Classifications

• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
  • A23J3/00—Working-up of proteins for foodstuffs
  • A23J3/14—Vegetable proteins
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
  • A23J3/00—Working-up of proteins for foodstuffs
  • A23J3/30—Working-up of proteins for foodstuffs by hydrolysis
  • A23J3/32—Working-up of proteins for foodstuffs by hydrolysis using chemical agents
  • A23J3/34—Working-up of proteins for foodstuffs by hydrolysis using chemical agents using enzymes
  • A23J3/346—Working-up of proteins for foodstuffs by hydrolysis using chemical agents using enzymes of vegetable proteins
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
  • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
  • C12N9/14—Hydrolases (3)
  • C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
  • C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
  • C12N9/52—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from bacteria or Archaea
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
  • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
  • C12N9/14—Hydrolases (3)
  • C12N9/78—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5)
  • C12N9/80—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5) acting on amide bonds in linear amides (3.5.1)
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
  • C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
  • C12P1/04—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using bacteria
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
  • C12P13/00—Preparation of nitrogen-containing organic compounds
  • C12P13/04—Alpha- or beta- amino acids
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12Y—ENZYMES
  • C12Y305/00—Hydrolases acting on carbon-nitrogen bonds, other than peptide bonds (3.5)
  • C12Y305/01—Hydrolases acting on carbon-nitrogen bonds, other than peptide bonds (3.5) in linear amides (3.5.1)
  • C12Y305/01044—Protein-glutamine glutaminase (3.5.1.44)
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
  • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
  • A23L33/17—Amino acids, peptides or proteins
  • A23L33/175—Amino acids

Definitions

• the present invention relates to a processing technique for modifying a protein-containing composition.
• Protein is one of the important nutrients that makes up the body.
• the efficiency of protein intake is synonymous with the efficiency of digestion into amino acids. In other words, efficient digestion is essential to effectively take in protein into the body.
• Non-Patent Document 1 Plant proteins are less digestible than animal proteins (Non-Patent Document 1), and the gap between food intake and nutrient absorption is also an issue.
• modifications include not only improving the digestibility of vegetable protein materials, but also improving the properties of protein materials, including animal protein materials, such as foaming ability, foam stability, and emulsifying properties.
• the present invention aims to provide a technique for producing a modified protein-containing composition.
• vegetable proteins are less digestible than animal proteins, and therefore a first object of the present invention is to provide a technique for producing a vegetable protein-containing composition with an improved amount of free essential amino acids.
• a second object of the present invention is to provide a technique for producing a protein-containing composition with improved foaming properties, foam stability, or emulsifying properties.
• treating a vegetable protein material with protein deamidase and a protease derived from the genus Chryseobacterium improves the amount of free essential amino acids in the resulting vegetable protein-containing food or drink, and that treating a protein material with protein deamidase and a protease derived from the genus Chryseobacterium improves the foaming properties, foam stability, or emulsifying properties of the resulting protein material.
• the present invention was completed based on this knowledge and through further research.
• the present invention provides the following aspects of the invention:
• a modifier for a protein-containing composition comprising protein deamidase and a protease derived from the genus Chryseobacterium, wherein the modification of the protein-containing composition is to improve the amount of free essential amino acids of a plant protein-containing composition, the foaming property of the protein-containing composition, the foam stability of the protein-containing composition, and/or the emulsifiability of the protein-containing composition.
• a combination of protein deamidase and a protease derived from the genus Chryseobacterium for modifying a protein-containing composition, wherein the modification of the protein-containing composition is to improve the amount of essential amino acid release from a plant protein-containing composition, the foaming property of the protein-containing composition, the foam stability of the protein-containing composition, and/or the emulsifying property of the protein-containing composition.
• Item 1 An agent for enhancing the release amount of essential amino acids from a vegetable protein-containing composition, comprising protein deamidase and a protease derived from the genus Chryseobacterium.
• the agent for enhancing an essential amino acid release amount in a vegetable protein-containing composition according to Item 1 wherein the protein deamidase is protein glutaminase.
• Item 3 The agent for enhancing the release amount of essential amino acids in a vegetable protein-containing composition according to Items 1 and 2, wherein the vegetable protein is a protein of a plant selected from the group consisting of pulses, cereals, and nuts and seeds.
• Item 4. A modifier for a protein-containing composition, comprising protein deamidase and a protease derived from the genus Chryseobacterium, wherein the modification is an improvement in foaming property, foam stability, and/or emulsifying property.
• (B) A method for modifying a protein-containing composition, comprising the step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on the protein-containing composition, wherein the modification of the protein-containing composition is to improve the amount of free essential amino acids of a plant protein-containing composition, the foaming property of the protein-containing composition, the foam stability of the protein-containing composition, and/or the emulsifiability of the protein-containing composition.
• the modification of the protein-containing composition is to improve the amount of free essential amino acids of a plant protein-containing composition, the foaming property of the protein-containing composition, the foam stability of the protein-containing composition, and/or the emulsifiability of the protein-containing composition.
• a method for increasing the amount of free essential amino acids in a vegetable protein-containing composition comprising the step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on the vegetable protein-containing composition.
• Item 6 A method for modifying a protein-containing composition, comprising a step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on the protein-containing composition, wherein the modification is to improve foaming property, foam stability, and/or emulsifying property.
• a method for producing a processed protein-containing composition comprising a step of reacting a protein-containing composition with protein deamidase and a protease derived from the genus Chryseobacterium, wherein the processed protein-containing composition is either a processed vegetable protein-containing composition having an improved amount of essential amino acid release, or a processed protein-containing composition having improved foaming property, foam stability, and/or emulsifying property.
• the processed protein-containing composition is either a processed vegetable protein-containing composition having an improved amount of essential amino acid release, or a processed protein-containing composition having improved foaming property, foam stability, and/or emulsifying property.
• Item 7 A method for producing a processed vegetable protein-containing composition having an improved amount of free essential amino acids, comprising the step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on a vegetable protein-containing composition.
• a method for producing a processed protein-containing composition having improved foaming properties, foam stability, and/or emulsifying properties comprising a step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on the protein-containing composition.
• a processed protein-containing composition obtained by a method comprising the step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on a protein-containing composition.
• Item 9 A processed vegetable protein-containing composition having an improved amount of released essential amino acids, obtained by a method comprising the step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on a vegetable protein-containing composition.
• a processed protein-containing composition having improved foaming properties, foam stability, and/or emulsifying properties obtained by a method comprising a step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to act on a protein-containing composition.
• the present invention provides a technique for producing a vegetable protein-containing composition with an improved amount of free essential amino acids, or a protein-containing composition with improved foaming properties, foam stability, or emulsifying properties.
• Modifier for protein-containing composition the agent for improving the amount of free essential amino acids in a vegetable protein-containing composition (hereinafter also referred to as the "first modifier") is characterized by containing protein deamidase and a protease derived from the genus Chryseobacterium.
