shown in Table 6, when the protein content of the untreated oat milk (Comparative Example 8) was compared with the protein content of the enzyme-treated oat milk (Comparative Examples 9 to 13, Example 9), the processed oat milk obtained by the combined treatment with PG, cellulase 1, and protease (Example 9) showed a synergistic increase in the protein content in the supernatant, compared with the processed oat milk obtained by the protein glutaminase (PG) treatment alone (Comparative Example 9), the processed oat milk obtained by the cellulase 1 treatment alone (Comparative Example 11), and the processed oat milk obtained by the protease treatment alone (Comparative Example 12), thereby confirming that the solubility of the protein was improved.

[0127] [Test Example 4] (1) Manufacturing of processed oat milk In "(1) Production of Processed Oat Milk" of Test Example 1, the enzymes added to the oat milk were set as shown in Table 7, and the enzyme reaction time was set to 4 hours, thereby preparing processed oat milk (Comparative Examples 2, 4, 14, Example 2, and 4). The same procedures were followed except that no enzyme was added, and this is referred to as Comparative Example 3.

[0128] (2) Experiment on processing oat milk (2-1) Digestibility test In vitro digestion studies were performed using 5 g of the processed oat milk according to the INFOGEST method (Brodkorb et al. 2019 Nat. Protoc.).

[0129] 5g of processed oat milk was mixed with 3.5mL of artificial saliva and 0.5mL of 0.77mg / mL amylase, and the mixture was stirred at 160rpm in a 37°C water bath for 2 minutes to create an oral digestive model. The composition of artificial saliva was 0.28M KCl, 0.07mM KH2PO4, 0.26mM NaHCO3, 2.8mM MgCl2, 1.1mM (NH4)2CO3, and 1.5mM CaCl2 (pH 7.0).

[0130] Next, 7.5 mL of artificial gastric juice and 1.6 mL of a 0.227 g / 19.2 mL pepsin solution (Sigma-Aldrich) were mixed with 10 mL of the oral digestive material model and stirred at 160 rpm in a 37°C water bath for 2 hours to prepare the gastric digestive material model. The composition of the artificial gastric juice was 0.10 M KCl, 0.01 M KH2PO4, 0.38 M NaHCO3, 0.71 M NaCl, 1.82 mM MgCl2, 7.58 mM (NH4)2CO3, and 0.165 mM CaCl2 (pH 3.0).

[0131] Furthermore, for 20 mL of the gastric digesta model, 11 mL of artificial intestinal fluid, 16.4 mg / 5 mL of pancreatic enzyme solution (Sigma-Aldrich), 215 mg / 2.5 mL of sodium taurocholate, 40 μ A 0.3 M calcium chloride solution was added to the flask and stirred at 160 rpm in a 37°C water bath for 2 hours to create a small intestinal digestion model. The composition of the artificial intestinal fluid was 8.5 mM KCl, 1 mM KH2PO4, 106.3 mM NaHCO3, 48 mM NaCl, and 0.4 mM MgCl2 (pH 7.0).

[0132] [Determination of free amino nitrogen content] To 1.0 mL of the appropriately diluted supernatant from the oral digestive model, gastric digestive model, or small intestinal digestive model was added 0.5 mL of ninhydrin (pH 6.7) (10% Na₂HPO₄ / 12H₂O, 6% KH₂PO₄, 0.50% ninhydrin, 0.30% fructose). The solution was then boiled for 16 minutes, and the free amino nitrogen content (mg / L) was measured using absorbance at 570 nm. A higher free amino nitrogen content indicates a higher amount of amino acids released from the protein during digestion, indicating a higher digestibility of the protein. The results are shown in Table 7.

[0133] [Amino acid analysis] Using the filter (0.45 μ m) Filter the supernatant of the small intestinal digestion model to prepare a sample for amino acid analysis. In addition, the supernatant of the small intestinal digestion model was prepared by performing the above-mentioned digestibility test except that PG and / or cellulase were not added, and the supernatant was further filtered (0.45 μ m) was filtered to prepare a control amino acid analysis sample. The amino acid analysis sample and the control amino acid analysis sample were analyzed for free amino acid levels using an amino acid analyzer (using an Agilent 1260 InfinityII LC system for amino acid analysis) according to the instrument's protocol. Specifically, free essential amino acids (EAAs) [specifically, branched-chain amino acids (BCAAs); valine, isoleucine, and leucine, aromatic amino acids (AAAs); phenylalanine and tryptophan, and other essential amino acids (methionine, lysine, histidine, arginine, and threonine)] and non-essential amino acids (NEAAs); glycine, alanine, serine, tyrosine, cysteine, asparagine, glutamine, proline, aspartic acid, and glutamic acid] were first measured. The ratio of the free amount of EAA (the total free amount of BCAA, AAA, and other EAA) to the free amount of NEAA in the amino acid analysis sample (EAA / NEAA) was derived. The relative amount of the ratio EAA / NEAA for each Example and Comparative Example was derived, assuming the ratio EAA / NEAA in Comparative Example 3 was set to 1. If this ratio exceeded 1, it means that the amount of EAA absorbed through digestion is higher than that of NEAA. The results are shown in Table 7.

