Compositions that promote intestinal health

Abstract

To provide compositions for promoting intestinal health and a method of treating a condition or disorder related to gastrointestinal health.SOLUTION: Compositions for promoting intestinal health can include a combination of cyanidins and delphinidins, in an amount sufficient to treat intestinal hyperpermeability. The compositions can further have a prebiotic blend and fructooligosaccharides. Further presented herein is a method of treating a condition or disorder related to gastrointestinal health in a subject. The method can include maximizing tight junction integrity in epithelial cells of gastrointestinal tract of the subject.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] Part of the research of the present invention was carried out with the support of the government under a grant awarded by the National Institute of Food and Agriculture. The government has certain rights in the present invention.

Background Art

[0002] The gastrointestinal system is a complex network of tissues, organs, host cells, and bacterial cells. Aging, diet, drugs, disruption of the gut microbiota, and an individual's lifestyle conditions can have an adverse impact on this network. These effects may include disruption of the gut microbiota balance, increased inflammation of gastrointestinal tissues, and breakdown of the epithelial cell barrier of the gastrointestinal system. These effects can ultimately reach systemic tissues. Formulations and methods for promoting and restoring gut health may provide benefits.

Summary of the Invention

Means for Solving the Problems

[0003] The features and advantages of the present invention will become apparent from the following detailed description, which describes the features of the present invention in conjunction with the accompanying drawings. It is understood that the drawings are merely illustrative of typical embodiments and results and are not to be considered limiting in scope.

Brief Description of the Drawings

[0004] [Figure 1] [[ID=3(0]]Figure 1 schematically shows the human digestive tract and related structures. [Figure 2] Figure 2 schematically shows the small intestine and epithelial cell layer lining the intestine. [Figure 3] Figure 3 schematically shows the tight junctions of epithelial cells and the systemic effects when the tight junction barrier collapses. [Figure 4]Figure 4 schematically shows the intestinal epithelial cell layer, tight junctions, paracellular pathways, and intercellular pathways. [Figure 5] Figure 5 shows a schematic representation of the chemical structure of cyanidin. [Figure 6] Figure 6 shows a schematic representation of the chemical structure of delphinidin. [Figure 7] Figure 7 shows a schematic representation of the chemical structure of petunidine. [Figure 8] Figure 8 shows a schematic representation of the chemical structure of peonidine. [Figure 9] Figure 9 shows a schematic representation of the chemical structure of malvidin. [Figure 10] Figure 10 schematically shows a transepithelial electrical resistance (TEER) method according to one embodiment of the present disclosure. [Figure 11] Figure 11 schematically shows the TEER resistance of various extracts according to one embodiment of the present disclosure. [Figure 12] Figure 12 schematically illustrates the paracellular transport of various extracts according to one embodiment of the present disclosure. [Figure 13] Figure 13 schematically shows the TEER resistance of various extracts according to one embodiment of the present disclosure. [Figure 14] Figure 14 schematically shows PCP FITC-dextran of various extracts according to one embodiment of the present disclosure. [Figure 15] Figure 15 schematically shows the TEER resistances of various cyanidins according to one embodiment of the present disclosure. [Figure 16] Figure 16 schematically shows the TEER resistances of various delphinidins according to one embodiment of the present disclosure. [Figure 17] Figures 17 and 18 schematically show the TEER resistance of epicatechin according to one embodiment of the present disclosure. [Figure 18] Figures 17 and 18 schematically show the TEER resistance of epicatechin according to one embodiment of the present disclosure. [Figure 19] Figure 19 schematically shows the TEER resistance of catechin according to one embodiment of the present disclosure. [Figure 20] Figure 20 schematically shows the TEER resistance of total catechin according to one embodiment of the present disclosure. [Figure 21] Figure 21 schematically shows the average colon length of mice fed different diets according to one embodiment presented herein. [Figure 22] Figure 22 schematically shows the average colon weight of mice fed different diets, according to one embodiment presented herein. [Figure 23] Figure 23 schematically shows the average colon weight / length of mice fed different diets, according to one embodiment presented herein. [Figure 24] Figure 24 schematically shows the weight gain and FITC-dextran permeability of mice fed different diets, according to one embodiment presented herein. [Figure 25] Figure 25 schematically shows FITC-DX paracellular transport in mice fed different diets, according to one example presented herein. [Figure 26] Figure 26 schematically shows the endotoxin concentrations measured in mice fed different diets, according to one example presented herein. [Figure 27] Figure 27 schematically shows the GTT(AUC) concentrations measured in mice fed different diets, according to one example presented herein. [Figure 28] Figure 28 schematically shows the ITT(AUC) concentrations measured in mice fed different diets, according to one example presented herein. [Figure 29] Figure 29 schematically shows the endotoxin and glucose loading test concentrations measured according to one example presented herein. [Figure 30] Figure 30 schematically shows endotoxin and fasting insulin concentrations measured according to one example presented herein. [Figure 31]Figure 31 schematically shows endotoxin and IL-6 test concentrations measured in accordance with an embodiment presented herein. [Figure 32] Figure 32 schematically shows endotoxin and IL-1β concentrations measured in accordance with an embodiment presented herein. [Figure 33] Figure 33 schematically shows endotoxin and IL-1α test concentrations measured in accordance with an embodiment presented herein. [Figure 34] Figure 34 schematically shows HOMA-IR concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 35] Figure 35 schematically shows adiponectin concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 36] Figure 36 schematically shows leptin concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 37] Figures 37 and 38 schematically show triglyceride concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 38] Figures 37 and 38 schematically show triglyceride concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 39] Figures 39 and 40 schematically show cholesterol concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 40] Figures 39 and 40 schematically show cholesterol concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 41] Figure 41 schematically shows hepatic triglyceride concentrations measured in mice fed different diets in accordance with an embodiment presented herein. [Figure 42]Figure 42 shows a photograph of a mouse liver extracted from a mouse that was fed a different diet before being euthanized according to one embodiment presented herein. [Figure 43] Figure 43 shows photographs of mouse feces from mice that consumed different diets according to one embodiment presented herein. [Figure 44] Figure 44 illustrates the mean Firmicutes:Bacteroides ratio of the gut microbiome after supplementation with the compositions presented herein. [Figure 45] Figure 45 illustrates the calprotectin concentration after supplementation with the compositions presented herein. [Figure 46] Figure 46 illustrates the baseline BSS score and changes in bowel movements after supplementation with the compositions presented herein. [Figure 47] Figure 47 illustrates the changes in baseline scores for abdominal distension, abdominal pain, and flatulence after supplementation with the compositions presented herein. [Modes for carrying out the invention]

