Tyrosine phenol-lyase inhibitor

Abstract

To provide a novel tyrosine phenol-lyase inhibitor derived from food components.SOLUTION: A tyrosine phenol-lyase inhibitor comprises a phenolic hydroxyl group-containing compound represented by the following formula (I), a salt or ester thereof, or a glycoside thereof. In the formula, X1 is carbon or nitrogen. R1 is a carboxy group, hydrogen, a 4-vinylphenyl group, or the like. R2 is a hydroxyl group, hydrogen, or an alkoxy group. R3 is a hydroxyl group or hydrogen. R4 is hydrogen or the like.SELECTED DRAWING: None

Description

Technical Field

[0001] The present invention relates to a tyrosine phenol lyase inhibitor, and more particularly to a tyrosine phenol lyase inhibitor containing a predetermined phenolic hydroxyl group-containing compound, a salt or an ester thereof, or a glycoside thereof.

Background Art

[0002] Tyrosine phenol lyase (TPL) is an enzyme that decomposes tyrosine into phenol, pyruvic acid, and ammonia. The present inventors have discovered that administration of a high-fat diet to diabetic model mice increases the phenol in feces by 108-fold (Non-Patent Document 1). The phenol in this feces is produced from tyrosine by intestinal bacteria expressing tyrosine phenol lyase. The phenol produced by the metabolism of these intestinal bacteria is metabolized in the liver and converted to phenyl sulfate. It is known that this phenyl sulfate damages glomerular epithelial cells and induces albuminuria, and is involved in the progression of diabetic nephropathy. Therefore, by inhibiting tyrosine phenol lyase, it may be possible to suppress the production of phenol, which is a precursor of phenyl sulfate, and prevent or improve diabetic nephropathy. Therefore, when 2-Aza-Tyr, which is a structural analog molecule of tyrosine and a tyrosine phenol lyase inhibitor, was orally administered to diabetic model mice (db / db mice), it was reported that the blood phenyl sulfate concentration and the urinary albumin / creatinine ratio decreased, and the progression of diabetic nephropathy was suppressed (Non-Patent Document 2).

[0003] On the other hand, it has been pointed out that phenol produced by intestinal bacteria expressing tyrosine phenol lyase is absorbed from the digestive tract and then migrates to the skin, interfering with epidermal formation and causing epidermal barrier dysfunction and skin dryness (Non-Patent Document 3). Furthermore, since phenol is a strong denaturing agent, it also causes diarrhea and constipation by damaging the digestive tract nerves and affecting intestinal peristaltic movement.

Prior Art Documents

Non-Patent Documents

[0004]

Non-Patent Document 1

Non-Patent Document 2

Non-Patent Document 3

Summary of the Invention

Problems to be Solved by the Invention

[0005] As described above, phenol produced by intestinal bacteria expressing tyrosine phenol lyase may be involved in various symptoms and diseases. Therefore, it is important to suppress phenol production in the intestinal tract using a tyrosine phenol lyase inhibitor. In addition, when reaching the intestinal tract with a tyrosine phenol lyase inhibitor, oral ingestion, which is a simple and safe method of intake, is preferable. Therefore, a natural product-derived material having high safety for the human body, for example, a tyrosine phenol lyase inhibitor derived from a food component, is expected.

[0006] Therefore, the present invention has been made in view of the above points, and its object is to provide a novel tyrosine phenol lyase inhibitor derived from food components and the like.

Means for Solving the Problems

[0007] For the purpose of searching for food components having tyrosine phenol lyase (TPL) inhibitory activity, the inventors conducted TPL inhibitory activity tests on various food components, and found that specific phenolic hydroxyl group-containing compounds exhibit stronger TPL inhibitory activity than the known TPL inhibitor 2-Aza-Tyr. Based on this finding, the present invention has been completed.

[0008] In order to solve the above problems, the tyrosine phenol lyase inhibitor of the present invention includes a phenolic hydroxyl group-containing compound represented by the following formula (I), a salt or an ester thereof, or a glycoside thereof. In formula (I), X1 represents carbon or nitrogen; R1 represents a group represented by the following formula (II), a carboxy group, hydrogen or a 4-vinylphenol group; R2 represents a hydroxyl group, hydrogen or an alkoxy group having 1 to 3 carbon atoms; R3 represents a hydroxyl group or hydrogen, and when X1 is nitrogen, R3 does not exist; R4 represents hydrogen or a group represented by the following formula (III). However, when R1 is a group represented by formula (II), X1 is carbon, and R3 and R4 are hydrogen; when R1 is a carboxy group, R4 is hydrogen; when R1 is hydrogen, X1 is carbon, R2 and R3 are hydroxyl groups, and R4 is hydrogen, or X1 is carbon, R2 and R3 are hydrogen, and R4 is a group represented by formula (III); when R1 is a 4-vinylphenol group, X1 is carbon, R2 and R4 are hydrogen, and R3 is a hydroxyl group.

[0009]

Chemical formula

[0010]

Chemical formula

[0011] By using the above-mentioned compounds, it is possible to inhibit tyrosine phenol lyase. Thereby, the production of phenol by tyrosine phenol lyase using tyrosine as a substrate can be suppressed. These compounds are mainly components contained in edible plants such as tea, spices, vegetables, and fruits, and have high safety. In addition, these compounds can be used in functional foods, pharmaceutical compositions, etc. for preventing, improving, or treating symptoms and diseases in which phenol produced by tyrosine phenol lyase is involved, such as in the intestinal tract.

[0012] In addition, in the tyrosine phenol lyase inhibitor of the present invention, in the above-mentioned formula (I), X1 represents carbon; R1 represents a group represented by formula (II) or a carboxy group; R2 represents a hydroxyl group or hydrogen; R3 represents a hydroxyl group or hydrogen; R4 represents hydrogen; provided that when R1 is a group represented by formula (II), it is also preferable that R3 is hydrogen. Thereby, preferable compounds are selected as compounds having an excellent tyrosine phenol lyase inhibitory action.

[0013] In addition, it is also preferable that the tyrosine phenol lyase inhibitor of the present invention is at least one compound selected from the group consisting of quercetin, gallic acid, 3-hydroxybenzoic acid, 3,4-dihydroxybenzoic acid, 3,5-dihydroxybenzoic acid, 4-methoxy-3-hydroxybenzoic acid, 5-hydroxynicotinic acid, pyrogallol, resveratrol, and daidzein, which is a phenolic hydroxyl group-containing compound represented by the above-mentioned formula (I). Thereby, specific compounds having an excellent tyrosine phenol lyase inhibitory action are selected.

[0014] Further, in the tyrosine phenol lyase inhibitor of the present invention, the ester of the phenolic hydroxyl group-containing compound represented by the above formula (I) is an ester of gallic acid, and it is also preferable that this ester of gallic acid is epicatechin gallate or epigallocatechin gallate. Thereby, specific compounds having an excellent tyrosine phenol lyase inhibitory action are selected. Further, both epicatechin gallate and epigallocatechin gallate are components contained in green tea and are particularly excellent in safety.

