Methods of producing preparations, methods of producing food and beverages, and fermented preparations.

Abstract

The invention relates to at least one method for producing a preparation containing equol and / or equol derivatives that does not brown even when heated in its original form or in the presence of an added amino acid or analog. The problem is solved by a method for producing the preparation which includes step (a); step (b1) and / or step (b2); and step (c): (a) producing glucose and isoflavone aglycon from isoflavone glycoside; (b1) for a microorganism capable of producing equol to produce equol from isoflavone aglycon; (b2) for a microorganism capable of producing equol derivatives to produce equol derivatives from isoflavone aglycon; and (c) for the degradation of glucose to obtain a glucose content of 0 g / L or more and 2.0 g / L or less in the preparation, or 30 g / kg or less based on the solids content of the preparation.

Description

Method for producing a composition, method for producing a food or drink, and fermented composition

[0001] The present disclosure relates to a method for producing a composition, a method for producing a food or drink, and a fermented composition.

[0002] Isoflavone glycosides contained in soybean hypocotyls and soybean hypocotyl extracts are converted to aglycones by β-glucosidase and then further metabolized to equol and equol derivatives. Because equol has strong physiological effects similar to those of female hormones, its use has been proposed for preventing and ameliorating menopausal symptoms and osteoporosis (Patent Document 1), preventing and treating skin aging and wrinkles (Patent Document 2), and alleviating allergic symptoms (Patent Document 3). Equol derivatives, particularly 5-hydroxyequol, are known to have antioxidant effects (Non-Patent Document 1) and lifespan-extending effects (Non-Patent Document 2).

[0003] Special Table of Contents No. 2001-523258 Publication of Special Publication No. 2002-511860 Publication of Patent No. 4479505

[0004] Archives Biochem. Biophys. vol.356, pp133-141 (1998) J. Chin. Pharm. Sci. vol.23, pp378-384 (2014)

[0005] The inventors of the present invention prepared a composition containing equol and / or equol derivatives from isoflavone glycosides and found that the composition browned when heated. Furthermore, when the composition was mixed with ingredients containing proteins and other substances to produce food, the composition browned when heated. This was thought to be due to the Maillard reaction, a reaction between reducing sugars and amino acids. Furthermore, the Maillard reaction may have produced by-products such as acrylamide, which are known to be carcinogenic.

[0006] An objective of the present disclosure is to provide at least a method for producing a composition containing equol and / or an equol derivative that does not brown when heated, either directly or in the presence of additional amino acids, etc.

[0007] As a result of intensive research, the present inventors have found that when isoflavone glycosides are converted into aglycones, glucose, a reducing sugar, is generated, and since this glucose remains in the produced composition, the Maillard reaction proceeds upon heating, causing the composition to brown. Furthermore, they have found that the above-mentioned problems can be solved by carrying out a step of decomposing the glucose contained in the composition. That is, the present disclosure includes at least the following.

[0008] [1] A method for producing a composition, comprising the following steps: (a) step (b1) and / or step (b2); and step (c): (a) producing glucose and isoflavone aglycone from isoflavone glycoside; (b1) causing a microorganism capable of producing equol to produce equol from the isoflavone aglycone; (b2) causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycone; and (c) degrading the glucose so that the glucose content in the composition is 0 g / L or more but less than 2.0 g / L, or 30 g / kg or less per solid content of the composition. [2] The method of [1], wherein step (c) is a step of degrading the glucose by fermentation using a microorganism capable of sugar utilization. [3] The method of [2], wherein the microorganism capable of sugar utilization is one or more microorganisms selected from the group consisting of butyric acid bacteria, lactic acid bacteria, and yeast. [4] The production method according to [3], wherein the microorganism capable of sugar utilization is one or more microorganisms selected from the group consisting of microorganisms belonging to the genus Clostridium, microorganisms belonging to the genus Lactobacillus, microorganisms belonging to the genus Lactococcus, microorganisms belonging to the genus Leuconostoc, and microorganisms belonging to the genus Saccharomyces. [5] The production method according to [1], wherein step (c) is a step of decomposing glucose using a glucose-degrading enzyme. [6] The production method according to [5], wherein the glucose-degrading enzyme is one or more selected from the group consisting of glucose oxidase and glucose dehydrogenase. [7] The production method according to any of [1] to [6], wherein step (b1) and / or step (b2) and step (c) are carried out simultaneously. [8] The method according to any one of [1] to [7], wherein the isoflavone glycoside is daidzin and the isoflavone aglycone is daidzein. [9] The method according to any one of [1] to [8], wherein the step (a) is a step of reacting β-glucosidase with the isoflavone glycoside to produce glucose and the isoflavone aglycone.

[10] A method for producing a food or beverage, comprising the following steps: (a) step (b1) and / or (b2); (c) step; and (d) step: (a) producing glucose and isoflavone aglycone from isoflavone glycoside; (b1) causing a microorganism capable of producing equol to produce equol from the isoflavone aglycone; (b2) causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycone; (c) decomposing the glucose so that the glucose content in a composition produced by a method comprising steps (a), (b1), and / or (b2), and step (c) is 0 g / L or more and less than 2.0 g / L, or 30 g / kg or less per solid content of the composition; and (d) producing a food or beverage using a composition produced by a method comprising steps (a), (b1), and / or (b2), and step (c).

[11] A fermented composition comprising equol and / or an equol derivative and substantially no glucose.

[12] The fermentation composition according to

[11] , wherein being substantially free of glucose means containing 0 g / L or more and less than 2.0 g / L of glucose.

[13] A fermentation composition comprising equol and / or an equol derivative and glucose, wherein the glucose content is 30 g / kg or less per solid content of the fermentation composition.

[14] The fermentation composition according to any of

[11] to

[13] , which is used in foods and beverages.

[0009] The present disclosure may have the effect of providing at least a method for producing a composition containing equol and / or an equol derivative, which has a reduced amount of glucose and does not undergo the Maillard reaction or brown when heated, either directly or in the presence of additional amino acids, etc.

[0010] Each configuration and combination thereof in each embodiment is merely an example, and additions, omissions, substitutions, and other modifications of configurations are possible as appropriate within the scope of the present disclosure. The present disclosure is not limited by the embodiments, but only by the scope of the claims. Furthermore, each aspect disclosed in this specification can be combined with any other feature disclosed in this specification. Furthermore, a numerical range expressed using "to" means a range that includes the numerical values ​​before and after "to" as the lower and upper limits, and "A to B" means greater than or equal to A and less than or equal to B.