• the modifier used for improving the foaming property, foam stability, and/or emulsifying property of the protein-containing composition (hereinafter also referred to as the "second modifier”) is also characterized by containing protein deamidase and a protease derived from the genus Chryseobacterium.
• the first modifier and the second modifier are collectively referred to as the "modifiers”.
• the agent for improving the free amount of essential amino acids in a vegetable protein-containing composition which is the first modifier of the present invention, is used for improving the free amount of essential amino acids contained in a vegetable protein-containing composition.
• the first modifier of the present invention is specifically used for increasing the amount of essential amino acids more than the amount of non-essential amino acids among the released amino acids.
• the improvement in the amount of released essential amino acids can be confirmed by the fact that the ratio of the amount of essential amino acids to the total amount of free amino acids is increased compared to when a combination of protein deamidase and a protease other than that derived from the genus Chryseobacterium is not used (i.e., when a vegetable protein-containing composition is treated under the same conditions except that a combination of protein deamidase and a protease other than that derived from the genus Chryseobacterium is not used), particularly when protein deamidase and a protease other than that derived from the genus Chryseobacterium are used (i.e., when a vegetable protein-containing composition is treated under the same conditions except that a combination of protein deamidase and a protease other than that derived from the genus Chr
• the second modifier of the present invention is used to improve the foaming properties, foam stability, and/or emulsifying properties of a protein-containing composition.
• Improved foaming properties can be confirmed by an increase in the total volume, including the foam, of the protein-containing composition foamed by homogenization compared to when the combination of protein deamidase and a protease other than that derived from the genus Chryseobacterium is not used (i.e., when the protein-containing composition is treated under the same conditions except that the combination of protein deamidase and a protease other than that derived from the genus Chryseobacterium is not used); improved foam stability can be confirmed by a suppressed degree of decrease over time in the total volume of the foamed protein-containing composition compared to the above case; improved emulsifying properties can be confirmed by an increase in the turbidity of the emulsion composition prepared using the protein-containing composition and an oily base compared to the above case.
• Protein deamidase is an active ingredient in the modifying agent of the present invention together with the protease derived from the genus Chryseobacterium. That is, protein deamidase is an active ingredient that contributes to the improvement of the essential amino acid release amount in the agent for improving the essential amino acid release amount of a vegetable protein-containing composition according to the first modifying agent of the present invention, and is an active ingredient that contributes to the improvement of foaming property, foam stability, and/or emulsifying property in the second modifying agent of the present invention.
• protein deamidating enzyme there are no particular limitations on the type or origin of the protein deamidating enzyme, so long as it is an enzyme that degrades amide group-containing side chains of proteins without cleaving peptide bonds or cross-linking proteins.
• protein deamidating enzymes include protein deamidating enzymes derived from the genus Chryseobacterium, Flavobacterium, Empedobacter, Sphingobacterium, Aureobacterium, or Myroides, as disclosed in JP 2000-50887 A, JP 2001-218590 A, and WO 2006/075772. These protein deamidating enzymes may be used alone or in combination.
• protein deamidating enzymes examples include protein glutaminase and protein asparaginase, and in a broader sense also include protein arginine deiminase.
• protein glutaminase is preferred from the viewpoint of further increasing the amount of essential amino acid release, foaming properties, foam stability, and/or emulsifying properties.
• protein deamidating enzymes from the viewpoint of further increasing the amount of essential amino acid release, foaming properties, foam stability, and/or emulsifying properties, more preferred are protein deamidating enzymes derived from the genus Chryseobacterium, even more preferred are protein glutaminases derived from the genus Chryseobacterium, and even more preferred are protein glutaminases derived from the species Chryseobacterium proteolyticum.
• Protein deamidase can be prepared from the culture medium of the microorganism from which the protein deamidase is derived. Specific preparation methods include a method of recovering protein deamidase from the culture medium or cells of the microorganism. For example, when a protein deamidase-secreting microorganism is used, the cells can be recovered from the culture medium in advance by filtration, centrifugation, or the like as necessary, and the enzyme can be separated and/or purified. When a protein deamidase-nonsecreting microorganism is used, the cells can be recovered from the culture medium in advance by pressure treatment, ultrasonic treatment, or the like to expose the enzyme, and the enzyme can be separated and/or purified.
• a known protein separation and/or purification method can be used without any particular limitation, and examples of the method include centrifugation, UF concentration, salting out, various chromatography methods using ion exchange resins, etc.
• the separated and/or purified enzyme can be powdered by a drying method such as freeze-drying or vacuum drying, and can also be powdered using an appropriate excipient and/or drying aid in the drying method.
• the separated and/or purified enzyme can also be liquefied by adding appropriate additives and sterilizing by filtration. Commercially available protein deamidating enzymes can also be used.
• Chryseobacterium-derived protease is an active ingredient in the modifying agent of the present invention together with protein deamidase. That is, Chryseobacterium-derived protease is an active ingredient that contributes to the improvement of the essential amino acid release amount in the essential amino acid release amount enhancer for a vegetable protein-containing composition according to the first modifying agent of the present invention, and is an active ingredient that contributes to the improvement of foaming property, foam stability, and/or emulsifying property in the second modifying agent of the present invention.
• the protease derived from the genus Chryseobacterium is not particularly limited as long as it is an enzyme derived from the genus Chryseobacterium that hydrolyzes peptide bonds in proteins.
• proteases derived from the genus Chryseobacterium include Chryseobacterium nematophagum, Chryseobacterium cucumeris, Chryseobacterium lactis, Chryseobacterium rhizoplane, and Chryseobacterium nematophagum.
• the proteases include those derived from Seobacterium joostei, Chryseobacterium shigense, Chryseobacterium proteolyticum, Chryseobacterium gleum, and Chryseobacterium soil. These Chryseobacterium proteases may be used alone or in combination.
• proteases derived from Chryseobacterium proteolyticum species are preferred from the viewpoint of further enhancing the effect of improving the amount of essential amino acid release, foaming properties, foam stability, and/or emulsifying properties.
• the protease derived from the genus Chryseobacterium can be prepared by known methods. For example, it can be easily prepared by culturing bacteria of the genus Chryseobacterium and isolating the protease using known means, or by using genetic recombination technology.
• the modifier of the present invention may be an enzyme composition containing protein deamidase and a protease derived from the genus Chryseobacterium, and may or may not contain additives and/or bases acceptable for the formulation of enzyme preparations as components other than the enzyme.
• additives and bases include excipients, buffers, antioxidants, UV protection agents, preservatives, antiseptics, pH adjusters, dispersants, emulsifiers, solubilizers, carriers, solvents (water, etc.), and the like.
• additives and bases may be used alone or in combination of two or more. The content of these additives and bases may be appropriately set depending on the types of the components and/or the formulation form, etc.