[0015] In addition, the composition for suppressing phenol production in the intestinal tract of the present invention contains the above-described tyrosine phenol lyase inhibitor. Thereby, it is possible to suppress the production of phenol in the intestinal tract by intestinal bacteria that express tyrosine phenol lyase. Therefore, this composition can be used for functional foods and pharmaceutical compositions for preventing, improving, or treating symptoms and diseases in which phenol in the intestinal tract is involved.

[0016] In addition, the food or drink for reducing the phenol concentration in feces of the present invention contains the above-described tyrosine phenol lyase inhibitor. Thereby, since it is possible to suppress the production of phenol in the intestinal tract by intestinal bacteria that express tyrosine phenol lyase, the phenol concentration in feces can be reduced, and the fecal odor derived from phenol can also be reduced.

[0017] In addition, the food or drink for suppressing skin dryness of the present invention contains the above-described tyrosine phenol lyase inhibitor. Thereby, since it is possible to suppress the production of phenol in the intestinal tract by intestinal bacteria that express tyrosine phenol lyase, the amount of phenol transferred from the intestinal tract to the skin is also reduced. Therefore, the hindrance to epidermal formation by phenol is reduced, epidermal formation is promoted, and skin dryness can be suppressed.

[0018] In addition, the food or drink for improving skin barrier function of the present invention contains the above-described tyrosine phenol lyase inhibitor. Thereby, since it is possible to suppress the production of phenol in the intestinal tract by intestinal bacteria that express tyrosine phenol lyase, the amount of phenol transferred from the intestinal tract to the skin is also reduced. Therefore, the hindrance to epidermal formation by phenol is reduced, epidermal formation is promoted, and the skin barrier function can be improved.

[0019] In addition, the composition for preventing, improving or treating symptoms, pathological conditions or diseases involving phenol in the intestinal tract of the present invention contains the above-described tyrosine phenol lyase inhibitor. Thereby, since it is possible to suppress the production of phenol in the intestinal tract by intestinal bacteria that express tyrosine phenol lyase, it can be used as a functional food or pharmaceutical composition for preventing, improving or treating symptoms and diseases involving phenol in the intestinal tract.

[0020] In addition, it is also preferable that the above-described symptoms, pathological conditions or diseases involving phenol in the intestinal tract are at least one symptom, pathological condition or disease selected from the group consisting of skin aging, diarrhea, constipation, uremia, fecal odor and diabetic nephropathy. Thereby, specific symptoms, pathological conditions or diseases in which phenol produced and accumulated in the intestinal tract is involved and can be prevented, improved or treated by the tyrosine phenol lyase inhibitor of the present invention are selected.

Advantages of the Invention

[0021] According to the present invention, it is possible to provide a tyrosine phenol lyase inhibitor having the following excellent effects. (1) It can inhibit tyrosine phenol lyase. (2) It is a component mainly contained in edible plants such as tea, spices, fruits, etc., and has high safety. (3) It can suppress the production of phenol in the intestinal tract. (4) It can be used in foods, pharmaceutical compositions, etc. for preventing, improving or treating symptoms and diseases involving phenol produced by tyrosine phenol lyase.

Brief Description of the Drawings

[0022]

Figure 1

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Figure 9

Figure 10

Figure 11

Mode for Carrying Out the Invention

[0023] Hereinafter, the tyrosine phenol lyase inhibitor of the present invention will be described in detail. The tyrosine phenol lyase (TPL) in the present invention, also called β-tyrosinase, is an enzyme that uses pyridoxal 5-phosphate (PLP) as a coenzyme and hydrolyzes tyrosine into phenol, pyruvic acid, and ammonia. Although D-tyrosine has low reactivity, tyrosine can be decomposed using either the L-form or the D-form as a substrate. Tyrosine phenol lyase is an enzyme identified as being involved in phenol production in the intestine and is distributed in intestinal bacteria genera such as the genus Erwinia, Escherichia, Citrobacter, and Aerobacter. The tyrosine phenol lyase expressed by these intestinal bacteria hydrolyzes tyrosine ingested from food and produces phenol in the intestinal tract. Phenol produced in the intestinal tract by the action of tyrosine phenol lyase is converted to phenylsulfuric acid in the liver and has been reported to be involved in the progression of kidney diseases such as diabetic nephropathy. In addition, it has been pointed out that phenol produced in the intestinal tract migrates to the skin and interferes with epidermal formation, causing epidermal barrier dysfunction and skin dryness. Moreover, since phenol is a strong denaturing agent, it can also cause diarrhea and constipation by damaging the gastrointestinal nerves and affecting intestinal peristaltic movement.

[0024] The tyrosine phenol lyase inhibition in the present invention refers to inhibiting tyrosine phenol lyase, that is, suppressing or reducing the enzyme activity of tyrosine phenol lyase, which means that the enzyme activity of tyrosine phenol lyase is suppressed or reduced as compared with a control in a state where the tyrosine phenol lyase inhibitor of the present invention is not added or administered.

[0025] The tyrosine phenol lyase inhibitor of the present invention includes a phenolic hydroxyl group-containing compound represented by the following formula (I), a salt or an ester thereof, or a glycoside thereof.

[0026]

Chemical formula

[0027] In formula (I) representing the above-described compound, examples of the atom represented by X1 include carbon or nitrogen. Among these, from the viewpoint of the inhibitory effect, carbon is preferable as the atom represented by X1. Further, in formula (I), when the atom or molecule represented by R1, which will be described later, is a group represented by formula (II), hydrogen, or a 4-vinylphenol group, the atom represented by X1 is carbon.

[0028] Next, in formula (I) representing the above-described compound, examples of the atom or molecule represented by R2 include a hydroxyl group, hydrogen, or an alkoxy group having 1 to 3 carbon atoms. Examples of the alkoxy group having 1 to 3 carbon atoms include a methoxy group, an ethoxy group, or a propoxy group, etc., and from the viewpoint of the inhibitory effect, a methoxy group is preferable. Among these, from the viewpoint of particularly excellent inhibitory effect, a hydroxyl group or hydrogen is particularly preferable as the atom or molecule represented by R2 described above. Further, in the compound according to formula (I), when the atom or molecule represented by R1, which will be described later, is hydrogen, the atom or molecule represented by R2 is a hydroxyl group or hydrogen. On the other hand, when the atom or molecule represented by R1 is a 4-vinylphenol group, the atom or molecule represented by R2 is hydrogen.

[0029] Next, in formula (I) representing the above-described compound, examples of the atom or molecule represented by R3 include a hydroxyl group or hydrogen. Note that when X1 in formula (I) is nitrogen, R3 does not exist. Further, in the compound according to formula (I), when the atom or molecule represented by R1, which will be described later, is a group represented by formula (II), the atom or molecule represented by R3 is hydrogen. On the other hand, when the atom or molecule represented by R1 is hydrogen, the atom or molecule represented by R3 is a hydroxyl group or hydrogen. Furthermore, when the atom or molecule represented by R1 is a 4-vinylphenol group, the atom or molecule represented by R3 is a hydroxyl group.

