Kestose improves atopic dermatitis and regulates the intestinal microbiome

Abstract

The present invention relates to a composition containing kestose as an active ingredient, and to uses of the composition for enhancing the production and / or content of intestinal short chain fatty acids (SCFAs), for promoting short chain fatty acid-producing bacteria, for preventing, treating or improving inflammatory diseases, such as atopic dermatitis or intestinal inflammation, and for regulating the intestinal microbiome, and the composition may be a food, pharmaceutical or health functional food composition.

Description

[Technical Field]

[0001] The present invention relates to a composition containing kestose as an active ingredient, and to uses of the composition for enhancing the production and / or content of intestinal short-chain fatty acids (SCFAs), promoting short-chain fatty acid-producing bacteria, preventing, treating, or improving inflammatory diseases such as skin inflammation or intestinal inflammation, and regulating the intestinal microbiome. [Background technology]

[0002] Recently, probiotics have been attracting attention as a health supplement due to research results showing that they are effective in alleviating atopic dermatitis symptoms and strengthening the immune system, thanks to their ability to increase beneficial bacteria in the intestines and suppress harmful bacteria.

[0003] However, there are reports that probiotics may cause side effects in some patients with intestinal membrane damage or immunosuppression, and caution is advised when taking them.

[0004] Recently, in addition to the term probiotics, there has been active research into prebiotics, which refers to nutrients that serve as food for probiotics and can maximize the function of intestinal probiotics by activating them.

[0005] Most prebiotics are in the form of oligosaccharides or polysaccharides, and typical examples include insulin, lactulose, and lactitol, which were previously used primarily as food for beneficial bacteria such as bifidobacteria, and more recently, AX (arabinoxylan) and other prebiotics have been used in a more diverse range of applications. Summary of the Invention [Problem to be solved by the invention]

[0006] One example of the present invention relates to an intestinal microbiome regulation application containing kestose as an active ingredient, and more specifically to an application for promoting / enhancing the growth of beneficial intestinal microorganisms and / or suppressing / reducing the growth of harmful microorganisms.

[0007] A further example of the present invention relates to the use of a composition containing kestose as an active ingredient to promote / enhance the growth of intestinal microorganisms, for example, one or more microorganisms selected from the group consisting of Lactobacillus microorganisms and Alistipes microorganisms.

[0008] A further example of the present invention relates to the use of a composition containing kestose as an active ingredient to inhibit / reduce the growth of intestinal microorganisms, such as microorganisms of the genus Prevotella.

[0009] A further example of the present invention relates to the use of kestose as an active ingredient for promoting / enhancing the growth of intestinal microorganisms that produce intestinal short-chain fatty acids, such as propionic acid.

[0010] Another example of the present invention relates to a composition for enhancing the production and / or content of intestinal short-chain fatty acids, such as propionic acid, comprising kestose as an active ingredient. The increase in the production and / or content of intestinal short-chain fatty acids, such as propionic acid, may be achieved by intestinal microorganisms.

[0011] A further example of the present invention relates to the use of kestose as an active ingredient for the prevention, treatment or amelioration of inflammatory diseases, such as skin inflammation (inflammatory skin diseases) and / or intestinal inflammation (inflammatory bowel diseases). [Means for solving the problem]

[0012] One example of the present invention relates to an intestinal microbiome regulation application containing kestose as an active ingredient, and more specifically to an application for promoting the growth of beneficial intestinal microorganisms, inhibiting the growth of harmful intestinal microorganisms, and / or promoting the growth of intestinal microorganisms that produce short-chain fatty acids, such as propionic acid, in the intestine.

[0013] Yet another example of the present invention relates to a composition for enhancing the production and / or content of intestinal short chain fatty acids (SCFAs), such as propionic acid, which comprises kestose as an active ingredient.

[0014] A further example of the present invention relates to the use of kestose as an active ingredient for the prevention, treatment or amelioration of inflammatory diseases, such as skin inflammation and / or intestinal inflammation. [Effects of the Invention]

[0015] The present invention has the advantage of improving the imbalance of the intestinal microbiome by increasing beneficial intestinal microorganisms and reducing harmful microorganisms in humans and animals simply and effectively, with little concern about side effects or safety.

[0016] Furthermore, the present invention has the advantage of improving the intestinal environment by increasing propionic acid or propionic acid-producing bacterial strains among the short-chain fatty acids in the intestines of humans and animals simply and effectively, with almost no concern about side effects or safety. [Brief explanation of the drawings]

[0017] [Figure 1] 1 is an experimental protocol for analyzing the effects of kestose on intestinal microorganisms using an animal model, according to one example of the present invention. [Figure 2] 1 shows the results of measuring changes in body weight in an experimental animal model according to an example of the present invention. [Figure 3] 1 is a graph showing the species diversity of intestinal microorganisms as determined by alpha-diversity analysis of intestinal microorganisms in an experimental animal model, according to an example of the present invention. [Figure 4] 1 is a graph showing differences between groups of intestinal microorganisms in an experimental animal model based on beta-diversity analysis of the intestinal microorganisms, according to an example of the present invention. [Figure 5]According to one example of the present invention, the results of phylum profiling of the gut microbial community are shown using fecal samples of an animal model collected at weeks 8, 12, and 16 of the study period. [Figure 6] According to one example of the present invention, the results of phylum profiling of the gut microbial community are shown using fecal samples from an animal model collected at week 16 of the study period. [Figure 7] According to one example of the present invention, the results of top 30 genera profiling by heatmap analysis of the gut microbial community are shown using fecal samples of an animal model collected at weeks 8, 12, and 16 of the study period. [Figure 8] According to one example of the present invention, the results of top 30 genera profiling by heatmap analysis of the intestinal microbial community are shown using fecal samples of an animal model collected at week 16 of the study period. [Figure 9] An example of the present invention shows a significant increase in Lactobacillus spp. with treatment of the experimental group. [Figure 10] An example of the present invention shows a significant increase in Alistipe spp. with treatment of the experimental groups. [Figure 11] An example of the present invention shows a significant reduction in Prevetella spp. with treatment of the experimental groups. [Figure 12] According to one example of the present invention, the change in butyric acid content due to administration of the experimental group was measured using fecal samples of an animal model collected at 16 weeks of the study period. [Figure 13] According to one example of the present invention, the results show that changes in acetic acid content due to administration of the experimental group were measured using fecal samples of an animal model collected at 16 weeks of the study period. [Figure 14] According to one example of the present invention, the results show that the change in propionic acid content due to administration of the experimental group was measured using fecal samples of the animal model collected at 16 weeks of the study period. [Figure 15]1 shows the results of an evaluation of the improvement in severity of atopic dermatitis based on total score (itching and severity) according to an example of the present invention. [Figure 16] 1 shows the results of an analysis of serum IgE secretion levels in experimental animals according to an example of the present invention. [Figure 17] 1 shows the results of an analysis of serum TNF-α, IFN-γ, and IL-12 secretion levels in experimental animals according to an example of the present invention. [Figure 18] 1 shows the results of an analysis of serum IL-4, IL-5, IL-13, TARC, and Eotaxin secretion levels in experimental animals according to an example of the present invention. [Figure 19] 1 shows the results of an analysis of the amounts of serum IL-1β and IL-10 secreted from experimental animals according to an example of the present invention. [Figure 20] FIG. 1 is a diagram showing the expression level of CD86 in splenocytes analyzed by flow cytometry according to one example of the present invention. [Figure 21] FIG. 1 shows the expression level of CD274 in splenocytes analyzed by flow cytometry according to one example of the present invention. [Figure 22] 1 shows the results of measuring the mast cell count in the skin tissue of an experimental animal according to an example of the present invention. [Figure 23] 1 shows the results of measuring the eosinophil count in the skin tissue of an experimental animal according to an example of the present invention. [Figure 24] 1 shows the results of measuring the mast cell count in the intestinal tissue of an experimental animal according to one example of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0018] The present invention will now be described in further detail. The present invention relates to a composition, use, or method for regulating the intestinal microbiome, comprising kestose as an active ingredient, to improve imbalance of intestinal microorganisms, and more particularly to a composition for use in regulating one or more intestinal microbiomes selected from the group consisting of inhibiting the growth of harmful intestinal microorganisms, promoting the growth of beneficial intestinal microorganisms, and promoting the growth of propionic acid-producing bacteria. One example of the present invention relates to a method for regulating the intestinal microbiome or a method for improving imbalance of intestinal microorganisms, comprising the step of administering kestose or a composition comprising kestose to a subject in need thereof.

[0019] According to the present invention, the active ingredient 1-kestose can be orally administered to a subject, for example, a mammal, including a human, as a pharmaceutical or food product to regulate the intestinal microbial community of the recipient and improve intestinal microbial imbalance. Specifically, by orally ingesting the composition for improving intestinal microbial imbalance and the agent for promoting the proliferation of beneficial intestinal microorganisms according to the present invention, the intestinal microbiome can be regulated in one or more ways selected from the group consisting of inhibiting the growth of harmful intestinal microorganisms, promoting the growth of beneficial intestinal microorganisms, and promoting the growth of propionic acid-producing bacteria.