[0134] (2-2) Results [Table 7]

[0135] As shown in Table 7, compared to processed oat milk treated with protein glutaminase (PG) alone (Comparative Example 2), processed oat milks treated with PG in combination with either cellulase 1 or cellulase 2 (Examples 2 and 4) showed improved protein digestibility and a higher efficiency of essential amino acid release through digestion. In particular, the processed oat milk treated with PG in combination with cellulase 2 (Example 4) showed a further improvement in protein digestibility and essential amino acid release through digestion.

[0136] The same procedure was also repeated for processed oat milk produced in Comparative Example 4 and Example 2 by changing the amount of Cellulase 1 added to 74.6 U / g-fiber (Comparative Example 4a, Example 2a), and for processed oat milk produced in Comparative Example 14 and Example 4 by changing the amount of Cellulase 2 added to 14 U / g-fiber (Comparative Example 14a, Example 4a). The results, as shown in Table 7, showed that compared to processed oat milk treated with protein-glutaminase (PG) alone (Comparative Example 2), the processed oat milks treated with either PG or Cellulase 1 or 2 (Examples 2a and 4a) showed improved protein digestibility and enhanced efficiency of essential amino acid release through digestion. In particular, the processed oat milk treated with PG and Cellulase 2 (Example 4a) showed further enhanced protein digestibility and essential amino acid release efficiency.

[0137] [Test Example 5] (1) Manufacturing of processed oat milk In the "(1) Production of Processed Oat Milk" of Test Example 1, the oat milk used in the production of oat milk was α -Amylase addition was set at 0.01U / g-starch. α The amylase treatment temperature was set to 60°C, the enzymes added to the oat milk were as shown in Table 9, and the enzyme treatment conditions were set to 50°C for 4 hours to prepare processed oat milk (Comparative Examples 9 to 11, Examples 6, 7, and 10 to 12).

[0138] (2) Experiment on processing oat milk (2-1) Mineral quality determination The processed oat milk was centrifuged (15,000 rpm, 5 minutes) to obtain a supernatant, and the amounts of calcium, iron, zinc, and magnesium in the supernatant were measured using the following mineral assay kit.

[0139] [Table 8]

[0140] (2-2) Results [Table 9]

[0141] As shown in Table 9, the processed oat milk obtained by combining protein glutaminase (PG) with cellulase 1 and / or cellulase 2 (Examples 7, 10-12, and 6) showed increased levels of soluble minerals, including calcium, compared to processed oat milk obtained by treating PG alone (Comparative Example 9). While the levels of soluble minerals were minimal when treating cellulase 1 or 2 alone, they were significantly increased when combined with PG. Furthermore, a comparison of the soluble mineral enhancement effect of combining PG and cellulase 1 (Example 7) relative to cellulase 1 alone (Comparative Example 11) with the soluble mineral enhancement effect of combining PG and cellulase 2 (Example 6) relative to cellulase 2 alone (Comparative Example 10) shows that the soluble mineral enhancement effect (per unit activity) achieved by combining PG with cellulase 2 was superior to that of PG. Calcium and iron were further enhanced by combining cellulase 1 and cellulase 2 at a predetermined ratio. When all minerals are comprehensively evaluated, Example 10 has the best effect of increasing the amount of soluble minerals.

Claims

1. A method for producing a processed liquid composition containing vegetable protein, characterized in that: The method comprises the following steps: treating a liquid composition containing vegetable protein with protein deamidase and cellulase.

2. The method for producing a processed liquid composition containing vegetable protein according to claim 1, wherein: The plant protein is cereal protein.

3. The method for producing a processed liquid composition containing vegetable protein according to claim 1, wherein: The vegetable protein is oat protein.

4. The manufacturing method according to claim 1, wherein The cellulase is a cellulase derived from Aspergillus and / or a cellulase derived from Trichoderma.

5. The manufacturing method according to claim 1, wherein Among the cellulases, the activity of exoglucanase is higher than that of endoglucanase.

6. The method for producing a processed liquid composition containing vegetable protein according to claim 1, wherein: 2 to 5000 U of cellulase are used per 1 U of protein deamidase.

7. The method for producing a processed liquid composition containing vegetable protein according to claim 1, wherein: 0.01 to 200 U of the protein deamidase is used per 1 g of the vegetable protein.

8. A method for improving protein solubility, digestibility and / or mineral solubility of a liquid composition containing vegetable protein, characterized in that: The method comprises the following steps: treating a liquid composition containing vegetable protein with protein deamidase and cellulase.

9. An agent for improving protein solubility, digestibility and / or mineral solubility of a liquid composition containing vegetable protein, characterized in that: Contains protein deamidase and cellulase.

10. The agent for improving protein solubility, digestibility and / or mineral solubility according to claim 9, wherein Contains 2 to 5000 U of cellulase per 1 U of protein deamidase.

11. An agent for improving protein solubility, digestibility and / or mineral solubility of a liquid composition containing vegetable protein treated with protein deamidase, characterized in that: Contains cellulase.

12. A food or beverage containing plant protein, characterized in that: A liquid composition containing vegetable protein obtained by the production method according to claim 1.

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