Serotonin secretion-promoting composition, food or drink, cosmetic, and medicine containing said composition, and methods for producing same

Abstract

[Problem] The purpose of the present invention is to provide: a composition capable of promoting the secretion of serotonin; food or drink, a cosmetic, and a medicine containing the composition; and methods for producing the same. [Solution] Provided is a serotonin secretion-promoting composition containing at least two types of enteric bacteria. In particular, the enteric bacteria are preferably selected from enteric bacteria belonging to the genus Eubacterium, Clostridium, or Bifidobacterium.

Description

Serotonin secretion-promoting composition, foods, beverages, cosmetics and pharmaceuticals containing the composition, and methods for producing the same

[0001] The present disclosure relates to a composition for promoting serotonin secretion using at least two types of intestinal bacteria that promote serotonin secretion, foods and beverages, cosmetics and pharmaceuticals made therefrom, and methods for producing these.

[0002] Recent advances in neuroscience, molecular biology, and genome analysis have revealed that neurotransmitters, such as nervous system and humoral factors (hormones and cytokines), form a bidirectional network between the central nervous system and peripheral organs via receptors, and signals are transmitted from the periphery to the brain via the vagus nervous system. Disturbances in the gut microbiota have been shown to be associated with sleep quality, psychiatric disorders such as depression, lifestyle-related diseases such as metabolic syndrome and diabetes, cancer, and aging. Research is also being conducted to establish mental well-being through the gut-brain connection by targeting the gut microbiota and analyzing its interactions with the central nervous system (Non-Patent Document 1). Additionally, numerous reviews have examined the effects of probiotics and fermented foods on mental disorders (Non-Patent Documents 2 and 3). Their effectiveness is thought to be due to their influence on the monoamine system, stress response, and inflammation.

[0003] Furthermore, microbiome drug discovery (LBPs), which combines multiple intestinal bacteria to produce effective drugs, is being conducted worldwide, and there has been great interest in the development of LBPs targeting inflammatory bowel disease (IBD) (Non-Patent Document 4). LBPs are a technology that encapsulates multiple live bacteria and delivers them to the patient's intestines. However, there are few pipelines using LBPs to target psychiatric disorders.

[0004] To prevent depression-like symptoms, it is important to reduce excessive stress, but in today's stressful society due to information overload, it is necessary to increase resilience to stress in addition to good eating habits, adequate sleep, and exercise (Non-Patent Documents 5 and 6). It is known that certain types of bifidobacteria contribute to the reduction of depression-like symptoms (Patent Document 1), and probiotics and synbiotics, including LBPs, are desirable.

[0005] Japanese Patent Application Laid-Open No. 2022-88787

[0006] Tochitani S et al. Neurosci Res 168:83-94, 2021Carlos R et al. Neurosci & Biobehav Rev158:105561, 2024 Ramya B et al. Neurosci & Biobehav Rev 158:105562, 2024 Lloyd-Price J. et al. Nature 569:655-662, 2019 Juntaro M et al. Gut Pathogens 16:8, 2024 Nathaniel L. R et al. PNAS 121(1): e2308706120, 2023

[0007] Incidentally, if the secretion of neurotransmitters (such as serotonin) can be promoted and controlled, it is expected that this will contribute to the normalization of mental state and the promotion of health. However, this is not limited to the above substances, but some people may find it difficult to achieve the desired effect due to their constitution or other factors. Therefore, there is a continuing need for a highly safe method that can effectively promote and control the secretion of neurotransmitters (such as serotonin).

[0008] Therefore, an object of the present disclosure is to provide a composition capable of promoting serotonin secretion, a food or drink product, a cosmetic product, and a pharmaceutical product thereof, as well as methods for producing these.

[0009] To solve the above-mentioned problems, the present inventors focused on the effects of intestinal bacteria on brain and mental function. In particular, based on the monoamine hypothesis that depression is caused by a decrease in monoamine transmission function, they focused on the fact that serotonin is essential for the biosynthesis of brain-derived neurotrophic factor (BDNF). They then found that a composition containing at least two types of intestinal bacteria can effectively promote serotonin secretion. Therefore, a first aspect of the present disclosure is: (1) a composition for promoting serotonin secretion, comprising at least two types of intestinal bacteria.

[0010] A preferred embodiment of the present disclosure is (2) the composition according to (1) above, wherein the intestinal bacteria are intestinal bacteria belonging to the genus Eubacterium, Clostridium, Ruthenibacterium, Ruminococcus, Parabacteroides, Enterocloster, or Bifidobacterium.

[0011] A preferred embodiment of the present disclosure is (3) the composition according to (1) above, wherein the intestinal bacteria are intestinal bacteria belonging to the genus Eubacterium, Clostridium, or Bifidobacterium.

[0012] A preferred embodiment of the present disclosure is (4) the composition according to (1) above, wherein the intestinal bacteria are selected from Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve.

[0013] A preferred embodiment of the present disclosure is (5) the composition according to (3) above, which contains 0.5 to 30% of the Eubacterium limosum, 10 to 80% of the Clostridium butyricum, and 10 to 80% of the Bifidobacterium breve.

[0014] A preferred embodiment of the present disclosure is (6) the composition according to (1) above, wherein the intestinal bacteria is selected from Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160).

[0015] The present inventors have found that culturing enterobacteria in a medium containing one or more naturally occurring saccharides promotes their growth. Therefore, a preferred embodiment of the present disclosure is (7) the composition according to (1) above, in which enterobacteria are cultured in a medium containing one or more naturally occurring saccharides.

[0016] A preferred embodiment of the present disclosure is (8) the composition according to (1) above, wherein the enterobacteria are cultured in a medium containing at least brown sugar.

[0017] A preferred embodiment of the present disclosure is the composition described in (8) above, wherein the culture medium further contains, in addition to brown sugar, granulated sugar, white sugar, cane sugar, brown sugar, unrefined sugar, invert sugar, trehalose, fructose, glucose, or a combination thereof.

[0018] In order to solve the above-mentioned problems, the present inventors have found that a food or drink containing a composition comprising at least two types of intestinal bacteria can effectively promote the secretion of serotonin. Therefore, a second aspect of the present disclosure is (10) a food or drink containing the composition described in any of (1) to (9) above.

[0019] In order to solve the above-mentioned problems, the present inventors have found that a cosmetic containing a composition including at least two types of intestinal bacteria can effectively promote the secretion of serotonin. Therefore, a third aspect of the present disclosure is (11) a cosmetic containing the composition described in any one of (1) to (9) above.

[0020] In order to solve the above problems, the present inventors have found that a pharmaceutical containing a composition comprising at least two types of intestinal bacteria can effectively promote serotonin secretion. Therefore, a fourth aspect of the present disclosure is (12) a pharmaceutical containing the composition described in any of (1) to (9) above.