[0005] Before disclosing and describing embodiments of the invention, it is understood that there is no intention to limit the specific structures, processes, or materials disclosed herein, but that their equivalents are included as would be recognized by a person ordinarily skilled in the art. It is also understood that the technical terms used herein are used to describe only specific embodiments and are not intended to limit them. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as that generally understood by a person skilled in the art to which this disclosure belongs. As used herein and in the appended claims, the singular form also includes plural references unless it is clearly indicated otherwise in the content.

[0006] As used in this explanation, the singular forms "a," "an," and "the" express support for multiple references, in particular, unless the context clearly indicates otherwise. For example, "prebiotic fiber" supports one or more prebiotic fibers.

[0007] As used herein, the term "approximately" refers to a certain degree of deviation. This means roughly, around, approximately, or before or after. When the term "approximately" is used in conjunction with a numerical range, it modifies this range by exceeding a boundary that is slightly above or below the indicated number. It is understood that the support provided herein for a number used in conjunction with the term "approximately" is also provided for an exact number as if "approximately" were not used.

[0008] Concentration, quantity, and other numerical data may be expressed or presented in range form as specified herein. Since such range form is used simply for convenience and conciseness, it should be understood that it includes all individual numerical values ​​and / or partial ranges that fall within that range, as well as each numerical value and partial range (including partial ranges) that is specified, not just the numerical value specified as the limit or endpoint of the range. For illustrative purposes, the numerical range "approximately 1 to approximately 5 g" should be interpreted as including not only the specified value of approximately 1 g to approximately 5 g, but also the individual values ​​and partial ranges within the range indicated above. Thus, included in this numerical range of the examples are individual numerical values ​​such as 2, 2.6, 3, 3.8, and 4, and partial ranges such as 1 to 3, 2 to 4, and 3 to 5, as well as 1, 2, 3, 4, and 5 individually.

[0009] As used herein, “concentrate” refers to a raw material extract containing at least the same amount of active fractions, compounds, or other components as the raw material itself, but in a smaller quantity. In one embodiment, the “concentrate” may be a dry powder derived from components that do not involve the use of any solvent during the concentration process.

[0010] Comparative terms such as "more efficient," "better," "improved," and "enhanced" can be used to describe the results achieved or properties present in a formulation or process that are, to some extent, better or more positive than those being compared. In some cases, this may be compared to the prior art or a method that exhibits a more positive outcome than the state of the formulation before administration.

[0011] As used herein, “having,” “possessing,” “containing,” and “holding” may have the meanings assigned to them under U.S. Patent Law, and may mean “including,” “containing,” etc., and are generally interpreted as unrestricted terms. The terms “~is” or “~become” are restricted terms and include only components, structures, processes, etc., that are specifically enumerated together with such terms and are subject to U.S. Patent Law. “Basically ~is” or “basically ~become” has the meaning generally attributed under U.S. Patent Law. In particular, such terms are generally restricted terms, except when they include additional items, materials, components, processes, or elements that do not materially affect the basic and novel characteristics or functions of the items used in connection therewith. For example, trace elements are present in a composition but are not explicitly enumerated in the list of items following such technical terms, but the properties or characteristics of the composition are permissible if they exist under the language of “basically ~become.” When used in terms that do not restrict the description, such as "possess" or "contain," it is also understood that direct support should be given to the language of "basically becomes" and the language of "becomes," as explicitly stated and vice versa.