• properties of the modifier for the protein-containing composition of the present invention are not particularly limited, and examples thereof include dry preparations in the form of powder, fine particles, or granules, and liquid preparations.
• Method for modifying a protein-containing composition As described above, the combination of protein deamidase and Chryseobacterium protease contributes to improving the amount of essential amino acid release from a vegetable protein-containing composition, and also contributes to improving the foaming property, foam stability, and/or emulsifying property of the protein-containing composition.
• the present invention further provides a method for improving the amount of essential amino acid release from a vegetable protein-containing composition, comprising the step of acting protein deamidase and Chryseobacterium protease on the vegetable protein-containing composition (hereinafter also referred to as the "first modification method"), and a method for modifying a protein-containing composition, comprising the step of acting protein deamidase and Chryseobacterium protease on the protein-containing composition, in which the modification is to improve the foaming property, foam stability, and/or emulsifying property (hereinafter also referred to as the "second modification method").
• the first modification method and the second modification method are collectively referred to as the "modification method”.
• the "improvement of the amount of free essential amino acids,” “protein deamidase,” and “protease derived from the genus Chryseobacterium” are as described in detail above in “1.
• Modifiers for protein-containing compositions In addition, in the second modification method of the present invention, the "improvement of foaming properties, foam stability, and/or emulsifying properties,” “protein deamidase,” and “protease derived from the genus Chryseobacterium” are as described in detail above in “1. Modifiers for protein-containing compositions.”
• Step of acting protein deamidase and Chryseobacterium genus-derived protease on a protein-containing composition in the first modification method, in the step of acting protein deamidase and Chryseobacterium genus-derived protease on a vegetable protein-containing composition, a vegetable protein mixture containing the vegetable protein-containing composition, protein deamidase and Chryseobacterium genus-derived protease is appropriately prepared, and a treatment is performed to advance the enzyme reaction.
• the second modification method in the step of acting protein deamidase and Chryseobacterium genus-derived protease on a protein-containing composition, a protein mixture containing the protein-containing composition, protein deamidase and Chryseobacterium genus-derived protease is appropriately prepared, and a treatment is performed to advance the enzyme reaction.
• the protein includes vegetable protein and animal protein.
• the vegetable protein-containing composition is an example of an enzyme treatment target in the first modification method and an application target of the first modifying agent.
• the vegetable protein-containing composition is an example of an enzyme treatment target in the second modification method and an application target of the second modifying agent.
• the vegetable protein-containing composition is not particularly limited, as long as it contains vegetable protein and is in a form that can be ingested by the living body.
• the vegetable protein-containing composition used in the present invention is a composition for consumption or drinking, containing vegetable protein and water.
• the vegetable protein-containing composition includes fluidity such as liquid, slurry, and paste (hereinafter, these fluidity-containing vegetable protein-containing compositions are collectively referred to as "liquid, etc.”), and solid.
• the vegetable protein-containing composition may be fluid or solid.
• the vegetable protein-containing composition is fluid.
• vegetable protein-containing compositions prepared from unstructured protein materials include (i) liquids obtained by dispersing a dry powder of a vegetable protein material (specifically, at least one of a plant organ of a plant from which the vegetable protein is derived and a material obtained by removing at least a portion of components other than protein from the plant organ to increase the protein content; the same applies below) in water; and (ii) liquids obtained by crushing and dispersing a vegetable protein material in water and, if necessary, removing insoluble matter derived from the skins of the plant material, etc., by any means such as centrifugation, filtration, a filter bag, or a sieve.
• a liquid obtained by removing components other than vegetable protein from the liquid (i) or (ii) above to increase the protein content (iv) a liquid obtained by mixing a dried powder prepared from any of the liquids (i) to (iii) above with water; and, preferably when cereals are used as the vegetable protein material, (v) a liquid obtained by further treating any of the liquids (i) to (iv) above with amylase, and (vi) a structured vegetable protein material swollen with water is an example of a material prepared from a structured protein material.
• the textured vegetable protein material used in the preparation of the specific example of (vi) above is a food material generally known as a meat substitute (imitation meat).
• a typical example of a textured vegetable protein material is a material that is made by extruding a raw material mixture containing vegetable protein and water with an extruder or the like, and then drying or freezing it to create a meat-like texture.
• the "meat” that the textured vegetable protein material mimics refers to the muscle of an animal that is consumed, and the term “meat” is used to include not only the muscle of mammals and birds, but also the flesh of seafood.
• a preferred example of a liquid vegetable protein-containing composition is so-called vegetable milk substitute (also called vegetable milk).
• the "content of plant protein material" in a plant protein-containing composition refers to the proportion of the dry weight of the components constituting the plant organ contained in the plant protein-containing composition in the case of a plant protein-containing composition prepared using a non-structured plant protein material, and refers to the proportion of the dry weight of the structured plant protein material in the case of a plant protein-containing composition prepared using a structured plant protein material.
• the plant protein-containing composition is a liquid etc. composed only of components derived from a plant organ and water, as in the specific examples (i) to (v) above, the “content of plant protein material” refers to the proportion of the dry weight of the liquid etc.
• the plant protein-containing composition is a liquid etc.
• the “content of plant protein material” refers to the proportion of the dry weight of the liquid etc. excluding the added component. Furthermore, when the vegetable protein-containing composition is a swollen product composed only of a textured vegetable protein material and water, as in the specific example of (vi) above, the “content of vegetable protein material” refers to the percentage of the dry weight of the composition. When the vegetable protein-containing composition is a swollen product containing the specific example of (vi) above and an added ingredient, the “content of vegetable protein material” refers to the percentage of the dry weight of the swollen product excluding the added ingredient.
• the vegetable protein content contained in the vegetable protein material is not particularly limited, but may be, for example, 5% by weight or more, 10% by weight or more, 13% by weight or more, 15% by weight or more, 20% by weight or more, 25% by weight or more, or 30% by weight or more. From the viewpoint of further enhancing the effect of improving the amount of free essential amino acids, the content is preferably 35% by weight or more, more preferably 40% by weight or more, and even more preferably 45% by weight or more.
• the upper limit of the content range is not particularly limited, but may be, for example, 90% by weight or less, preferably 85% by weight or less, 80% by weight or less, more preferably 70% by weight or less, even more preferably 60% by weight or less, and even more preferably 55% by weight or less.
• Specific ranges of the vegetable protein content in the vegetable protein material include, for example, 5 to 90% by weight, preferably 10 to 85% by weight; when the vegetable protein is derived from pulses, for example, 5 to 90% by weight, preferably 30 to 85% by weight, more preferably 40 to 85% by weight, 45 to 85% by weight, or 50 to 80% by weight; when the vegetable protein is derived from cereals, for example, 5 to 90% by weight, preferably 10 to 85% by weight, 10 to 60% by weight, 10 to 40% by weight, 10 to 20% by weight, or 13 to 20% by weight.