[0021] In order to solve the above problems, the present inventors have found that a composition that can effectively promote serotonin secretion can be produced using at least two types of intestinal bacteria. Therefore, a fifth aspect of the present disclosure is (13) a method for producing a composition for promoting serotonin secretion, comprising a culture step of culturing at least two types of intestinal bacteria in a medium inoculated with the at least two types of intestinal bacteria.

[0022] It is possible to provide a composition capable of promoting serotonin secretion, a food or drink, a cosmetic, or a pharmaceutical product thereof, and methods for producing these.

[0023] Figure 1A shows the calculated secretion amounts of the top five combinations with the highest serotonin secretion levels among any combinations selected from 59 RD strains. Figure 1B shows the calculated secretion amounts of the top five combinations with the highest serotonin secretion levels among any combinations selected from 28 NBRC strains and 8 JCM strains. Figure 2 shows the secretion amounts of serotonin for each composition listed in Table 7.

[0024] The following provides a detailed description of the present disclosure of a composition capable of promoting serotonin secretion, its food and beverage products, cosmetics, and pharmaceutical products, as well as methods for producing these products. However, the embodiments are merely examples for explaining the present disclosure, and the present disclosure is not limited to these embodiments. Although the term "plurality" is used in the present disclosure, this term is not intended to be limited to a specific number (e.g., two), but may include any number, such as three or four.

[0025] 1. Production of a Composition for Promoting Serotonin Secretion Serotonin (5-hydroxytryptamine) is known as a type of neurotransmitter. Serotonin is primarily produced in the raphe nucleus of the brainstem by the hydroxylation of tryptophan, an essential amino acid, by tryptophan hydroxylase, resulting in its conversion to 5-hydroxytryptophan. The raphe nucleus of the brainstem influences brain regions such as the cerebral cortex, limbic system, hypothalamus, brainstem, and spinal cord. Therefore, serotonin produced in the raphe nucleus is involved in the control of various brain functions, including physiological and mental functions such as mood, sleep, appetite, sexual desire, and stress resistance. For example, a decrease in serotonin secretion can lead to mental problems such as depression and anxiety disorders, or physical symptoms such as sleep disorders, loss of appetite, and fatigue.

[0026] Here, for example, short-chain fatty acids produced by intestinal bacteria attach to free fatty acid receptors, which affect neurotransmitters. Furthermore, gamma-aminobutyric acid receptors, which are widely distributed in the central nervous system, can be produced or consumed by intestinal bacteria. Furthermore, specific tryptamine receptors produced by intestinal bacteria can activate neural signaling pathways. Thus, neurotransmitters such as serotonin can be affected by intestinal bacteria. The present inventors have discovered that intestinal bacteria can be used to control the secretion of serotonin, a type of neurotransmitter, and to appropriately control physiological functions, pathologies such as mood disorders and drug addiction, or emotions and mental conditions. In particular, by forming a composition using a combination of specific intestinal bacteria rather than using intestinal bacteria alone, the serotonin secretion-promoting effect is enhanced, and the composition may be able to exert excellent control over various brain functions, including physiological functions, pathologies such as mood disorders and drug addiction, and mental functions.

[0027] As mentioned above, approximately 90% of serotonin produced in the brainstem is secreted in the intestine in humans and other animals. Specifically, serotonin is secreted by enteric neurons and intestinal wall cells, including enteroendocrine cells. Neurotransmitters such as serotonin secreted by intestinal wall cells bind to receptors on the enteric nerve plexus, activating intracellular signaling pathways and regulating gastrointestinal motility, secretion, blood flow, and other functions.

[0028] Therefore, in the present disclosure, a composition for promoting serotonin secretion containing at least two types of intestinal bacteria is produced, and this composition promotes the secretion of serotonin in animals, including humans and non-human animals, thereby improving various brain functions such as physiological functions, pathological conditions such as mood disorders and drug addiction, or mental functions.

[0029] Here, intestinal bacteria refer to a type of microorganism that inhabits the digestive tract of animals, including humans and non-human animals. These intestinal bacteria are primarily found in the small intestine, cecum, and large intestine, and there are an extremely large number of types. According to one theory, 1,000 types and 100 trillion intestinal bacteria inhabit the human body. Intestinal bacteria form a complex ecosystem known as the intestinal flora, or intestinal flora, and are involved in various physiological functions, such as food breakdown and absorption, immune system regulation, and vitamin and hormone synthesis. Intestinal bacteria also function as a barrier to prevent the proliferation of harmful microorganisms and influence the development of the immune system.

[0030] The composition for promoting serotonin secretion of the present disclosure contains, as such intestinal bacteria, a combination of at least two types of intestinal bacteria selected from the group consisting of intestinal bacteria belonging to the genera Eubacterium, Clostridium, Ruthenibacterium, Ruminococcus, Parabacteroides, Enterocloster, and Bifidobacterium.

[0031] Furthermore, the composition for promoting serotonin secretion of the present disclosure preferably contains a combination of at least two types of intestinal bacteria selected from the group consisting of intestinal bacteria belonging to the genus Eubacterium, intestinal bacteria belonging to the genus Clostridium, and intestinal bacteria belonging to the genus Bifidobacterium. From the viewpoint of promoting serotonin secretion, the composition more preferably contains a combination of at least two types of intestinal bacteria selected from the group consisting of Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve. Furthermore, from the viewpoint of promoting serotonin secretion, the composition more preferably contains a combination of at least two intestinal bacteria selected from the group consisting of Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160). Furthermore, from the viewpoint of promoting serotonin secretion, the composition particularly preferably contains a combination of three intestinal bacteria selected from the group consisting of Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160).

[0032] Furthermore, in the present disclosure, the at least two types of enterobacteria used in the composition for promoting serotonin secretion may be bacteria having a 16S rDNA sequence that has 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 98% or more, 99% or more, or 99.5% or more base sequence identity with the 16S rDNA sequences of the representative serotypes of Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve used as described above.

[0033] As described above, the composition for promoting serotonin secretion containing a combination of at least two types of intestinal bacteria is produced by a culture step and, if necessary, post-treatment steps such as concentration and drying steps.

[0034] (A) Culturing Step In the present disclosure, the method for producing a composition for promoting serotonin secretion includes culturing the target intestinal bacteria in a predetermined medium. Any medium, such as a liquid medium, a semi-liquid medium, or a solid medium, can be used as desired. Such a medium also includes a nitrogen source added to the medium, an extract added as a growth promoter, a carbon source added to support growth, a buffer for adjusting the pH to an optimal range for lactic acid bacteria growth, water, nucleic acids, inorganic salts, etc.