[0011] In this specification, the accession number of a bacterial strain beginning with the word "DSM" is the number assigned to microorganisms preserved at DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH). The accession number of a bacterial strain beginning with the word "JCM" is the number assigned to microorganisms preserved at Japan Collection of Microorganisms (Microbial Materials Development Division, RIKEN BioResource Research Center, National Research and Development Agency, Postal Code: 305-0074, Address: 3-1-1 Takanodai, Tsukuba City, Ibaraki Prefecture, Japan), and can be obtained from the same institution. The accession number of a bacterial strain beginning with the word "FERM" is the number assigned to microorganisms preserved at the National Institute of Advanced Industrial Science and Technology (AIST Patent Organism Depositary Center, Independent Administrative Institution, National Institute of Technology and Evaluation, Postal Code: 292-0818, Address: Room 120, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan), and can be obtained from the same institution. The accession numbers of strains beginning with the words KCCM are numbers assigned to microorganisms preserved at the Korean Culture Center of Microorganisms (KCCM) and can be obtained from the institution.

[0012] As used herein, "microorganism" refers to eubacteria, archaea, or fungi, and does not include viruses, animals, or plants.

[0013] In this specification, "decomposing glucose" means inactivating the reducing end of glucose so that glucose and amino acids or the like do not undergo a Maillard reaction, and includes oxidation of glucose and conversion to another compound by metabolism. Furthermore, the amino acids or the like are not particularly limited as long as they are compounds that can undergo a Maillard reaction with glucose, and examples thereof include compounds containing an amino group. Examples of compounds containing an amino group include amino acids, peptides, and proteins.

[0014] One embodiment of the present disclosure is a method for producing a composition, comprising the following steps: (a) producing glucose and isoflavone aglycones from isoflavone glycosides; (b1) causing a microorganism capable of producing equol to produce equol from the isoflavone aglycones; (b2) causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycones; and (c) decomposing the glucose so that the glucose content in the composition is 0 g / L or more and less than 2.0 g / L, or 30 g / kg or less per solid content of the composition.

[0015] [Step (a)] The production method according to this embodiment includes the step (a) of producing glucose and isoflavone aglycone from isoflavone glycoside.

[0016] Isoflavone glycosides refer to glycosides of isoflavones. Isoflavones are a type of polyphenol, and are flavonoids with an isoflavone as the basic skeleton. Isoflavones are found in large amounts in legumes such as soybeans, kudzu, red clover, and licorice. Examples of isoflavones in this embodiment include isoflavones derived from legumes containing isoflavones. Specifically, these include soybean-derived isoflavones (which may be referred to as "soybean isoflavones" or "soybean isoflavones" in the relevant technical field and market, and are treated as synonyms in the present disclosure), kudzu-derived isoflavones (which may be referred to as "kudzu isoflavones" or "kudzu isoflavones" in the relevant technical field and market, and are treated as synonyms in the present disclosure), red clover-derived isoflavones (which may be referred to as "red clover isoflavones" or "red clover isoflavones" in the relevant technical field and market, and are treated as synonyms in the present disclosure), and licorice-derived isoflavones (which may be referred to as "licorice isoflavones" or "licorice isoflavones" in the relevant technical field and market, and are treated as synonyms in the present disclosure).

[0017] Examples of isoflavone glycosides include genistin, glycitin, and daidzin. In this embodiment, a mixture of two or more of these may be used. For example, a mixture of daidzin, glycitin, and genistin may be used. These are contained in soybean hypocotyls, soybean hypocotyl extracts, etc., so for example, soybean hypocotyl extracts can be used as a mixture of isoflavone glycosides.

[0018] Examples of isoflavone aglycones include daidzein, 6-hydroxydaidzein, dihydroxydaidzein, genistein, glycitein, biochanin A, formononetin, orobol, and coumestrol. In this embodiment, a mixture of two or more isoflavone aglycones may be used. Note that the term "isoflavone aglycone" in this disclosure is sometimes referred to as "aglycone of isoflavones" in the technical field and market, and is treated as having the same meaning in this disclosure.

[0019] In this embodiment, preferably, the isoflavone glycoside is daidzin and the isoflavone aglycone is daidzein, or the isoflavone glycoside is genistin and the isoflavone aglycone is genistein, and more preferably, the isoflavone glycoside is daidzin and the isoflavone aglycone is daidzein.

[0020] Methods for producing glucose and isoflavone aglycone from isoflavone glycosides include a method of allowing β-glucosidase to act on isoflavone glycosides and a method of culturing a microorganism that produces β-glucosidase in a medium containing isoflavone glycosides.

[0021] β-Glucosidase is an enzyme that catalyzes the hydrolysis of β-glycosidic bonds in sugars. The β-glucosidase is not limited as long as it has the above-mentioned enzymatic activity, and examples include those derived from microorganisms belonging to the genera Aspergillus, Penicillium, Rhizopus, Pseudomonas, Pichia, and Lactococcus, as well as those derived from higher plants and animals. Commercially available enzyme preparations containing β-glucosidase, such as Pectinase G (manufactured by Amano Enzyme Co., Ltd.), can also be used.

[0022] To allow β-glucosidase to act on isoflavone glycosides, β-glucosidase may be added to a solution containing the isoflavone glycosides. The amount of β-glucosidase added can be adjusted as appropriate, but is typically 10 U to 3000 U, preferably 20 U to 1000 U, and more preferably 50 U to 500 U per gram of isoflavone glycoside.

[0023] The conditions for allowing β-glucosidase to act on isoflavone glycosides are not particularly limited as long as β-glucosidase acts on the isoflavone glycosides and catalyzes the release of glucose, but the temperature is usually 15° C. or higher and 65° C. or lower, preferably 20° C. or higher and 55° C. or lower, and more preferably 30° C. or higher and 45° C. or lower. The pH is usually 2 or higher and 9 or lower, preferably 3 or higher and 7 or lower, and more preferably 4 or higher and 6 or lower.

[0024] The microorganism that produces β-glucosidase is not particularly limited, and one or more microorganisms can be used regardless of genus, species, or strain. Examples include microorganisms belonging to the genera Aspergillus, Lactococcus, Streptococcus, Lactobacillus, Bifidobacterium, Penicillium, Pichia, Pseudomonas, and Rhizopus. Preferred examples include microorganisms belonging to the genera Aspergillus, Lactococcus, Streptococcus, Lactobacillus, and Bifidobacterium. More preferred examples include microorganisms belonging to Aspergillus niger, Aspergillus oryzae, Aspergillus aculeatus, Lactococcus lactis, Streptococcus thermophilus, Lactobacillus acidophilus, and Bifidobacterium bifidum.

[0025] As the culture conditions for the microorganisms that produce β-glucosidase, culture conditions that are generally used for culturing the above-mentioned microorganisms or culture conditions that are appropriately modified therefrom can be used.

