Composition for promoting TPH2 gene expression

Abstract

To provide an oral composition for promoting serotonin synthesis.SOLUTION: An oral composition for promoting serotonin synthesis comprises 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid (GMC) as an active ingredient. This oral composition exhibits superior activity in promoting serotonin synthesis and is applicable as a functional food or a supplement.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to an oral composition for promoting serotonin synthesis. [Background technology]

[0002] Serotonin, along with dopamine and noradrenaline, is one of the three major brain substances known to enhance neutrophil migration, balance the autonomic nervous system, and relieve tension. Diseases in which serotonin is involved include depression, panic disorder, insomnia, anxiety, social anxiety disorder, and obsessive-compulsive disorder.

[0003] Cortisol is a hormone secreted from the renal cortex, and its secretion from the adrenal cortex is known to increase in response to external environmental factors such as psychological stress and ultraviolet rays. Diseases in which cortisol is involved include mental disorders such as depression and insomnia, and stress-related diseases such as lifestyle-related diseases.

[0004] As such, a deficiency of serotonin and an increase in cortisol levels can cause various diseases, and therefore research into the components related to the secretion of serotonin and cortisol is ongoing. For example, lactoferrin (Patent Document 1), lactic acid bacteria belonging to the genus Lactobacillus other than Lactobacillus delbrueckii, and lactic acid bacteria belonging to the genus Pediococcus, or processed products thereof (Patent Document 2) have been reported to have the effect of promoting serotonin synthesis. Furthermore, lactic acid bacteria belonging to the genus Lactococcus (Patent Document 3) and black reishi mushroom extract (Patent Document 4) have been reported to have the effect of suppressing cortisol production. [Prior art documents] [Patent documents]

[0005] [Patent Document 1] Japanese Patent Application Publication No. 2019-156814 [Patent Document 2] Japanese Patent Application Laid-Open No. 2017-061567 [Patent Document 3] Patent Publication No. 2021-153461 [Patent Document 4] Japanese Patent Application Laid-Open No. 2004-307425 Summary of the Invention [Problem to be solved by the invention]

[0006] As mentioned above, various ingredients have been reported to have the effect of promoting serotonin synthesis and suppressing cortisol production, but the search for more effective ingredients is ongoing.

[0007] Under these circumstances, an object of the present invention is to provide a novel oral composition for promoting serotonin synthesis. [Means for solving the problem]

[0008] The present inventors have conducted extensive research to solve the above problems, and as a result have found that the following invention meets the above object, thereby completing the present invention.

[0009] That is, the present invention relates to the following inventions. <1> An oral composition for promoting serotonin synthesis, containing 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid as an active ingredient. <2> It is a functional food or supplement <1> The oral composition according to claim 1. [Effects of the Invention]

[0010] According to the present invention, an oral composition for promoting serotonin synthesis can be provided. [Brief explanation of the drawings]

[0011] [Figure 1]1 is a graph showing the results of an evaluation test of 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid (cis isomer, trans isomer) on serotonin synthesis. [Figure 2] 1 is a graph ((a) cis isomer, (b) trans isomer) showing the results of an evaluation test of 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid (cis isomer, trans isomer) on cortisol production. DETAILED DESCRIPTION OF THE INVENTION

[0012] The present invention will be described in detail below with reference to examples, but the present invention is not limited to the examples below and can be practiced with any modifications within the scope of the gist of the present invention. In this specification, the symbol "to" is used as an expression including the numerical value or physical quantity before and after it.

[0013] The present invention relates to an oral composition for promoting serotonin synthesis containing 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid (hereinafter sometimes referred to as "GMC") as an active ingredient (hereinafter referred to as "the oral composition for promoting serotonin synthesis of the present invention").

[0014] The present invention relates to an oral composition for inhibiting cortisol production, which contains 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid as an active ingredient (hereinafter referred to as "the oral composition for inhibiting cortisol production of the present invention").

[0015] In the following description, when there is no need to distinguish between the "oral composition of the present invention for promoting serotonin synthesis" and the "oral composition of the present invention for suppressing cortisol production," they will be collectively referred to as the "oral composition of the present invention."

[0016] 2-β-D-Glucopyranosyloxy-4-methoxycinnamic acid (GMC) is classified into cis and trans isomers based on the relative arrangement of the carboxyl and phenyl groups attached to the double bond in the cinnamic acid skeleton. GMC (cis and trans isomers) will be discussed later.