• the content of the vegetable protein material in the vegetable protein-containing composition is not particularly limited, but may be, for example, 0.05 to 80% by weight.
• the content may be, for example, 0.05 to 80% by weight, 0.1 to 60% by weight, or 1 to 40% by weight, preferably 2 to 30% by weight, or 3 to 20% by weight, and more preferably 5 to 15% by weight, or 8 to 12% by weight.
• Vegetable Proteins There are no particular limitations on the vegetable proteins, so long as they are derived from plants. Examples of such proteins include: soybeans, peas, lentils, chickpeas, black beans, broad beans, mung beans, lupine beans, kidney beans, and other cereals; wheat, barley, oats, sorghum, rice, rye, buckwheat, barnyard millet, millet, teff, quinoa, corn, potatoes, and other cereals; almonds, coconuts, peanuts, cashew nuts, hazelnuts, pecan nuts, macadamia nuts, pistachios, walnuts, Brazil nuts, pili nuts, chestnuts, sesame seeds, pine nuts, hemp seeds (industrial hemp), chia seeds, chia seeds, amaranth, canary seeds, linseeds, and other nuts and seeds; and natural proteins contained in algae.
• the vegetable protein of the present invention may be any protein derived from a plant, and may be, in addition to the above-mentioned natural proteins, proteins obtained by chemically partially hydrolyzing the above-mentioned natural proteins with acid, alkali, etc., proteins obtained by enzymatically partially hydrolyzing the above-mentioned natural proteins with proteases, etc., proteins chemically modified with various reagents, or proteins obtained by artificial peptide synthesis.
• the above-mentioned vegetable proteins may be used alone or in combination of two or more.
• preferred are cereal or grass proteins more preferred are soybean, pea, lentil, chickpea, black bean, broad bean, mung bean, lupin bean, kidney bean, and oat proteins, and even more preferred are soybean and pea proteins.
• preferred are cereal proteins more preferred are oat proteins.
• the content of the vegetable protein in the vegetable protein-containing composition is not particularly limited, but may be, for example, 0.01 to 50% by weight.
• the content of the vegetable protein in the vegetable protein-containing composition may be, for example, 0.01 to 50% by weight
• the content of the vegetable protein in the vegetable protein-containing composition may be, for example, 0.01 to 50% by weight
• the content may be, for example, 0.01 to 50% by weight, 0.1 to 40% by weight, or 0.5 to 30% by weight, preferably 1 to 25% by weight, or 2 to 20% by weight, more preferably 2.5 to 15% by weight, or 3 to 10% by weight.
• the content of the vegetable protein in the vegetable protein-containing composition may be, for example, 0.01 to 50% by weight, preferably 0.1 to 40% by weight, more preferably 0.5 to 30% by weight, even more preferably 1 to 25% by weight, 1 to 20% by weight, 1 to 15% by weight, 1 to 10% by weight, or 1 to 5% by weight.
• the vegetable protein-containing composition may or may not contain any other components in addition to the vegetable protein.
• Other components include components contained in the plant from which the vegetable protein is derived (carbohydrates, lipids, etc.) and added components (other food ingredients, food additives, etc.).
• Food additives include thickeners, binders, seasonings, pH adjusters, buffers, colorants, flavors, etc.
• the animal protein-containing composition is an example of an object to be treated with the enzyme in the second modification method, and is an example of an object to which the second modifying agent is applied. There are no particular limitations on the animal protein-containing composition, as long as it is in a form that can be ingested by the living body.
• the animal protein-containing composition used in the present invention contains animal protein and has fluidity such as liquid, slurry, and paste.
• animal protein-containing compositions include animal milk, liquids obtained by dispersing dried animal milk in water, and liquids obtained by dispersing crudely refined or purified protein from animal milk in water.
• the "content of animal protein material" in the animal protein-containing composition refers to the proportion of the dry weight of the animal protein-containing composition (excluding the weight of added ingredients).
• the content of the animal protein material in the animal protein-containing composition is not particularly limited, but may be, for example, 0.05 to 40% by weight, preferably 0.1 to 30% by weight, or 1 to 10% by weight, preferably 2 to 7% by weight.
• animal Protein There are no particular limitations on the animal protein, so long as it is derived from an animal, but milk protein is preferred, whey and casein are more preferred, and whey is even more preferred.
• the content of animal protein in the animal protein-containing composition is not particularly limited, but may be, for example, 0.03 to 30% by weight, preferably 0.08 to 20% by weight, or 0.8 to 8% by weight, preferably 1 to 6% by weight.
• the animal protein-containing composition may or may not contain any other components in addition to the animal protein.
• Other components include components contained in the material from which the animal protein is derived (carbohydrates, lipids, etc.) and added components (other food ingredients, food additives, etc.).
• Food additives include thickeners, binders, seasonings, pH adjusters, buffers, colorants, flavors, etc.
• the amount of protein deamidase used is not particularly limited, but in the first modification method, the amount used per 1 g of vegetable protein material contained in the vegetable protein-containing composition is, for example, 0.01 U or more, or 0.1 U or more, and from the viewpoint of further enhancing the effect of improving the amount of free essential amino acids, preferably 1 U or more, or 5 U or more, more preferably 7.5 U or more, even more preferably 10 U or more, and even more preferably 13 U or more.
• the upper limit of the above-mentioned range of the amount of protein deamidase used per 1 g of vegetable protein material is not particularly limited, but for example, 400 U or less, 300 U or less, 200 U or less, 150 U or less, 100 U or less, or 75 U or less, preferably 60 U or less, or 50 U or less, more preferably 40 U or less, or 30 U or less, and even more preferably 25 U or less, 20 U or less, or 17 U or less.
• the amount of protein deamidase used per 1 g of vegetable protein material contained in the protein-containing composition may be, for example, 0.01 U or more, or 0.1 U or more, and from the viewpoint of further enhancing the effects of improving foamability, foam stability, and/or emulsifiability, preferably 1 U or more.
• the upper limit of the range of the amount of protein deamidase used per 1 g of vegetable protein material is not particularly limited, and may be, for example, 100 U or less, 50 U or less, or 20 U or less, preferably 10 U or less, more preferably 5 U or less, and even more preferably 3 U or less.
• the amount of protein deamidase used per 1 g of the animal protein material contained in the protein-containing composition may be, for example, 0.1 U or more, or 1 U or more, and from the viewpoint of further enhancing the effects of improving foamability, foam stability, and/or emulsifiability, preferably 3 U or more, more preferably 5 U or more, and even more preferably 7 U or more or 8 U or more.
• the upper limit of the range of the amount of protein deamidase used per 1 g of the animal protein material is not particularly limited, and may be, for example, 400 U or less, 200 U or less, 100 U or less, 50 U or less, or 30 U or less, preferably 20 U or less, more preferably 15 U or less, and even more preferably 12 U or less.