[0035] In addition to the above, it is possible to add desired sugars to such a medium. Examples of such sugars include monosaccharides, disaccharides, oligosaccharides, polysaccharides, and combinations thereof. More specifically, examples of monosaccharides include glucose (grape sugar), fructose (fruit sugar), xylose, sorbose, galactose, invert sugar, and combinations thereof. Examples of disaccharides include sucrose (cane sugar), maltose (malt sugar), isomaltose, lactose (milk sugar), palatinose, trehalose, cellobiose, and combinations thereof. Examples of oligosaccharides include those in which 3 to 10 monosaccharides are bonded together, such as fructooligosaccharides, galactooligosaccharides, lactofructose oligosaccharides, xylooligosaccharides, maltooligosaccharides, isomaltooligosaccharides, and combinations thereof. Examples of polysaccharides include starch, cellulose, pectin, inulin, glycogen, chitin, xanthan gum, textlan, and combinations thereof.

[0036] The sugars exemplified above do not necessarily contain each component alone, and can be sugars derived from natural products such as sugarcane, sugar beet, palms, maple, corn, agave, potato, tapioca, or a combination thereof. Such sugars can be obtained by appropriately combining various processes such as squeezing, clarification, concentration, crystallization, decomposition, synthesis, and separation of the natural products, and can be classified into types such as granulated sugar, white sugar, cane sugar, brown sugar, unrefined sugar, invert sugar, trehalose, fructose, and glucose depending on the molasses and mineral content, the degree of purification, or the production method.

[0037] Among the sugars exemplified above, the medium preferably contains at least brown sugar as a naturally occurring sugar from the viewpoint of promoting the growth of enterobacteria. Furthermore, from the viewpoint of promoting the growth of enterobacteria, the medium desirably further contains, in addition to the brown sugar, one or more naturally occurring sugars. Examples of such naturally occurring sugars include granulated sugar, white sugar, cane sugar, brown sugar, unrefined sugar, invert sugar, trehalose, fructose, glucose, or a combination thereof.

[0038] In the above-mentioned medium, brown sugar or a combination of brown sugar and other naturally-derived sugars can be contained in the culture medium at a ratio of 0.001% to 20% by weight, preferably 0.01% to 10% by weight, and more preferably 0.1 to 5% by weight, from the viewpoint of promoting the growth of enterobacteria. Furthermore, the brown sugar or a combination of brown sugar and other naturally-derived sugars contained as described above can be contained in the final composition at a ratio of 0.001% to 20% by weight, preferably 0.01% to 10% by weight, and more preferably 0.1 to 5% by weight, in the culture medium.

[0039] The medium prepared as described above is inoculated with at least two types of enterobacteria. As such enterobacteria, as described above, from the viewpoint of promoting serotonin secretion, the genera Eubacterium, Clostridium, Ruthenibacterium, Ruminococcus, Parabacteroides, Enterocloster, and Bifidobacterium are selected. A combination of at least two intestinal bacteria selected from the group consisting of intestinal bacteria belonging to the genus Eubacterium, intestinal bacteria belonging to the genus Clostridium, and intestinal bacteria belonging to the genus Bifidobacterium is more preferably a combination of at least two intestinal bacteria selected from the group consisting of intestinal bacteria belonging to the genus Eubacterium, intestinal bacteria belonging to the genus Clostridium, and intestinal bacteria belonging to the genus Bifidobacterium, and even more preferably a combination of at least two intestinal bacteria selected from the group consisting of intestinal bacteria belonging to the genus Eubacterium, intestinal bacteria belonging to the genus Clostridium, and intestinal bacteria belonging to the genus Bifidobacterium, and A combination of at least two intestinal bacteria selected from the group consisting of Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve is particularly preferred, and a combination of at least two intestinal bacteria selected from the group consisting of Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160) is particularly preferred, and a combination of three species of Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160) is particularly preferred.

[0040] Furthermore, the proportion of each intestinal bacterium to be blended in the composition is, from the viewpoint of promoting serotonin secretion, 0.5 to 30% of intestinal bacterium belonging to the genus Eubacterium, 10 to 80% of intestinal bacterium belonging to the genus Clostridium, and 10 to 80% of intestinal bacterium belonging to the genus Bifidobacterium, preferably 1 to 20% of intestinal bacterium belonging to the genus Eubacterium, 10 to 80% of intestinal bacterium belonging to the genus Clostridium, and 10 to 80% of intestinal bacterium belonging to the genus Bifidobacterium, and more preferably 1 to 20% of intestinal bacterium belonging to the genus Eubacterium, 10 to 80% of intestinal bacterium belonging to the genus Clostridium, and 10 to 80% of intestinal bacterium belonging to the genus Bifidobacterium. The composition preferably contains 20 to 60% enterobacteria belonging to the genus Clostridium and 20 to 60% enterobacteria belonging to the genus Bifidobacterium, more preferably 8 to 20% enterobacteria belonging to the genus Eubacterium, 30 to 50% enterobacteria belonging to the genus Clostridium, and 30 to 50% enterobacteria belonging to the genus Bifidobacterium.

[0041] In the above medium, the at least two species of enterobacteria selected as described above are cultured for a culture period of 1 hour to 10 days, preferably 6 hours to 5 days, and more preferably 12 hours to 1 day. The culture temperature is, for example, 30°C to 40°C, but can be adjusted appropriately within these ranges depending on the strain used. Culture conditions can be selected appropriately depending on the strain used, from among static culture, aeration, stirring, shaking, etc.

[0042] Although not specifically described, two types of enterobacteria may be pre-cultured in any pre-culture medium.

[0043] (B) Post-treatment step In the method for producing a composition for promoting serotonin secretion disclosed herein, the culture medium obtained by the culture step can be used as a composition for promoting serotonin secretion as is, but it is also possible to produce a composition for promoting serotonin secretion in the desired form by subjecting the culture medium to any post-treatment as needed.

[0044] For example, the bacterial concentration in the culture solution obtained by the above-mentioned culture step is adjusted to a desired concentration, and then a dry bacterial cell powder is obtained by a drying step such as freeze-drying, spray-drying, or drum-drying. Note that the form of the dry bacterial cell powder is merely an example, and it is also possible to produce any solid, semi-solid, or liquid formulation by further subjecting it to a shaping step such as tableting.

[0045] Furthermore, any additives may be added to the culture solution obtained by the culture step to prepare a composition for promoting serotonin secretion. Examples of such additives include humectants, sweeteners, stabilizers, buffers, solubilizers, UV inhibitors, antioxidants, surfactants, preservatives, and flavorings.

[0046] Furthermore, from the viewpoint of ease of administration, the composition for promoting serotonin secretion produced as described above contains each enterobacterium so that the weight of the enterobacterium cells per dose is 1 to 500 mg, more preferably 10 to 250 mg, and even more preferably 50 to 200 mg. In this case, each enterobacterium contained can be used as live bacteria, killed bacteria, or a combination thereof.