[0017] In this specification, GMC (cis isomer) and GMC (trans isomer) are collectively referred to as "GMC", and when distinguishing between them, they are referred to as cis-GMC and trans-GMC, respectively.

[0018] GMC promotes the synthesis of serotonin by promoting the expression of genes related to serotonin synthesis, and therefore can be used as an active ingredient in oral compositions for promoting serotonin synthesis.

[0019] The serotonin synthesis promoting effect can be determined by evaluating the expression of the TPH2 gene, a serotonin synthase, and specifically, can be evaluated by the evaluation method described in the Examples below, <1. Evaluation of serotonin synthesis promoting effect>. As shown in the Examples, both cis-GMC and trans-GMC have excellent effects on promoting serotonin synthesis, with cis-GMC having a particularly excellent effect.

[0020] Serotonin is a neurotransmitter in the brain that balances the autonomic nervous system and relieves tension. A serotonin deficiency can lead to irritability, loss of ambition, loss of motivation at work, depression, and insomnia.

[0021] Furthermore, GMC has the effect of suppressing cortisol production by suppressing the expression of genes related to cortisol production, and therefore GMC can be used as an active ingredient in oral compositions for suppressing cortisol production.

[0022] The cortisol production inhibitory effect can be determined by evaluating the expression of the 11β-HSD1 gene, a cortisol-producing enzyme, and specifically, can be evaluated by the evaluation method described in the Examples below, <2. Evaluation of cortisol production inhibitory effect>. As shown in the Examples, cis-GMC has an excellent effect on cortisol production inhibition.

[0023] Cortisol is a hormone secreted by the adrenal cortex, and its secretion increases due to external environmental factors such as psychological stress and ultraviolet rays. Chronically elevated cortisol levels can contribute to stress-related disorders such as insomnia and depression.

[0024] The oral composition for promoting serotonin synthesis and the oral composition for suppressing cortisol production of the present invention exert a serotonin synthesis promoting effect and a cortisol production inhibiting effect when orally taken, and therefore can be used as a functional food or supplement for promoting serotonin synthesis or inhibiting cortisol production. The oral composition for promoting serotonin synthesis and the oral composition for suppressing cortisol production of the present invention are useful for at least one of preventing and ameliorating symptoms (stress, depression, insomnia, low mood, etc.) caused by a deficiency of serotonin or an excess secretion of cortisol, based on their serotonin synthesis-promoting effect and cortisol production-suppressing effect. "Prevention" includes suppressing and delaying the onset of the symptoms.

[0025] The oral composition of the present invention will be described in detail below.

[0026] (2-β-D-glucopyranosyloxy-4-methoxycinnamic acid) 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid (GMC) is a compound represented by the following chemical formula (a) or (b):

[0027] The name of the compound (cis-GMC) represented by the following chemical formula (a) is (Z)-2-glucopyranosyloxy-4-methoxycinnamic acid (English name: cis-2-β-D-glucopyranosyloxy-4-methoxycinnamic acid).

[0028] [ka]

[0029] The name of the compound (trans-GMC) represented by the following chemical formula (b) is (E)-2-glucopyranosyloxy-4-methoxycinnamic acid (English name: trans-2-β-D-glucopyranosyloxy-4-methoxycinnamic acid).

[0030] [ka]

[0031] The above chemical formula (a) (cis-GMC) represents the Z-isomer in which the carboxyl group and phenyl group bonded to the double bond of the cinnamic acid skeleton are both in the cis position relative to each other, and the above chemical formula (b) (trans-GMC) represents the E-isomer in which the carboxyl group and phenyl group bonded to the double bond of the cinnamic acid skeleton are both in the trans position relative to each other.

[0032] The GMC contained as an active ingredient in the oral composition of the present invention may be chemically synthesized or derived from natural sources such as plants containing GMC. Alternatively, commercially available GMC may be used.

[0033] The oral composition for promoting serotonin synthesis of the present invention may contain either cis-GMC or trans-GMC, or may contain both, as long as it exhibits the effect of promoting serotonin synthesis. When both are contained, the content of cis-GMC and trans-GMC can be set to any ratio as long as the serotonin synthesis promoting effect is exhibited.

[0034] Furthermore, the total content of GMC in the oral composition for promoting serotonin synthesis of the present invention is appropriately set within a range that achieves the effects of the present invention, and may be appropriately selected depending on the use of the oral composition for promoting serotonin synthesis of the present invention.