• the amount of protein deamidase used per 1 g of vegetable protein is, for example, 0.01 U or more, or 0.1 U or more, and from the viewpoint of further enhancing the effect of improving the amount of free essential amino acids, preferably 1 U or more, or 5 U or more, more preferably 7.5 U or more, or 10 U or more, even more preferably 13 U or more, or 16 U or more, and even more preferably 18 U or more, 20 U or more, or 22 U or more.
• the upper limit of the above range of the amount of protein deamidase used per 1 g of vegetable protein is not particularly limited, and examples thereof include 500 U or less, 400 U or less, 300 U or less, 200 U or less, 150 U or less, 100 U or less, or 80 U or less, preferably 70 U or less, or 60 U or less, more preferably 50 U or less, or 45 U or less, and even more preferably 40 U or less, 35 U or less, or 32 U or less.
• specific ranges of the amount of protein deamidating enzyme used per gram of vegetable protein include, for example, 0.01 to 500 U, 0.01 to 400 U, or 0.1 to 300 U, preferably 1 to 200 U, 5 to 150 U, or 5 to 100 U, more preferably 7.5 to 80 U, or 10 to 70 U, even more preferably 13 to 60 U, 16 to 50 U, 18 to 45 U, or 20 to 45 U, and even more preferably 22 to 40 U, 22 to 35 U, or 22 to 32 U.
• the amount of protein deamidase used per 1 g of vegetable protein is, for example, 0.01 U or more, or 0.1 U or more, and from the viewpoint of further enhancing the effect of improving foamability, foam stability, and/or emulsifiability, preferably 1 U or more or 3 U or more, more preferably 5 U or more or 7 U or more, and even more preferably 9 U or more.
• the upper limit of the above-mentioned range of the amount of protein deamidase used per 1 g of vegetable protein is not particularly limited, and examples thereof include 100 U or less, 50 U or less, or 20 U or less, and preferably 15 U or less.
• the specific range of the amount of protein deamidase used per 1 g of vegetable protein is, for example, 0.01 to 100 U, or 0.1 to 50 U, 1 to 20 U, 3 to 20 U, more preferably 5 to 15 U, or 7 to 15 U, and even more preferably 9 to 15 U.
• the amount of protein deamidase used per 1 g of animal protein is, for example, 0.01 U or more, or 0.1 U or more, and from the viewpoint of further enhancing the effect of improving foamability, foam stability, and/or emulsifiability, preferably 1 U or more or 3 U or more, more preferably 5 U or more or 7 U or more, and even more preferably 9 U or more.
• the upper limit of the range of the amount of protein deamidase used per 1 g of animal protein is not particularly limited, and examples thereof include 100 U or less, 50 U or less, or 20 U or less, and preferably 15 U or less.
• the specific range of the amount of protein deamidase used per 1 g of animal protein is, for example, 0.01 to 100 U, or 0.1 to 50 U, 1 to 20 U, 3 to 20 U, more preferably 5 to 15 U, or 7 to 15 U, and even more preferably 9 to 15 U.
• the amount of enzyme that liberates 1 ⁇ mol of ammonia per minute using benzyloxycarbonyl-L-glutaminylglycine (Z-Gln-Gly) as a substrate is defined as 1 unit (1 U).
• the amount of Chryseobacterium-derived protease used is not particularly limited, but in the first modification method, the amount used per 1 g of the vegetable protein material contained in the vegetable protein-containing composition is, for example, 0.05 U or more, and from the viewpoint of further enhancing the effect of improving the amount of free essential amino acids, the amount is preferably 0.1 U or more, or 1 U or more, more preferably 5 U or more, or 7.5 U or more, even more preferably 10 U or more, or 12 U or more, and even more preferably 15 U or more, or 18 U or more.
• the upper limit of the above-mentioned range of the amount of Chryseobacterium-derived protease used per 1 g of vegetable protein material is not particularly limited, but may be, for example, 400 U or less, 300 U or less, 200 U or less, 150 U or less, 100 U or less, or 90 U or less, preferably 80 U or less, 70 U or less, or 60 U or less, more preferably 50 U or less, or 40 U or less, and even more preferably 30 U or less, 25 U or less, or 22 U or less.
• the amount of Chryseobacterium-derived protease used per 1 g of vegetable protein material contained in the protein-containing composition is, for example, 0.1 U or more or 1 U or more, and from the viewpoint of further enhancing the effect of improving foamability, foam stability, and/or emulsification, preferably 2 U or more or 2.5 U or more, more preferably 5 U or more, 5.5 U or more, 6 U or more, 7 U or more, or 8 U or more, and even more preferably 10 U or more or 12 U or more.
• the upper limit of the above-mentioned range of the amount of Chryseobacterium-derived protease used per 1 g of vegetable protein material is not particularly limited, and examples thereof include 400 U or less, 200 U or less, 100 U or less, 50 U or less, 30 U or less, or 20 U or less.
• the amount of Chryseobacterium-derived protease used per 1 g of animal protein material contained in the protein-containing composition is, for example, 0.005 U or more, and from the viewpoint of further enhancing the effect of improving foamability, foam stability, and/or emulsification, preferably 0.05 U or more, 0.5 U or more, or 1 U or more, more preferably 5 U or more or 10 U or more, even more preferably 15 U or more or 20 U or more, even more preferably 35 U or more, 40 U or more, or 50 U or more, and even more preferably 70 U or more, 80 U or more, or 85 U or more.
• the upper limit of the above-mentioned range of the amount of Chryseobacterium-derived protease used per 1 g of animal protein material is not particularly limited, but examples include 400 U or less, 200 U or less, or 100 U or less.
• the amount of Chryseobacterium-derived protease used per 1 g of plant protein can be, for example, 0.05 U or more, and from the viewpoint of further enhancing the effect of improving the amount of free essential amino acids, the amount can be preferably 0.1 U or more, or 1 U or more, more preferably 5 U or more, or 7.5 U or more, even more preferably 10 U or more, 12.5 U or more, 15 U or more, or 20 U or more, and even more preferably 22 U or more, or 24 U or more.
• the upper limit of the range of the amount of Chryseobacterium-derived protease used per gram of plant protein is not particularly limited, but may be, for example, 1000 U or less, 500 U or less, 400 U or less, 300 U or less, 200 U or less, 150 U or less, 100 U or less, or 90 U or less, preferably 80 U or less, 70 U or less, or 60 U or less, more preferably 55 U or less, or 50 U or less, and even more preferably 45 U or less, or 40 U or less.
• specific ranges of the amount of Chryseobacterium-derived protease used per 1 g of vegetable protein include, for example, 0.05 to 1000 U, 0.05 to 500 U, or 0.05 to 400 U, preferably 0.1 to 300 U, 0.1 to 200 U, or 1 to 150 U, more preferably 5 to 100 U, or 7.5 to 90 U, even more preferably 10 to 80 U, 12.5 to 70 U, 15 to 60 U, or 20 to 55 U, and even more preferably 22 to 50 U, 22 to 45 U, or 24 to 40 U.