[0047] <Food and Drink> In the present disclosure, the culture medium obtained by each of the above steps, i.e., the composition for promoting serotonin secretion, can be used as a food or drink as is. Furthermore, when using the composition for promoting serotonin secretion as a food or drink, it is also possible to prepare the composition for promoting serotonin secretion by adding an additive appropriately selected from the various additives listed below to the culture medium. Furthermore, the composition for promoting serotonin secretion can also be provided as a food additive that imparts a secretion-promoting effect to existing food compositions. - Sugar alcohols such as sorbitol, erythritol, maltitol and xylitol, - High-intensity sweeteners such as aspartame, stevioside, sucralose and acesulfame K, - Organic acids such as citric acid, tartaric acid, malic acid, succinic acid and lactic acid, - Vitamins such as L-ascorbic acid, dl-α-tocopherol, B vitamins, nicotinamide and calcium pantothenate, - Surfactants such as glycerin fatty acid esters, polyglycerin fatty acid esters, sucrose fatty acid esters, sorbitan fatty acid esters and propylene glycol fatty acid esters, - Thickeners such as gum arabic, carrageenan, pectin and agar, - Stabilizers such as casein and gelatin, minerals such as amino acids and calcium salts, - Additives such as sodium ascorbate, sodium erythorbate, glycerin and propylene glycol, - Other additives such as colorings, flavorings and preservatives

[0048] Furthermore, the serotonin secretion-promoting composition obtained by each of the above steps may be provided in either liquid or solid form, after being subjected to a concentration step or drying step as necessary.

[0049] Furthermore, the serotonin secretion-promoting composition produced as described above can be used as an additive in foods and beverages. Examples of such foods and beverages include processed grain products (noodles, bread, cooked rice, rice cakes, etc.), processed fats and oils (mayonnaise, margarine, etc.), processed meat products (ham, sausage, etc.), processed seafood products (kamaboko, chikuwa, satsumaage, etc.), dairy products (butter, cheese, yogurt, etc.), processed fruit products (jam, marmalade, etc.), confectioneries (chocolate, cookies, cakes, jellies, etc.), various beverages (juice, coffee, black tea, green tea, carbonated drinks, lactic acid bacteria drinks, energy drinks, alcoholic beverages, etc.), seasonings (soy sauce, sauces, mirin, etc.), nutritional supplements, functional supplements, supplements, confectioneries (butter cake, etc.), and quasi-drugs.

[0050] When the composition for promoting serotonin secretion of the present disclosure is provided as a food or beverage, it can be provided in the form of a general food or as a health functional food that claims a function related to the promotion of serotonin secretion. Such claims include claims related to the improvement of mood disorders, drug addiction, sleep, loss of appetite, decreased libido, stress resistance, fatigue, etc.

[0051] When the composition for promoting serotonin secretion of the present disclosure is provided as a food or beverage, the intake amount is appropriately selected depending on the type, sex, species, age, and desired degree of effect of the individual to be ingested. The intake amount is typically set appropriately within the range of 0.001 to 500 mg / day. Specifically, for a human weighing 60 kg, the recommended daily intake amount can be set appropriately so that the weight of enterobacteria cells is 1 to 500 mg, more preferably 10 to 250 mg, and even more preferably 50 to 200 mg, but is not limited thereto. Furthermore, the recommended number of intakes may be the above amount taken once a day or in divided doses.

[0052] Here, the serotonin secretion-promoting composition of the present disclosure is used in foods and beverages, assuming that the subject of intake is a human. However, the serotonin secretion-promoting composition of the present disclosure can also be provided as feed. In such cases, the subject of intake is not particularly limited, but non-limiting examples include mammals such as mice, rats, guinea pigs, rabbits, hamsters, dogs, cats, weasels, cows, pigs, horses, deer, wild boars, sheep, and goats; birds such as poultry such as chickens, ducks, turkeys, and quails; reptiles such as snakes and turtles; amphibians such as frogs; salmon, fishes, yellowtail and cobia, freshwater trout, sweetfish, sea bass, sea bream, and meagre fish; flounder, pufferfish, and crayfish. Examples of fish that can be used include yellowtail, amberjack, bluefin tuna, horse mackerel, striped jack, tiger pufferfish, yellowtail amberjack, stonefish, filefish, rockfish, scorpion fish, black porgy, rockfish, sea bass, crimson sea bream, grunt, sea bream, chub mackerel, grouper, grouper, eel, catfish, loach, sturgeon, tropical fish, tilapia, pangasius, bluefin tuna, bighead carp, silver carp, grass carp, and carp; bivalve shells (oysters, scallops, clams, mussels, pearl oysters), abalone, shrimp, and seaweed.

[0053] <Cosmetics> In the present disclosure, the culture medium obtained by each of the above steps, i.e., the composition for promoting serotonin secretion, can be used as a cosmetic product as is, or can be added to a cosmetic product. Such cosmetics may be in any form, such as a solubilized system, an emulsion system, a powder dispersion system, a powder system, a hard capsule system, or a soft capsule system, and can be used for a variety of purposes, such as basic cosmetics such as lotions, emulsions, creams, and packs, makeup cosmetics such as foundations, toiletry products such as shampoos, conditioners, soaps, and body soaps, and bath additives such as bath additives.

[0054] When the composition for promoting serotonin secretion is used as a cosmetic, it is also possible to form the composition for promoting serotonin secretion by adding to the culture solution an additive selected from a variety of pharmaceutically acceptable additives, such as oils, humectants, sweeteners, stabilizers, buffers, solubilizers, UV protection agents, antioxidants, surfactants, preservatives, moisturizers, fragrances, water, alcohol, thickeners, cell activators, sebum secretion regulators, anti-inflammatory agents, astringents, antioxidants, whitening agents, active oxygen inhibitors, antiallergic agents, animal and plant extracts having physiologically active effects, and extract fractions thereof.

[0055] <Pharmaceuticals> In the present disclosure, the culture medium obtained by each of the above steps, i.e., the composition for promoting serotonin secretion, can be used as a pharmaceutical directly or added to a pharmaceutical. Such pharmaceuticals may be in any form, including solid oral preparations such as tablets, chewable tablets, effervescent tablets, troches, drops, hard capsules, soft capsules, granules, orally disintegrating tablets, powders, pills, dry syrups, infusions, and decoctions; semisolid oral preparations such as electuaries, syrups, whipped creams, and chewing gums (nicotine preparations); liquid preparations such as syrups, drinks, suspensions, and spirit preparations; solid external preparations such as suppositories, poultices, and plasters; semisolid external preparations such as ointments, creams, and mousses; and liquid external preparations such as liquids, eye drops, aerosols, sprays, and aerosols.