[0026] Examples of culture solutions (medium) that can be used include BHI medium manufactured by Difco, ANAEROBE BASAL BROTH (ABB medium) manufactured by Oxoid, Wilkins-Chalgren Anaerobe Broth (CM0643) manufactured by Oxoid, and GAM medium and modified GAM medium manufactured by Nissui Pharmaceutical Co., Ltd. In this specification, the term "medium" refers to a solution in which microorganisms can grow, including a minimal medium, and does not include solutions in which microorganisms cannot grow, such as water, salt solutions, and buffer solutions.

[0027] The medium is supplemented with isoflavone glycosides. The isoflavone glycosides can be added to the medium before or during the culture of the microorganism. The content of the isoflavone glycosides is not particularly limited, but is usually 0.5 g / L to 100 g / L, preferably 1 g / L to 50 g / L, and more preferably 2 g / L to 20 g / L.

[0028] Water-soluble organic substances can be added to the medium as a carbon source. Examples of water-soluble organic substances include sugars such as glucose, arabinose, sorbitol, fructose, mannose, sucrose, trehalose, and xylose; alcohols such as glycerol; organic acids such as valeric acid, butyric acid, propionic acid, acetic acid, formic acid, fumaric acid, and succinic acid; and polysaccharides such as dextrin.

[0029] The concentration of the organic substance added to the medium as a carbon source can be adjusted appropriately for efficient growth. Generally, the amount added can be selected from the range of 0.1 to 10 wt / vol %.

[0030] In addition to the carbon sources described above, a nitrogen source can be added to the medium. Various nitrogen compounds commonly used in conventional fermentation can be used as the nitrogen source. Examples include inorganic and organic nitrogen sources. Examples of inorganic nitrogen sources include ammonium salts and nitrates. Preferred inorganic nitrogen sources are ammonium sulfate, ammonium chloride, ammonium phosphate, ammonium hydrogen phosphate, potassium nitrate, and sodium nitrate. Organic nitrogen sources include amino acids (glutamic acid, arginine, ornithine, etc.), fats and oils such as oleic acid, yeast extract, peptones (e.g., polypeptone N, soybean peptone, etc.), meat extracts (e.g., Ehrlich bonito extract, Lab-Remco powder, bouillon, etc.), seafood extracts, liver extracts, digested serum powders, and fish oils. More preferred are arginine, cysteine, citrulline, lysine, yeast extract, and peptones (e.g., polypeptone N, etc.).

[0031] Furthermore, in addition to carbon and nitrogen sources, the growth and activity of microorganisms can sometimes be enhanced by adding cofactors such as vitamins and inorganic compounds such as various salts to the medium. For example, the following are examples of microbial growth cofactors derived from plants and animals, such as inorganic compounds and vitamins:

[0032] Inorganic compounds, vitamins, potassium dihydrogen phosphate, biotin, magnesium sulfate, folic acid, manganese sulfate, pyridoxine, sodium chloride, thiamine, cobalt chloride, riboflavin, calcium chloride, nicotinic acid, zinc sulfate, pantothenic acid, copper sulfate, vitamin B12, alum, thiooctoic acid, sodium molybdate, p-aminobenzoic acid, potassium chloride, vitamin K, boric acid, etc., nickel chloride, sodium tungstate, sodium selenate, ferrous ammonium sulfate, sodium acetate trihydrate, magnesium sulfate heptahydrate, manganese sulfate tetrahydrate

[0033] Methods for producing a culture medium by adding growth cofactors derived from animals or plants, such as inorganic compounds and vitamins, are known. The medium can be liquid, semi-solid, or solid. The preferred form of the medium is a liquid medium.

[0034] The medium of the present disclosure may also contain dextrins. By culturing anaerobic microorganisms in a medium containing dextrins, a solution containing a functional substance and dextrins can be obtained without adding dextrins, even if dextrins are required in the culture solution after culture. Dextrins can be added to the medium before and during the culture of the microorganisms.

[0035] Furthermore, when culturing anaerobically, growth may be improved by adding reducing agents such as cysteine, cystine, sodium sulfide, sulfite, ascorbic acid, glutathione, thioglycolic acid, and rutin, or enzymes that decompose reactive oxygen species such as catalase and superoxide mutase to the medium.

[0036] When culturing anaerobically, the gas phase and aqueous phase during culturing preferably do not contain air or oxygen. For example, they may contain nitrogen and / or hydrogen at any ratio, or nitrogen and / or carbon dioxide at any ratio. These may be supplied in gaseous form. The proportion of hydrogen in the gas phase is not particularly limited, but is usually 0.5% to 100%, preferably 1.0% to 20%, and more preferably 2.0% to 10%.

[0037] The pH of the medium is preferably 5.0 or higher, more preferably 6.0 or higher, and even more preferably 6.5 or higher, while it is preferably 8.0 or lower, more preferably 7.5 or lower. The culture temperature is preferably 20°C to 45°C, more preferably 25°C to 40°C, and even more preferably 30°C to 37°C. The pressure conditions for the culture vessel are not particularly limited as long as they allow growth, but can be in the range of 0.001 to 1 MPa, preferably 0.01 to 0.5 MPa. The culture time is usually 8 to 340 hours, preferably 12 to 170 hours, and more preferably 16 to 120 hours.

[0038] [Step (b1)] The production method according to this embodiment includes (b1) a step of causing a microorganism capable of producing equol to produce equol from the isoflavone aglycone, and / or (b2) described below.

[0039] Examples of microorganisms capable of producing equol include microorganisms belonging to the genus Adlercreutzia, microorganisms belonging to the genus Atopobium, microorganisms belonging to the genus Bacteroides, microorganisms belonging to the genus Collinsella, microorganisms belonging to the genus Coriobacterium, microorganisms belonging to the genus Cryptobacterium, microorganisms belonging to the genus Denitrobacterium, and microorganisms belonging to the genus Eggerthella. Examples of microorganisms include microorganisms belonging to the genus Enterorhabdus, microorganisms belonging to the genus Eubacterium, microorganisms belonging to the genus Gordonibacter, microorganisms belonging to the genus Lactococcus, microorganisms belonging to the genus Olsenella, microorganisms belonging to the genus Paraeggerthella, microorganisms belonging to the genus Ruminococcus, microorganisms belonging to the genus Slackia, and microorganisms belonging to the genus Streptococcus.

[0040] Preferably, the microorganisms include microorganisms belonging to Adlercreutzia equolifaciens subsp. equolifaciens, microorganisms belonging to Adlercreutzia equolifaciens subsp. celatus, microorganisms belonging to Bacteroides ovatus, microorganisms belonging to Eggerthella sp., microorganisms belonging to Eubacterium sp., microorganisms belonging to Lactococcus garvieae, microorganisms belonging to Paraeggerthella sp., microorganisms belonging to Ruminococcus productus, and Slackia isoflavonicombetens. isoflavoniconvertens), Slackia equolifaciens, Slackia sp., and Streptococcus intermedius.