[0020] One example of the present invention relates to the use of a composition containing kestose as an active ingredient to promote the growth or increase the content of beneficial intestinal microorganisms, such as one or more microorganisms selected from the group consisting of Alistipes microorganisms and Lactobacillus microorganisms.

[0021] One example of the present invention relates to the use of a composition containing kestose as an active ingredient to inhibit the growth of or reduce the content of harmful microorganisms, such as microorganisms of the genus Prevotella.

[0022] One example of the present invention relates to the use of a composition containing kestose as an active ingredient to promote the growth of propionic acid-producing bacteria. The composition can increase the content of propionic acid in the intestines.

[0023] In one example of the present invention, the composition may be a pharmaceutical composition or a food composition. The composition may be a prebiotic composition or may be used as a synbiotic by additionally including probiotics. Probiotics may refer to microorganisms that have a positive effect on health in the body. Prebiotics may refer to ingredients that are used by microorganisms, including beneficial bacteria, to promote the growth and activity of microorganisms, thereby resolving imbalances in the intestinal microbiome and providing a positive effect on the health of the host. The probiotic strain or probiotic microorganism may be in the form of living cells or non-viable cells.

[0024] The microorganisms applicable to the present invention as the probiotic microorganisms can include one or more probiotics selected from the group consisting of microorganisms of the genus Bifidobacterium, microorganisms of the genus Streptococcus, microorganisms of the genus Saccharomyces, microorganisms of the genus bacillus, microorganisms of the genus Alistipe, and microorganisms of the genus Lactobacillus.

[0025] The active ingredient of the present invention promotes the proliferation of one or more microorganisms from the group consisting of Lactobacillus and Alistipes microorganisms in the intestines of a subject in need of improvement of the intestinal microbiome, thereby increasing the proliferation of bacteria that have beneficial effects in the intestines, and as a result, can bring about an improvement effect of the intestinal microbiome.

[0026] The composition of the present invention contains kestose, which can increase the number of Lactobacillus microorganisms and Alistipes microorganisms and reduce Prevotella microorganisms, thereby increasing the production of SCFAs. Therefore, according to the present invention, it is possible to simply and effectively increase Lactobacillus microorganisms and / or Alistipe microorganisms in a subject's body with little concern about side effects or safety. Furthermore, according to the present invention, kestose has the advantage of improving the intestinal environment by increasing Lactobacillus microorganisms and / or Alistipe microorganisms in a subject's body. Furthermore, kestose can improve the intestinal environment and achieve a balance of intestinal microorganisms by reducing Prevotella microorganisms in the human or animal body.

[0027] The genus Alistipe is classified as a gram-negative, rod-shaped, anaerobic, and non-spore-forming organism, and includes, but is not limited to, Alistipes finegoldii, Alistipes putredinis, Alistipes onderdonkii, Alistipes shahii, Alistipes indistinctus, Alistipes senegalensis, Alistipes timonensis, Alistipes obesi, Alistipes ihumii, Alistipes inops, Alistipes megaguti, and Alistipes provencensis.

[0028] Lactobacillus microorganisms include L.mucosae, L.salivarius, L.acidophilus, L.plantarum, L.pentosus), L.arizonensis, L.rhamnosus, L.brevis, L.caseii, L.paracasei, Including, but not limited to, L.bulgaricus, L.delbruckii, L.arabinosus, L.caucasicus, L.leishmanni, L.musicus, L.themophilus, L.fermentum, and L.helveticus.

[0029] As used herein, the terms "microbiota," "microflora," and "microbiome" refer to the living community of microorganisms that typically inhabit a body organ or part, particularly the gastrointestinal tract of a subject, e.g., an animal or human.

[0030] The growth promotion or inhibition of the microorganisms is expressed in terms of the content of intestinal microorganisms, the content of intestinal microbial metabolites, and / or the relative abundance (%) of intestinal microorganisms. As used herein, the "relative abundance" of a bacterial species refers to the relative abundance of a specific bacterium relative to other bacterial species excluding the specific bacterial species in the human gastrointestinal microbiota.

[0031] The modulation of gut microorganisms by the compositions of the present invention can include increasing the relative abundance (%) of Lactobacillus and / or Alistipe microorganisms in the gut, and decreasing the relative abundance (%) of Prevotella microorganisms in the gut.

[0032] The composition of the present invention may increase the relative abundance (%) of fecal or intestinal Lactobacillus microorganisms or fecal or intestinal Alistipe microorganisms by 5% or more, or 7% or more, for example, 5% to 100%, which may be 105% or more or 107% or more, for example, 105% to 200%, based on a pre-ingestion level of 100% for the active ingredient.

[0033] Specifically, the composition of the present invention may be a composition containing kestose as an active ingredient, which achieves an increase in the relative abundance (%) of Lactobacillus microorganisms in feces or intestines of 50-70%, 50-65%, or 50-60%, and / or an increase in the relative abundance (%) of Alistipe microorganisms in feces or intestines of 5-25%, 5-20%, or 5-15%. The relative abundance (%) of Lactobacillus microorganisms in feces or intestines may be 150-170% based on a pre-ingestion level of the active ingredient (100%), and the relative abundance (%) of Alistipe microorganisms in feces or intestines may be 105-125% based on a pre-ingestion level of the active ingredient (100%).

[0034] The composition of the present invention may contain kestose as an active ingredient, which reduces the decrease in the relative abundance (%) of Prevotella microorganisms in feces to 0.5% to 2.5%, 0.5 to 2%, or 0.5 to 1.5%. The relative abundance (%) of Prevotella microorganisms in the feces or intestines may be 97.5 to 99.5% based on a pre-ingestion level of 100% for the active ingredient.

[0035] Yet another example of the present invention relates to a composition containing kestose as an active ingredient for increasing the content of short-chain fatty acids, such as propionic acid, in the intestine, or a composition containing kestose as an active ingredient for growing or increasing propionic acid-producing bacteria in the intestine.

[0036] The present invention also relates to a method for increasing the intestinal propionic acid content or a method for growing intestinal propionic acid-producing bacteria, comprising the step of administering a composition containing kestose to a subject in need thereof.

[0037] The composition may be a food composition, a health functional food, or a pharmaceutical composition.

[0038] The increased fecal or intestinal propionic acid content may be due to proliferation of intestinal propionic acid-producing microorganisms, which proliferation is expressed as an increased content of intestinal microorganisms, an increased propionic acid content, and / or an increased relative abundance (%).

[0039] The intestinal propionic acid-producing microorganisms may include microorganisms known in the art, for example, one or more microorganisms selected from the group consisting of intestinal Lactobacillus spp. and Alistipe sspp.

[0040] Specifically, it was confirmed that the short-chain fatty acid-producing bacteria Lactobacillus spp. and Alistipes spp. significantly increased in both groups administered Advanced Fos (powder and syrup - medium / high concentrations), while Prevotella spp. significantly decreased in the groups administered Advanced Fos (powder and syrup - medium and high concentrations) (see Test Example 4). Prevotella spp. are associated with autoimmune diseases, insulin resistance, diabetes, and the induction of intestinal inflammation, and are known to play a role in reducing the production of short-chain fatty acids.

[0041] One example of the present invention provides a kestose-containing composition characterized by a propionic acid content in feces of 100±30 mg / L, 150±30 mg / L, 200±30 mg / L, 250±30 mg / L or 300±30 mg / L or more.

[0042] One example of the present invention provides a kestose-containing composition characterized by an increase in fecal propionic acid content of 100 to 700%, 100 to 500%, 100 to 300%, 130 to 500%, 130 to 700%, 150 to 500%, 150 to 700%, 150 to 300%, or 130 to 300% based on the pre-ingestion level.

[0043] One example of the present invention provides a kestose-containing composition characterized in that the weight ratio of butyric acid to propionic acid in the feces or intestines is 4:1 to 1:3, 3:1 to 1:3, 3:1 to 1:2, 2:1 to 1:2, or more preferably 1:1 to 1:2. Increasing the ratio of propionic acid to butyric acid in the feces or intestines to this ratio can maintain an acidic pH in the intestinal tract, calm immune cells, and maximize the anti-inflammatory effect.

[0044] Another example of the present invention relates to a composition containing kestose for preventing, ameliorating, or treating intestinal diseases and / or inflammatory diseases. The diseases may be caused by immune hypersensitivity reactions, such as inflammatory skin diseases and / or inflammatory bowel diseases caused by immune hypersensitivity reactions. The composition may be a food composition, a health functional food, or a pharmaceutical composition.

[0045] The composition disclosed herein not only improves bowel function and intestinal function by promoting the proliferation or growth of beneficial intestinal bacteria and suppressing the proliferation or growth of harmful intestinal bacteria, thereby improving the intestinal flora, but also acts as a prebiotic, thereby enhancing the efficacy of probiotics when used in combination with probiotics, resulting in a synergistic effect. In an exemplary embodiment, the composition may be a composition for preventing, ameliorating, or treating, for example, constipation, diarrhea, or intestinal-related diseases.