[0030] Next, in formula (I) representing the above-described compound, examples of the atom or molecule represented by R4 include hydrogen or a group represented by the following formula (III). Among these, as the atom or molecule represented by R4, hydrogen is particularly preferred from the viewpoint of particularly excellent inhibitory effects. Further, in the compound according to formula (I), when the atom or molecule represented by R1, which will be described later, is a group represented by formula (II), a carboxy group, or a 4-vinylphenol group, the atom or molecule represented by R4 is hydrogen. On the other hand, when the atom or molecule represented by R1 is hydrogen, the atom or molecule represented by R4 is hydrogen or a group represented by formula (III).

[0031]

Chemical formula

[0032] Next, in formula (I) representing the above-described compound, examples of the atom or molecule represented by R1 include a group represented by the following formula (II), a carboxy group, hydrogen, or a 4-vinylphenol group. Among the above-described atoms or molecules represented by R1, a group represented by the following formula (II) or a carboxy group is particularly preferred from the viewpoint of particularly excellent inhibitory effects.

[0033]

Chemical formula

[0034] In the compound according to formula (I), when the atom or molecule represented by R1 is a group represented by formula (II), X1 in formula (I) is carbon, R2 is a hydroxyl group, hydrogen, or an alkoxy group having 1 to 3 carbon atoms, and R3 and R4 are hydrogen. At this time, it is preferable that X1 in formula (I) is carbon, R2 is a hydroxyl group, hydrogen, or a methoxy group, and R3 and R4 are hydrogen, and it is particularly preferable that X1 in formula (I) is carbon, R2 is a hydroxyl group, and R3 and R4 are hydrogen.

[0035] In the compound according to formula (I), when the atom or molecule represented by R1 is a carboxy group, X1 in formula (I) is carbon or nitrogen, R2 is a hydroxyl group, hydrogen or an alkoxy group having 1 to 3 carbon atoms, R3 is a hydroxyl group or hydrogen, and when X1 is nitrogen, R3 does not exist, and R4 is hydrogen. At this time, X1 in formula (I) is preferably carbon, R2 is a hydroxyl group, hydrogen or a methoxy group, R3 is a hydroxyl group or hydrogen, and R4 is hydrogen. Particularly preferably, X1 in formula (I) is carbon, R2 is a hydroxyl group or hydrogen, R3 is a hydroxyl group or hydrogen, and R4 is hydrogen.

[0036] In the compound according to formula (I), when the atom or molecule represented by R1 is hydrogen, X1 in formula (I) is carbon, R2 is a hydroxyl group, R3 is a hydroxyl group, and R4 is hydrogen.

[0037] On the other hand, in the compound according to formula (I), when the atom or molecule represented by R1 is hydrogen, X1 in formula (I) is carbon, R2 is hydrogen, R3 is hydrogen, and R4 is a group represented by formula (III).

[0038] In the compound according to formula (I), when the atom or molecule represented by R1 is a 4-vinylphenol group, X1 in formula (I) is carbon, R2 is hydrogen, R3 is a hydroxyl group, and R4 is hydrogen.

[0039] Moreover, specific compounds represented by the above formula (I) include compounds represented by the following formulas, namely, quercetin, gallic acid (GA), 3-hydroxybenzoic acid (3-HBA), 3,4-dihydroxybenzoic acid (3,4-DHBA), 3,5-dihydroxybenzoic acid (3,5-DHBA), 4-methoxy-3-hydroxybenzoic acid (4M,3-HBA), 5-hydroxynicotinic acid (5-HNA), pyrogallol, resveratrol, and daidzein.

[0040]

Chemical formula

[0041] Among the compounds represented by various formulas, preferred compounds from the viewpoint of enzyme inhibitory effect include quercetin, gallic acid, 3-hydroxybenzoic acid, 3,4-dihydroxybenzoic acid, and 3,5-dihydroxybenzoic acid. As shown in the examples described later, quercetin and 3,5-dihydroxybenzoic acid (3,5-DHBA) are particularly preferred.

[0042] The compound contained in the tyrosine phenol lyase inhibitor according to the present invention may be a salt of a phenolic hydroxyl group-containing compound represented by formula (I), and is preferably a pharmacologically acceptable salt. The pharmacologically acceptable salt of this compound may be a salt formed with an acid or a base, and is not particularly limited.

[0043] Further, the compound contained in the tyrosine phenol lyase inhibitor according to the present invention may be an ester of a phenolic hydroxyl group-containing compound represented by formula (I). For example, in the compound according to formula (I), when the atom or molecule represented by R1 is a carboxy group, the compound according to the present invention can be a carboxylic acid ester. More specifically, as shown in the examples described later, an ester with epicatechin or epigallocatechin is preferred, and an ester of gallic acid with epicatechin or epigallocatechin, that is, epicatechin gallate or epigallocatechin gallate is particularly preferred. Both epicatechin gallate and epigallocatechin gallate are components contained in green tea and are particularly excellent in safety.

[0044] Furthermore, the compound contained in the tyrosine phenol lyase inhibitor according to the present invention may be a glycoside of a phenolic hydroxyl group-containing compound represented by formula (I), and can be a glycoside formed by binding to a sugar via the hydroxyl group of these compounds. Examples of the sugar to be bound include monosaccharides, disaccharides, trisaccharides, oligosaccharides, and polysaccharides. Although not particularly limited, examples of the monosaccharide include glucose, galactose, rhamnose, arabinose, etc., and examples of the disaccharide include rutinose, etc.

[0045] In addition, for the compound contained in the tyrosine phenol lyase inhibitor according to the present invention, that is, the phenolic hydroxyl group-containing compound represented by the formula (I), its salt, its ester, or its glycoside, there may be stereoisomers due to the bonding position of the substituent or the asymmetric carbon atom, but all stereoisomers can be included in the present invention.

[0046] For the phenolic hydroxyl group-containing compound represented by the above formula (I) and its salt, its ester or its glycoside contained in the tyrosine phenol lyase inhibitor according to the present invention, those produced by known synthetic methods can be used, and it is also possible to use commercially available reagent raw materials. In addition, it is also possible to extract each component from edible plant bodies such as tea, spices, fruits, etc. containing these compounds, purify them, etc. and then use them.

[0047] The tyrosine phenol lyase inhibitor of the present invention contains the phenolic hydroxyl group-containing compound represented by the above formula (I) and its salt, its ester or its glycoside, and has an action of inhibiting tyrosine phenol lyase. These compounds are mainly components contained in edible plants such as tea, spices, fruits, etc., and have high safety. In addition, the inhibition constant Ki of the inhibitor is not particularly limited, but for example, the inhibition constant Ki value is preferably 50 μM or less, more preferably 30 μM or less, and even more preferably 20 μM or less. This inhibition constant Ki value can be measured by a known method, for example, it can be obtained by the kinetic analysis of the enzyme activity according to the examples described later. By using the tyrosine phenol lyase inhibitor of the present invention, the production of phenol by tyrosine phenol lyase using tyrosine as a substrate can be suppressed. In addition, these compounds can be used in functional foods, pharmaceutical compositions, etc. for preventing, improving or treating symptoms and diseases in which phenol produced by tyrosine phenol lyase is involved in the intestinal tract, etc.