[0041] More preferably, Adlercreutzia equolifaciens subsp. equolifaciens strain DSM 19450, Adlercreutzia equolifaciens subsp. celatus strain DSM 18785, Eggerthella sp. strain KCCM 10490, Lactococcus garvieae strain DSM 6783, Slackia isoflavoniconvertens strain DSM 22006, Slackia equolifaciens strain DSM 24851, Slackia sp. sp.) strain FERM AP-20729. The above microorganisms may be used singly or in combination of two or more, regardless of genus, species or strain.

[0042] Furthermore, in this embodiment, the Adlercreutzia equolifaciens subsp. equolifaciens DSM 19450 strain is not limited to being the same strain as the deposited strain, but may also be a strain substantially equivalent to the deposited strain. A substantially equivalent strain refers to a strain that belongs to the same genus or species as the deposited strain and has the ability to produce equol. Furthermore, a substantially equivalent strain refers to a microorganism whose 16S rRNA gene nucleotide sequence has 97% or more, preferably 97.5% or more, more preferably 98% or more, even more preferably 98.7% or more, and even more preferably 99% or more homology with the 16S rRNA gene nucleotide sequence of the deposited strain. Furthermore, the equol-producing microorganism may be a strain bred from the above-mentioned deposited strain or a strain substantially equivalent thereto by mutation treatment, genetic recombination, selection of natural mutants, etc., so long as the effects of the present disclosure are not impaired. This also applies to the other deposited strains in this specification.

[0043] When the equol-producing microorganism produces β-glucosidase, the microorganism producing β-glucosidase may be the same as the equol-producing microorganism. In this case, steps (a) and (b1) are carried out consecutively in the same system.

[0044] The culture conditions may be those generally used for culturing the microorganisms or modified as appropriate, and may include the same conditions as those for culturing microorganisms that produce β-glucosidase.

[0045] [Step (b2)] The production method according to this embodiment includes the step (b1) and / or the step (b2) of causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycone. Examples of equol derivatives include 5-hydroxyequol.

[0046] When the equol derivative is 5-hydroxyequol, examples of microorganisms capable of producing 5-hydroxyequol include microorganisms belonging to the genera Adlercreutzia, Eggerthella, and Slackia. As microorganisms belonging to the genus Adlercreutzia, microorganisms belonging to Adlercreutzia equolifaciens are preferred, and among these, Adlercreutzia equolifaciens subsp. equolifaciens DSM 19450 strain or Adlercreutzia equolifaciens subsp. celatus DSM 18785 strain is preferred.

[0047] Examples of microorganisms belonging to the genus Eggerthella include microorganisms belonging to Eggerthella sp., and particularly preferably Eggerthella sp. KCCM 10490 strain. Examples of microorganisms belonging to the genus Slackia include microorganisms belonging to Slackia isoflavoniconvertens, Slackia equolifaciens, and Slackia sp.

[0048] When the microorganism capable of producing equol derivatives produces β-glucosidase, the microorganism producing the β-glucosidase may be the same as the microorganism capable of producing equol derivatives. In this case, steps (a) and (b2) are carried out consecutively in the same system.

[0049] The culture conditions may be those generally used for culturing the microorganisms or modified as appropriate, and may include the same conditions as those for culturing microorganisms that produce β-glucosidase.

[0050] [Step (c)] The production method according to this embodiment includes a step (c) of decomposing the glucose so that the glucose content in the composition is 0 g / L or more but less than 2.0 g / L, or 30 g / kg or less per solid content of the composition. Decomposing glucose refers to converting glucose into a compound with a smaller number of carbon atoms. Step (c) may be a step of decomposing the glucose by fermentation using a microorganism capable of sugar assimilation, or may be a step of decomposing the glucose using a glucose-degrading enzyme.

[0051] When step (c) utilizes fermentation using microorganisms capable of sugar utilization, the microorganisms capable of sugar utilization are preferably one or more microorganisms selected from the group consisting of butyric acid bacteria, lactic acid bacteria, and yeast, and more preferably one or more microorganisms selected from the group consisting of microorganisms belonging to the genus Clostridium, Lactobacillus, Lactococcus, Leuconostoc, and Saccharomyces.

[0052] Examples of microorganisms belonging to the genus Clostridium include Clostridium asparagiforme DSM 15981 strain and Clostridium bolteae JCM 12243 strain. Examples of microorganisms belonging to the genus Lactobacillus include Lactobacillus brevis DSM 20054 strain. Examples of microorganisms belonging to the genus Lactococcus include Lactococcus lactis DSM 20481 strain. Examples of microorganisms belonging to the genus Leuconostoc include Leuconostoc mesenteroides subsp. mesenteroides DSM 20343 strain.

[0053] When a microorganism capable of sugar utilization produces β-glucosidase, the microorganism producing the β-glucosidase may be the same as the microorganism capable of sugar utilization. In this case, steps (a) and (c) may be carried out continuously in the same system.

[0054] The microorganism capable of sugar utilization may be of the same species as the microorganism capable of equol production. That is, the microorganism capable of sugar utilization may be a microorganism capable of both equol production and sugar utilization. In this case, steps (b1) and (c) are carried out consecutively in the same system.

[0055] The microorganism capable of sugar utilization may be of the same species as the microorganism capable of equol derivative production. That is, the microorganism capable of sugar utilization may be a microorganism capable of both equol derivative production and sugar utilization. In this case, steps (b2) and (c) are carried out consecutively in the same system.

[0056] The culture conditions may be those generally used for culturing the microorganisms or modified as appropriate, and may include the same conditions as those for culturing microorganisms that produce β-glucosidase.

[0057] When step (c) utilizes an enzyme that decomposes glucose, examples of the enzyme that decomposes glucose include glucose oxidase and glucose dehydrogenase. These may be used alone or in combination of two or more. Glucose oxidase and glucose dehydrogenase are enzymes that catalyze the reaction of oxidizing β-D-glucose to D-glucono-1,5-lactone.

[0058] The glucose oxidase and glucose dehydrogenase are not limited as long as they have the above-mentioned enzymatic activity, and examples thereof include those derived from Aspergillus niger and Gluconobacter oxydans.

[0059] To decompose glucose using a glucose-degrading enzyme, the enzyme is added to a solution (culture medium) containing glucose. The amount of the enzyme added can be adjusted as appropriate, but is usually 1 U to 10,000 U, preferably 10 U to 5,000 U, and more preferably 100 U to 2,000 U per 1 g of glucose.