[0046] In addition, it is known that intestinal microbial dysbiosis is highly associated with inflammatory diseases, such as inflammatory bowel disease (IBD) and inflammatory skin diseases. Therefore, the composition containing kestose according to the present invention can provide a composition for preventing, ameliorating, or treating inflammatory diseases.

[0047] While various possibilities, including genetic and environmental factors, have been discussed regarding the pathogenesis of inflammatory bowel disease, recent research has suggested that abnormalities in intestinal immunity (gastrointestinal mucosal immunity) may be a contributing factor. Recent studies have also suggested that abnormal immune responses to certain resident bacteria may manifest as chronic inflammatory reactions. The mechanism by which intestinal inflammation and allergies develop due to abnormalities in intestinal immunity is supported by analyses of the function and differentiation of T cells in patients and the cytokine production patterns in lesions and serum. Furthermore, analyses of various recently developed animal models of inflammatory bowel disease (Gastroenterology, Vol. 109, pp. 1344-1367, 1995) have also revealed that abnormalities in mucosal immunity cause chronic inflammation in the intestine.

[0048] As used herein, the terms "subject," "individual," "host," and "patient" are used interchangeably herein and refer to any animal subject, including humans, laboratory animals, livestock, and household pets, preferably humans. A subject can host a diverse range of microorganisms. A subject can be diagnosed or suspected of being at high risk for disease and can have a microbiome condition (i.e., a microbial imbalance) that contributes to disease.

[0049] The inflammatory disease according to the present invention may preferably be an inflammatory skin disease or inflammatory bowel disease caused by an immune hypersensitivity reaction, and the inflammatory skin disease may be atopic dermatitis, contact dermatitis, or allergic contact dermatitis, and the inflammatory bowel disease may be Crohn's disease (CD) or ulcerative colitis (UC), with UC being known as a chronic inflammatory disease that primarily targets the colon.

[0050] The inflammatory disease may be an inflammatory skin disease or inflammatory bowel disease caused by an immune hypersensitivity reaction, and the experimental group (Advanced Fos: powder and syrup - medium and high concentrations) was found to contribute to restoring the immune system damaged by the immune reaction, suppressing excessive immune cell activity, and improving atopic symptoms (see Test Example 8).

[0051] Inflammatory skin diseases, such as atopic dermatitis, are caused by an imbalance between the gut microbiome and Th1 / Th2, and it is known that the symptoms of AD are exacerbated by a decrease in Th1 IFN-r, which is typically involved in IgE expression, and an increase in Th2 IL-13. Furthermore, IL-5, IL-13, TARC, and eotaxin, which are associated with eosinophils in histological analysis, induce an increase in eosinophil numbers, skin influx, and degranulation, leading to inflammation and tissue damage.

[0052] Specifically, in this study, the Advanced Fos group found an increase in Lactobacillus spp. and Alistipes spp. and a decrease in Prevotella spp., which was confirmed to be associated with an increase in SCFA secretion, and confirmed that these results contribute to the decrease in Th2 expression, which is increased in patients with atopic dermatitis. Furthermore, by upregulating the expression of Th1-related factors and downregulating the expression of Th2-related factors to balance Th1 / Th2 not only in the blood but also in the intestine and skin, AD symptoms are improved.

[0053] The present invention confirmed the efficacy of kestose-containing fructooligosaccharides in improving atopic dermatitis (AD) through experiments on different dosage forms (powder, syrup) and concentrations.

[0054] Specifically, in an animal model with induced atopic dermatitis, administration of the composition of the present invention reduced serum IgE levels, IL-13 secretion, which is involved in IgE production, TARC secretion, which is associated with increased eosinophil counts, Eotaxin secretion, which is involved in eosinophil influx and degranulation, and IL-4 and IL-5 secretion, which cause Th1 / Th2 imbalance. Therefore, administration of the experimental group (Advanced Fos: powder and syrup - medium / high concentrations) improved atopic symptoms by regulating cytokine secretion, suppressing Th2 immune responses, and restoring Th1 / Th2 balance.

[0055] Furthermore, when the composition of the present invention was administered to an animal model with induced atopic dermatitis, there was a relative increase in the secretion of TNF-α, Th1-related IFN-γ, and Th1-related IL-12. Administration of the experimental group (Advanced Fos: powder and syrup - medium concentration / high concentration) improved atopic symptoms by regulating the secretion of Th1-related cytokines and restoring the Th1 / Th2 balance (see Figure 17).

[0056] When the composition of the present invention was administered to an animal model in which atopic dermatitis was induced, the administration suppressed excessive Treg immune responses, suppressed inflammatory responses, and restored the balance between Th1 and Th2, thereby improving atopic symptoms, by reducing the secretion of Treg-related cytokines, such as IL-1β, and IL-10, which causes an imbalance between Th1 and Th2.

[0057] In addition, the composition of the present invention was administered to an animal model in which atopic dermatitis was induced, and the effect of improving the severity of atopic dermatitis was evaluated using itching score and severity score. As a result, it was confirmed that dermatitis was alleviated in the experimental group (see Figure 15).

[0058] The kestose of the present invention can be used alone or as a sugar composition containing the same, and can be used in liquid or powder form, and the powder may be amorphous or crystalline. The kestose may be purchased as a commercially available product or may be produced using predetermined raw materials.

[0059] The kestose that can be used in the present invention can be in any form, liquid or powder, and can be contained in the composition of the present invention in various amounts.

[0060] The kestose may be used as a single component or as a mixed composition containing other sugars.

[0061] In the composition containing kestose, the liquid or powdered kestose content is 50% (w / w) or more, 60% (w / w) or more, 70% (w / w) or more, 80% (w / w) or more, or 85% (w / w) or more, based on the total solid content of sugars, for example, fructooligosaccharides. Also, the crystalline kestose content may be 90% (w / w) or more, 95% (w / w) or more, or 98% (w / w) or more, based on the total solid content of sugars.

[0062] The kestose-containing composition may be a fructooligosaccharide (FOS). For example, it may contain a high content of 1-kestose (GF2) and additionally contain one or more saccharides selected from the group consisting of nystose (GF3) and 1-F-fructosyl nystose (GF4). FOS consists of a linear chain having 1 to 9 fructose residues linked to sucrose molecules by β2→1 bonds. The kestose-containing fructooligosaccharide preferably has a high 1-kestose content, and more preferably, 1-kestose is the main component. For example, it may contain a high content of 1-kestose (GF2) in an amount of 50% by weight or more based on the solid content of the saccharides.

[0063] Some commercially available FOS products contain small amounts of kestose (1-kestose) at approximately 20-35% (w / w) of the total solids. However, in the present invention, the active ingredient kestose (1-kestose) exhibiting beneficial effects is contained in a content of 50% (w / w) or more of the total solids of sugars. This allows for effective regulation of the daily intake (dosage) of kestose, thereby inducing a balance in body metabolism through regulation of the intestinal microbial community, the microbiome, and microbial metabolites. When consuming a sugar composition with a low kestose content, such as fructooligosaccharides, the intake of fructooligosaccharides increases due to the effective amount of kestose consumed according to the present invention. This can lead to discomfort in the digestive tract (diarrhea, abdominal distension, borborygmus, etc.), which may result in undesirable effects. Therefore, by ingesting a sugar composition having a high kestose content, such as fructooligosaccharides, the undesirable problems caused by excessive intake of fructooligosaccharides can be reduced, and further, the intake of an effective content of kestose according to the present invention can be easily achieved to further increase efficacy.

[0064] Kestose or a sugar composition containing kestose that can be used in the present invention is not particularly limited, and can generally be produced using an enzyme having kestose conversion activity or a microorganism that produces the enzyme, using sugar as a substrate. The kestose-containing product thus produced may be used as is, or may be used as a sugar composition having an increased kestose content after being subjected to extraction and purification processes.

[0065] The enzyme having kestose conversion activity is an enzyme having the activity of converting a substrate containing sugar into a fructooligosaccharide containing kestose, and may be, for example, an enzyme derived from one or more strains selected from the group consisting of Aspergillus niger strains, Pichia farinosa strains, Yarrowia lipolytica, Millerozyma farinosa, and Aspergillus oryzae strains.

[0066] The dosage form of medicines, quasi-drugs, and supplements containing kestose is not particularly limited, and a dosage form suitable for the administration method can be appropriately selected. For example, in the case of oral administration, it may be in the form of a solid or liquid dosage form such as powder, tablet, sugar-coated agent, capsule, granule, dry syrup, liquid, syrup, drop, or drink.

[0067] The total daily intake of kestose as an active ingredient in the kestose composition may be 2 to 30 g, 2 to 27 g, 2 to 26 g, 2 to 25 g, 2 to 24 g, 2 to 20 g, 2 to 15 g, 2 to 10 g, or 2 to 8 g for a 60 kg adult. This intake amount is not limited to a single dose per day, but can be divided into multiple doses. The content can be set appropriately, taking into consideration the effects of ingestion of kestose or fructooligosaccharides containing kestose on intestinal regulation, improvement of intestinal function, and enhancement of calcium absorption, as well as the side effect of "heavy stomach feeling."