[0048] In addition, the composition for suppressing phenol production in the intestinal tract of the present invention contains the above-described tyrosine phenol lyase inhibitor as an active ingredient. As a result, since the tyrosine phenol lyase expressed by intestinal bacteria is inhibited, the production of phenol in the intestinal tract can be suppressed. Therefore, this composition for suppressing phenol production can be used as a functional food or pharmaceutical composition for preventing, improving, or treating symptoms and diseases in which phenol in the intestinal tract is involved.

[0049] In addition, the food or drink for reducing the phenol concentration in feces of the present invention contains the above-described tyrosine phenol lyase inhibitor. As a result, since the tyrosine phenol lyase expressed by intestinal bacteria is inhibited, the production of phenol in the intestinal tract is suppressed, and the phenol concentration in feces is reduced. When the phenol in feces is reduced, the fecal odor derived from phenol is also reduced, so the fecal odor can be improved.

[0050] In addition, the food or drink for suppressing skin dryness of the present invention contains the above-described tyrosine phenol lyase inhibitor. As a result, since the tyrosine phenol lyase of intestinal bacteria is inhibited, the production of phenol in the intestinal tract is suppressed, and the amount of phenol transferred from the intestinal tract to the skin is also reduced. Therefore, the hindrance to epidermal formation by phenol is reduced, epidermal formation is promoted, and skin dryness can be suppressed.

[0051] In addition, the food or drink for improving skin barrier function of the present invention contains the above-described tyrosine phenol lyase inhibitor. As a result, since the tyrosine phenol lyase of intestinal bacteria is inhibited, the production of phenol in the intestinal tract is suppressed, and the amount of phenol transferred from the intestinal tract to the skin is also reduced. Therefore, the hindrance to epidermal formation by phenol is reduced, epidermal formation is promoted, and the skin barrier function can be improved.

[0052] In addition, the composition for preventing, improving, or treating symptoms, pathological conditions, or diseases involving phenol in the intestinal tract of the present invention contains the above-described tyrosine phenol lyase inhibitor as an active ingredient. Symptoms, pathological conditions, or diseases involving phenol in the intestinal tract include skin aging, skin dryness, reduced skin barrier function, diarrhea, constipation, fecal odor, uremia (caused by amino acid-derived urinary toxins), and diabetic nephropathy, etc. The composition according to the present invention is used for the treatment, improvement, suppression, or prevention of one or more of these symptoms, pathological conditions, or diseases. The composition for preventing, improving, or treating symptoms, pathological conditions, or diseases involving phenol in the intestinal tract according to the present invention can be used as pharmaceuticals, quasi-drugs, and foods for humans or animals. Foods also include supplements, health foods, functional foods, foods for specified health use, etc.

[0053] When the tyrosine phenol lyase inhibitor of the present invention and the composition containing the same are used as pharmaceuticals or quasi-drugs, they can be prepared in various forms by conventionally used methods. In this case, pharmaceutically acceptable carriers, excipients, lubricants, dispersants, disintegrants, buffers, solvents, extenders, preservatives, fragrances, or stabilizers, etc., additives that are acceptable as pharmaceutical additives, can be used for formulation. Furthermore, in order to improve the bioavailability and stability of this compound, drug delivery systems including formulation technologies such as microcapsules, liposome formulations, micronization, or inclusion using cyclodextrin, etc., can also be used. Also, the tyrosine phenol lyase inhibitor of the present invention and the composition containing the same are preferably made into oral administration formulations and can be used in various forms such as tablets, powders, granules, capsules, or oral liquids. Also, when providing a formulation in a desired form for reasons such as flowability and storage stability, conventional formulation technologies can be used. The dosage or effective intake amount of the tyrosine phenol lyase inhibitor according to the present invention is not particularly limited because it varies depending on the targeted preventive, improving, or therapeutic effect, the administration subject, the dosage form, etc.

[0054] Furthermore, the tyrosine phenol lyase inhibitor of the present invention and the composition containing the same can be used as food and drink. The food and drink composition according to the present invention can be in various forms such as supplements in the form of tablets, capsules, granules, syrups, etc., beverages such as soft drinks, fruit juice drinks, alcoholic beverages, confectioneries such as candies, gums, cookies, biscuits, chocolates, etc., bread, porridge, cereals, noodles, jelly, soup, dairy products, seasonings, etc. When used as food and drink in this way, within the range that does not affect the efficacy of the active ingredient of the present invention, it is also possible to variously combine other active ingredients, probiotics or prebiotics such as vitamins, minerals, oligosaccharides, etc. The food and drink of the present invention includes supplements, health foods, functional foods, foods for specified health use, etc. Also, the daily intake of the food and drink of the present invention is preferably usually 1 μg to 500 mg, more preferably 5 μg to 200 mg, and even more preferably 10 μg to 100 mg as the phenolic hydroxyl group-containing compound and its salts, its esters or their glycosides shown in the above formula (I).

[0055] Next, the present invention will be described in more detail with reference to examples, but the present invention is not limited by these examples.

Examples

[0056] [Example 1] 1. Screening of compounds that inhibit tyrosine phenol lyase (TPL) In this example, a search for compounds that inhibit TPL was conducted. As screening targets, phenolic hydroxyl group-containing compounds including polyphenols contained in edible plants, spices, tea, coffee beverages, etc. were selected. The screening method is specifically as follows.

[0057] (Preparation of screening compounds) The screening compounds were obtained from Tokyo Chemical Industry Co., Ltd. and Sun Chemical Corporation. Also, 2-Aza-Tyr, a known TPL inhibitor used as a positive control, was prepared based on the method by Watkins et al. (Watkins, E. B. et al., Bioorganic & Medicinal Chemistry Letters, 2001, Vol.11, pp.2099-2100).

[0058] (Preparation of Recombinant TPL) For screening, recombinant protein of tyrosine phenol lyase (TPL) from Morganella morganii subsp. morganii, an intestinal bacterium expressing TPL, was used. First, the gene encoding TPL of Morganella morganii subsp. morganii (JCM1672) was inserted into the pET-15b vector. The constructed TPL expression vector was introduced into Escherichia coli BL21(DE3) strain for transformation to obtain a TPL-producing strain derived from Morganella morganii. The obtained TPL-producing strain was inoculated into LB liquid medium for large-scale culture. Specifically, under the condition of 37 °C, shaking culture was carried out until the optical density at 660 nm reached about 0.8, and IPTG (isopropyl-β-D-thiogalactopyranoside) was added to a final concentration of 0.1 mM to induce the expression of TPL protein. After the addition of IPTG, the culture temperature was lowered to 25 °C and shaking culture was carried out overnight. The cultured cells were collected, and the TPL protein was recovered from the cells. The recovery of TPL protein was specifically carried out by the following method. The culture solution was centrifuged to remove the LB liquid medium. After suspension in Buffer I (10 mM HEPES-NaOH pH 8.0, 50 mM NaCl), the cells were disrupted with an ultrasonic homogenizer. This disrupted solution was centrifuged at 4 °C, 20,800×g for 10 minutes to recover the supernatant, and then filtered through a 0.45 μm filter to recover the filtrate. The purification of the filtrate was carried out under the condition of 4 °C. The filtrate was adsorbed onto a HisTrapTM HP column (product of GE Healthcare Japan Corporation), and then linear gradient elution was carried out using a buffer containing 20 mM to 500 mM imidazole. The eluate was applied to an anion exchange column (HiTrap Q column, product of GE Healthcare Japan Corporation), and then linear gradient elution was carried out using a buffer containing 50 mM to 1 M NaCl.The dissolved protein was passed through a gel filtration chromatography column (HiLoad 16 / 600 Superdex200, a product of GE Healthcare Japan Co., Ltd.) and equilibrated with Buffer II (50 mM potassium phosphate buffer pH 8.0, 1 mM dithiothreitol, and 1 mM ethylenediaminetetraacetic acid). This was concentrated to 10 - 15 mg / mL using a centrifugal concentration tube (VIVASPIN, a product of Sartorius Japan K.K.). Thereby, the TPL protein derived from Morganella morganii was obtained. The purified protein was frozen in liquid nitrogen and stored at -80 °C until use.