[0060] The temperature during glucose decomposition is not particularly limited, but is usually 15° C. to 60° C., preferably 25° C. to 50° C., more preferably 30° C. to 40° C., and particularly preferably 35° C. The pH during glucose decomposition is not particularly limited, but is usually 4 to 9, preferably 4 to 7, and more preferably 5 to 6.

[0061] When steps (b1) and (b2) are performed, the order of these steps is not limited, and step (b2) may be performed after step (b1), step (b2) may be performed after step (b1), or step (b1) and step (b2) may be performed simultaneously.

[0062] The order of steps (b1) and / or (b2) and step (c) is not limited, and step (c) may be performed after step (b1) and / or (b2), step (b1) and / or (b2) may be performed after step (c), or steps (b1) and / or (b2) and step (c) may be performed simultaneously. Specific examples of simultaneous performance of steps (b1) and / or (b2) and step (c) include adding a glucose-degrading enzyme to a medium containing glucose, isoflavone aglycone, and a microorganism capable of producing equol, co-culturing a microorganism capable of producing equol and a microorganism capable of sugar utilization in a medium containing glucose and isoflavone aglycone, and culturing a microorganism capable of producing equol and a microorganism capable of sugar utilization in a medium containing glucose and isoflavone aglycone.

[0063] In step (c), the glucose is decomposed so that the glucose content in the composition is 0 g / L or more but less than 2.0 g / L, or 30 g / kg or less per solid content of the composition. The glucose content can be adjusted to the above range by appropriately adjusting the culture conditions and the treatment conditions for the glucose-decomposing enzyme. For example, the culture time or treatment time can be extended.

[0064] The glucose content in the composition is preferably 0 g / L or more but less than 2.0 g / L, more preferably 0 g / L or more but less than 1.0 g / L, even more preferably 0 g / L or more but less than 0.5 g / L, and particularly preferably 0 g / L or more but less than 0.2 g / L. Furthermore, the glucose content per solid content of the composition is preferably 30 g / kg or less, more preferably 10 g / kg or less, and even more preferably 4 g / kg or less. The solid content is determined by measuring the residual weight after drying the composition. Drying methods include reduced pressure heat drying, normal pressure heat drying, and freeze drying. The glucose content can be measured by an electrode method. When the composition is solid and does not contain water, water can be added to the composition, glucose can be dissolved in water, and then the glucose content per solid content can be measured.

[0065] [Other Steps] The production method of this embodiment may include, for example, a step of quantifying the obtained equol and / or equol derivative. This can be done according to a conventional method. For example, a portion of the culture medium may be sampled, appropriately diluted, thoroughly stirred, and then filtered using a membrane such as a polytetrafluoroethylene (PTFE) membrane to remove insoluble matter, followed by quantification by high-performance liquid chromatography (HPLC).

[0066] The production method of this embodiment may also include a step of recovering the resulting equol and / or equol derivative. The recovery step may include a purification step, a concentration step, or the like. Purification treatments in the purification step include sterilization of microorganisms by heat or the like; sterilization by microfiltration (MF), ultrafiltration (UF), or the like; removal of solids and polymeric substances; extraction using organic solvents, ionic liquids, or the like; and adsorption and decolorization using hydrophobic adsorbents, ion exchange resins, activated carbon columns, or the like. Concentration treatments in the concentration step include concentration using an evaporator, reverse osmosis membrane, or the like.

[0067] Furthermore, the resulting solution containing equol and / or an equol derivative can be powdered by freeze-drying, spray-drying, etc. In the powdering process, excipients such as lactose, dextrin, cornstarch, etc. can also be added.

[0068] [Method for producing food and drink] The composition produced by the production method according to this embodiment, whether directly or in the presence of additional amino acids, does not undergo the Maillard reaction and does not brown when heated, so it can be suitably included in food and drink. In this specification, food and drink includes supplements. That is, another embodiment of the present disclosure is a method for producing food and drink, comprising the following steps (a); (b1) and / or (b2); (c); and (d). (a) producing glucose and isoflavone aglycones from isoflavone glycosides; (b1) causing a microorganism capable of producing equol to produce equol from the isoflavone aglycones; (b2) causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycones; (c) decomposing the glucose so that the glucose content in a composition produced by a method comprising steps (a), (b1), and / or (b2), and step (c) is 0 g / L or more and less than 2.0 g / L, or 30 g / kg or less per solid content of the composition; and (d) producing a food or beverage using a composition produced by a method comprising steps (a), (b1), and / or (b2), and step (c).

[0069] The foods and beverages produced by the manufacturing method of this embodiment contain equol and / or equol derivatives. These foods and beverages can be used as general foods and beverages, as well as foods for specified health uses, nutritional supplements, functional foods, foods for the sick, food additives, and the like (including beverages). The foods and beverages can be prepared, for example, by adding appropriate additives and then forming them into edible forms, such as granules, tablets, capsules, and pastes, using conventional means. The equol and / or equol derivatives can also be added to various foods, such as processed meat foods such as ham and sausage, processed seafood foods such as kamaboko and chikuwa, bread, confectionery, butter, powdered milk, and fermented dairy products, or beverages such as water, fruit juice, milk, and soft drinks.

[0070] The above-mentioned foods and beverages may contain water, proteins, carbohydrates, lipids, vitamins, minerals, organic acids, organic bases, fruit juice, flavors, etc. Examples of proteins include animal and vegetable proteins such as whole milk powder, skim milk powder, partially skim milk powder, casein, soy protein, egg protein, and meat protein, as well as hydrolysates thereof, and butter. Examples of carbohydrates include sugars, modified starch (dextrin, soluble starch, British starch, oxidized starch, starch ester, starch ether, etc.), and dietary fiber. Examples of lipids include vegetable oils and fats such as lard, safflower oil, corn oil, rapeseed oil, palm oil, fractionated oils thereof, hydrogenated oils, and interesterified oils. Examples of vitamins include vitamin A, carotenes, B vitamins, vitamin C, D vitamins, vitamin E, K vitamins, vitamin P, vitamin Q, niacin, nicotinic acid, pantothenic acid, biotin, inositol, choline, and folic acid. Examples of minerals include calcium, potassium, magnesium, sodium, copper, iron, manganese, zinc, selenium, and whey minerals. Examples of organic acids include malic acid, citric acid, lactic acid, and tartaric acid. Two or more of these components may be used in combination, and synthetic products and / or foods and beverages containing large amounts of these may also be used.

[0071] The explanations for step (a), step (b1) and / or step (b2), and step (c) in the method for producing a composition according to an embodiment of the present disclosure are incorporated herein by reference. The method for producing a food or beverage according to this embodiment may also include the other steps described above.