[0056] When the composition for promoting serotonin secretion is used as a pharmaceutical, it is also possible to form the composition for promoting serotonin secretion by adding to the culture medium an additive selected from various additives such as oils, humectants, sweeteners, stabilizers, buffers, solubilizers, UV protection agents, antioxidants, surfactants, preservatives, moisturizers, fragrances, water, alcohol, thickeners, cell activators, sebum secretion regulators, anti-inflammatory agents, astringents, antioxidants, whitening agents, active oxygen inhibitors, antiallergic agents, animal and plant extracts having physiologically active effects, and extract fractions thereof.

[0057] By providing the composition for promoting serotonin secretion of the present disclosure as a pharmaceutical, it can be used to improve diseases and conditions related to the promotion of serotonin secretion, such as mood disorders, drug addiction, sleep, loss of appetite, decreased libido, stress resistance, or fatigue.

[0058] When the composition for promoting serotonin secretion of the present disclosure is provided as a pharmaceutical product, the subjects to which it is to be administered are not particularly limited, and non-limiting examples include mammals such as humans, mice, rats, guinea pigs, rabbits, hamsters, dogs, cats, weasels, cows, pigs, horses, deer, wild boars, sheep, and goats; birds such as poultry, chickens, ducks, turkeys, and quails; reptiles such as snakes and turtles; amphibians such as frogs; salmonid fish; yellowtail and cobia; freshwater trout; sweetfish; sea bass; sea bream; and meagre. Examples of fish that can be caught include flounder, pufferfish, yellowtail, amberjack, bluefin tuna, horse mackerel, striped jack, tiger pufferfish, yellowtail amberjack, stonefish, filefish, rockfish, scorpion fish, black porgy, rockfish, sea bass, crimson sea bream, grunt, black sea bream, chub mackerel, yellowtail grouper, grouper, eel, catfish, loach, sturgeon, tropical fish, tilapia, pangasius, bluefin tuna, bighead carp, silver carp, grass carp, and carp; bivalve shells (oysters, scallops, clams, mussels, pearl oysters), abalone, shrimp, and seaweed.

[0059] When the serotonin secretion-promoting composition of the present disclosure is provided as a pharmaceutical, its dosage is appropriately selected depending on the type of disease, sex, species, age, general condition, severity of the disease, and the desired level of effect of the individual to be administered. The dosage is typically set appropriately within the range of 0.001 to 500 mg / day. Specifically, for a human weighing 60 kg, the daily dosage can be set appropriately so that the weight of enterobacterial cells is 1 to 500 mg, more preferably 10 to 250 mg, and even more preferably 50 to 200 mg, but is not limited thereto. The dosage frequency can also be set as a single dose or in several divided doses per day. Furthermore, the dosage interval can be daily, every other day, weekly, biweekly, every 2 to 3 weeks, monthly, bimonthly, or every 2 to 3 months, as long as the dosage has a therapeutic effect against the disease or condition.

[0060] Furthermore, when the serotonin secretion-promoting composition of the present disclosure is provided as a pharmaceutical, its administration method can be various methods such as oral administration, administration by injection, sublingual administration, enteral administration, vaginal administration, ocular administration, nasal administration, inhalation administration, transdermal administration, or transdermal absorption, but among these, oral administration is preferred.

[0061] Furthermore, when the serotonin secretion-promoting composition of the present disclosure is provided as a pharmaceutical product, it is naturally possible to use a combination of multiple compositions according to the present disclosure or to use them in combination with other pharmaceutical products. In this case, these drugs may be contained in a single formulation or may be contained in separate formulations. When these are provided as separate formulations, they may be provided in the form of a kit containing the combination of formulations, or each may be provided as a single formulation to be combined with another formulation. These formulations may be administered simultaneously or sequentially. In one aspect, the pharmaceutical composition of the present disclosure is provided in the form of a raw material composition for the final pharmaceutical product.

[0062] 2. Method of Using the Composition for Promoting Serotonin Secretion The composition for promoting serotonin secretion produced through the above-described production process may be formed into any form, such as a solid, semi-solid, or liquid, as described above. In this case, as described above, the composition for promoting serotonin secretion contains each enterobacterium so that the weight of the enterobacterium cells per dose is 1 to 500 mg, more preferably 10 to 250 mg, and even more preferably 50 to 200 mg. For example, when taken by an adult human, ingesting the composition for promoting serotonin secretion containing the above-described amount of bacteria once a day is expected to favorably promote serotonin secretion.

[0063] Such a composition for promoting serotonin secretion can be ingested by any method. One example of such a method includes administering the composition for promoting serotonin secretion to a subject, including humans and non-human animals. This administration method can be exemplified by various methods such as oral administration, injection, sublingual administration, rectal administration, vaginal administration, ocular administration, nasal administration, inhalation administration, transdermal administration, or transdermal absorption, among which oral administration is preferred.

[0064] Thus, by using the serotonin secretion-promoting composition according to the present disclosure, and the food, beverage, cosmetic, and pharmaceutical products containing the composition, it is possible to effectively promote serotonin secretion. In particular, by using the secretion-promoting composition containing at least two types of intestinal bacteria, and the food, beverage, cosmetic, and pharmaceutical product containing the composition, it is possible to more effectively promote serotonin secretion. Furthermore, as a result, the serotonin secretion-promoting composition, and the food, beverage, cosmetic, and pharmaceutical product containing the composition are expected to exert excellent control functions over various brain functions, including physiological functions, pathologies such as mood disorders and drug addiction, and mental functions.

[0065] In one aspect, the composition for promoting serotonin secretion according to the present disclosure is provided as a composition for use in improving diseases or conditions such as mood disorders, drug addiction, sleep, loss of appetite, decreased libido, stress resistance, or fatigue. In particular, the composition can be effectively utilized by using it in the following method.

[0066] <Method for promoting serotonin secretion or improving a disease or condition> In one aspect, the present disclosure provides a method for promoting serotonin secretion in a subject in need of such promotion, the method comprising administering to the subject a composition comprising at least two types of intestinal bacteria and a pharmaceutically acceptable additive.

[0067] As the composition containing at least two types of intestinal bacteria and pharmaceutically acceptable additives used in the method of the present disclosure, the compositions described above in the <Food and Drink> section, the <Cosmetics> section, or the <Pharmaceuticals> section can be used.

[0068] Furthermore, in one aspect, the present disclosure provides a method for promoting serotonin secretion in a subject in need of such promotion, the method comprising administering to the subject a composition comprising at least two types of intestinal bacteria and a pharmaceutically acceptable additive, wherein the intestinal bacteria are intestinal bacteria belonging to the genus Eubacterium, Clostridium, or Bifidobacterium.

[0069] Furthermore, in one aspect, the present disclosure provides a method for promoting serotonin secretion in a subject in need of such promotion, the method comprising administering to the subject a composition comprising at least two types of intestinal bacteria and a pharmaceutically acceptable additive, wherein the intestinal bacteria are selected from Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve.