[0012] The term “dosage unit” is understood to mean a unit of a composition that can be administered to a subject or patient and provides a sufficient amount of active drug to achieve or contribute to a therapeutic effect. In some embodiments, “dosage unit” remains a unit that is easily handled and packaged and is a physically and chemically stable unit dose having the active ingredient in itself or as a pharmaceutically solid or liquid medium or a mixture thereof. Furthermore, “dosage” and “dose” may refer to such a dosage unit. Alternatively, “dosage” or “dose” may include multiple dosage units that collectively provide a desired amount of active drug to be administered to a subject at a single point in time. Multiple doses or dosages may be available in a schedule to establish a method of administration.

[0013] The terms “effective dose,” “therapeutic dose,” or “therapeutic rate” of an active ingredient refer to a non-toxic but sufficient amount or delivery rate of the active ingredient to achieve a therapeutic outcome in the treatment of the disease or condition to which the active drug is delivered. It is understood that various biological factors may influence a substance’s ability to perform its intended function. Therefore, the “effective dose,” “therapeutic dose,” or “therapeutic rate” may, in some cases, depend on such biological factors. Furthermore, while the achievement of therapeutic effect may be measured by a physician or other qualified medical staff by assessments known in the field, it is recognized that individual variability and response to treatment may make the achievement of therapeutic effect a subjective judgment. The determination of therapeutic doses or delivery rates is well known to those skilled in the art of the pharmaceutical and medical fields.

[0014] The term “extract” refers to a substance prepared with a solvent, such as ethanol, water, steam, superheated water, methanol, hexane, chloroform solution, liquid CO2, liquid N2, propane, supercritical CO2, or any combination thereof. As used herein, an extract may refer to an extract in liquid form, or a product obtained by further processing the liquid form, such as a dry powder or other solid form. Extracts can take many forms, including but not limited to solids, liquids, particles, fragments, distillates, etc., and can be prepared by numerous techniques or protocols, such as cutting, grinding, pulverizing, boiling, steaming, immersion, soaking, infusion, gas injection, etc., and may utilize appropriate reagents such as water, alcohol, steam, or other organic materials. Extracts typically have a certain purity percentage and may be of relatively high purity. In some embodiments, the extract may be a phytoextract made from specific parts of the raw material, such as the outer layer, pulp, leaves, flowers, fruits, grains, seeds, etc., or it may be made from the whole raw material. In some embodiments, the extract may contain one or more active fractions or active drugs. In some embodiments, multiple extracts can be controlled by fractionation of the extraction process or protocol, or can be a function of the extraction process or protocol.

[0015] As used herein, “formulation” and “composition” are used synonymously and can refer to a combination of at least two components. In some embodiments, at least one component may be an active drug, or if not, may have properties that exhibit physiological activity when administered to a subject.

[0016] The formulations or components included in or cited herein are presumed to be expressed in wt% unless otherwise specified. Furthermore, the amounts of components presented in the form of ratios are presumed to be in wt% (e.g., %w / w) proportions.

[0017] "Intestines" pastThe term "permeability" refers to a level higher than the normal permeability of the gastrointestinal system (i.e., higher than the average subject's permeability), and in some cases, can refer to increased permeability of the stomach, small intestine, and / or large intestine.

[0018] As used herein, “linear inhibitory effect” or “dose-response” refers to a linear decrease in secretion or biosynthesis obtained from all concentrations of the inhibitory material in a dose-response curve. For example, if inhibition occurs at low concentrations, followed by failure or increase in secretion at high concentrations, it indicates that there is no linear inhibitory effect.

[0019] The term "or" is used in the "inclusive" sense of "and / or," not in the "exclusive" sense of "either / or."

[0020] As used herein, “pharmaceutically acceptable” generally refers to a material suitable for administration to a subject in combination with an active drug or component. For example, a “pharmaceutically acceptable carrier” is a substance or material that can be appropriately used in combination with an active drug and can provide a composition or formulation suitable for administration to a subject. Excipients, diluents, and other components used in or used to prepare a formulation or composition administered to a subject may be used in such terms.

[0021] The term "prevent" and its variations refer to prevention against a specific undesirable physiological condition. Such preventive measures may be partial or complete. In partial preventive measures, the onset of the physiological condition may be delayed. Individuals skilled in the art will recognize that delaying the onset of a physiological condition is desirable and will know that the compositions of the present invention are administered to subjects at risk of a specific physiological condition in order to delay the onset of these conditions. For example, a person skilled in the art will recognize that obese subjects have an increased risk of coronary artery disease. Therefore, a person skilled in the art would administer the compositions of the present invention to improve the gut microbiota of obese individuals.

[0022] As used herein, “subject” refers to the individual being administered the drug. In one embodiment, the subject may be a mammal. In another embodiment, the subject may be a human. In yet another embodiment, the subject may be livestock or domesticated animals.