[0068] The kestose composition may contain kestose, nystose, and fructofuranosyl nystose in a combined amount of 400 mg / g or more, 500 mg / g or more, 800 mg / g or more, or 900 mg / g or more. The kestose composition may have a weight ratio of kestose to the total weight of nystose and fructofuranosyl nystose of 1:1 to 10:1, 1:1 to 5:1, 1:1 to 3:1, or 1:1 to 2:1.

[0069] The subjects or individuals to which the composition of the present invention containing kestose as an active ingredient is administered or ingested are animals including humans, and include, for example, humans, mice, rats, monkeys, etc.

[0070] The composition of the present invention containing kestose as an active ingredient can be ingested before, after, or simultaneously with a meal, and the conditions for ingestion are not particularly limited. The composition containing kestose as an active ingredient can be ingested for a period effective for achieving improvement of inflammatory diseases, improvement of intestinal function, or change in the intestinal microbial community or environment, for example, for a period of 4 weeks or more, 8 weeks or more, or 15 weeks or more, for example, 4 to 20 weeks. Although effective effects can be obtained by ingesting the composition of the present invention only once, it may also be ingested two or more times.

[0071] A dosage unit for ingestion of the composition containing kestose as an active ingredient can be prepared to contain, for example, 1, 2, 3, or 4 times the amount of an individual dose, or 1 / 2, 1 / 3, or 1 / 4 of the amount. An individual dose preferably contains an amount of the active ingredient to be administered once a day, which is usually formulated in an amount equivalent to the whole, 1 / 2, 1 / 3, or 1 / 4 of the daily dose and can be administered in divided doses.

[0072] The composition containing kestose according to the present invention may be a food composition or a pharmaceutical composition.

[0073] The appropriate dosage of the composition of the present invention can be variously formulated depending on factors such as the formulation method, administration method, age, weight, sex, pathological condition, diet, administration time, administration route, excretion rate, and reaction sensitivity of the patient.

[0074] The composition of the present invention containing kestose as an active ingredient may be a food, food additive, beverage, beverage additive, health food, or functional food. In the present invention, "health functional food" refers to a food manufactured (including processed) using raw materials or ingredients that have functional properties beneficial to the human body under the Act on Health Functional Foods, and "functionality" refers to obtaining beneficial effects for health purposes, such as maintaining the normal functions of the human body or activating physiological functions to maintain and improve health. Kestose can be added to general foods or produced as an encapsulated, powdered, or suspension. When ingested, it provides specific health benefits, and, unlike general medicines, it has the advantage of being made from food ingredients, eliminating the side effects that can occur with long-term drug use.

[0075] When the kestose of the present invention is used as a food additive, it can be added as is, used together with other foods or food ingredients, or used appropriately by other conventional methods. The amount of active ingredient to be mixed can be suitably determined depending on the purpose of use (prevention, health, or therapeutic treatment).

[0076] The kestose or a composition containing kestose is administered to a subject, such as a human or animal, by oral ingestion either as is or in the form of a food, drink, or pharmaceutical.

[0077] Kestose (1-kestose) can be added to various foods, beverages, food additives, and animal feed during the normal manufacturing process. 1-kestose has a sweetness index of 30, and its quality of taste, physical properties, and processability are similar to those of sucrose. Therefore, it can be used in the same way as sugar, such as by substituting 1-kestose for part or all of the sugar in the manufacturing process of various foods and beverages, and in various foods, beverages, food additives, medicines, and feed.

[0078] Specific embodiments of the composition according to the present invention include, for example, beverages, dairy products, edible granules, pastes, seasonings, retort foods, baby foods, fermented foods, preserved foods, processed seafood products, processed meat products, processed grain products and other processed foods, food additives, health foods, animal feeds, and the like. [Example]

[0079] The present invention will be described in more detail with reference to the following examples, but the scope of the present invention is not intended to be limited to the following examples.

[0080] Example 1: Construction of an animal model The general schedule for constructing the animal model and the dietary administration experiment in this experiment is shown in FIG. Specifically, to construct an animal model of atopic dermatitis, 5-week-old female Balb / c mice were separated into groups of six and allowed to adapt for one week. For all groups except the negative control group (Neg), an OVA / Al(OH)3 mixture was prepared by dissolving OVA and Alum (Sigma-Aldrich) in PBS. This mixture was administered intraperitoneally four times (once every two weeks, for a total of four administrations) on days 7, 21, 35, and 49 from the start of atopic dermatitis induction for eight weeks. Additionally, the backs of the Balb / c mice were continuously shaved for eight weeks, and AD was induced by continuous contact with OVA / Al(OH)3 on the back skin. These experimental animals with atopic dermatitis were used as experimental animal models in subsequent experiments. For the negative control group, 5-week-old female Balb / c mice were separated into groups of 6 mice and allowed to adapt for 1 week. The procedure was similar to that described in (1) above, except that atopy was not induced for 8 weeks using the OVA / Al(OH)3 mixture, and instead, PBS was orally administered at the same dose as the experimental group. For the positive control group, 5-week-old female Balb / c mice were separated into groups of 6 mice and allowed to adapt for 1 week. Then, atopy was induced with OVA / Al(OH)3 for 8 weeks, similar to the construction of the atopic animal model. During the administration period, PBS was orally administered at the same dose as the experimental group. Fecal samples were collected from the experimental, negative control, and positive control animals at weeks 8, 12, and 16 for subsequent microbiome and SCFA analysis. Cecal fecal samples were also collected at the end of the experiment for microbiome profiling and metabolite analysis.

[0081] Comparative Examples 1 and 2: Maltose feeding experiments The maltose-fed animal model was an experimental animal model in which atopy was induced with OVA / Al(OH)3 for 8 weeks in Example 1 and then continued on the same diet. During the administration period, maltose (Daezo Co., Ltd., product name: Maltose D) was dissolved in 200 μl of PBS at a concentration per mouse and orally administered at the same concentration. Specifically, in Comparative Example 1-1, Maltose was applied at a medium concentration of 8 g / day, and in Comparative Example 1-2, Maltose was applied at a high concentration of 16 g / day.

[0082] Comparative Examples 3 and 4: Experiments with FOS powder diets containing low amounts of kestose For the experimental group administered FOS, atopy was induced with OVA / Al(OH)3 for 8 weeks, and then FOS was dissolved in 200 μl of PBS per mouse with a kestose content (33%) during the administration period to prepare a sample for oral administration. The low kestose-containing FOS powder has a fructooligosaccharide content of 95% based on the total solids, and the kestose content of the fructooligosaccharides is 33% by weight, so the dietary powder contains 31.35% by weight of kestose based on the total solids. Each sample was dissolved in PBS and the solution was divided into two doses for the experiment, administered twice a day for an adult (60 kg). Comparative Example 3 was administered to the medium concentration group at 8 g / day, and Comparative Example 4 was administered to the high concentration group at 16 g / day for 8 weeks. The high-concentration and medium-concentration diets were divided into two groups based on daily dose of the same diet composition, with the medium-concentration group administered at a dose of 8 g / day and the high-concentration group administered at a dose of 16 g / day.

[0083] [Table 1]

[0084] Examples 1 and 2: Experiments with high-content kestose-containing FOS powder diets For the experimental group administered high-content kestose-containing FOS powder, atopy was induced with OVA / Al(OH)3 for 8 weeks, and then the high-content kestose-containing FOS powder was dissolved in 200 μl of PBS at a concentration per mouse during the administration period to prepare a sample for oral administration. The high kestose-containing FOS powder used in this example had a fructooligosaccharide content of 91.2% based on the total solids, and the kestose content of the fructooligosaccharides was 85.5% by weight. Therefore, the dietary powder contained 77.98% by weight of kestose based on the total solids. Each sample was dissolved in PBS and administered in two doses, twice daily, for an adult (60 kg). Example 1 was administered at a dose of 8 g for the medium concentration group, and Example 2 was administered at a dose of 16 g for the high concentration group, for an adult (60 kg) for 8 weeks. The animal model was fed a diet in substantially the same manner as in Comparative Example 3, except that the FOS powder of Comparative Example 3 was replaced with the FOS powder containing a high amount of kestose produced according to the present invention.

[0085] Examples 3 and 4: High-kestose-containing FOS syrup dietary experiment In the experimental group administered high-content kestose FOS syrup, atopy was induced with OVA / Al(OH)3 for 8 weeks, and then the high-content kestose FOS syrup was dissolved in 200 μl of PBS at a concentration based on one mouse, with the total solids (89.9%), kestose content (85.6%), and solids of the FOS syrup (75.3%). This was then used to prepare a sample for oral administration. The high-kestose-containing FOS syrup used in this example had a fructooligosaccharide content of 91.2% based on the total solids, and the kestose content of the fructooligosaccharides was 89.9% by weight. Therefore, the feed powder contained 81.998% by weight of kestose based on the total solids. Each sample was dissolved in PBS and administered in two doses, twice daily, for an adult (60 kg). Example 3 was administered to the medium-dose group with a dose of 8 g, and Example 4 was administered to the high-dose group with a high dose of 16 g for 8 weeks. The animal model was fed a diet in substantially the same manner as in Comparative Example 3, except that the FOS powder of Comparative Example 3 was replaced with the high kestose-containing FOS syrup produced according to the present invention.