[0059] (Measurement of TPL inhibitory activity) Using the TPL protein prepared as described above, the TPL inhibitory activity of each screening compound was examined by quantifying the amount of phenol produced when each screening compound was added. This TPL assay was a method that was partially modified from the methods reported by Chen et al. (Chen, H. et al., Eur. J. Biochem., 1995, Vol.229, pp.540 - 549) or Oikawa et al. (Oikawa, D. et al., Biochem. Biophys. Res. Commun. 2022, Vol.590, pp.158 - 162), and specifically was carried out as follows. Among the components of the reaction solution for measuring TPL activity shown in Table 1 below, potassium phosphate, L - tyrosine, and the screening compound were mixed and incubated at 37 °C for 15 minutes. Then, PLP and TPL were added to the reaction solution to initiate the reaction. After incubating at 37 °C for 3 minutes, 50 μL of the reaction stop solution (1N HCl / acetonitrile) was added to terminate the reaction. In addition, as a control, the same reaction was also carried out for a reaction solution containing only the screening compound.

[0060]

Table 1

[0061] The reaction solution was centrifuged at 20,800×g for 10 minutes, and then the supernatant was collected. The phenol concentration in the collected supernatant was quantified using a mass spectrometer (LC-MS; 1290 Infinity II LC system (Agilent Technologies, Inc.), 6410 triple quadrupole mass spectrometer (Agilent Technologies, Inc.)). More specifically, the measurement conditions by the mass spectrometer were as follows.

[0062] · Column... ODS-120H column (1.9 μm, 50 mm×2.0 mm, manufactured by Tosoh Corporation) · Mobile phase... Mobile phase A: water, Mobile phase B: methanol · Flow rate... 0.4 mL / min · Column temperature... 40 °C · Ionization method... Atmospheric pressure chemical ionization (APCI), negative mode · Measurement mode... Single ion monitoring (SIM) · Monitor ion... m / z = 93.0 (phenol), 135.0 (4-isopropylphenol)

[0063] The results are shown in the graph of Figure 1. The vertical axis represents the phenol concentration, and each value is shown as a relative value when the phenol concentration of the control (reaction solution without adding the screening compound) is set to 100. The values are the mean ± standard error of the mean (SEM) (n = 3). According to this result, most of the compounds did not significantly inhibit the TPL of Morganella morganii (JCM1672), but it was confirmed that daidzein, resveratrol, quercetin, epicatechin gallate, epigallocatechin gallate, and gallic acid inhibited TPL. The TPL inhibition rate of 2-Aza-Tyr, which is known as a TPL inhibitor, was 43.4 ± 0.2%, but daidzein, resveratrol, quercetin, epicatechin gallate, epigallocatechin gallate, and gallic acid showed higher TPL inhibition rates than this. Among them, the TPL inhibition rates of gallic acid and quercetin were both about 80%, showing strong TPL inhibitory activity among the screened compounds.

[0064] [Example 2] 2. Examination of the TPL inhibitory activity of quercetin and quercetin analogs In this example, in Example 1, since strong TPL inhibitory activity was confirmed in quercetin, the structure-activity relationship was examined using the following quercetin analogs (taxifolin, quercetol, myricetin, and rutin). These quercetin analogs were obtained from Tokyo Chemical Industry Co., Ltd. Using these, the TPL inhibitory activity was measured in the same manner as in Example 1.

[0065]

Chemical formula

[0066] The results are shown in the graph of Figure 2. The vertical axis indicates the phenol concentration, and each value is shown as a relative value when the phenol concentration of the control is set to 100. The values are the mean ± standard error of the mean (SEM) (n = 3). According to this result, quercetin significantly inhibited TPL, but taxifolin, quercetol, myricetin, and rutin did not show significant inhibition of TPL. Therefore, it became clear that the structure of quercetin specifically inhibits TPL. From this, it was suggested that the hydroxyl group at the 3'-position and the hydrogen at the 5'-position in the flavonol skeleton are important.

[0067] [Example 3] 3. Examination of the inhibition mode and inhibition constant Ki value of the TPL inhibitor (1) In this example, in order to investigate the TPL inhibition mode of quercetin, kinetic analysis of enzyme activity was performed. The measurement of TPL inhibitory activity was carried out in the same manner as in Examples 1 and 2, except that the concentrations of quercetin, which is a TPL inhibitor, and L-tyrosine, which is a substrate, were changed. The inhibition mode was determined by double reciprocal plots (Lineweaver - Burk plots) of the initial velocity data in the presence and absence of the inhibitor. The Ki value was determined using statistical analysis software (GraphPad Prism 9, GraphPad Software). Also, as a comparative control, the same test was conducted for 2 - Aza - Tyr, which is known as a TPL inhibitor, to determine the inhibition mode and the inhibition constant Ki value.

[0068] The Lineweaver - Burk plot of quercetin is shown in Fig. 3. The values are mean ± standard error of the mean (SEM) (n = 3). According to this result, it was shown that quercetin inhibits TPL in a dose - dependent manner. Also, from the kinetic analysis of quercetin, its inhibition mode was competitive inhibition, and the Ki value was 19.9 μM. On the other hand, when 2 - Aza - Tyr, which is known as a TPL inhibitor, was analyzed, as reported previously, it showed dose - dependent inhibition and competitive inhibition, and the Ki value was 42.0 μM (not shown). From this result, it became clear that quercetin has a smaller Ki value of the inhibition constant than 2 - Aza - Tyr, and quercetin has stronger TPL inhibitory activity than 2 - Aza - Tyr.

[0069] [Example 4] 4. Examination of the effect of quercetin on phenol production by TPL - expressing bacteria (1) Using Morganella morganii subsp. morganii (JCM1672) and Citrobacter koseri (JCM1658), which are intestinal bacteria expressing TPL, the effect of quercetin on phenol production by these TPL - expressing bacteria was examined. The specific test method is as follows.