[0023] As used herein, “substantial” or “substantially” means, when used in reference to the amount of a material, effect, or specific feature of a composition, a sufficient amount to provide the effect intended to provide the material or the features of the material. The exact degree of acceptable deviation depends in some cases on the specific content. Similarly, “substantially absent,” etc., means the absence of an identified element or drug in a composition. In particular, an element identified as “substantially absent” is either completely absent from the composition or present in such small an amount that it does not have a measurable effect on the composition.

[0024] As used herein, the terms “not important” or “not clinically important” refer to the degree of effect of administering the composition to a subject. For example, if the degree of the result does not produce a clinical change in the subject, the result is not clinically important.

[0025] The terms “to treat,” “treating,” or “treatment” mean, as used herein and as is well understood in the art, an approach to obtain beneficial or desired outcomes, including clinical outcomes in the subject being treated. Beneficial or desired outcomes may include, but are not limited to, reduction or recovery of one or more signs or symptoms of a condition, whether detectable or not; reduction of disease severity; stabilization of the disease or condition (i.e., no worsening); delay or slowing of disease progression; recovery or remission of the disease condition; reduction of disease recurrence; and (partial or complete) remission. “To treat,” “treating,” and “treatment” may also mean an extension of survival compared to the survival expected without treatment, and may be preventive. Such preventive treatment may also be referred to as prevention or a preventive measure of disease or condition. Such preventive measures may be partial or complete. In partial preventive measures, the onset of the physiological condition may be delayed.

[0026] As used herein, the term “solvent” refers to a gaseous, aqueous, or organic liquid having the characteristics required for the extracted solid material of a plant preparation. Examples of solvents include, but are not limited to, water, water vapor, superheated water, methanol, ethanol, ethyl acetate, hexane, chloroform, liquid CO2, liquid N2, propane, or any combination of such materials.

[0027] For convenience, as used herein, a common list may contain multiple items, structural elements, compositional elements, and / or materials. However, these lists should be interpreted as each number in the list being individually identified as distinct and unique. Therefore, individual contents of such lists should not be interpreted as being substantially equivalent to, or to the contrary, other contents of the same list based on the presentation of the common group.

[0028] Unless otherwise stated, the steps listed in any claim for any method or process may be performed in any order, and are not limited to the order presented in the claim.

[0029] The gastrointestinal system is a tubular system of interconnected structures involved in the digestion of food and the absorption of energy and nutrients. The gastrointestinal system is shown in Figure 1 and consists of all structures from the mouth to the anus. The total length of the gastrointestinal system is approximately 9 meters and can be divided into the upper and lower digestive tracts. The lower digestive tract includes the small intestine and the large intestine. The small intestine is approximately 20 feet (6.1 meters) long and has a highly folded structure that includes finger-like projections called villi. Within the villi are a monolayer of epithelial cells and a capillary layer. The main function of the small intestine is to absorb digestive products. Nutrients pass through the epithelial cell layer and enter the capillaries below. These nutrients eventually enter larger blood vessels and travel to the liver, where they are processed and controlled for release into the body. The large intestine is approximately 3 feet (91 centimeters) long and is used to collect solid matter that was not digested in the small intestine and to absorb water. The large intestine is parasitized by bacteria that synthesize vitamins.

[0030] As already mentioned, the small intestine has a barrier composed of a single layer of epithelial cells. These epithelial cells are sealed at tight junctions. These tight junctions can regulate intestinal permeability by controlling paracellular transport and intercellular pathways of water and ions. See Figure 3. In addition to its involvement in nutrient absorption, this cell layer is also involved in maintaining mucosal immune homeostasis and preventing inflammation, forming a first line of defense against the invasion of harmful bacteria / bacterial toxins and / or other antigens that can initiate chronic inflammation. All bacteria, bacterial toxins, antigens, water, and ions that pass through this cell layer can enter the bloodstream and potentially affect other organs, potentially having systemic effects on the entire organism. See Figure 4. Lifestyle and dietary factors such as strenuous exercise, high-fat diets, and overnutrition can also affect intestinal permeability and may be involved in increased toxin permeability to tight junctions and blood circulation.

[0031] The collapse of the aforementioned tight junctions can lead to increased intestinal permeability, as the tight junction barrier becomes more vulnerable. Increased permeability may be a significant factor in the pathophysiology of several inflammatory conditions and obesity-related conditions. Low levels of chronic inflammation may negatively impact increased intestinal barrier permeability.

[0032] Tumor necrosis factor alpha (TNFα) can also be a central basal mediator. TNFα induces apoptosis, and these changes in distribution occur through its ability to promote increased barrier permeability and the expression of specific tight junction proteins. Ultimately, TNFα may also be involved in promoting tight junction barrier dysfunction. Loss of tight junction function and increased intestinal permeability may contribute to allergic conditions (e.g., celiac disease), inflammatory bowel diseases (Crohn's disease and ulcerative colitis), food intolerances, dyspepsia, low levels of chronic enteritis (e.g., inflammation associated with obesity and type 1 and type 2 diabetes), insulin resistance, autism, multiple sclerosis, malnutrition, metabolic syndromes, cancer, asthma, atopic dermatitis, and rheumatoid arthritis.