[0070] Furthermore, in one aspect, the present disclosure provides a method for promoting serotonin secretion in a subject in need of such promotion, comprising the step of administering to said subject a composition comprising at least two types of intestinal bacteria and a pharmaceutically acceptable additive, wherein the intestinal bacteria comprise 0.5 to 30% Eubacterium limosum, 10 to 80% Clostridium butyricum, and 10 to 80% Bifidobacterium breve, respectively.

[0071] In one aspect, the present disclosure provides a method for promoting serotonin secretion in a subject in need of such promotion, the method comprising administering to the subject a composition comprising at least two species of intestinal bacteria and a pharmaceutically acceptable additive, wherein the intestinal bacteria are selected from Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160).

[0072] Subjects requiring the promotion of serotonin secretion or the improvement of a disease or condition associated with serotonin secretion using the methods of the present disclosure are not particularly limited, and non-limiting examples include mammals such as humans, mice, rats, guinea pigs, rabbits, hamsters, dogs, cats, weasels, cows, pigs, horses, deer, wild boars, sheep, and goats; poultry birds such as chickens, ducks, turkeys, and quails; reptiles such as snakes and turtles; amphibians such as frogs; salmonids, yellowtail and cobia; freshwater trout, sweetfish, and sea bass. - Fish such as sea bream, meagre, flounder, pufferfish, yellowtail, amberjack, bluefin tuna, horse mackerel, striped jack, tiger pufferfish, yellowtail amberjack, stonefish, filefish, rockfish, scorpion fish, black porgy, rockfish, sea bass, crimson sea bream, grunt, sea bream, mackerel, yellowtail grouper, grouper, eel, catfish, loach, sturgeon, tropical fish, tilapia, pangasius, bluefin tuna, bighead carp, silver carp, grass carp, carp, bivalve shells (oysters, scallops, clams, mussels, pearl oysters), abalone, shrimp, and seaweed.

[0073] In the method of the present disclosure, the method of ingesting or administering the composition is not particularly limited, and the ingesting or administering methods described in the <Foods and Beverages> section, the <Cosmetics> section, or the <Pharmaceuticals> section can be used.

[0074] In the methods disclosed herein, methods for promoting serotonin secretion in a subject include methods for improving diseases or conditions associated with serotonin secretion, such as mood disorders, drug addiction, sleep, loss of appetite, decreased libido, stress resistance, or fatigue.

[0075] The present invention will be explained in more detail below with reference to examples, but the present invention is not limited to these examples.

[0076] <Screening of intestinal bacteria> (A) Culturing of intestinal bacteria (single organisms) To select intestinal bacteria useful for promoting serotonin secretion, known intestinal bacteria were obtained, cultured, and their activity was examined. Specifically, 59 RD strains listed in Tables 1-1 and 1-2 were obtained from the National Institute of Technology and Evaluation (NITE), 28 NRBC strains listed in Table 2 were obtained from the National Institute of Technology and Evaluation, and 8 JCM strains listed in Table 3 were obtained from the Microbe Division of the RIKEN BioResource Research Center.

[0077]

[0078]

[0079]

[0080]

[0081] (B) Screening of cultured enterobacteria (single organism) Each of the enterobacteria listed in Tables 1 to 3 obtained as described above was subjected to primary screening based on the following points: (1) whether colonies are visible on plate culture media, (2) whether growth is confirmed when cultured overnight in liquid culture media, (3) whether the number of bacteria after liquid culture is sufficiently large, (4) whether there is no significant decrease in expression of GAPDH and β-actin, which are internal standards, when co-cultured with cells, and (5) whether serotonin secretion is sufficiently promoted.

[0082] For the visual confirmation of colonies in (1) above, first, a GAM medium for culturing anaerobic bacteria was prepared with the composition shown in Table 4. Then, each strain was added to the GAM medium and cultured at 37°C for 18 hours. After the culture, colonies were visually observed, and colonies that were visible were recorded as "+" and those that were not visible as "-", and only "+" colonies were used.

[0083] To confirm growth in the liquid medium (2) above, first, a GAM medium for anaerobic bacterial culture with the composition shown in Table 4 was prepared. Each strain was then added to the GAM medium, and cultured at 37°C for 18 hours. The turbidity (OD value: Optical Density value) of the culture medium after culture was measured. Only those cultures that showed an OD value of 0.5 or higher were used after diluting the culture medium to 1 / 10. The culture medium after the test was washed with D-PBS(-) reagent (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) for use as a specimen, and the resulting pellet was adjusted with 15% glycerol-containing D-PBS(-) reagent so that the bacterial concentration was OD = 0.6 based on the turbidity (OD value: Optical Density value), and used as the test specimen for each test.

[0084] The number of bacteria after the liquid culture in (3) above was estimated from the OD value obtained above.

[0085]

[0086] Furthermore, the promotion of serotonin secretion in (5) above was confirmed by calculating the amount of serotonin secreted from rat-derived cells in culture solutions containing each of the intestinal bacteria prepared as test substances as described above. Specifically, RPMI 1640 medium with the composition shown in Table 5 was prepared, and rat-derived RIN14B cells were seeded in the medium. The RIN14B cells were then incubated at 37°C for 24 hours in a 5% CO atmosphere. ２ After the culture, the RIN14B cells contained in the dish were detached by incubating at 37°C for 10 minutes using a cell detachment enzyme (Innovative Cell Technologies), and then 2 × 10 ５ 500 μL of RIN14B cells at 1000 μL / mL were added to a 24-well plate. A hemocytometer (manufactured by Nanoentek) was used to calculate the cell count. The culture solution of each enterobacteria prepared in (A) above was then diluted with 15% glycerol-containing D-PBS(-) reagent to a bacterial concentration of OD = 0.6, and the diluted solution was added to each well to a final concentration of 1%. The RIN14B cells containing each enterobacteria in each well were incubated at 37°C for 24 hours in a 5% CO atmosphere. ２After the culture, the supernatant was collected and the serotonin content in the supernatant was measured using a commercially available serotonin immunoassay kit (Enzo Life Sciences) according to the manual attached to the kit.

[0087]

[0088] The decrease in expression of (4) above was confirmed by qPCR after co-culturing each enterobacteria with RIN14B cells as described above.

[0089] (C) Preparation of a Composition Containing At Least Two Intestinal Bacteria. For the strains selected by primary screening from the 59 RD strains described above, the culture solution of each strain was diluted in a random combination with 15% glycerol-containing D-PBS(-) reagent to a bacterial concentration of OD = 0.6, and then mixed in equal amounts to prepare a composition containing at least two types of bacteria for investigation. Then, using the same method as in (5) above, serotonin secretion levels were calculated using rat-derived RIN14B cells. Note that 15% glycerol-containing D-PBS(-) reagent was used as a control. Similarly, for the strains selected by primary screening from the 28 NBRC strains and 8 JCM strains described above, equal amounts of each strain were mixed in a random combination to prepare a composition containing at least two types of bacteria for investigation. Then, using the same method as in (5) above, serotonin secretion levels were calculated using rat-derived RIN14B cells.