[0086] Test Example 1: Confirmation of mouse weight gain and food intake During the administration period, the changes in the body weight of the mice in each experimental group were recorded, and the weights of the mice were measured at the same time every week before oral administration. To confirm whether or not the intake of fructooligosaccharides caused obesity, the food intake of mice in each experimental group was checked during the administration period, and the food intake was checked at the same time every week. Figure 2 shows the results of weight changes in experimental animal models following administration of the experimental groups, and shows that weight gain was reduced due to the induction of atopy in the positive control group and the maltose-administered groups of Comparative Examples 1 and 2. In the experimental groups of Examples 1 to 4, weight gain was similar to that of the negative control group, indicating that weight gain was restored due to the improving effect of Advanced FOS of Examples 1 to 4 on atopy.

[0087] Test Case 2: Analysis of species diversity in the gut microbiome (1) Microbial community analysis of fecal samples For the samples of Comparative Examples 1 to 4 and Examples 1 to 4, fecal samples were collected at 8 weeks, 12 weeks, and 16 weeks throughout the entire study period from the experimental animals of Production Example 1. DNA was extracted from the collected fecal samples at 8 weeks, 12 weeks, and 16 weeks using a DNA soil kit. The 180 DNA samples were sequenced using the Illumina Miseq platform, yielding a total of 13,117,845 16S rRNA sequence read counts, with an average of 85,737 reads per sample, and all groups yielding reads greater than 2500. 16S rRNA sequences were analyzed using the qiime2 pipeline, and missing or unknown 5' or 3' sequences were removed. Trimmed sequences less than 250 bp were removed, and chimera sequences were removed using the chimera in the qiime2 pipeline. This allowed us to identify each sequencing read in the sample and confirm the microbial community structure. The Shannon index, Simpson index, and inverse Simpson index were calculated for alpha-diversity analysis of the gut microbiome. Beta-diversity analysis was performed using Adonis analysis with Bray-Curtis distance metrics. Graphs were visualized using R software (version 4.0.3).

[0088] (2) Analysis of gut microbiome species diversity and intergroup differences To analyze the species diversity of the gut microbiome and differences between groups, fecal samples obtained from the experimental animals in Comparative Examples 1-4 and Examples 1-4 at week 16 of the study were subjected to low-quality sequencing to confirm an average sequence count of 85,737. To examine the similarity between the positive and negative control groups (atopic mice), the species diversity (alpha-diversity) of gut microbes was examined. The alpha diversity results showed no differences between the groups in the Shannon, Simpson, and InvSimpson diversity indices (Figure 3). However, microbial beta-diversity confirmed that the communities in each group were separated. In particular, the positive control group and maltose group are on the right, but the experimental group showed similar changes to the negative control group (Figure 4).

[0089] Test Example 3: Profiling of the intestinal microbiome Fecal samples were collected from the experimental animals in Comparative Examples 1 to 4 and Examples 1 to 4 at weeks 8, 12, and 16 throughout the study period. DNA was extracted from the collected fecal samples at weeks 8, 12, and 16 using a DNA soil kit. The DNA samples were used to perform profiling analysis of the gut microbiome between the control group and the experimental group at weeks 8, 12, and 16. The microbiome can be designated by taxonomy, which groups organisms according to their lineage and subordination. The phylum level was confirmed for samples taken at 8, 12, and 16 weeks for each group. At the phylum level, Bacteroidetes and Firmicutes were generally found to be predominant in all groups for samples taken at 8, 12, and 16 weeks (Figure 5).

[0090] Additionally, when comparing the phylum level of the 16-week samples with the positive control group, the Firmicutes phylum increased and the Bacteroidetes phylum decreased in the Advanced Fos powder high concentration group (Example 2) and the Advanced Fos syrup medium and high concentration groups (Example 4). More specifically, when compared with the positive control group (1.83% ± 0.05), the relative abundance ratio of Firmicutes:Bacteroidetes was found to be significantly increased in the high-concentration kestose-containing FOS powder group (Example 2) (2.72% ± 0.15, p = 0.0048), the medium-concentration kestose-containing FOS syrup group (Example 3), and the high-concentration kestose-containing FOS syrup group (Example 4) (medium concentration: 2.42% ± 0.15, p = 0.0245; high concentration: 3.34% ± 0.15, p < 0.0001). There was no significant difference between the positive control group and the maltose groups of Comparative Examples 1 and 2 (medium concentration of Comparative Example 1: 2.11% ± 0.07, p = 0.2184, high concentration of Comparative Example 2: 1.58% ± 0.10, p = 0.26) (Figure 6). In summary, it was confirmed that Bacteroidetes and Firmicutes predominate at the phylum level, and at week 16, Firmicutes increased relative to Bacteroidetes in the experimental groups of Examples 1 to 4, unlike the positive control group and the Maltose groups of Comparative Examples 1 and 2. In the medium-concentration group containing high-content kestose-containing Fos powder of Example 1, the proportion of Bacteroidetes did not decrease, but genus profiling confirmed a significant increase in Alistipes and other genus species.

[0091] Heatmap analysis of the top 30 genus levels was used to identify the intestinal microorganisms in the 16-week samples by group. At the genus level, Lactobacillus, Alistipes, Prevotella, Muribaculum, and Bacteroides were identified as the dominant species in the intestine. More specifically, four genera were dominant: Lactobacillus, Muribaculum, Alistipes, Bacteroides, and Prevotella. Lactobacillus was found to have a higher occupancy rate in the normal control group and the group administered the high-concentration Advanced Fos syrup (Example 4) (63.70% ± 2.31, p = 0.0022). In contrast, the group administered with medium concentration Advanced Fos powder in Example 2 visually confirmed a greater increase in Alistipes than the groups administered with medium concentration Advanced Fos syrup in Example 3 and high concentration Advanced Fos syrup in Example 4 (Figures 7 and 8).

[0092] Test Example 4: Analysis of SCFA-producing bacteria (1) Increase in SCFA-producing bacteria due to administration to the experimental group When fecal samples obtained from the experimental animals in Comparative Examples 1 to 4 and Examples 1 to 4 at week 16 of the study period were compared with those from the positive control group and the maltose group, it was confirmed that the short-chain fatty acid-producing bacteria Lactobacillus spp. and Alistipes spp. were significantly increased in all experimental groups (Advanced Fos: powder and syrup - medium concentration / high concentration) administered in Examples 1 to 4.

[0093] (2) Lactobacillus spp. It was confirmed that Lactobacillus spp. increased significantly in the groups administered Advanced Fos powder high concentration in Examples 1 and 2 and Advanced Fos syrup medium concentration / high concentration in Examples 3 and 4. It was confirmed that Lactobacillus spp. gradually increased over 8, 12, and 16 weeks. In particular, at 16 weeks, compared with the positive control group (39.61% ± 4.61), significant increases were observed in the Advanced Fos powder high concentration group (Example 2) (63.70% ± 2.31, p = 0.0022) and the Advanced Fos syrup medium / high concentration groups (Medium concentration: 58.91% ± 3.28, p = 0.0043; High concentration: 66.31% ± 4.35, p = 0.0022) administered in Examples 3 and 4. There was no significant difference between the positive control group and the maltose groups (Medium concentration: 40.71% ± 5.48, p = 0.8571; High concentration: 35.85% ± 0.61, p = 0.0905) administered in Comparative Examples 1 and 2. Lactobacillus spp. (family Lactobacillaceae, phylum Firmicutes) are short-chain fatty acid-producing bacteria and are known to be beneficial to the intestine.

[0094] (3) Alistipes spp. There was no significant difference in Alistipes spp. between weeks 8 and 12, but a significant increase was observed at week 16. A greater increase in Alistipes spp. was observed in the Advanced Fos powder medium concentration group (Example 1) than in the Advanced Fos syrup medium / high concentration groups (Examples 3 and 4). In particular, at week 16, significant increases were observed in the Advanced Fos powder medium / high concentration groups (Examples 1 and 2) (medium concentration: 14.73% ± 1.43, p = 0.0043; high concentration: 14.38% ± 1.97, p = 0.0043) and the Advanced Fos syrup medium / high concentration groups (Examples 3 and 4) (medium concentration: 8.74% ± 0.40, p = 0.0095; high concentration: 6.59% ± 0.25, p = 0.0152), compared to the positive control group (3.47% ± 0.99). There was no significant difference between the positive control group and the maltose groups of Comparative Examples 1 and 2 (medium concentration of Comparative Example 1: 4.22% ± 1.51, p = 0.4429; high concentration of Comparative Example 2: 1.76% ± 0.43, p = 0.6095). Alistipes spp. (family Rikenellaceae, phylum Bacteroidetes) are short-chain fatty acid-producing bacteria known to be beneficial to the intestines.