[0072] (d) The step of producing a food or beverage using a composition produced by a method including steps (a), (b1), and / or (b2), and step (c) can be carried out according to conventional methods. The amount, method, and timing of incorporation of equol and / or equol derivatives can be selected as appropriate. Furthermore, the food or beverage can be packaged in a container such as a bottle, bag, can, box, or pack, as needed.

[0073] [Fermented Composition] A composition production method according to one embodiment of the present disclosure can produce a composition containing equol and / or an equol derivative that has a reduced amount of glucose and does not undergo the Maillard reaction when heated, either directly or in the presence of additional amino acids, etc. That is, another embodiment of the present disclosure is a fermented composition that contains equol and / or an equol derivative and is substantially free of glucose. Alternatively, another embodiment of the present disclosure is a fermented composition that contains equol and / or an equol derivative and has a glucose content of 30 g / kg or less per solid content of the fermented composition. An example of an equol derivative is 5-hydroxyequol. Furthermore, the fermented composition according to this embodiment does not undergo the Maillard reaction or brown when heated, either directly or in the presence of additional amino acids, etc., and therefore can be suitably incorporated into foods and beverages. That is, the fermented composition can be used in foods and beverages.

[0074] "Substantially free of glucose" means that the glucose content is low enough that browning of the composition due to the Maillard reaction does not occur, and specifically includes cases where the glucose content is 0 g / L or more but less than 2.0 g / L.

[0075] The glucose content in the fermentation composition is preferably 0 g / L or more but less than 2.0 g / L, more preferably 0 g / L or more but less than 1.0 g / L, even more preferably 0 g / L or more but less than 0.5 g / L, and particularly preferably 0 g / L or more but less than 0.2 g / L. It is preferably 30 g / kg or less, more preferably 10 g / kg or less, and even more preferably 4 g / kg or less per solid content of the fermentation composition. The solid content is determined by measuring the residual weight after drying the fermentation composition. Drying methods include reduced pressure heat drying, atmospheric pressure heat drying, and freeze drying. The glucose content can be measured by an electrode method. When the fermentation composition does not contain water, such as in a solid state, water can be added to the fermentation composition, glucose dissolved in the water, and then the glucose content per solid content can be measured.

[0076] The content of equol and / or equol derivatives is preferably 1 g / kg or more, more preferably 5 g / kg or more, and even more preferably 10 g / kg or more, based on the solid content of the fermented composition. The content of equol and / or equol derivatives can be measured by HPLC as described above.

[0077] The fermentation composition may contain other components that may be included in the medium described above.

[0078] Foods and beverages containing the fermented composition can be used as general foods and beverages, as well as foods for specified health uses, nutritional supplements, functional foods, foods for the sick, food additives, etc. (These include beverages.) As for the form of the food and beverage, for example, after adding appropriate auxiliary agents, the food and beverage may be formed into an edible form, such as granules, particles, tablets, capsules, paste, etc., using conventional means and then provided for consumption. The fermented composition may also be added to various foods, for example, processed meat foods such as ham and sausage, processed seafood foods such as kamaboko and chikuwa, bread, confectionery, butter, powdered milk, fermented dairy products, or beverages such as water, fruit juice, milk, and soft drinks.

[0079] The above-mentioned foods and beverages may contain water, proteins, carbohydrates, lipids, vitamins, minerals, organic acids, organic bases, fruit juice, flavors, etc. Examples of proteins include animal and vegetable proteins such as whole milk powder, skim milk powder, partially skim milk powder, casein, soy protein, egg protein, and meat protein, as well as hydrolysates thereof, and butter. Examples of carbohydrates include sugars, modified starch (dextrin, soluble starch, British starch, oxidized starch, starch ester, starch ether, etc.), and dietary fiber. Examples of lipids include vegetable oils and fats such as lard, safflower oil, corn oil, rapeseed oil, palm oil, fractionated oils thereof, hydrogenated oils, and interesterified oils. Examples of vitamins include vitamin A, carotenes, B vitamins, vitamin C, D vitamins, vitamin E, K vitamins, vitamin P, vitamin Q, niacin, nicotinic acid, pantothenic acid, biotin, inositol, choline, and folic acid. Examples of minerals include calcium, potassium, magnesium, sodium, copper, iron, manganese, zinc, selenium, and whey minerals. Examples of organic acids include malic acid, citric acid, lactic acid, and tartaric acid. Two or more of these components may be used in combination, and synthetic products and / or foods and beverages containing large amounts of these may also be used.

[0080] The above-mentioned foods and beverages can be produced according to conventional methods. The amount, method, and timing of incorporation of equol and / or equol derivatives can be selected as appropriate. Furthermore, the above-mentioned foods and beverages can be packaged in containers such as bottles, bags, cans, boxes, and packs, as needed.

[0081] The present disclosure will be described in more detail below with reference to specific examples, but the present disclosure is not limited to these examples.

[0082] (Method for measuring glucose concentration) 1 mL of the liquid to be measured was taken and the bacterial cells were precipitated by centrifugation. The supernatant was filtered through a 0.45 μm filter, and the filtrate was analyzed under the following conditions. If the concentration exceeded the measurement range (10 mg / dL to 600 mg / dL), the filtrate was diluted with Milli-Q water and analyzed. Testing device: Glutest Neo Alpha, manufactured by Sanwa Kagaku Kenkyusho Co., Ltd. Chip: Glutest Neo Sensor, manufactured by Sanwa Kagaku Kenkyusho Co., Ltd. The content per solid content (g / kg) was calculated by measuring the solid content (kg / L) of the liquid to be measured using an electronic moisture meter (MOC-120H, manufactured by Shimadzu Corporation) and dividing the glucose concentration (g / L) by the solid content.

[0083] (Method for measuring equol concentration and equol derivatives) 20 μL of the liquid to be measured was taken and diluted 50-fold with a diluent consisting of ethanol and Milli-Q water at a ratio of 70:30 (v / v). The diluted liquid was filtered through a 0.45 μm filter, and the filtrate was analyzed under the following HPLC conditions. (S)-equol manufactured by Fujifilm Wako Pure Chemical Industries, Ltd. was used as the equol standard, and 5-hydroxyequol manufactured by Toronto Research Chemicals was used as the 5-hydroxyequol standard. {HPLC conditions} Column: Phenomenex SYNERGI, 4 μm, POLAR-R, 150 mm x 4.6 mm Eluent: distilled water / methanol at a ratio of 55 / 45 (v / v) Temperature: 40°C Detection wavelength: 280 nm Flow rate: 1.0 mL / min Injection: 10 μL Time: 30 min

[0084] Test Example 1 (Preparation of Pre-culture Medium) A pre-culture medium was prepared by dissolving 35.4 g of Anaerobe Basal Broth (ABB) medium (manufactured by Thermo Scientific) in 1 L of water. 10 mL of the pre-culture medium was dispensed into a test tube and capped with a butyl rubber stopper. After replacing the gas with nitrogen gas, the tube was sterilized in an autoclave.