[0090] Figure 1A shows the calculated secretion levels of the top five combinations with the highest serotonin secretion levels among any combinations selected from 59 RD strains. Figure 1B shows the calculated secretion levels of the top five combinations with the highest serotonin secretion levels among any combinations selected from 28 NBRC strains and 8 JCM strains. The strain combinations contained in each composition in Figures 1A and 1B are shown in Table 6.

[0091]

[0092] 1A and 1B , it was confirmed that addition of each of the compositions containing the 10 combinations of bacterial strains shown in Table 6 to RIN14B cells resulted in the secretion of serotonin from RIN14B cells equivalent to or greater than that of the control. In particular, it was confirmed that addition of a composition containing a combination of three bacterial strains, Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve, resulted in the secretion of extremely large amounts of serotonin. In other words, it was confirmed that compositions containing these combinations, particularly the composition containing the combination of three bacterial strains, Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve, have a high serotonin secretion-promoting effect and may be able to exert excellent control over various brain functions, including physiological functions, pathologies such as mood disorders and drug addiction, and mental functions.

[0093] Next, the optimal blend ratio was confirmed for a combination of at least two intestinal bacteria confirmed to have a particularly high serotonin secretion-promoting effect in FIGS. 1A and 1B. Specifically, a composition (MC127) containing a combination of at least two intestinal bacteria (Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve) confirmed to have a particularly high serotonin secretion-promoting effect in FIGS. 1A and 1B was diluted with 15% glycerol-containing D-PBS(-) reagent at the blend ratio shown in Table 7 to a bacterial concentration of OD = 0.6, and the resulting solution was prepared to a final concentration of 1%, to obtain each composition. Then, the amount of serotonin secretion was calculated using rat-derived RIN14B cells using the same method as in (5) above. As a control, a 15% glycerol-containing D-PBS(-) reagent was used.

[0094]

[0095] Fig. 2 is a diagram showing the amount of serotonin secreted in each of the compositions listed in Table 7. According to Fig. 2, the blending ratio of the three types of combination of Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve was 12.8% for Composition C (i.e., 12.8% Eubacterium limosum (NBRC114520), 41.1% Clostridium butyricum (NBRC3858), and 11.5% Bifidobacterium breve (NBRC1151). In the composition A (i.e., a composition containing 46.1% of Eubacterium limosum (NBRC114520), 42.8% of Clostridium butyricum (NBRC3858), and 47.8% of Bifidobacterium breve (NBRC115160)), secretion of serotonin was confirmed to be 1.8 times higher than that of the control. This confirms that a composition containing a combination of three strains, Eubacterium limosum (NBRC114520), Clostridium butyricum (NBRC3858), and Bifidobacterium breve (NBRC115160), in the above-mentioned ratio, has an extremely high effect of promoting the secretion of serotonin and may be able to exert excellent control functions over various brain functions, including physiological functions, pathological conditions such as mood disorders and drug addiction, and mental functions.

[0096] (D) Growth-promoting effect on intestinal bacteria Among the strains confirmed to have the effect of promoting serotonin secretion, for each of Bifidobacterium breve (NBRC115160), Clostridium butyricum (NBRC3858), and Eubacterium limosum (NBRC114520), predetermined sugars were further added to the liquid medium described in (2) above, and the growth-promoting effect on intestinal bacteria when each sugar was added was confirmed. The medium was the GAM medium shown in Table 4, to which brown sugar (product name "Powdered Sugar (Brown Sugar)": manufactured by Tomizawa Shoten Co., Ltd.) was added, in addition to white sugar (product name "White Sugar": manufactured by Fuji Nisshin Sugar Co., Ltd.), fructose (product name "Fructose Fruit Sugar": manufactured by Nisshin Sugar Co., Ltd.), granulated sugar (product name "Granulated Sugar": manufactured by Fuji Nisshin Sugar Co., Ltd.), invert sugar (product name "Trimorene (Made in France)": manufactured by Tomizawa Shoten Co., Ltd.), trehalose (product name "Trehalose": manufactured by Tomizawa Shoten Co., Ltd.), oligosaccharides (product name "Fructooligosaccharides": manufactured by Tomizawa Shoten Co., Ltd.), glucose (product name "Glucose": manufactured by Tomizawa Shoten Co., Ltd.), and unrefined sugar (product name "Unrefined Sugar": manufactured by Tomizawa Shoten Co., Ltd.). The medium was prepared by adding equal amounts of sugars such as "Sudakito" (manufactured by Daito Sugar Co., Ltd.), cane sugar (product name "Kimiyoshi" (manufactured by Alm Co., Ltd.), Yaeyama Hon-Kokuto (product name "Yaeyama Hon-Kokuto" (manufactured by Tomizawa Shoten Co., Ltd.)), or a combination thereof, so that the amount of sugars in the medium was 2% by weight. While there are 1,024 possible combinations, in this study, 128 combinations shown in Table 8 were added from among brown sugar, white sugar, fructose, granulated sugar, invert sugar, trehalose, oligosaccharides, glucose, unrefined sugar, cane sugar, and Yaeyama Hon-Kokuto. In Table 8, the symbols A to K represent the following sugars, respectively. A: White sugar B: Fructose C: Granulated sugar D: Invert sugar (tremorine) E: Trehalose F: Oligosaccharide (granules) G: Pure glucose H: Powdered sugar (brown sugar) I: Sudaki sugar J: Kimira (domestic cane sugar) K: Yaeyama brown sugar

[0097]

[0098] Bifidobacterium breve (NBRC115160), Clostridium butyricum (NBRC3858), and Eubacterium limosum (NBRC114520) strains were added to GAM medium supplemented with the above-described combinations of sugars, and each strain was cultured at 37°C for 24 hours. After the culture, the turbidity (OD value: Optical Density value) of the culture solution was measured.

[0099] First, the turbidity (OD value) of the culture medium (MSC1 to MSC128) in which Bifidobacterium breve (NBRC115160) was cultured, which showed excellent growth effects, is shown in Tables 9A and 9B. Each turbidity (OD value) indicates a multiple of the OD value of the culture medium containing no sugar combination as the positive control (PC), which is set to 1.0000. Each turbidity (OD value) indicates the average value of the turbidity measured after preparing eight GAM media with the same composition and adding each strain and culturing them under the same conditions. The positive control (PC) was the turbidity (OD value) of the culture medium in which Bifidobacterium breve (NBRC115160) was similarly cultured in the GAM medium described in Table 4 without any added sugars, and the negative control (NC) was the GAM medium described in Table 4 without any added sugars.