[0095] (4) Prevotella spp. When compared with the positive control group and the maltose groups of Comparative Examples 1 and 2, a significant reduction in Prevotella spp. was confirmed in the experimental groups administered Advanced Fos (powder and syrup - medium and high concentrations) according to Examples 1 to 4. In particular, at 16 weeks, when compared with the positive control group (5.37% ± 1.40), a significant reduction was confirmed in the Advanced Fos powder medium / high concentrations according to Examples 1 and 2 (medium concentration in Example 1: 1.56% ± 0.37, p = 0.0043, high concentration in Example 2: 0.90% ± 0.35, p = 0.0043) and the Advanced Fos syrup medium / high concentration groups according to Examples 3 and 4 (medium concentration in Example 3: 1.58% ± 0.36, p = 0.0087, high concentration in Example 4: 1.69% ± 0.33, p = 0.0043). There was no significant difference between the positive control group and the maltose group (medium concentration: 2.80% ± 0.16, p = 0.1077, high concentration: 4.31% ± 0.47, p = 0.8571). Prevotella spp. are associated with autoimmune diseases, insulin resistance, diabetes, and intestinal inflammation, and are known to play a role in reducing the production of short-chain fatty acids.

[0096] Test Example 5: Analysis of intestinal metabolites (SCFA) (1) Measurement of fecal butyrate content Stool samples (100 mg) were homogenized in NaOH containing 5 μg / ml caproic acid (Sigma-Aldrich) and centrifuged for 20 minutes at 12,000 g and 4°C. The supernatant was transferred to a tube (Corning, USA) and stored at -80°C until HPLC analysis. SCFAs were analyzed using an Aminex 87H column (300 × 10 mm; Bio-Rad, USA), and butyric acid was detected using an RI-detector (ERC, RefractoMax520, Japan). In the experimental groups administered low-kestose-containing Fos (powder) from Comparative Examples 3 and 4, high-kestose-containing Advanced Fos (powder) from Examples 1 and 2, and high-kestose-containing Advanced Fos syrup from Examples 3 and 4, high butyrate content was measured in feces at 16 weeks, showing values ​​similar to those of the negative control group (Neg). Figure 12 is a graph comparing the butyric acid content in fecal samples of mice administered the experimental groups. As a result, at week 16, the butyric acid content in the positive control group and the groups administered maltose (medium concentration / high concentration) of Comparative Examples 1 and 2 was 32.2±4.9 mg / L, 45.5±3.9 mg / L, and 29.5±3.5 mg / L, respectively, while the experimental groups of Examples 1 to 4 (Advanced Fos: powder and syrup - medium concentration / high concentration) showed a significant increase of 172.8±5.8 mg / L, 232.6±43.2 mg / L, 188.1±27.4 mg / L, or 221.9±23.9 mg / L, respectively. Butyric acid is important for maintaining endogenous immunity and intestinal health and is involved in suppressing inflammation. Therefore, intake of the experimental group (Advanced Fos: powder and syrup - medium / high concentration) according to Examples 1 to 4 improved atopic disease by restoring immune response and intestinal health to normal levels.

[0097] (2) Measurement of fecal acetate content The content of acetic acid in the fecal samples was measured in the same manner as in Test Example 5-(1). Figure 13 is a graph comparing the acetic acid content in fecal samples of mice administered the experimental group. As a result of the comparison, at 16 weeks, the acetic acid content of the positive control group and the group administered maltose (medium concentration / high concentration) was 296.1±40.4mg / L, 304.9±46.3mg / L, and 277.1±20.8mg / L, respectively, while the experimental groups according to Examples 1 to 4 (Advanced Fos: powder and syrup - medium concentration / high concentration) showed a significant increase of 471.1±38.0mg / L, 561.2±75.4mg / L, 561.0±29.7mg / L, and 653.4±103.7mg / L, respectively.

[0098] (3) Measurement of fecal propionic acid content The propionic acid content of the fecal samples was measured in the same manner as in Test Example 5-(1). Figure 14 is a graph comparing the propionic acid content in fecal samples of mice administered the experimental group. As a result, at 16 weeks, the propionic acid content of the positive control group and the group administered maltose (medium concentration / high concentration) was 28.1±2.7mg / L, 96.1±17.2mg / L, and 42.8±6.2mg / L, respectively, while the experimental groups according to Examples 1 to 4 (Advanced Fos: powder and syrup - medium concentration / high concentration) showed a significant increase of 112.1±19.2mg / L, 158.2±35.6mg / L, 143.2±12.7mg / L, and 211.5±19.6mg / L, respectively.

[0099] Test Example 6: Evaluation of improvement in severity of atopic dermatitis The number of scratching incidents was measured for 15 minutes at the end of atopic dermatitis induction with OVA / Al(OH)3 (week 8) and at the end of experimental group administration (week 16). The severity of dermatitis was quantified using a severity score of 0 (none), 1 (mild), 2 (normal), and 3 (severe) based on the symptoms of dryness, erythema, and keratin. The final score was calculated by adding the number of scratching incidents and the severity score. Figure 15 is a graph comparing the effect of administration of the experimental groups on the severity of atopic dermatitis, quantified as itching score and severity score. It was confirmed that dermatitis was alleviated in the experimental groups administered Advanced Fos (powder and syrup) according to Examples 1 to 4 compared to the low-content kestose-containing Fos (powder) according to Comparative Examples 3 and 4, and that the improvement effect was particularly excellent in the experimental groups administered Advanced Fos according to Examples 1 to 4.

[0100] Test Example 7: Serological analysis of laboratory animals On the final day of administration, blood was obtained from the sacrificed mice, stored at 4°C for 1 hour, and then centrifuged at 5,000 x g for 1 hour to separate the serum. Serum IgE and cytokine (TNF-α, IFN-γ, IL-12, IL-4, IL-5, IL-13, TARC, eotaxin, IL-1β, IL-10) analyses were performed according to the manufacturer's guidelines for ELISA kits (R&D systems, USA), and absorbance was measured at 450 nm using a microplate reader (Tecan, Mönchsberg, Switzerland). FIG. 16 is a graph showing the analysis of serum IgE secretion levels following AD induction and administration to the experimental groups. As a result of comparing the secretion levels of IgE, which stimulates mast cells and worsens the symptoms of AD, the positive control group and the group administered maltose (medium concentration / high concentration) showed IgE secretion levels of 16.7±1.8 pg / ml, 16.4±1.1 pg / ml, and 13.13±1.3 pg / ml, respectively. Meanwhile, the Fos (powder) according to Comparative Examples 3 and 4 showed levels of 10.9±1.1 pg / ml and 10.7±0.64 pg / ml at medium and high concentrations, respectively. The Advanced Fos (powder) according to Examples 1 and 2 showed levels of 10.6±1.1 pg / ml and 9.9±1.3 pg / ml at medium and high concentrations, respectively. The Advanced Fos (powder) according to Examples 3 and 4 showed levels of 10.9±1.1 pg / ml and 10.7±0.64 pg / ml at medium and high concentrations, respectively. For the medium and high concentrations of Fos (syrup), the secretion of IgE was 9.2±1.6pg / ml and 8.3±0.6pg / ml, respectively, and a relative decrease in the secretion of IgE was confirmed in the experimental groups administered Fos (powder) and Advanced Fos (powder and syrup).

[0101] Figure 17 shows the results of analyzing the secretion of Th1-related cytokines following AD induction and administration to the experimental groups. In a graph comparing the secretion of TNF-α, the positive control group and the groups administered maltose (medium and high concentrations) showed TNF-α secretion levels of 23.8±2.6 pg / ml, 23.9±2.22 pg / ml, and 23.1±1.3 pg / ml, respectively, while the Fos (powder) of Comparative Examples 3 and 4 showed levels of 29.8±1.2 pg / ml and 31.7±0.9 pg / ml at medium and high concentrations, respectively. Furthermore, the levels of TNF-α secretion were relatively increased in the experimental groups administered with Advanced Fos (powder) from Examples 1 and 2 at medium and high concentrations of 32.4±1.3 pg / ml and 35.8±2.4 pg / ml, respectively, and the levels of Advanced Fos (syrup) from Examples 3 and 4 at medium and high concentrations of 32.3±1.1 pg / ml and 34.9±2.8 pg / ml, respectively, confirming that the levels of TNF-α secretion were relatively increased in the experimental groups administered with Advanced Fos (powder) from Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) from Examples 1 to 4. As a result of comparing the secretion levels of Th1-related IFN-γ, the positive control group and the group administered with maltose (medium concentration / high concentration) showed secretion levels of IFN-γ of 12.7±0.9pg / ml, 13.1±0.3pg / ml, and 13.7±0.3pg / ml, respectively. In contrast, the secretion levels of Fos (powder) according to Comparative Examples 3 and 4 at medium concentration / high concentration were 16.4±0.6pg / ml and 17.3±0.7pg / ml, respectively, of Advanced Fos (powder) according to Examples 2 and 3 at medium concentration / high concentration, 21.1±1.4pg / ml and 23.3±0.9pg / ml, respectively, of Advanced Fos (syrup) according to Examples 3 and 4 at medium concentration / high concentration, and 17.3±0.5pg / ml and 18.3±1.3pg / ml, respectively, of Advanced Fos (syrup) according to Comparative Examples 3 and 4 and Examples 1 to 4 at medium concentration / high concentration. It was confirmed that the amount of IFN-γ secreted was relatively increased in the experimental group administered with Fos (powder and syrup). As a result of comparing the secretion levels of Th1-related IL-12, the secretion levels of IL-12 in the positive control group and the group administered with maltose (medium concentration / high concentration) were 103.3±5.5pg / ml, 101.5±4.6pg / ml, and 106.3±9.6pg / ml, respectively. In contrast, the secretion levels of Fos (powder) according to Comparative Examples 3 and 4 at medium concentration / high concentration were 115.3±5.6pg / ml and 122.5±1.6pg / ml, respectively. The secretion levels of Advanced Fos (powder) according to Examples 1 and 2 at medium concentration / high concentration were 128.5±2.9pg / ml and 129.2±2.7pg / ml according to Examples 3 and 4. The secretion levels of Advanced Fos (syrup) according to Comparative Examples 3 and 4 and Advanced Fos (syrup) according to Examples 1 to 4 at medium concentration / high concentration were 126.5±3.5pg / ml and 125.8±3.0pg / ml, respectively. It was confirmed that the amount of IL-12 secreted was relatively increased in the experimental group administered Fos (powder and syrup). Therefore, administration of the experimental group according to Examples 1 to 4 (Advanced Fos: powder and syrup - medium / high concentration) improved atopic symptoms by regulating the secretion of Th1-related cytokines and restoring the Th1 / Th2 balance.