[0033] Another factor in an individual's gastrointestinal health is the gut microbiota. This microbiota is a complex network of bacteria, archaea, viruses, and eukaryotes that influence an individual's health and physiological function. The gastrointestinal microbiota has a microbial population of approximately 3.9 x 10⁶ 13 It was estimated to reach [a certain level]. The composition of an individual's gut microbiome can vary greatly in number, diversity, histological features, and activity. The richness and diversity of the gut microbiome have been shown to be greatly influenced by antibiotics, age, diet, ethnicity, geographical location, physical stress, mental stress, and sex.

[0034] The aforementioned changes in the gut microbiota may increase endotoxin production. Microbiota with few beneficial bacteria and low diversity, or with many beneficial bacteria and high diversity, may occur in conditions associated with aging and / or accelerated aging such as obesity or a high-fat diet. Microbiota imbalances may increase inflammation of the gastrointestinal epithelium, alter the integrity of the intestinal cell wall, and increase intestinal permeability. These changes may contribute to the development of gastrointestinal infections, asthma / atopy, obesity, metabolic syndromes, cancer, rheumatoid arthritis, Crohn's disease, and ulcerative colitis. In contrast, a balanced and diverse microbiota may provide resistance to infection, enable healthy aging, prevent intestinal diseases, contribute to the metabolism of polyhydric phenols, and produce absorbable bioactive substances. The gut microbiota may also be involved in metabolism, immune development, endocrine signaling, and neurological signaling.

[0035] Flavonoids may play a crucial role in preventing and improving increased intestinal barrier permeability. The gut microbiota can metabolize flavonoid-releasing, bioactive substances. These bioactive substances can maintain and / or restore nutritional balance. Therefore, flavonoids have the ability to suppress inflammation, modulate the selection of signaling cascades, and control the cellular redox state. However, the numerous existing flavonoids with different chemical / spatial structures, and the multiple metabolites produced in the large intestine by the microbiota, prevent simple generalization of the health benefits and mechanisms of action of flavonoids as a class.

[0036] Anthocyanins (ACs) are one of the major flavonoid subgroups and are found in many fruits and vegetables (e.g., berries, red cabbage, black rice) to provide color. Several different types of ACs exist, differing in the number and substitution of hydroxyl groups, the bonded sugar structure, the type of carboxylate salt, and the bond to sugar. Some of the different classes of anthocyanins include cyanidin, delphinidin, peonidin, petunidine, and malvidin.

[0037] Recent evidence indicates that the parent compound AC is present throughout the entire gastrointestinal tract. In the colon, AC may be metabolized into several metabolites by intestinal microorganisms. Therefore, theoretically, the small intestine, and to a lesser extent the large intestine, may be exposed to large amounts of the parent compound AC from food. Dietary AC may be beneficial to the gastrointestinal system, but little is known about the details under conditions of increased intestinal permeability. AC can exert beneficial effects in the gastrointestinal tract through direct and indirect effects. The indirect effects relate to AC's ability to modulate the aforementioned microbiome and influence AC metabolism via the microbiome. Both a "healthier" microbiome and the formation of specific active metabolites may be involved in the indirect effects of AC on gut health, potentially leading to systemic effects.

[0038] Specific embodiments of the present invention are described below in detail. While the present invention is described in conjunction with these specific embodiments, it is understood that there is no intention to limit the invention to such specific embodiments. On the contrary, other forms, modifications, and equivalent forms are intended to be covered as they fall within the spirit and scope of the invention, as defined in the appended claims. Numerous specific details are given below in order to fully understand the invention. The invention can be operated without some or all of these specific details. In other cases, well-known process operations have not been described in detail so as not to unnecessarily complicate the invention.

[0039] This disclosure relates to compositions and methods for improving the intestinal health of subjects. In one embodiment, a composition that promotes intestinal health is introduced. The composition may contain cyanidin and delphinidin (a class of anthocyanins) in an amount sufficient to treat intestinal hyperpermeability. In one embodiment, the cyanidin and delphinidin may be present collectively in an amount that maintains the integrity of tight junctions of intestinal epithelial cells. In another embodiment, the cyanidin and delphinidin may be present collectively in an amount that restores the integrity of tight junctions of intestinal epithelial cells.

[0040] Cyanidin and delphinidin can protect intestinal epithelial cells from TNFα by reducing monolayer permeability in transepithelial electrical resistance (TEER) and enhancing paracellular transport of FITC-dextran. In contrast, malvidin, peonidin, and petunidine are not thought to provide protection to intestinal epithelial cells against TNFα-induced TEER permeability and are not thought to enhance paracellular transport of FITC-dextran. While not bound by theory, the protective effects of cyanidin and delphinidin are thought to be due to the presence of a catechol group in the B ring of cyanidin and delphinidin. Anthocyanins without a B ring, which were investigated by the inventors, did not exhibit these protective effects. In one embodiment, this activity can be selective. In another embodiment, the protective function can be dose-dependent.