[0070] First, pre-cultures of Morganella morganii (JCM1672) and Citrobacter koseri (JCM1658) were performed. Each bacterium was pre-cultured in trypticase soy broth for 24 hours, and the optical density OD 600 of the culture solution was measured. This pre-culture solution was centrifuged at 5000×g at 4°C for 10 minutes, and the supernatant was removed. To wash the cell pellet, 500 μL of M9 salts medium was added to the cell pellet and resuspended, and the step of centrifuging at 5000×g at 4°C for 10 minutes to remove the supernatant was repeated twice.

[0071] Next, quercetin was added to the M9 salts medium so that the final concentration of quercetin was 20 μM or 100 μM to prepare a test M9 medium. Also, as a solvent control group, DMSO was added to the M9 salts medium to form a 0 μM group. Each cell pellet washed after pre-culture was resuspended in these test M9 media, respectively, to reach a predetermined optical density OD 600 . This suspension was incubated at 37°C, and after 30 minutes, 1 hour, and 3 hours, the suspension was collected respectively. To the collected suspension, an equivalent amount of 50 μM 4-isopropylphenol solution, twice the amount of 2N HCl, and six times the amount of ethyl acetate were added, and vortexed for 10 minutes. The mixture was centrifuged at 5000×g at 4°C for 10 minutes, and the organic phase was collected in a new tube. After removing the organic solvent from the organic phase, the residue was dissolved in 20% methanol, and the phenol concentration in the solution was quantified by a mass spectrometer. The measurement conditions by the mass spectrometer were the same as in Example 1.

[0072] The results are shown in the graph of Figure 4. The vertical axis represents the phenol concentration (μM), and each value is the mean ± standard error of the mean (SEM) (n = 3). According to this result, in the test group with a quercetin concentration of 20 μM in the medium, for both bacteria, Morganella morganii (JCM1672) and Citrobacter koseri (JCM1658), at any culture time (30 minutes, 1 hour, 3 hours), the phenol concentration did not decrease significantly. However, in the test group with a quercetin concentration of 100 μM in the medium, the phenol concentration decreased significantly at all culture times.

[0073] [Example 5] 5. Examination of the Effect of Quercetin on TPL Expression in TPL-Expressing Bacteria (1) In Example 4, it was shown that the addition of quercetin decreased the phenol concentration in the culture broth of TPL-expressing bacteria. However, it is also conceivable that this decrease in phenol concentration might have been caused by the inhibition of TPL expression. Therefore, in order to clarify whether the decrease in phenol concentration due to the addition of quercetin was due to the inhibition of the TPL enzyme by quercetin or due to the inhibition of TPL expression, the TPL expression level in TPL-expressing bacteria was examined. The specific test method is as follows.

[0074] In Example 4, Western blot analysis was performed on the suspension obtained by incubating each bacterium in the test M9 medium for 3 hours to analyze the TPL expression level. Specifically, the suspension was mixed with SDS sample buffer, and this mixture was heated at 100°C for 5 minutes, followed by SDS-polyacrylamide gel electrophoresis. The proteins separated in the gel were transferred to a polyvinylidene fluoride (PVDF) membrane. This PVDF membrane was incubated with a blocking agent (Blocking one, manufactured by Nacalai Tesque, Inc.) at room temperature for 60 minutes. Thereafter, the anti-TPL primary antibody was added and the treatment was carried out overnight at 4°C, and the corresponding horseradish peroxidase-labeled secondary antibody was added and incubated at 4°C for 1 hour. The anti-TPL primary antibody used was a monoclonal antibody prepared from a hybridoma obtained by immunizing BALB / c mice with the recombinant TPL of Morganella morganii (JCM1672) prepared in Example 1, fusing the spleen and lymph nodes obtained therefrom with P3U1 cells. The protein bands were visualized using a peroxidase luminescent substrate (ImmunoStar (registered trademark) LD, manufactured by Fujifilm Wako Pure Chemical Corporation) and detected with a luminescence imaging device (LuminoGraph I, manufactured by Atto Corporation). The intensity of the specific bands was determined using image analysis software (Image J, National Institutes of Health, USA).

[0075] The results are shown in the graph of Fig. 5. The vertical axis indicates the level of TPL expression, shown as the relative value when the TPL expression level of the control (0 μM group with DMSO added) is set to 100. The values are the mean ± standard error of the mean (SEM) (n = 3). This result shows the TPL expression level in bacteria cultured for 3 hours with the addition of quercetin. Regarding Morganella morganii (JCM1672), inhibition of TPL expression by the addition of quercetin was not confirmed. Furthermore, in the case of Citrobacter koseri (JCM1658), it was confirmed that the addition of quercetin rather promoted TPL expression. Additionally, it was examined whether quercetin affects the growth of each bacterium, but it was found that it did not affect the growth of any of the strains. From these results, it became clear that the decrease in phenol concentration by the addition of quercetin confirmed in Example 4 is due to the inhibition of the TPL enzyme by quercetin.

[0076] [Example 6] 6. Examination of the TPL inhibitory activity of gallic acid and gallic acid analogs In this example, in Example 1, since high TPL inhibitory activity was confirmed for gallic acid, the structure-activity relationship was examined using analogs of gallic acid shown in the following chemical formula. These gallic acid analogs were obtained from Tokyo Chemical Industry Co., Ltd. Using these, the TPL inhibitory activity was measured in the same manner as in Example 1.

[0077] [Chemical formula]

[0078] [Chemical formula]

[0079] The results are shown in the graph of Fig. 6. The vertical axis indicates the phenol concentration, and each value is shown as a relative value when the phenol concentration of the control is set to 100. The values are the mean ± standard error of the mean (SEM) (n = 3). According to these results, it was found that gallic acid (GA: 3,4,5-trihydroxybenzoic acid), 3-hydroxybenzoic acid (3-HBA), 3,4-dihydroxybenzoic acid (3,4-DHBA), 3,5-dihydroxybenzoic acid (3,5-DHBA), 4-methoxy-3-hydroxybenzoic acid (4M,3-HBA), 5-hydroxynicotinic acid (5-HNA), and pyrogallol significantly inhibited TPL. Among these, 3,5-dihydroxybenzoic acid (3,5-DHBA) showed the strongest inhibitory activity. From these results, it was shown that among the structures of hydroxybenzoic acids, especially the hydroxyl group at the 3-position is important for TPL inhibition.

[0080] [Example 7] 7. Examination of the Inhibition Mode and Inhibition Constant Ki Value of the TPL Inhibitor (2) In this example, in order to examine the TPL inhibition mode of 3,5-dihydroxybenzoic acid, which showed the strongest inhibitory activity in Example 6, a kinetic analysis of the enzyme activity was performed. While changing the concentrations of 3,5-dihydroxybenzoic acid, which is a TPL inhibitor, and L-tyrosine, which is a substrate, the TPL inhibitory activity was measured in the same manner as in Example 3. The inhibition mode was determined by a double-reciprocal plot (Lineweaver-Burk plot) of the initial velocity data in the presence and absence of the inhibitor. The Ki value was determined using statistical analysis software (GraphPad Prism 9, GraphPad Software).