[0102] 18 shows the results of comparing the secretion levels of Th2-related cytokines following AD induction and administration to the experimental groups. Specifically, the secretion levels of IL-4, which suppresses Th1 cell differentiation and promotes Th2 cell differentiation, resulting in a Th1 / Th2 imbalance, were compared. The positive control group and the group administered maltose (medium / high concentrations) showed IL-4 secretion levels of 46.9±1.0 pg / ml, 43.5±1.6 pg / ml, and 42.1±2.2 pg / ml, respectively. Meanwhile, the Fos (powder) of Comparative Examples 3 and 4 showed 26.5±1.7 pg / ml and 23.1±3.5 pg / ml at medium and high concentrations, respectively. The Advanced Fos (powder) of Examples 1 and 2 showed 12.8±0.9 pg / ml and 9.3±0.6 pg / ml at medium and high concentrations, respectively. The Advanced Fos (powder) of Examples 3 and 4 showed 12.8±0.9 pg / ml and 9.3±0.6 pg / ml at medium and high concentrations, respectively. The levels of IL-4 secretion in the medium and high concentrations of Fos (syrup) were 13.1±2.5pg / ml and 11.3±1.1pg / ml, respectively, and it was confirmed that the amount of IL-4 secretion was relatively reduced in the experimental groups administered with Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4. Similar to IL-4, the secretion amount of IL-5, which causes Th1 / Th2 imbalance, was compared. As a result, the secretion amount of IL-5 in the positive control group and the group administered maltose (medium concentration / high concentration) was 136.9±6.8pg / ml, 130.9±8.3pg / ml, and 136.1±14.7pg / ml, respectively. In contrast, the secretion amount of IL-5 in the positive control group and the group administered maltose (medium concentration / high concentration) was 122.5±10.9pg / ml and 79.8±20.3pg / ml at medium concentration / high concentration, respectively. The ... Advanced Fos (powder) according to Examples 1 and 2 was 54.2±17.1pg / ml and 44.7±15.5pg / ml at medium concentration / high concentration, respectively. The secretion amount of IL-5 in the Advanced Fos (powder) according to Examples 3 and 4 was 54.2±17.1pg / ml and 44.7±15.5pg / ml at medium concentration / high concentration, respectively. The medium and high concentrations of Fos (syrup) were 76.3±12.4pg / ml and 68.9±12.7pg / ml, respectively, and it was confirmed that IL-5 secretion was relatively reduced in the experimental groups administered with Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4. As a result of comparing the secretion levels of IL-13, which is involved in the production of IgE, the positive control group and the group administered maltose (medium concentration / high concentration) showed IL-13 secretion levels of 324.8±31.4pg / ml, 316.6±58.4pg / ml, and 316.6±32.6pg / ml, respectively. In contrast, the secretion levels of Fos (powder) according to Comparative Examples 3 and 4 at medium concentration / high concentration were 225.6±5.7pg / ml and 168.7±7.1pg / ml, respectively, the secretion levels of Advanced Fos (powder) according to Examples 1 and 2 at medium concentration / high concentration were 143.0±11.3pg / ml and 128.4±10.6pg / ml, respectively, and the secretion levels of Advanced Fos (powder) according to Examples 3 and 4 at medium concentration / high concentration were 143.0±11.3pg / ml and 128.4±10.6pg / ml, respectively. The levels of IL-13 secretion were 178.4±14.5pg / ml and 138.8±6.9pg / ml at the medium and high concentrations of Fos (syrup), respectively, and it was confirmed that the amount of IL-13 secretion was relatively reduced in the experimental groups administered with Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4. As a result of comparing the secretion of TARC, which is related to an increase in the number of eosinophils, the TARC secretion levels of the positive control group and the group administered maltose (medium concentration / high concentration) were 27.8±1.95pg / ml, 26.6±1.1pg / ml, and 22.8±1.2pg / ml, respectively. In contrast, the secretion levels of Fos (powder) according to Comparative Examples 3 and 4 at medium concentration / high concentration were 22.4±1.5pg / ml and 21.2±2.0pg / ml, respectively, of the Advanced Fos (powder) according to Examples 1 and 2 at medium concentration / high concentration, 18.0±0.9pg / ml and 17.7±1.8pg / ml, respectively, of the Advanced Fos (syrup) according to Examples 3 and 4 at medium concentration / high concentration, and 19.7±1.1pg / ml and 19.1±0.4pg / ml, respectively, of the Advanced Fos (syrup) according to Comparative Examples 3 and 4 and Examples 1 to 4 at medium concentration / high concentration. It was confirmed that the amount of TARC secretion was relatively reduced in the experimental group administered Fos (powder and syrup). As a result of comparing the secretion amount of Eotaxin, which is involved in the influx of eosinophils into the skin and degranulation, the secretion amounts of Eotaxin in the positive control group and the group administered maltose (medium concentration / high concentration) were 144.7±3.7pg / ml, 141.9±3.5pg / ml, and 137.0±5.8pg / ml, respectively. In contrast, the secretion amounts of Fos (powder) according to Comparative Examples 3 and 4 were 125.3±6.1pg / ml and 122.3±1.2pg / ml at medium concentration / high concentration, respectively, and the secretion amounts of Advanced Fos (powder) according to Examples 1 and 2 were 117.9±5.7pg / ml and 113.6±1.5pg / ml at medium concentration / high concentration, respectively, and the secretion amounts of Advanced Fos (powder) according to Examples 3 and 4 were 117.9±5.7pg / ml and 113.6±1.5pg / ml at medium concentration / high concentration, respectively. The levels of Fos (syrup) at medium and high concentrations were 123.0±2.5pg / ml and 118.6±7.4pg / ml, respectively, and it was confirmed that the amount of eotaxin secretion was relatively reduced in the experimental groups administered Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4.

[0103] Administration of the experimental group (Advanced Fos: powder and syrup - medium / high concentration) according to Examples 1 to 4 improved atopic symptoms by regulating cytokine secretion, suppressing Th2 immune responses, and restoring the Th1 / Th2 balance. FIG. 19 is a graph showing a comparative analysis of the secretion amount of Treg-associated cytokines by AD induction and administration to the experimental groups. First, the secretion amount of IL-1β, which induces systemic inflammation by overexpression, was compared. As a result, the secretion amount of IL-1β in the positive control group and the group administered maltose (medium concentration / high concentration) was 50.7±2.3 pg / ml, 49.8±1.4 pg / ml, and 45.1±3.2 pg / ml, respectively. Meanwhile, the secretion amounts of IL-1β in the medium concentration / high concentration groups of Fos (powder) according to Comparative Examples 3 and 4 were 34.2±4.2 pg / ml and 21.1±3.0 pg / ml, respectively, in the medium concentration / high concentration groups of Advanced Fos (powder) according to Examples 1 and 2 were 14.0±3.8 pg / ml and 11.1±2.6 pg / ml, respectively, in the medium concentration / high concentration groups of Advanced Fos (powder) according to Examples 3 and 4. The levels of IL-1β secretion were 7.3±2.3pg / ml and 10.3±3.6pg / ml at the medium and high concentrations of Fos (syrup), respectively, and it was confirmed that the amount of IL-1β secretion was relatively reduced in the experimental groups administered with Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4. In the case of the amount of IL-10 secreted, which causes the imbalance between Th1 and Th2, the positive control group and the group administered with maltose (medium concentration / high concentration) showed the levels of IL-10 secreted of 173.7±20.8pg / ml, 171.8±20.7pg / ml, and 170.4±18.3pg / ml, respectively, whereas the Fos (powder) according to Comparative Examples 3 and 4 showed the levels of 126.2±25.9pg / ml and 90.9±8.7pg / ml at medium concentration / high concentration, respectively, the Advanced Fos (powder) according to Examples 1 and 2 showed the levels of 39.9±12.8pg / ml and 27.6±4.04pg / ml at medium concentration / high concentration, respectively, and the Advanced Fos (powder) according to Examples 3 and 4 showed the levels of 173.7±20.8pg / ml, 171.8±20.7pg / ml, and 170.4±18.3pg / ml at medium concentration / high concentration, respectively. The levels of IL-10 secretion were 41.7±16.8pg / ml and 38.1±10.4pg / ml at the medium and high concentrations of Fos (syrup), respectively, and it was confirmed that the amount of IL-10 secretion was relatively reduced in the experimental groups administered with Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4. Administration of the experimental groups according to Examples 1 to 4 (Advanced Fos: powder and syrup - medium concentration / high concentration) suppresses excessive Treg immune responses, suppresses inflammatory responses, and improves atopic symptoms by restoring the Th1 / Th2 balance. The high kestose content not only achieves even better effects, but also reduces the intake of fructooligosaccharides despite consuming a high kestose content, thereby reducing discomfort in the digestive tract (diarrhea, abdominal distension, borborygmus, etc.) caused by excessive intake of lacto-oligosaccharides, making it even more preferable.