[0041] In another embodiment, the Disclosure provides a method for treating a condition or disease associated with the gastrointestinal health of a subject, comprising maximizing the integrity of tight junctions of the subject's gastrointestinal epithelial cells. In yet another embodiment, a method for maximizing the integrity of tight junctions of a subject's gastrointestinal epithelial cells is presented. In yet another embodiment, the Disclosure provides a method for treating intestinal hyperpermeability. In some embodiments, these methods may target (i) maintaining and / or creating a healthy microbiome of the gastrointestinal system, (ii) maintaining and / or creating an inflammatory balance within the gastrointestinal system, and / or (iii) maintaining and / or creating integrity of the intestinal cellular barrier.

[0042] In some embodiments, a composition that promotes intestinal health may contain cyanidin, delphinidin, or a combination thereof, and in some embodiments, these drugs may be present in therapeutically effective amounts. In one embodiment, the composition may contain cyanidin and delphinidin in combination in an amount sufficient to treat intestinal hyperpermeability. In one embodiment, the cyanidin and delphinidin may be present collectively in an amount that maintains intestinal permeability. In another embodiment, the cyanidin and delphinidin may be present collectively in an amount that inhibits intestinal hyperpermeability. In yet another embodiment, the cyanidin and delphinidin may affect the microbiome and provide anti-inflammatory properties.

[0043] The amounts of cyanidin and delphinidin may vary in the composition. In one embodiment, the cyanidin and delphinidin, individually or collectively, may constitute about 5 wt% to about 50 wt% of the composition, or form an active fraction of the composition. In another embodiment, the cyanidin and delphinidin, individually or collectively, may constitute about 12 wt% to about 45 wt% of the composition, or form an active fraction of the composition. In yet another embodiment, the cyanidin and delphinidin, individually or collectively, may constitute about 12 wt% to about 25 wt% of the composition, or form an active fraction of the composition.

[0044] The cyanidin and delphinidin can be derived from a variety of raw materials. In one embodiment, at least one of the raw materials for the cyanidin and delphinidin may be derived from black rice components, blueberry components, blackcurrant components, crowberry components, bilberry components, black chokeberry components, or a combination thereof. In another embodiment, the raw materials for the cyanidin and delphinidin may be derived from the black rice components, blueberry components, and blackcurrant components. In yet another embodiment, the raw materials for the cyanidin and delphinidin may be derived from the black rice components and bilberry components. In yet another embodiment, the raw materials for the cyanidin and delphinidin may be derived from the blackcurrant components and blueberry components. In yet another embodiment, the cyanidin and delphinidin can be artificially created or synthesized (e.g., "synthetic products").

[0045] In one embodiment, the composition may contain a black rice component. The black rice component may be derived from black rice grains, black rice concentrate, black rice extract, black rice powder, or a combination thereof. In one embodiment, the black rice component may be a black rice extract. For example, the rice extract is obtained by concentrating black rice grains and passing the concentrated black rice grains through resin adsorption (chromatographic column). In one embodiment, the solvent may be water and ethanol. In one embodiment, the column may be eluted with 70 wt% ethanol and 30 wt% aqueous solution. In another embodiment, the column may be eluted with 75 wt% ethanol and 25 wt% aqueous solution. The eluent may be concentrated into a dried and packaged concentrated extract. In another embodiment, the black rice extract may be derived from black rice grains. In one embodiment, the black rice may be derived from Oryza sativa L.

[0046] In one embodiment, the black rice extract may constitute about 2.5 wt% to about 20 wt% of the composition, or be the active fraction of the composition. In another embodiment, the black rice component may constitute about 10 wt% to about 15 wt% of the composition, or be the active fraction of the composition. In yet another embodiment, the black rice extract may constitute about 2.5 wt% to about 5 wt% of the composition, or be the active fraction of the composition. In yet another embodiment, the black rice extract may constitute about 2.5 wt% to about 7.5 wt% of the composition, or be the active fraction of the composition.

[0047] In one embodiment, the black rice component may have a standardized anthocyanin content. In one embodiment, the black rice component may have a standardized anthocyanin content of about 10 wt% to about 30 wt%. In another embodiment, the black rice component may have a standardized anthocyanin content of about 20 wt%. In a further embodiment, the black rice component may have a standardized anthocyanin content of about 25 wt%. In one embodiment, the standard anthocyanin content can be measured by HPLC. In another embodiment, the standard anthocyanin content can be measured by UV.

[0048] In another embodiment, the composition may contain the blueberry component. The blueberry component may include components selected from the group consisting of blueberry fruit, blueberry extract, blueberry concentrate, blueberry juice, blueberry powder, or a combination thereof. In one embodiment, the blueberry component may be blueberry powder. In another embodiment, the blueberry component may be blueberry juice. In yet another embodiment, the blueberry component may be blueberry powder derived from blueberry juice. In a further embodiment, the blueberry component may be blueberry extract. For example, a blueberry fruit extract can be obtained by extracting blueberry fruit with water. The extract can be filtered, the filtrate can be washed with water, and the extract can be analyzed with 75% ethanol by resin adsorption. The extract can then be concentrated under reduced pressure and spray-dried. The concentrated extract can be crushed, sieved, and packaged. In one embodiment, the blueberry component may be bog blueberry component. In one embodiment, the black rice may be derived from Vaccinium uliginosum L.