[0085] (Enzyme Treatment) 0.16 g / L of pectinase G Amano (manufactured by Amano Enzyme Inc.) was added to 16 g / L of soybean hypocotyl extract (containing 80% isoflavone glycosides such as daidzin, glycitin, and genistin), and the mixture was kept overnight at 50°C with stirring to liberate sugars from the isoflavone glycosides.

[0086] (Preparation of main culture medium) ABB medium, arginine, and β-cyclodextrin were added to the enzyme treatment solution so that the final concentrations were 35.4 g / L, 1 g / L, and 16 g / L, respectively, and the mixture was dispensed into 2 L mini jars in 1 L portions. After replacing the atmosphere with nitrogen gas, the mixture was sterilized in an autoclave.

[0087] (Pre-culture) Microorganisms were inoculated into the pre-culture medium in the combinations shown in Table 1, then the atmosphere was replaced with anaerobic gas and the medium was cultured at 37°C and 200 spm for 2 days. As the microorganisms capable of producing equol, either Adlercreutzia equolifaciens subsp. celatus DSM 18785 strain (represented as AC in the table) or Adlercreutzia equolifaciens DSM 19450 strain (represented as AE in the table) was used. These microorganisms also have the ability to produce equol derivatives (particularly 5-hydroxyequol). The microorganisms capable of sugar utilization used were Clostridium asparagiforme DSM 15981 strain (represented as CA in the table), Clostridium bolteae JCM 12243 strain (represented as CB in the table), Lactobacillus brevis DSM 20054 strain (represented as LB in the table), Lactococcus lactis DSM 20481 strain (represented as LL in the table), and Leuconostoc mesenteroides subsp. mesenteroides DSM 20343 strain (represented as LM in the table).

[0088] (Main Culture) After inoculating the main culture medium with the above-mentioned pre-culture medium, the main culture medium was cultured at 37°C and 500 rpm for 2 days while aerating anaerobic gas through a 0.22 µm filter.

[0089] (Results) After cultivation, the equol concentration, 5-hydroxyequol concentration, and glucose concentration were measured. The results are shown in Table 1. The glucose concentration expressed in "g / kg" indicates the glucose content per solid content.

[0090]

[0091] It is clear from Comparative Examples 1 and 2 and Examples 1 to 6 that equol and 5-hydroxyequol were produced by culturing microorganisms capable of producing equol. Furthermore, it is clear from Examples 1 to 6 that co-culturing microorganisms capable of sugar assimilation resulted in sugar assimilation, with no glucose remaining in the culture medium.

[0092] Test Example 2 (Preparation of Pre-culture Medium) A pre-culture medium was prepared in the same manner as in Test Example 1.

[0093] (Enzyme Treatment) 0.16 g / L of pectinase G Amano was added to 16 g / L of soybean hypocotyl extract (containing 80% isoflavone glycosides such as daidzin, glycitin, and genistin), and the mixture was kept overnight at 50°C with stirring to liberate sugars from the isoflavone glycosides.

[0094] (Preparation of main culture medium) ABB medium was added to the enzyme treatment solution so that the final concentrations were 35.4 g / L, arginine 1 g / L, and β-cyclodextrin 16 g / L, and 50 mL of the mixture was dispensed into vials and sealed with butyl rubber stoppers. After replacing the atmosphere with nitrogen gas, the mixture was sterilized in an autoclave.

[0095] (Pre-culture) Microorganisms capable of producing equol shown in Table 2 were inoculated into a pre-culture medium, and the atmosphere was replaced with anaerobic gas, followed by culture at 37°C and 200 spm for 2 days.

[0096] (Main Culture) After inoculating the main culture medium with the above-mentioned pre-culture, the atmosphere was replaced with anaerobic gas and the medium was cultured at 37°C and 200 spm for 3 days. The cultured medium was designated as Comparative Examples 3 and 4, and the equol concentration, 5-hydroxyequol concentration, and glucose concentration were measured.

[0097] (Yeast Treatment) To Comparative Examples 3 and 4, 1 g of baker's yeast (Super Camellia Dry Yeast, manufactured by Nisshin Seifun Welna Co., Ltd.) was added per 100 mL of culture solution, and the mixture was stored overnight at 30° C. to prepare Examples 7 and 8, respectively.

[0098] (Results) The measurement results of the equol concentration, 5-hydroxyequol concentration, and glucose concentration in Comparative Examples 3 and 4 and Examples 7 and 8 are shown in Table 2.

[0099]

[0100] The glucose concentration in Examples 7 and 8, which were cultured after the yeast treatment, was significantly lower than that in Comparative Examples 3 and 4, which were cultured after the main culture. It is believed that glucose was assimilated by the yeast. In addition, the yeast-treated samples had a distinctive yeast smell and a good flavor.

[0101] Test Example 3 The solutions of Comparative Example 3 and Example 7 were centrifuged to remove bacterial cells and insoluble components, and the supernatants were recovered. The absorbance of the supernatants was measured at 420 nm. After heating at 80°C for 2 hours, the absorbance was measured in the same manner. The results are shown in Table 3.

[0102]

[0103] In Comparative Example 3, the absorbance increased upon heating. This is thought to be due to the Maillard reaction caused by the remaining glucose, resulting in browning. On the other hand, in Example 7, the glucose concentration was reduced to 0.2 g / L or less due to the action of yeast, so no increase in absorbance was observed even after heat treatment.

[0104] Test Example 4: Glycine was added to the solutions of Comparative Example 3 and Example 7 to a concentration of 2 g / L, followed by centrifugation to remove bacterial cells and insoluble components, and the supernatant was recovered. The absorbance of the supernatant was measured at 420 nm. Similarly, glycine was added to the solutions of Comparative Example 3 and Example 7 to a concentration of 2 g / L, followed by heating at 80°C for 2 hours, and the absorbance was measured in the same manner. The results are shown in Table 4.

[0105]

[0106] In Comparative Example 3, the absorbance increased upon heating. This is thought to be due to the Maillard reaction caused by the remaining glucose and the added glycine, resulting in browning. On the other hand, in Example 7, the glucose concentration was reduced to 0.2 g / L or less due to the action of yeast, so no increase in absorbance was observed even after the addition of glycine.