[0104] Test Example 8: Flow cytometry analysis (1)Analysis method Spleens were obtained from sacrificed mice and homogenized in a 50-mm Petri dish (Nunclon, Denmark) using ACK buffer (Gibco, USA) to lyse red blood cells. The homogenized sample was filtered through a cell strainer (SPL, Korea) and washed with 2% RPMI (Hyclone, USA). After centrifugation at 2,000 × g for 10 minutes at 4°C, the supernatant was discarded and resuspended in 2% RPMI. All cells were diluted identically and treated with anti-mouse CD86 and CD274 antibodies (BD Pharmigen, USA) for 20 minutes at 4°C. Cell surface expression was confirmed using a flow cytometer (FACSCalibur; Becton Dickinson, USA). Mean fluorescence intensity (MFI) was analyzed using Cell Quest software (version 6.0) on the FACSCalibur instrument.

[0105] (2) Confirmation of CD86 expression levels after AD induction and experimental administration FIG. 20 is a graph showing the expression level of CD86, which is involved in the activation of T cells and B cells. When atopy is induced in splenocytes isolated from the spleen, the expression level of CD86, a costimulatory factor that activates T cells and B cells, was examined. As a result, the CD86 expression levels of the positive control group and the group administered with maltose (medium concentration / high concentration) were 25.4±2.2%, 25.8±0.4%, and 23.1±1.2%, respectively. Meanwhile, the expression levels of Fos (powder) according to Comparative Examples 3 and 4 were 21.7±1.9% and 20.9±1.1% at medium and high concentrations, respectively, the expression levels of Advanced Fos (powder) according to Examples 1 and 2 were 16.1±0.4% and 14.0±0.9% at medium and high concentrations, respectively, and the expression levels of Advanced Fos (powder) according to Examples 3 and 4 were 16.1±0.4% and 14.0±0.9% at medium and high concentrations, respectively. The expression levels of CD86 at the medium and high concentrations of Fos (syrup) were 16.9±1.0% and 17.7±0.8%, respectively, and the expression levels of CD86 were relatively reduced in the experimental groups administered Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4.

[0106] (3) Confirmation of CD274 expression levels after AD induction and experimental administration FIG. 21 is a graph showing the expression level of CD274 after AD induction and administration to the experimental groups. The expression level of CD274, a negative immune regulation factor involved in impaired immune regulation function, was confirmed in splenocytes isolated from the spleen. As a result, the CD274 expression levels of the positive control group and the group administered maltose (medium concentration / high concentration) were 14.1±1.5%, 14.9±0.2%, and 14.8±1.6%, respectively. Meanwhile, the expression levels of Fos (powder) according to Comparative Examples 3 and 4 at medium concentration / high concentration were 18.8±1.3% and 20.4±0.1%, respectively, the expression levels of Advanced Fos (powder) according to Examples 1 and 2 at medium concentration / high concentration were 22.1±1.2% and 23.1±0.2%, respectively, and the expression levels of Advanced Fos (powder) according to Examples 3 and 4 at medium concentration / high concentration were 24.1±1.5%, 24.9±0.2%, and 24.8±1.6%, respectively. The expression levels of CD274 were relatively increased in the experimental groups administered Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4 at the medium and high concentrations, respectively, by 20.9±1.0% and 21.0±0.9%. The experimental groups according to Examples 1 to 4 (Advanced Fos: powder and syrup - medium and high concentrations) were found to contribute to restoring the immune system damaged by immune reactions, suppressing excessive immune cell activity, and improving atopic symptoms.

[0107] Test Example 9: Tissue analysis of animal models (1) Experimental method The dorsal skin of sacrificed mice was fixed in 10% formalin solution and embedded in paraffin. Skin tissue sections were cut at a thickness of 4-5 μm and stained with Toluidine blue and Congo red stains. The numbers of mast cells and eosinophils were measured using a DM 4000B microscope (Leika) at 400x magnification.

[0108] (2) Measurement of mast cells in dorsal skin FIG. 22 is a graph showing the mast cells in dorsal skin. The number of mast cells, which produce histamine and induce inflammation and itching, was measured in the skin. The number of mast cells in the positive control group and the group administered with maltose (medium concentration / high concentration) was 328±19, 366±13, and 357±31, respectively. In contrast, the number of mast cells in the medium concentration / high concentration groups of Fos (powder) according to Comparative Examples 3 and 4 was 234±1 and 226±8, respectively, the number of Advanced Fos (powder) according to Examples 1 and 2 was 214±7 and 206±11, respectively, the number of Advanced Fos (syrup) according to Examples 3 and 4 was 231±9 and 221±18, respectively, and the number of mast cells in the experimental groups administered with Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4 was 231±9 and 221±18, respectively. The number of cells decreased relatively.

[0109] (3) Measurement of eosinophils in dorsal skin FIG. 23 is a graph showing the measurement of eosinophils in dorsal skin. The number of eosinophils, which act on hypersensitivity reactions and worsen atopic symptoms, was confirmed in skin tissue. The positive control group and the group administered maltose (medium concentration / high concentration) showed eosinophil counts of 6.5±2.6, 6±2.5, and 6.3±2.5, respectively. In contrast, the numbers of eosinophils in the medium concentration / high concentration groups of Fos (powder) according to Comparative Examples 3 and 4 were 4.3±1.7 and 4.6±1.9, respectively. The numbers of Advanced Fos (powder) according to Examples 1 and 2 were 4±1.6 and 2.8±1.5, respectively. The numbers of Advanced Fos (syrup) according to Examples 3 and 4 were 3±1.2 and 2.8±1.1, respectively. The experimental groups administered Fos (powder and syrup) showed a relative decrease in eosinophil counts.

[0110] (4) Measurement of mast cells in the intestinal tissue (ileum) FIG. 24 is a graph showing the measurement of mast cells in the intestinal tissue. The results of confirming the number of mast cells showed that the positive control group and the group administered maltose (medium concentration / high concentration) had mast cell counts of 48±6, 56±12, and 59±11, respectively. In contrast, the numbers of mast cells for the medium and high concentrations of Fos (powder) according to Comparative Examples 3 and 4 were 33±7 and 32±7, respectively, for the medium and high concentrations of Advanced Fos (powder) according to Examples 1 and 2 were 23±6 and 24±4, respectively, for the medium and high concentrations of Advanced Fos (syrup) according to Examples 3 and 4 were 25±6 and 19±8, respectively, indicating a relative decrease in the number of mast cells in the experimental groups administered Fos (powder) according to Comparative Examples 3 and 4 and Advanced Fos (powder and syrup) according to Examples 1 to 4.

Claims

1. A composition for promoting the growth of intestinal microorganisms, comprising kestose as an active ingredient, The intestinal microorganism is a microorganism of the genus Alistipes, A composition for promoting the growth of intestinal microorganisms, wherein the daily dose of the kestose is in the range of 2 to 30 g per day for an adult weighing 60 kg.

2. The composition of claim 1 , wherein the intestinal microorganisms are microorganisms of the genus Alistipes and microorganisms of the genus Lactobacillus.

3. 10. The composition of claim 1, wherein when administered to a subject, the composition increases the relative abundance (%) of the gut microbes in the subject.

4. 4. The composition of claim 3, wherein the relative abundance of the intestinal microorganisms is 105% or more based on a level of 100% before administration of the active ingredient to the subject.

5. The composition according to any one of claims 1 to 4, wherein the kestose is provided as a fructooligosaccharide composition containing 50% by weight or more of kestose based on the solid content of sugars.

6. The composition according to any one of claims 1 to 5, wherein the composition is a prebiotic.

7. The composition according to claim 6, further comprising one or more probiotic compositions selected from the group consisting of microorganisms of the genus Bifidobacterium, Streptococcus, Saccharomyces, bacillus, Alistipes, and Lactobacillus.

Images