[0049] In one embodiment, the blueberry component may constitute about 1 wt% to about 30 wt% of the composition, or it may be the active fraction of the composition. In another embodiment, the blueberry component may constitute about 1 wt% to about 10 wt% of the composition, or it may be the active fraction of the composition. In yet another embodiment, the blueberry component may constitute about 25 wt% to about 30 wt% of the composition, or it may be the active fraction of the composition.

[0050] In one embodiment, the blueberry component may have a standardized anthocyanin content. In one embodiment, the blueberry component may have a standardized anthocyanin content of about 0.5 wt% to about 30 wt%. In another embodiment, the blueberry component may have a standardized anthocyanin content of about 0.5 wt% to about 5 wt%. In yet another embodiment, the blueberry component may have a standardized anthocyanin content of about 20 wt% to about 30 wt%. In yet another embodiment, the blueberry component may have a standardized anthocyanin content of about 25 wt%. In yet another embodiment, the blueberry component may have standardized anthocyanins, which may be about 17% when measured by UV and about 25% when measured by HPLC.

[0051] In one embodiment, the composition may contain the blackcurrant component. In one embodiment, the blackcurrant component may include blackcurrant fruit, blackcurrant extract, blackcurrant concentrate, blackcurrant juice, blackcurrant powder, or a combination thereof. In another embodiment, the blackcurrant component may be blackcurrant powder. In yet another embodiment, the blackcurrant component may be blackcurrant juice. In yet another embodiment, the blackcurrant component may be blackcurrant extract. In one embodiment, the blackcurrant extract may be extracted using water, resin exchange, and ethanol. In one embodiment, the blackcurrant may be an ethylene oxide-free product. In one embodiment, the blackcurrant component may be derived from Ribes nigrum.

[0052] In one embodiment, the blackcurrant component may constitute about 0.5 wt% to about 15 wt% of the composition, or it may be an active fraction. In another embodiment, the blackcurrant component may constitute about 1 wt% to about 5 wt% of the composition, or it may be an active fraction of the composition. In yet another embodiment, the blackcurrant component may constitute about 10 wt% to about 15 wt% of the composition, or it may be an active fraction of the composition.

[0053] In one embodiment, the blackcurrant component may have a standardized anthocyanin content. In one embodiment, the blackcurrant component may have a standardized anthocyanin content of about 20 wt% to about 40 wt%. In another embodiment, the blackcurrant component may have a standardized anthocyanin content of about 30 wt%. In yet another embodiment, the blackcurrant component may have a standardized anthocyanin content of about 2.5 wt% to about 10 wt%. In one embodiment, the blackcurrant component may have a standardized anthocyanin content of about 5 wt%.

[0054] In one embodiment, the composition may contain a crowberry component. In one embodiment, the crowberry component may be crowberry fruit, crowberry extract, crowberry concentrate, crowberry juice, crowberry powder, or a combination thereof. In another embodiment, the crowberry component may be crowberry fruit. In yet another embodiment, the crowberry component may be crowberry extract. In one embodiment, the crowberry extract may be extracted using water and ethanol. In one embodiment, the crowberry component may be derived from Empetrum nigrum.

[0055] In one embodiment, the crowberry component may constitute about 1 wt% to about 30 wt% of the composition, or it may be the active fraction of the composition. In another embodiment, the crowberry component may constitute about 5 wt% to about 25 wt% of the composition, or it may be the active fraction of the composition. In yet another embodiment, the crowberry component may constitute about 5 wt% to about 15 wt% of the composition, or it may be the active fraction of the composition.

[0056] In one embodiment, the crowberry component may have a standardized anthocyanin content. In another embodiment, the crowberry component may have a standardized anthocyanin content of about 40 wt% to about 50 wt%. In yet another embodiment, the crowberry component may have a standardized anthocyanin content of about 46.7 wt%.

[0057] In one embodiment, the composition may contain a bilberry component. In one embodiment, the bilberry component may be bilberry fruit, bilberry extract, bilberry concentrate, bilberry juice, bilberry powder, or a combination thereof. In one embodiment, the bilberry component may be bilberry powder. In another embodiment, the bilberry component may be bilberry extract. In yet another embodiment, the bilberry extract may be extracted using water and ethanol. In one embodiment, the extract ratio of ethanol to water may be 150:1. In one embodiment, the bilberry component may be derived from Vaccinium myrtillus.

[0058] The amount of the bilberry component in the composition may vary. In one embodiment, the bilberry component may constitute about 0.5 wt% to about 30 wt% of the composition, or may be the active fraction of the composition. In another embodiment, the bilberry component may constitute about 2 wt% to about 20 wt% of the composition, or may be the active fraction of the composition. In a further embodiment, the bilberry component may constitute about 5 wt% to about 15 wt% of the composition, or may be the active fraction of the composition.