• the amount of Chryseobacterium-derived protease used per 1 g of vegetable protein contained in the protein-containing composition is, for example, 1 U or more or 10 U or more, and from the viewpoint of further enhancing the effects of improving foamability, foam stability, and/or emulsification, preferably 15 U or more or 20 U or more, more preferably 40 U or more, 50 U or more, 60 U or more, 70 U or more, and even more preferably 80 U or more, 90 U or more, or 100 U or more.
• the upper limit of the above-mentioned range of the amount of Chryseobacterium-derived protease used per 1 g of vegetable protein is not particularly limited, but examples thereof include 400 U or less, 300 U or less, 200 U or less, or 150 U or less.
• specific ranges of the amount of Chryseobacterium-derived protease used per gram of plant protein include, for example, 1 to 400 U, 10 to 400 U, preferably 15 to 400 U or 20 to 400 U, more preferably 40 to 400 U, 50 to 400 U, 60 to 400 U, 70 to 400 U, and even more preferably 80 to 400 U, 90 to 400 U, 90 to 300 U, 90 to 200 U, or 90 to 150 U.
• the amount of Chryseobacterium protease used may be, for example, 0.1 U or more or 1 U or more per gram of animal protein contained in the protein-containing composition; from the viewpoint of further enhancing the effect of improving foamability, foam stability, and/or emulsifying properties, the amount is preferably 7 U or more, 10 U or more, or 13 U or more, more preferably 17 U or more or 20 U or more; from the viewpoint of further enhancing the effect of improving foamability and/or emulsifying properties, the amount is even more preferably 30 U or more, 40 U or more, or 50 U or more; and from the viewpoint of further enhancing the effect of improving emulsifying properties, the amount is even more preferably 60 U or more, 80 U or more, 90 U or more, or 100 U or more.
• the upper limit of the above-mentioned range of the amount of Chryseobacterium-derived protease used per gram of animal protein is not particularly limited, but may be, for example, 400 U or less, 300 U or less, 200 U or less, or 150 U or less, and from the viewpoint of improving foam stability, preferably 100 U or less, 60 U or less, or 50 U or less, and more preferably 30 U or less.
• specific ranges of the amount of Chryseobacterium-derived protease used per 1 g of animal protein include, for example, 0.1 to 400 U, or 1 to 400 U, preferably 7 to 400 U, 10 to 400 U, 10 to 200 U, 10 to 150 U, 13 to 100 U, 17 to 60 U, 20 to 50 U, 20 to 30 U, 13 to 400 U, 17 to 400 U, 20 to 400 U, 30 to 400 U, 40 to 400 U, 50 to 400 U, 60 to 400 U, 80 to 400 U, 90 to 400 U, 90 to 300 U, 90 to 200 U, or 90 to 150 U.
• the ratio of protein deamidase to Chryseobacterium protease is determined based on the above-mentioned amounts of each enzyme used, but from the viewpoint of further enhancing the effect of improving the amount of released essential amino acids, the following ratio is preferable.
• the amount of protease derived from the genus Chryseobacterium used per 1 U of protein deamidase is preferably 0.005 U or more, more preferably 0.05 U or more or 0.1 U or more, even more preferably 0.25 U or more, 0.5 U or more, or 0.7 U or more, and even more preferably 0.8 U or more, 1 U or more, 1.1 U or more, or 1.2 U or more.
• the upper limit of the above-mentioned range of the amount of protease derived from the genus Chryseobacterium used per 1 U of protein deamidase is, for example, 20 U or less, and from the viewpoint of further enhancing the effect of improving the amount of essential amino acids released, the amount is preferably 15 U or less or 10 U or less, more preferably 8 U or less or 6 U or less, even more preferably 5 U or less or 4 U or less, and even more preferably 3 U or less, 2 U or less, or 1.5 U or less.
• the specific range of the amount of protease derived from the genus Chryseobacterium used per 1 U of protein deamidating enzyme is preferably 0.005 to 20 U, more preferably 0.05 to 15 U or 0.1 to 10 U, even more preferably 0.25 to 8 U, 0.5 to 6 U, or 0.7 to 5 U, and even more preferably 0.8 to 4 U, 1 to 3 U, 1.1 to 2 U, or 1.2 to 1.5 U.
• the amount of Chryseobacterium-derived protease used per 1 U of protein deamidating enzyme is preferably 0.005 U or more or 0.01 U or more, and from the viewpoint of further enhancing the effect of improving foamability, foam stability, and/or emulsifying properties, the amount is more preferably 0.05 U or more, 0.1 U or more, or 0.3 U or more, even more preferably 0.8 U or more, 1 U or more, or 1.3 U or more, and even more preferably 1.8 U or more or 2 U or more, and from the viewpoint of further enhancing the effect of improving foamability and/or emulsifying properties, the amount is even more preferably 2.5 U or more, 4 U or more, or 5 U or more, and from the viewpoint of further enhancing the effect of improving emulsifying properties, the amount is particularly preferably 6 U or more, 8 U or more, or 10 U or more.
• the upper limit of the above-mentioned range of usage amount per 1 U of protein deamidating enzyme derived from the genus Chryseobacterium may be, for example, 50 U or less, 30 U or less, 20 U or less, or 15 U or less, and from the viewpoint of further enhancing foam stability, preferably 10 U or less, more preferably 6 U or less, or 5 U or less, and even more preferably 3 U or less.
• the specific range of the amount of protease derived from the genus Chryseobacterium used per 1 U of protein deamidating enzyme is preferably 0.005 to 50 U, 0.01 to 50 U, more preferably 0.05 to 50 U, 0.1 to 50 U, 0.3 to 50 U, 0.8 to 50 U, 1 to 50 U, 1.3 to 50 U, 1.3 to 30 U, 1.3 to 20 U, 1.3 to 15 U, 1.3 to 10 U, 1.3 to 7 U, 1.3 to 5 U, 1.3 to 3 U, 1.8 to 50 U, 2 to 50 U, 2.5 to 50 U, 4 to 50 U, 5 to 50 U, 6 to 50 U, 8 to 50 U, 8 to 50 U, 8 to 30 U, 8 to 20 U, 8 to 15 U, or 8 to 10 U.
• Protease activity is measured by the Folin method using casein as a substrate.
• protease activity is measured by carrying out an enzyme reaction using casein as a substrate at a pH that is set according to the optimal pH of the protease being measured in a standard manner, and the amount of enzyme that causes an increase in the Folin test solution color substance equivalent to 1 ⁇ g of tyrosine per minute is defined as 1 unit (1 U).
• reaction operation and treatment conditions There is no particular limitation on the method for preparing the vegetable protein mixture in the first modification method and the protein mixture in the second modification method.