[0035] The GMC in the oral composition for inhibiting cortisol production of the present invention may contain only cis-GMC, which has a cortisol production inhibitory effect, or may contain both cis-GMC and trans-GMC. When both are contained, the contents of cis-GMC and trans-GMC can be set to any ratio as long as the cortisol production inhibitory effect is exhibited.

[0036] Furthermore, the total content of GMC in the oral composition for inhibiting cortisol production of the present invention is appropriately set within a range that achieves the effects of the present invention, and may be appropriately selected depending on the use of the oral composition for inhibiting cortisol production of the present invention.

[0037] The oral composition of the present invention may contain 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid (GMC) as an active ingredient, but is usually used in combination with other ingredients that do not impair the effects of the present invention. The other ingredients may be any ingredients that can be safely orally ingested by humans, and are appropriately determined taking into consideration the form of use (for example, supplements, functional foods, etc.) described below. The blending ratio of the other components can be appropriately selected and determined depending on the purpose as long as it does not impair the effects of the present invention.

[0038] The oral composition of the present invention has excellent serotonin synthesis promoting effects and cortisol production inhibiting effects, and therefore can be used in supplements, functional foods, etc. for promoting serotonin synthesis or inhibiting cortisol production.

[0039] When the oral composition of the present invention is used to promote serotonin synthesis, the amount of GMC contained therein is appropriately determined within a range that allows the necessary amount to be ingested, taking into consideration the type and severity of symptoms related to serotonin synthesis, the form of the oral composition of the present invention, and the method of use.

[0040] When the oral composition of the present invention is used to suppress cortisol production, the amount of GMC contained therein is appropriately determined within a range that allows the necessary amount to be ingested, taking into consideration the type and severity of symptoms related to cortisol production, the form of the oral composition of the present invention, and the method of use.

[0041] The intake amount of the oral composition of the present invention is not particularly limited as long as it is an amount that can exert the effects of the present invention, and is appropriately determined depending on the form of the oral composition of the present invention, and the sex, weight, health condition, etc. of the subject to be ingested. The intake amount may be taken in one dose or in divided doses several times a day. In addition, in order to obtain the action or efficacy of the oral composition of the present invention, it is preferable to take the oral composition of the present invention continuously.

[0042] The following describes suitable forms of the oral composition of the present invention, such as supplements, functional foods, and food additives, but the uses of the oral composition of the present invention are not limited to these.

[0043] The oral composition of the present invention can be mixed with various foods and beverages to form functional foods that can be easily taken orally on a daily basis, and can also be easily taken over a long period of time. The term "functional foods" as used here includes not only general foods but also foods and / or beverages that are taken for the purpose of maintaining or improving health, such as foods for special dietary uses, foods for specified health uses, foods with functional claims, foods with nutrient functions, nutritional supplements, health supplements, and supplements. Among these, foods with functional claims that are health functional foods are the preferred embodiment. Foods with functional claims include foods that claim functionality or effects related to one or more selected from the group consisting of stress, depression, insomnia, low mood, menopausal symptoms, PMS, and prevention and improvement of PMDD.

[0044] The form of the oral composition of the present invention is not particularly limited, and may be any of a tablet, hard capsule, soft capsule, powder, granules, particulate, liquid, rod, plate, block, solid, round, paste, cream, gel, sol, caplet, chewable, stick, jelly, etc.

[0045] When commercializing the oral composition of the present invention, various additives used in foods and beverages, specifically coloring agents, preservatives, thickening stabilizers, antioxidants, bleaching agents, antibacterial and antifungal agents, acidulants, seasonings, emulsifiers, strengthening agents, manufacturing agents, fragrances, etc. may be added.

[0046] The foods and beverages that can be used as functional foods are not particularly limited. Examples of foods include bread, candy, drops, jellies, gummies, chewing gum, and other confectioneries, seasonings, and retort pouch foods. Examples of beverages include various types of tea, soft drinks, and nutritional drinks. Examples of tea include packaged beverages filled in PET bottles, cans, jars, etc., instant beverages that can be dissolved in water or hot water, and tea bags.

[0047] The ratio of GMC to be added to functional foods is arbitrary, but the ratio is selected within a range that can contribute to the promotion of serotonin synthesis and the inhibition of cortisol production.