[0081] Figure 7 shows the Lineweaver - Burk plot of 3,5 - dihydroxybenzoic acid (3,5 - DHBA). The values are mean ± standard error of the mean (SEM) (n = 3). According to this result, it was shown that 3,5 - DHBA inhibits TPL in a dose - dependent manner. Also, from the enzyme kinetic analysis of 3,5 - DHBA, its inhibition mode was competitive inhibition, and the Ki value showed a very small value of 13.0 μM. From this result, it became clear that 3,5 - DHBA has a stronger TPL inhibitory activity than 2 - Aza - Tyr (Ki value: 42.0 μM), which is a known TPL inhibitor.

[0082] [Example 8] 8. Examination of the effect of 3,5 - dihydroxybenzoic acid (3,5 - DHBA) on phenol production by TPL - expressing bacteria (2) Using Morganella morganii subsp. morganii (JCM1672) and Citrobacter koseri (JCM1658), which are intestinal bacteria expressing TPL, the effect of 3,5 - dihydroxybenzoic acid (3,5 - DHBA) on phenol production by these TPL - expressing bacteria was examined. The experimental method was the same as in Example 4, except that the final concentration of 3,5 - DHBA added to the test M9 medium was formulated to be 100 μM or 500 μM.

[0083] The results are shown in the graph of Figure 8. The vertical axis indicates the phenol concentration (μM), and each value is mean ± standard error of the mean (SEM) (n = 3). According to this result, in the test group with a 3,5 - DHBA concentration of 500 μM in the medium, for both bacteria, Morganella morganii (JCM1672) and Citrobacter koseri (JCM1658), the phenol concentration significantly decreased at all culture times (30 minutes, 1 hour, 3 hours). Also, for Citrobacter koseri (JCM1658), the phenol concentration significantly decreased at all culture times even in the test group with a 3,5 - DHBA concentration of 100 μM in the medium.

[0084] [Example 9] 9. Examination of the effect on TPL expression in TPL-expressing bacteria (2) In Example 8, it was shown that the addition of 3,5-dihydroxybenzoic acid (3,5-DHBA) decreased the phenol concentration in the culture broth of TPL-expressing bacteria. However, it is also conceivable that this decrease in phenol concentration might have been caused by the inhibition of TPL expression. Therefore, to clarify whether the decrease in phenol concentration due to the addition of 3,5-DHBA was due to the inhibition of the TPL enzyme or the inhibition of TPL expression, the amount of TPL expression in TPL-expressing bacteria was examined. The experimental method was the same as that in Example 5 described above.

[0085] The results are shown in the graph of Fig. 9. The vertical axis indicates the level of TPL expression, and it is shown as the relative value when the TPL expression level of the control (0 μM group with DMSO added) is set to 100. The values are the mean ± standard error of the mean (SEM) (n = 3). This result shows the amount of TPL expression in bacteria cultured for 3 hours after the addition of 3,5-DHBA. Looking at Morganella morganii (JCM1672), no inhibition of TPL expression due to the addition of 3,5-DHBA was confirmed. Furthermore, in the case of Citrobacter koseri (JCM1658), it was confirmed that the addition of 3,5-DHBA rather increased the amount of TPL expression. From these results, it became clear that the decrease in phenol concentration due to the addition of 3,5-DHBA confirmed in Example 8 was due to the inhibition of the TPL enzyme by 3,5-DHBA.

[0086] [Example 10] 10. Examination of the phenol concentration in feces by administration of a TPL inhibitor In this example, as the TPL inhibitor of the present invention, quercetin or 3,5-dihydroxybenzoic acid (3,5-DHBA) was orally administered to mice, and the amount of phenol in mouse feces was quantified.

[0087] (Quercetin feeding test) The test method for quercetin is as follows. Twelve C57BL / 6 male mice (purchased from Japan SLC, Inc.) were used and individually housed at one mouse per cage. First, a one-week pre-breeding period was set before the start of the test, and each mouse was fed a standard purified diet (AIN-93G, a product of Nihon Clea, Inc.). Taking the start of the test as day 0, feces (Day0 feces) were collected from the cage until 24 hours had passed after the start of the test. For two days from the start of the test, a high-tyrosine diet with 5% L-tyrosine added to the diet fed during the pre-breeding period was fed. From the mice that had ingested the high-tyrosine diet for two days, feces (Day2) were collected until 24 hours had passed after two days of the test. Subsequently, from the third day after the start of the test, a test diet with 5% L-tyrosine and 0.2% quercetin added to the diet fed during the pre-breeding period was fed for two days. From the mice that had ingested the test diet for 48 hours (two days), feces (Day4) were collected until 24 hours had passed after four days of the test. 100 mg each of the collected Day0 feces, Day2 feces, and Day4 feces were taken and solvent-extracted, and the phenol concentration in the extract was quantified using a mass spectrometer. The measurement conditions using the mass spectrometer were the same as in Example 1.

[0088] (3,5-DHBA Feeding Test) For the other party, the test method for 3,5-DHBA is as follows. Eight C57BL / 6J male mice (purchased from Japan SLC, Inc.) were used and individually housed at one mouse per cage. First, a one-week pre-breeding period was set before the start of the test, and each mouse was fed with breeding feed CE-2 (product of Nihon Clea, Inc.). The start of the test was set as day 0, and feces (Day0 feces) were collected from the cage until 24 hours after the start of the test. From day 1 to 14 of the test, a high-tyrosine diet with 5% L-tyrosine added to the standard purified diet (AIN-93G, product of Nihon Clea, Inc.) was fed. From the mice that had ingested the high-tyrosine diet for 14 days, feces (Day14) were collected until 24 hours after 14 days of the test. Subsequently, from the 15th day after the start of the test, a test diet with 5% L-tyrosine and 0.2% quercetin added to the standard purified diet (AIN-93G) was fed for 2 days. From the mice that had ingested the test diet for 48 hours (2 days), feces (Day16) were collected until 24 hours after 16 days of the test. 100 mg each of the collected Day0 feces, Day14 feces, and Day16 feces were taken and solvent-extracted, and the phenol concentration in the extract was quantified using a mass spectrometer. The measurement conditions by the mass spectrometer were the same as in Example 1.

[0089] The results are shown in the graph of FIG. 10. FIG. 10(a) shows the phenol concentration in the feces of mice that ingested quercetin, and FIG. 10(b) shows the phenol concentration in the feces of mice that ingested 3,5-DHBA. The vertical axis shows the phenol concentration (nmol) per gram of feces. According to this result, it was revealed that the amount of phenol in feces significantly increases by ingesting a high-tyrosine diet, but the amount of phenol in feces is reduced to the level of the control (Day0) by ingesting quercetin or 3,5-DHBA together. From this, it became clear that by orally ingesting the TPL inhibitor according to the present invention, such as quercetin or 3,5-DHBA, TPL in intestinal bacteria is inhibited, and phenol production in the intestinal tract is suppressed.

[0090] [Example 11] 11. Examination of the effect of TPL inhibitor administration on skin function In this example, HR-1 hairless mice were used, and as the TPL inhibitor of the present invention, quercetin or 3,5-dihydroxybenzoic acid (3,5-DHBA) was orally administered to the mice to examine the effect on mouse skin function. The test method is specifically as follows.