• the treatment conditions (temperature, time, pH, etc.) of these mixtures are not particularly limited as long as the effects of the present invention can be obtained.
• the treatment temperature of the vegetable protein mixture in the first modification method and the protein mixture in the second modification method may be, for example, 4 to 80°C, preferably 8 to 70°C.
• the lower limit of the temperature range may be 15°C, 30°C, or 45°C
• the upper limit of the temperature range may be 65°C, 60°C, or 55°C.
• the treatment time in the first modification method is not particularly limited, but may be, for example, 0.1 to 72 hours, preferably 12 to 36 hours.
• the treatment time in the second modification method is also not particularly limited, but may be, for example, 0.1 to 12 hours, preferably 0.5 to 6 hours, more preferably 1 to 3 hours.
• the treatment pH (25°C) of the vegetable protein mixture may be, for example, 2 to 9, preferably 3 to 8, more preferably 5 to 7.6, even more preferably 6 to 7.4, and even more preferably 6.5 to 7.2.
• processing conditions are appropriately selected depending on the optimum temperature and optimum pH of the enzyme used, and/or the degree of the desired effect of the present invention (improvement of the amount of essential amino acid release, foaming properties, foam stability, and/or emulsifiability).
• the optimal processing conditions can be determined through preliminary experiments.
• the modification method of the present invention may or may not include other steps other than the step of acting with the above-mentioned protein deamidase and Chryseobacterium protease.
• Examples of the other steps include a step of preparing a vegetable protein-containing composition used in the first modification method or a protein-containing composition (vegetable protein-containing composition or animal protein-containing composition) used in the second modification method, an enzyme inactivation step, a cooling step, a filtration step, etc. These other steps may be performed individually or in combination of two or more steps.
• the step of preparing a vegetable or animal protein-containing composition can be carried out by any method for preparing a vegetable or animal protein-containing composition.
• a plant-based milk substitute plant-based milk
• it can be prepared by any method for preparing a plant-based milk substitute depending on the type of plant from which the plant protein is derived.
• any food additive such as a seasoning, pH adjuster, buffer, colorant, fragrance, etc. may be further blended, and/or any treatment other than the treatment with protein deamidating enzyme and Chryseobacterium genus-derived protease (for example, fermentation treatment such as lactic acid fermentation) may or may not be performed.
• the present invention also provides a method for producing a processed vegetable protein-containing composition having an improved amount of free essential amino acids, which comprises the step of allowing a vegetable protein-containing composition to react with protein deamidase and a protease derived from the genus Chryseobacterium (hereinafter also referred to as "first production method"), and a method for producing a processed protein-containing composition having improved foaming properties, foam stability, and/or emulsifying properties, which comprises the step of allowing a protein deamidase and a protease derived from the genus Chryseobacterium to react with a protein-containing composition (hereinafter also referred to as "second production method").
• first production method and the second production method are collectively referred to as "production methods”.
• the "improvement of the amount of free essential amino acids", “protein deamidase”, and “protease derived from the genus Chryseobacterium” are as described in detail in "1.
• Modifier for protein-containing composition” above, and the “step of acting protein deamidase and protease derived from the genus Chryseobacterium on the vegetable protein-containing composition” is as described in detail in "2.
• Method for modifying protein-containing composition above.
• the "improvement of foaming properties”, “improvement of foam stability”, “improvement of emulsifying properties”, “protein deamidase”, and “protease derived from the genus Chryseobacterium” are as described in detail in "1.
• Modifier for protein-containing composition” above, and the “step of acting protein deamidase and protease derived from the genus Chryseobacterium on the protein-containing composition” is as described in detail in "2. Method for modifying protein-containing composition” above.
• the manufacturing method of the present invention may or may not include other steps in addition to the step of acting on protein deamidase and Chryseobacterium protease.
• any cooking step may be performed to prepare the processed vegetable protein-containing composition to be produced in the final form of a food or drink.
• the cooking step for example, when a structured vegetable protein-containing composition is used in the first production method, the composition after being treated with protein deamidase and Chryseobacterium protease can be molded into a shape suitable for the desired form and cooked as necessary to obtain a processed vegetable protein-containing composition (food).
• the cooking method can be appropriately determined by a person skilled in the art depending on the type of food. Specifically, cooking methods include boiling, baking (roasting, toasting, baking, grilling, broiling), steaming, frying, etc. These cooking methods may be used alone or in combination.
• the present invention also provides a processed vegetable protein-containing composition with an improved amount of essential amino acid release, obtained by a method including a step of acting on a vegetable protein-containing composition with protein deamidase and a protease derived from the genus Chryseobacterium (hereinafter, "first processed protein-containing composition"), and a processed protein-containing composition with improved foaming properties, foam stability, and/or emulsifying properties, obtained by a method including a step of acting on a protein-containing composition with protein deamidase and a protease derived from the genus Chryseobacterium (hereinafter, "second processed protein-containing composition”).
• first processed protein-containing composition and the second processed protein-containing composition are also collectively referred to as "processed protein-containing compositions”.
• the first processed protein-containing composition of the present invention is a composition that has been processed with protein deamidase and Chryseobacterium-derived protease to improve the amount of essential amino acid release.
• the second processed protein-containing composition of the present invention is a composition that has been processed with protein deamidase and Chryseobacterium-derived protease to improve foaming properties, foam stability, and/or emulsifying properties.
• the "improvement of the amount of free essential amino acids", “protein deamidase”, and “Chryseobacterium-derived protease” are as described in detail in "1.
• Modifier for protein-containing composition above, and the “method including the step of acting protein deamidase and Chryseobacterium-derived protease on a vegetable protein-containing composition” is as described in detail in "2.
• Method for modifying protein-containing composition and "3. Method for producing processed protein-containing composition”.
• the "improvement of foaming properties”, “improvement of foam stability”, “improvement of emulsifying properties”, “protein deamidase”, and “Chryseobacterium-derived protease” are as described in detail in "1.
• Modifier for protein-containing composition” above, and the “step of acting protein deamidase and Chryseobacterium-derived protease on a protein-containing composition” is as described in detail in "2. Method for modifying protein-containing composition” and "3. Method for producing processed protein-containing composition".
• the specific form of the processed protein-containing composition of the present invention can be selected from any food or drink form.
• the form can conform to meat, poultry, and/or fish paste processed foods.
• the first processed protein-containing composition of the present invention includes meat-like processed foods (meaning foods that imitate meat, poultry, and/or fish paste processed foods). More preferably, the first processed protein-containing composition of the present invention includes meat and/or poultry-like processed foods (meaning foods that imitate meat and/or poultry processed foods).
• meat and/or poultry processed foods may be foods that are cooked by shaping and heating meat ingredients using meat and/or poultry, and specific examples thereof include hamburger steaks, meatballs, patties, meatloaf, minced meat cutlets, dim sum, etc.