[0048] The oral composition (GMC) of the present invention can also be used in the form of a supplement. The form of the supplement is not particularly limited, and may be any form such as a tablet, powder, powder, granule, capsule (hard capsule, soft capsule), sugar-coated tablet, film, lozenge, jelly, chewable tablet, paste, gummy, agar, solution, emulsion, suspension, etc. The supplement of the present invention may contain any ingredients commonly used in supplements in addition to the composition of the present invention. Such ingredients include, for example, amino acids, peptides, vitamins such as vitamin E, vitamin C, vitamin A, vitamin B, and folic acid, minerals, sugars, inorganic salts, citric acid or its salts, tea extract, oils and fats, tonic ingredients such as propolis, royal jelly, and taurine, herbal extracts such as ginger extract and ginseng extract, and herbs such as collagen.

[0049] The ratio of GMC to be mixed into a supplement is arbitrary, but the ratio is selected within a range that can contribute to the promotion of serotonin synthesis and the inhibition of cortisol production.

[0050] When the oral composition of the present invention is used as a health food, the action or efficacy brought about by the GMC of the present invention may be displayed on the product itself, on the contents such as the body, packaging, container, or package of the product of the present invention, or on its advertisements.

[0051] Furthermore, the oral composition (GMC) of the present invention can be used as a food additive either by itself or by adding other ingredients to it. There are no particular restrictions on the other ingredients as long as they can be used as food and beverage additives. There are also no particular restrictions on the beverages and foods to which the food additive can be added.

[0052] The oral composition of the present invention may be mixed with a food additive at any ratio, but the ratio is selected within a range that can contribute to the promotion of serotonin synthesis and the inhibition of cortisol production.

[0053] The oral composition of the present invention can also be applied to animals other than humans, and therefore can be used as an oral composition for animals such as pet food. [Example]

[0054] The present invention will be explained in more detail below with reference to examples, but the present invention is not limited to these examples. In addition, the "%" shown in the mixed solvents in the examples indicates "volume %" unless otherwise specified. Furthermore, when the extraction solvent is a mixed solvent, the balance of water is omitted. For example, "50% ethanol" means a mixed solvent of 50% ethanol by volume and 50% water by volume.

[0055] <1. Evaluation of serotonin synthesis promotion effect> The serotonin synthesis promoting effect of cis-GMC or trans-GMC was evaluated according to the following procedures (1A) to (3A). The samples (cis-GMC, trans-GMC) used for the evaluation were the following standard substances. ·cis-GMC:cis-2-beta-D-Glucopyranosyloxy-4-methoxy-cinnamic acid (AnalytiCon Discovery) ·trans-GMC: trans2-beta-D-Glucopyranosyloxy-4-methoxy-cinnamic acid (AnalytiCon Discovery)

[0056] (1A) Cell culture Mouse neuroblastoma Neuro2a cells were precultured in cell growth medium No. 104 (Eagle Medium + NEAA + 10% FBS) in a 10 cm dish until confluent. After washing with phosphate-buffered saline (PBS), the cells were resuspended in medium and plated at 1.0 × 10 cells per well in a 24-well plate. 5 The cells were seeded at a density of 1000 cells / well and cultured overnight in a CO2 incubator (37°C, 5% CO2).

[0057] (2A) Sample addition After overnight incubation, the medium was replaced with cell growth medium No. 104 containing the sample (cis-GMC or trans-GMC) at 100, 200, or 400 μg / mL, and the cells were incubated in a CO 2 incubator for 24 hours. Although not shown, cell viability was measured using an MTT staining solution (5 mg / mL in PBS), and it was confirmed that there was no effect on cell viability in all groups.

[0058] (3A) Comparison of TPH2 gene expression levels by real-time PCR Neuro2a cells were harvested 24 hours after addition of the sample (cis-GMC or trans-GMC), and total RNA was extracted using the PureLink RNA Mini kit (Invitrogen). cDNA was synthesized from the extracted total RNA using ReverTra Ace qPCR RT Master Mix with gDNA Remover (Toyobo). Real-time PCR was performed using the synthesized cDNA as a template on an AriaMX real-time PCR system (Agilent Technologies). THUNDERBIRD SYBR qPCR Mix (Toyobo) was used for the real-time PCR reaction. The primers for the TPH2 gene were 5'-CTACCCGACTCATGCTTGCC-3' and 5'-CAGGAAGTCTCTTGGGCTCAG-3', and the primers for the internal standard β-actin were 5'-CAAAAGCCACCCCCACTCCTAAGA-3' and 5'-GCCCTGGCTGCCTCAACACCTC-3'. The real-time PCR reaction conditions were initial denaturation at 95°C for 3 minutes, followed by 40 cycles of a two-step PCR reaction consisting of denaturation at 95°C for 3 seconds and annealing / extension at 60°C for 30 seconds. The expression level of the TPH2 gene was evaluated by adding cis-GMC and trans-GMC.