[0107] Test Example 5 Main cultivation was carried out in the same manner as in Test Example 2. After cultivation, equol and 5-hydroxyequol production was confirmed by HPLC analysis, and glucose concentrations were measured. After confirming that glucose remained, commercially available glucose oxidase (Amano Enzyme Inc.) and peroxidase (Fujifilm Wako Pure Chemical Industries, Ltd.) were added and the mixture was stored overnight at 30°C. The equol, 5-hydroxyequol, and glucose concentrations were then measured. The culture solutions before enzyme addition are shown as Comparative Examples 5 and 6, and the culture solutions after enzyme addition are shown as Examples 9 and 10 in Table 5.

[0108] It can be seen that the glucose is decomposed by the enzyme addition treatment and does not remain in the solution after treatment.

[0109]

[0110] Test Example 6 (Preparation of pre-culture medium) A pre-culture medium was prepared in the same manner as in Test Example 1. (Enzyme treatment) 0.16 g / L of pectinase G Amano (manufactured by Amano Enzyme Inc.) was added to 16 g / L of soybean hypocotyl extract (containing 44% isoflavone glycosides such as daidzin, glycitin, and genistin), and the mixture was kept overnight at 50°C with stirring to liberate sugars from the isoflavone glycosides.

[0111] (Preparation of main culture medium) ABB medium, arginine, and β-cyclodextrin were added to the enzyme treatment solution so that the final concentrations were 35.4 g / L, 1 g / L, and 16 g / L, respectively, and the mixture was dispensed into 2 L mini jars in 1 L portions. After replacing the atmosphere with nitrogen gas, the mixture was sterilized in an autoclave.

[0112] (Pre-culture) Microorganisms were inoculated into the pre-culture medium in the combinations shown in Table 5, and then the atmosphere was replaced with anaerobic gas and cultured for two days at 37°C and 200 spm. The microorganisms used were the same as in Test Example 1. (Main culture) Microorganisms were inoculated into the main culture medium from the pre-culture medium, and then cultured for two days at 37°C and 500 rpm while aerating anaerobic gas through a 0.22 μm filter. (Results) After culture, the equol concentration, 5-hydroxyequol concentration, and glucose concentration were measured. The results are shown in Table 6.

[0113]

[0114] It is clear from Comparative Examples 7 and 8 and Examples 11 to 16 that equol and 5-hydroxyequol were produced by culturing microorganisms capable of producing equol. Furthermore, it is clear from Examples 11 to 16 that co-culturing microorganisms capable of sugar assimilation resulted in sugar assimilation, with no glucose remaining in the culture medium.

[0115] Test Example 7 Commercially available glucose oxidase (Amano Enzyme Inc.) and peroxidase (Fujifilm Wako Pure Chemical Industries, Ltd.) were added to Comparative Examples 7 and 8, and the mixture was stored overnight at 30° C. Thereafter, the equol concentration, 5-hydroxyequol concentration, and glucose concentration were measured.

[0116] (Results) The culture solutions after the enzyme addition treatment were designated as Examples 17 and 18, and the measurement results of the equol concentration, 5-hydroxyequol concentration, and glucose concentration in Comparative Examples 7 and 8 and Examples 17 and 18 are shown in Table 7.

[0117]

[0118] It can be seen that the glucose is decomposed by the enzyme addition treatment and does not remain in the solution after treatment.

[0119] Test Example 8 The solutions of Comparative Example 7 and Example 17 were centrifuged to remove bacterial cells and insoluble components, and the supernatants were recovered. The absorbance of the supernatants was measured at 420 nm. After heating at 80°C for 2 hours, the absorbance was measured in the same manner. The results are shown in Table 8.

[0120]

[0121] In Comparative Example 7, the absorbance increased upon heating. This is thought to be due to the Maillard reaction caused by the remaining glucose, resulting in browning. On the other hand, in Example 17, the glucose concentration was reduced to 0.1 g / L or less due to the action of the enzyme, so no increase in absorbance was observed even after heat treatment.

Claims

1. A method for producing a composition, comprising the following steps (a); (b1) and / or (b2); and (c). (a) producing glucose and isoflavone aglycone from an isoflavone glycoside; (b1) causing a microorganism capable of producing equol to produce equol from the isoflavone aglycone; (b2) causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycone; and (c) decomposing the glucose so that the glucose content in the composition is 0 g / L or more and less than 2.0 g / L, or 30 g / kg or less per solid content of the composition.

2. The method according to claim 1, wherein the step (c) is a step of decomposing the glucose by fermentation using a microorganism having sugar assimilation ability.

3. The method according to claim 2, wherein the microorganism capable of utilizing sugar is one or more microorganisms selected from the group consisting of butyric acid bacteria, lactic acid bacteria, and yeast.

4. The method of claim 3, wherein the microorganism having the ability to utilize sugar is one or more microorganisms selected from the group consisting of microorganisms belonging to the genus Clostridium, microorganisms belonging to the genus Lactobacillus, microorganisms belonging to the genus Lactococcus, microorganisms belonging to the genus Leuconostoc, and microorganisms belonging to the genus Saccharomyces.

5. The method according to claim 1, wherein the step (c) is a step of decomposing the glucose using an enzyme that decomposes glucose.

6. The method according to claim 5, wherein the glucose-decomposing enzyme is one or more selected from the group consisting of glucose oxidase and glucose dehydrogenase.

7. The manufacturing method according to any one of claims 1 to 6, wherein the steps (b1) and / or (b2) and the step (c) are carried out simultaneously.

8. The method according to any one of claims 1 to 6, wherein the isoflavone glycoside is daidzin and the isoflavone aglycone is daidzein.

9. The method according to any one of claims 1 to 6, wherein the step (a) is a step of reacting β-glucosidase with an isoflavone glycoside to produce glucose and an isoflavone aglycone.

10. A method for producing a food or beverage, comprising the following steps: (a), (b1) and / or (b2), (c), and (d): (a) producing glucose and an isoflavone aglycone from an isoflavone glycoside; (b1) causing a microorganism capable of producing equol to produce equol from the isoflavone aglycone; (b2) causing a microorganism capable of producing an equol derivative to produce an equol derivative from the isoflavone aglycone; (c) decomposing the glucose so that the glucose content in a composition produced by the method comprising steps (a), (b1), and / or (b2), and (c) is 0 g / L or more and less than 2.0 g / L, or 30 g / kg or less per solid content of the composition; and (d) producing a food or beverage using a composition produced by the method comprising steps (a), (b1), and / or (b2), and (c).

11. A fermented composition comprising equol and / or an equol derivative, and being substantially free of glucose.

12. The fermentation composition according to claim 11, wherein said substantially free of glucose means containing between 0 g / L and less than 2.0 g / L of glucose.

13. A fermentation composition comprising equol and / or an equol derivative and glucose, wherein the glucose content is 30 g / kg or less based on the solid content of the fermentation composition.

14. The fermentation composition according to claim 11 or 13, for use in food or beverage products.