• specific forms of the first processed protein-containing composition of the present invention include vegetable milk substitutes, yogurt substitutes, cheese substitutes, ice cream substitutes, etc., and preferably vegetable milk substitutes and yogurt substitutes.
• the second processed protein-containing composition of the present invention include plant-based milk substitutes, cream substitutes, ice cream substitutes, and animal-based dairy products (animal milk, cream, ice cream), etc.
• Protein-Containing Compositions The protein materials shown in Table 1 were used.
• Protein deamidation enzyme protein glutaminase derived from Chryseobacterium proteolyticum (manufactured by Amano Enzyme Inc.) was used. Hereinafter, this protein deamidase will also be referred to as "PG.”
• the protein deamidase activity was measured by the following method. 0.1 mL of the sample solution containing protein deamidase was added to 1 mL of 0.2 M phosphate buffer (pH 6.5) containing 30 mM Z-Gln-Gly, and the mixture was left at 37° C. for 10 minutes, after which 1 mL of 0.4 M TCA solution was added to stop the reaction. As a blank, 1 mL of 0.4 M TCA solution was added to 1 mL of 0.2 M phosphate buffer (pH 6.5) containing 30 mM Z-Gln-Gly, and 0.1 mL of the sample solution containing protein deamidase was further added, followed by leaving the mixture at 37° C. for 10 minutes.
• the amount of ammonia produced in the reaction solution was measured using an Ammonia Test Wako (FUJIFILM Wako Pure Chemical Corporation).
• the ammonia concentration in the reaction solution was calculated from a calibration curve showing the relationship between ammonia concentration and absorbance (630 nm) created using an ammonia standard solution (ammonium chloride).
• the activity of protein deamidating enzyme was calculated from the following formula, with 1 unit (1U) being the amount of enzyme that produces 1 ⁇ mol of ammonia per minute.
• the reaction solution volume is 2.1
• the enzyme solution volume is 0.1
• Df is the dilution ratio of the enzyme solution.
• 17.03 is the molecular weight of ammonia.
• protease derived from Chryseobacterium genus As the protease derived from the genus Chryseobacterium, a protease derived from Chryseobacterium proteolyticum was used. The protease activity was measured by the following method. 5 mL of 0.6% (w/v) casein solution (0.05 mol/L sodium hydrogen phosphate, pH 8.0) was heated at 37°C for 10 minutes, and then 1 mL of a sample solution containing protease was added and immediately shaken.
• casein solution 0.05 mol/L sodium hydrogen phosphate, pH 8.0
• tyrosine standard stock solution 0.2mol/L hydrochloric acid 1mL, 2mL, 3mL and 4mL were measured, and 0.2mol/L hydrochloric acid test solution was added to each to make 100mL. 2mL of each solution was measured, 5mL of 0.55mol/L sodium carbonate test solution and 1mL of Folin test solution (1 ⁇ 3) were added, and the mixture was immediately shaken and left at 37°C for 30 minutes. For these solutions, 2mL of 0.2mol/L hydrochloric acid test solution was measured and the solution obtained by the same operation as above was used as a control, and the absorbances A1, A2, A3 and A4 at a wavelength of 660nm were measured.
• the absorbances A1, A2, A3 and A4 were plotted on the vertical axis and the amount of tyrosine ( ⁇ g) in 2mL of each solution was plotted on the horizontal axis, and a calibration curve was created to determine the amount of tyrosine ( ⁇ g) relative to the absorbance difference of 1.
• the amount of enzyme that causes an increase in the color substance of Folin's test solution equivalent to 1 ⁇ g of tyrosine per minute was defined as 1 unit (1 U).
• proteases a protease derived from Bacillus licheniformis and a protease derived from Bacillus amyloliquefaciens were used. Protease was measured by the method described in the section "B-2. Protease derived from the genus Chryseobacterium.”
• [C-2] Second protein-containing composition The vegetable protein material shown in [A. Protein-containing composition] above was suspended in deionized water (final concentration 10% by weight), boiled until completely gelatinized, and then 10 U/g-starch ⁇ -amylase was added and incubated at 60°C for 30 minutes to prepare a liquid vegetable protein-containing composition (pH about 7). A predetermined amount of enzyme shown in Table 3 was added to the vegetable protein-containing composition and incubated at 50°C for 2 hours. Then, the mixture was treated at 100°C for 5 minutes to perform an enzyme inactivation step. After the enzyme inactivation step, the mixture was cooled to room temperature to obtain a processed vegetable protein-containing composition (processed oat milk). In addition, the animal protein material shown in [A.
• Protein-containing composition] above was suspended in deionized water (final concentration 5 wt%) to prepare a liquid animal protein-containing composition (pH about 7).
• a predetermined amount of enzyme shown in Table 4 was added to the animal protein-containing composition and incubated at 50°C for 2 hours. Then, the mixture was treated at 100°C for 5 minutes to perform an enzyme inactivation process. After the enzyme inactivation process, the mixture was cooled to room temperature to obtain a processed animal protein-containing composition.
• essential amino acids essential AA: histidine, threonine, valine, methionine, tryptophan, phenylalanine, isoleucine, leucine, lysine
• non-essential amino acids non-essential AA: tyrosine, cysteine, aspartic acid, asparagine, serine, glutamic acid, glutamine, proline, glycine, alanine, arginine
• F. Foam Stability As in the test of [E. Foaming property] above, 50 mL of the processed protein-containing liquid composition obtained in [C-2] above was homogenized at 18,000 rpm for 30 minutes, and immediately transferred to a 100 mL graduated cylinder, and the total volume of the composition including the foam, VF0, was measured. After standing for 30 minutes, the total volume of the composition, VF30, was measured, and the "foam stability" was calculated using the following formula.
• a processed vegetable protein-containing composition was prepared by the method of "(C-1) First protein-containing composition" in the above [C. Production of processed protein-containing composition] using the vegetable protein material (soybean protein material or pea protein material) shown in Table 1 in the above [A. Protein-containing composition].
• the amount of PG added per 1 g of soybean protein was 30 U
• the amount of protease added was 39 U
• the amount of PG added per 1 g of pea protein was 18.8 U
• the amount of protease added was 24.4 U.
• the processed vegetable protein-containing composition was subjected to the above-mentioned [D. Amino acid analysis] test, and the ratio (%) of essential amino acids to the total amount of free amino acids was calculated. The results are shown in Table 2.
• a processed protein-containing composition was prepared by the method of "[C-2] Second protein-containing composition" in [C. Production of processed protein-containing composition] above, using a vegetable protein material (oat protein material) or an animal protein material (whey protein material) shown in Table 1 of [A. Protein-containing composition] above.
• the processed protein-containing composition was subjected to the above-mentioned tests for [E. Foamability], [F. Foam stability], and [G. Emulsifying ability]. The results are shown in Tables 3 and 4.

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• 2024
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