[0059] The expression level of the TPH2 gene in the control (DMSO) was set to 1, and the relative values ​​of the expression levels of the TPH2 gene in cis-GMC and trans-GMC were calculated. The results are shown in Figure 1. In Figure 1, a significant difference test was performed on samples to which DMSO was added instead of the sample (cis-GMC or trans-GMC), and * indicates p<0.01 (Student's t-test).

[0060] As shown in Figure 1, it was confirmed that both cis-GMC and trans-GMC increased the expression level of the TPH2 gene in Neuro2a cells compared to the control (DMSO).

[0061] <2. Evaluation of cortisol production inhibitory effect> The cortisol production inhibitory effect of cis-GMC or trans-GMC was evaluated according to the following procedures (1B) to (3B). The samples (cis-GMC or trans-GMC) used for the evaluation were the same as those described in <1. Evaluation of serotonin synthesis promoting effect> above.

[0062] (1B) Cell culture Human adrenal gland cells NCI-H295R cells were precultured in Dulbecco's Modified Eagle Medium: Nutrient Mixture F-12 (DMEM / F12) (containing 1% penicillin-streptomycin, 1% ITS+Premix, and 2.5% Nu-Serum) in a 10 cm dish until confluent. After washing with phosphate-buffered saline (PBS), the cells were resuspended in medium, and then plated at 1.0 × 10 cells per well in a 24-well plate. 5 The cells were seeded at a density of 1000 cells / well and cultured overnight in a CO2 incubator (37°C, 5% CO2).

[0063] (2B) Add sample After overnight incubation, the cells were cultured in serum-free DMEM medium (containing 1% penicillin-streptomycin) containing the sample (cis-GMC or trans-GMC) at concentrations of 100, 200, or 400 μg / mL in a CO 2 incubator for 24 hours. Although not shown, cell viability was measured using an MTT staining solution (5 mg / mL in PBS), and it was confirmed that there was no effect on cell viability in all groups.

[0064] (3B) Comparison of 11β-HSD1 gene expression levels by real-time PCR NCI-H295R cells were harvested 24 hours after addition of the sample (cis-GMC or trans-GMC), and total RNA was extracted using the PureLink RNA Mini kit (Invitrogen). cDNA was synthesized from the extracted total RNA using ReverTra Ace qPCR RT Master Mix with gDNA Remover (Toyobo). Real-time PCR was performed using the synthesized cDNA as a template on an AriaMX real-time PCR system (Agilent Technologies). THUNDERBIRD SYBR qPCR Mix (Toyobo) was used for the real-time PCR reaction. Primers for the 11β-HSD1 gene were 5'-GCAAAGGGATCGGAAGAGAGA-3' and 5'-GACCTCGCTGTCACCACCA-3', and primers for the internal standard GAPDH were 5'-GCACCGTCAAGGCTGAGAAC-3' and 5'-ATGGTGGTGAAGACGCCAGT-3'. The real-time PCR reaction conditions were initial denaturation at 95°C for 3 minutes, followed by 40 cycles of a two-step PCR reaction consisting of denaturation at 95°C for 3 seconds and annealing / extension at 60°C for 30 seconds. The expression level of the 11β-HSD1 gene was evaluated by adding cis-GMC and trans-GMC.

[0065] The expression level of the 11β-HSD1 gene in the control (DMSO) was set to 1, and the relative expression levels of the 11β-HSD1 gene in cis-GMC and trans-GMC were calculated. The results are shown in Figure 2(a) and (b). In Figure 2(a) and (b), significance tests were performed on samples to which DMSO was added instead of the sample (cis-GMC or trans-GMC), and * indicates p<0.01 (Student's t-test).

[0066] As shown in FIG. 2(a), it was confirmed that cis-GMC reduced the expression level of the 11β-HSD1 gene in NCI-H295R cells in a concentration-dependent manner. [Industrial Applicability]

[0067] According to the present invention, an oral composition for promoting serotonin synthesis containing 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid as an active ingredient is provided, and the oral composition of the present invention can be suitably used as a functional food, supplement, etc.

Claims

[Claim 1] A composition for promoting TPH2 gene expression, comprising 2-β-D-glucopyranosyloxy-4-methoxycinnamic acid as an active ingredient.

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