[0091] Eight female Hos:HR-1 mice (purchased from Japan SLC, Inc.) were used and individually housed, one mouse per cage. First, a one-week pre-breeding period was set before the start of the test, and each mouse was fed with breeding feed CE-2 (product of Nippon Clea Co., Ltd.). The eight mice were divided into four groups: a control group, an L-Tyr group, a 3,5-DHBA group, and a quercetin group, with two mice in each group. After the start of the test, the control group was fed a standard purified diet (AIN-93G, product of Nippon Clea Co., Ltd.), the L-Tyr group was fed a high-tyrosine diet with 5% L-tyrosine added to the same diet (AIN-93G), the 3,5-DHBA group was fed a test diet with 5% L-tyrosine and 0.2% 3,5-DHBA added to the same diet (AIN-93G), and the quercetin group was fed a test diet with 5% L-tyrosine and 0.2% quercetin added to the same diet (AIN-93G) for 7 days each. During the test period, each mouse was housed in a metabolic cage to minimize the impact of feces and urine adhering to the skin of the hairless mice.

[0092] (Measurement of stratum corneum water content) The stratum corneum water content refers to the amount of water contained at a depth of approximately 15 μm from the skin surface (mainly the stratum corneum), and it is evaluated that the drier the skin, the lower the stratum corneum water content. On the 8th day after the start of the test at the end of the test period, under isoflurane inhalation anesthesia, the stratum corneum water content near the left foot of the mouse was measured using a stratum corneum moisture meter (corneometer, manufactured by Courage+Khazaka). The stratum corneum water content measured in this test is the value measured as the amount of water contained at a depth of approximately 15 μm from the skin surface by the capacitance method (electrostatic capacitance method) using the above-mentioned stratum corneum moisture meter (corneometer, manufactured by Courage+Khazaka).

[0093] (Measurement of transepidermal water loss (TEWL)) The skin barrier function generally refers to the suppression of transepidermal water loss, and it is evaluated that the smaller the amount of transepidermal water loss (TEWL), the better the skin barrier function. On the 8th day from the start of the test after the end of the test period, under isoflurane inhalation anesthesia, the transepidermal water loss (TEWL) near the left hind leg of the mouse was measured using a transepidermal water loss meter (Tewameter, manufactured by Courage+Khazaka).

[0094] The results are shown in the graph of Fig. 11. Fig. 11(a) shows the measurement results of the stratum corneum water content, and Fig. 11(b) shows the measurement results of the transepidermal water loss. According to these results, compared with the control, it was confirmed that the stratum corneum water content of the skin of the mice fed a high-tyrosine diet decreased and became dry, and the transepidermal water loss (TEWL) increased, resulting in a decrease in the skin barrier function. On the other hand, by ingesting quercetin together, it was confirmed that the stratum corneum water content of the skin became higher than that of the control, the dryness was improved, the transepidermal water loss (TEWL) also became smaller than that of the control, and the skin barrier function was improved. In addition, by ingesting 3,5-DHBA together, it was confirmed that the stratum corneum water content of the skin became comparable to that of the control, the dryness was improved, and the transepidermal water loss (TEWL) also became smaller than that of the control, and the skin barrier function was improved. From this, it became clear that by orally ingesting the TPL inhibitor according to the present invention, such as quercetin or 3,5-DHBA, skin dryness can be suppressed and the skin barrier function can also be improved.

[0095] The present invention is not limited to the above-described embodiments or examples, and various forms with design changes within the scope not departing from the gist of the invention described in the claims are also included in the technical scope.

Industrial Applicability

[0096] The present invention provides a tyrosine phenol lyase inhibitor capable of inhibiting the production of phenol from tyrosine, and is widely useful in industries in fields such as functional foods and pharmaceuticals.

Claims

1. A tyrosine phenol lyase inhibitor comprising a phenolic hydroxyl group-containing compound represented by the following formula (I), a salt thereof, an ester thereof, or a glycoside thereof. 【Chemical 1】 [In formula (I), X1 represents carbon or nitrogen; R1 represents a group represented by the following formula (II), a carboxy group, hydrogen, or a 4-vinylphenol group; R2 represents a hydroxyl group, hydrogen, or an alkoxy group having 1 to 3 carbon atoms; R3 represents a hydroxyl group or hydrogen, and when X1 is nitrogen, R3 does not exist; R4 represents hydrogen or a group represented by the following formula (III).] [Chemical Formula 2] However, when R1 is a group represented by the formula (II), X1 is carbon, and R3 and R4 are hydrogen; when R1 is a carboxy group, R4 is hydrogen; when R1 is hydrogen, X1 is carbon, R2 and R3 are hydroxyl groups, and R4 is hydrogen, or X1 is carbon, R2 and R3 are hydrogen, and R4 is a group represented by the formula (III); when R1 is a 4-vinylphenol group, X1 is carbon, R2 and R4 are hydrogen, and R3 is a hydroxyl group.

2. In formula (I), X1 represents carbon; R1 represents a group represented by the formula (II) or a carboxy group; R2 represents a hydroxyl group or hydrogen; R3 represents a hydroxyl group or hydrogen; R4 represents hydrogen; However, when R1 is a group represented by the formula (II), R3 is hydrogen. The tyrosine phenol lyase inhibitor according to claim 1, characterized by this.

3. The phenolic hydroxyl group-containing compound represented by formula (I) is at least one compound selected from the group consisting of quercetin, gallic acid, 3-hydroxybenzoic acid, 3,4-dihydroxybenzoic acid, 3,5-dihydroxybenzoic acid, 4-methoxy-3-hydroxybenzoic acid, 5-hydroxynicotinic acid, pyrogallol, resveratrol, and daidzein. The tyrosine phenol lyase inhibitor according to claim 1, characterized by this.

4. The ester of the phenolic hydroxyl group-containing compound represented by formula (I) is an ester of gallic acid, and the ester of gallic acid is epicatechin gallate or epigallocatechin gallate. The tyrosine phenol lyase inhibitor according to claim 1, characterized by this.

5. A composition for inhibiting the production of phenol in the intestinal tract, characterized by containing the tyrosine phenol lyase inhibitor according to any one of claims 1 to 4.

6. A food or drink for reducing the phenol concentration in feces, characterized by containing the tyrosine phenol lyase inhibitor according to any one of claims 1 to 4.

7. A food or drink for suppressing skin dryness, characterized by containing the tyrosine phenol lyase inhibitor according to any one of claims 1 to 4.

8. A food or drink for improving the skin barrier function, characterized by containing the tyrosine phenol lyase inhibitor according to any one of claims 1 to 4.

9. A composition for preventing, improving or treating symptoms, conditions or diseases involving phenol in the intestinal tract, characterized by containing the tyrosine phenol lyase inhibitor according to any one of claims 1 to 4 as an active ingredient.

10. The composition according to claim 9, wherein the symptoms, conditions or diseases involving phenol in the intestinal tract are at least one symptom, condition or disease selected from the group consisting of skin aging, diarrhea, constipation, uremia, fecal odor and diabetic nephropathy.

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