[0100]

[0101] According to Tables 9A and 9B, for Bifidobacterium breve (NBRC115160), an excellent growth-promoting effect was confirmed in GAM medium supplemented with a sugar combination of MSC87, MSC89, MSC79, MSC81, MSC85, MSC77, MSC82, MSC71, MSC75, MSC70, MSC39, MSC6, MSC38, MSC47, MSC72, ​​MSC14, MSC15, or MSC23. Furthermore, among these, a particularly excellent growth-promoting effect was confirmed in GAM medium supplemented with a sugar combination of MSC77, MSC71, MSC75, MSC38, or MSC72. That is, by adding a sugar combination containing at least trehalose and molten sugar in addition to brown sugar to GAM medium, Bifidobacterium breve (NBRC115160) could be effectively grown.

[0102] Next, the turbidity (OD value) of the culture medium (MSC1 to MSC128) in which Eubacterium limosum (NBRC114520) was cultured, which showed excellent growth effects, is shown in Tables 10A and 10B. Each turbidity (OD value) indicates a multiple of the OD value of the culture medium not containing the sugar combination as the positive control (PC), which was set to 1.0000. Each turbidity (OD value) indicates the average value of the turbidity measured after preparing eight GAM media with the same composition and adding each strain and culturing them under the same conditions. The positive control (PC) was the turbidity (OD value) of the culture medium in which Eubacterium limosum (NBRC114520) was similarly cultured in the GAM medium described in Table 4 without any added sugars, and the negative control (NC) was the GAM medium described in Table 4 without any added sugars.

[0103]

[0104] According to Tables 10A and 10B, for Eubacterium limosum (NBRC114520), an excellent growth-promoting effect was confirmed in GAM medium supplemented with a sugar combination of MSC81, MSC82, MSC83, MSC85, MSC90, MSC65, MSC79, MSC91, MSC93, MSC72, ​​MSC73, MSC86, MSC80, MSC85, MSC69, MSC84 or MSC76. Furthermore, among these, a particularly excellent growth-promoting effect was confirmed in GAM medium supplemented with a sugar combination of MSC91, MSC93, MSC72, ​​MSC73, MSC80, or MSC76. That is, by adding a sugar combination containing at least white sugar in addition to brown sugar to GAM medium, Eubacterium limosum (NBRC114520) could be effectively grown.

[0105] Next, the turbidity (OD value) of the cultures (MSC1 to MSC128) in which Clostridium butyricum (NBRC3858) was cultured, which showed excellent growth effects, is shown in Tables 11A and 11B. Each turbidity (OD value) represents a multiple of the OD value of the culture medium containing no sugar combination as the positive control (PC), which was set at 1.0000. Each turbidity (OD value) represents the average value of the turbidity measured after eight GAM medium samples containing each strain were prepared with the same composition and cultured under the same conditions. The positive control (PC) was the turbidity (OD value) of the culture medium in which Clostridium butyricum (NBRC3858) was similarly cultured in the GAM medium described in Table 4 without any added sugars, and the negative control (NC) was the GAM medium described in Table 4 without any added sugars.

[0106]

[0107] According to Tables 11A and 11B, for Clostridium butyricum (NBRC3858), an excellent growth-promoting effect was confirmed in GAM medium supplemented with a saccharide combination of MSC14, MSC31, MSC6, MSC15, MSC72, ​​MSC7, MSC30, MSC79, MSC22, MSC4, MSC69, MSC80, MSC79, MSC88, MSC86, MSC87, MSC72, ​​MSC85, MSC77, or MSC74. Furthermore, among these, a particularly excellent growth-promoting effect was confirmed in GAM medium supplemented with a saccharide combination of MSC15, MSC7, MSC79, MSC69, MSC79, or MSC87. That is, by adding a sugar combination containing at least trehalose and an oligosaccharide in addition to brown sugar to the GAM medium, Clostridium butyricum (NBRC3858) could be effectively grown.

[0108] As described above, in Tables 9A, 9B, 10A, 10B, 11A, and 11B, it was confirmed that the addition of a combination of naturally occurring sugars in addition to brown sugar to the GAM medium in which each strain was cultured promoted the growth of each strain. This indicates that when culturing at least two types of enterobacteria, such as the combination of strains shown in Table 6, adding a combination of naturally occurring sugars in addition to brown sugar to the medium promoted the growth of these bacteria, and was found to be extremely effective in producing compositions containing these enterobacteria.

[0109] The present disclosure has confirmed that a composition containing at least two types of intestinal bacteria has a serotonin secretion-promoting effect. In other words, the composition, as well as foods, beverages, cosmetics, pharmaceuticals, and the like containing the composition, can be provided, and the compositions and the like according to the present disclosure can be used in these industries.

Claims

1. A composition for promoting serotonin secretion, comprising at least two types of intestinal bacteria, including Eubacterium limosum.

2. The composition according to claim 1, wherein the at least two types of intestinal bacteria are selected from intestinal bacteria belonging to the genera Eubacterium, Clostridium, Ruthenibacterium, Ruminococcus, Parabacteroides, Enterocloster, or Bifidobacterium, in addition to Eubacterium limosum.

3. The composition according to claim 1, wherein the at least two types of intestinal bacteria are selected from intestinal bacteria belonging to the genera Eubacterium, Clostridium, or Bifidobacterium, in addition to Eubacterium limosum.

4. The composition according to claim 1, wherein the at least two types of intestinal bacteria are Eubacterium limosum, Clostridium butyricum, and Bifidobacterium breve.

5. The composition according to claim 4, comprising 0.5 to 50% of Eubacterium limosum, 10 to 80% of Clostridium butyricum, and 10 to 80% of Bifidobacterium breve, respectively.

6. The composition according to claim 4, wherein the Eubacterium limosum is Eubacterium limosum (NBRC114520), the Clostridium butyricum is Clostridium butyricum (NBRC3858), and the Bifidobacterium breve is Bifidobacterium breve (NBRC115160).

7. The composition according to claim 1, wherein the at least two types of intestinal bacteria are cultured in a culture medium containing one or more sugars derived from natural products.

8. The composition according to claim 1, wherein the at least two types of intestinal bacteria are cultured in a culture medium containing at least brown sugar.

9. The composition according to claim 8, wherein the culture medium further comprises, in addition to brown sugar, granulated sugar, refined sugar, cane sugar, brown sugar, unrefined sugar, invert sugar, trehalose, fructose, glucose, or a combination thereof.

10. Food and beverages comprising the composition described in any one of claims 1 to 9.

11. A cosmetic comprising the composition described in any one of claims 1 to 9.

12. A pharmaceutical product comprising the composition described in any one of claims 1 to 9.

13. A method for producing a serotonin secretion-promoting composition, comprising a culture step of culturing at least two types of intestinal bacteria, including Eubacterium limosum, in a culture medium in which the at least two types of intestinal bacteria are seeded.