[0059] In one embodiment, the bilberry component may have a standardized anthocyanin content. In one embodiment, the bilberry component may have a standardized anthocyanin content of about 1 wt% to about 40 wt%. In another embodiment, the bilberry component may have a standardized anthocyanin content of about 5 wt% to about 25 wt%. In yet another embodiment, the bilberry component may have a standardized anthocyanin content of about 20 wt% to about 40 wt%. In a further embodiment, the bilberry component may have an anthocyanin content of about 36% when measured by HPLC and about 25% when measured by UV.

[0060] In one embodiment, at least one of the cyanidin and delphinidin components may be derived from black rice components, blueberry components, and blackcurrant components. In another embodiment, the composition may contain black rice components, blueberry components, and blackcurrant components in a ratio of about 1:1:1. In yet another embodiment, the black rice components, blueberry components, and blackcurrant components may have a ratio of about 1:1.4:4.3. In yet another embodiment, the black rice components, blueberry components, and blackcurrant components may have a ratio of about 1:2:4.

[0061] In one embodiment, the composition may further include prebiotic components or mixtures. Prebiotics can be substances that occur naturally from fruits, vegetables, and grains. Prebiotics can support the microbiome and gastrointestinal system by stimulating the growth or activity of at least one type of bacteria in the colon.

[0062] In one embodiment, the prebiotics or prebiotic mixture may contain inulin, fructooligosaccharides, or a combination thereof. In another embodiment, the prebiotics or prebiotic mixture may contain inulin. In yet another embodiment, the prebiotics or prebiotic mixture may contain fructooligosaccharides. In a further embodiment, the prebiotics or prebiotic mixture may contain inulin and fructooligosaccharides. In some embodiments, the prebiotics or prebiotic mixture may promote a healthy microbiome by increasing the fuel available to beneficial bacteria.

[0063] In one embodiment of the composition, the prebiotics or prebiotic mixture may contain inulin. In one embodiment, the inulin may be in powder form. In one embodiment, the inulin may be chicory inulin. In another embodiment, the inulin may be chicory inulin having oligosaccharides and polysaccharides with fructose units linked by β(2-1) bonds. In another embodiment, the raw material for the inulin may be provided by Orafti® GR. (Beneo-Orafti, Belgium). In some embodiments, the raw material for the inulin may be derived from banana, onion, wheat flour, garlic, asparagus, wheat, rye, chives, chicory root, sugar beet, or a combination thereof. In one embodiment, the inulin may be recovered from the raw material by diffusion with hot water. In some embodiments, the inulin may be hydrolyzed or partially hydrolyzed by enzymatic treatment.

[0064] In one embodiment, the inulin can be present in the composition in various amounts. In one embodiment, the inulin can constitute about 15 wt% to about 60 wt% of the composition, or it can be the active fraction of the composition. In another embodiment, the inulin can constitute about 15 wt% to about 25 wt% of the composition, or it can be the active fraction of the composition. In yet another embodiment, the inulin can constitute about 40 wt% to about 60 wt% of the composition, or it can be the active fraction of the composition.

[0065] In one embodiment, the composition may contain fructooligosaccharides (FOS). In one embodiment, the FOS may be short-chain FOS (having a degree of polymerization (DP) of 5 or less). The fructooligosaccharides may be derived from a variety of raw materials, including grains, fruits, and vegetables. In one embodiment, the short-chain FOS may be derived from sucrose. In another embodiment, the short-chain FOS may be derived from sugarcane. In yet another embodiment, the short-chain FOS may be derived from non-GMO raw materials. In yet another embodiment, the FOS may be galactooligosaccharides (GOS).

[0066] In one embodiment, the FOS can be present in the composition in various amounts. In one embodiment, the FOS can constitute about 10 wt% to about 40 wt% of the composition, or it can be the active fraction of the composition. In another embodiment, the FOS can constitute about 10 wt% to about 20 wt% of the composition, or it can be the active fraction of the composition. In yet another embodiment, the fructooligosaccharide can constitute about 25 wt% to about 40 wt% of the composition, or it can be the active fraction of the composition.

[0067] In one embodiment, the composition may contain inulin and fructooligosaccharides. In one embodiment, if present, the inulin and fructooligosaccharides may collectively constitute about 55 wt% to about 95 wt% of the composition, or constitute the active fraction of the composition. In another embodiment, the inulin and fructooligosaccharides may collectively constitute about 70 wt% to about 90 wt% of the composition, or constitute the active fraction of the composition.

[0068] The above components can be adapted to various compositions in different ways. Some typical compositions are summarized in the table below. In the table below, some of these examples include excipients such as silicon dioxide, while others show only the active fraction of the aforementioned composition. [Table 1] [Table 2] [Table 3] [Table 4]

[0069] In some embodiments, the composition may be formulated to further include additional excipients.

[0070] In one embodiment, the composition may further contain epicatechin, catechin, or a combination thereof. In some formulations, epicatechin and catechin can act as NADPH oxidase inhibitors.

[0071] In one embodiment, the composition may include a pharmaceutically acceptable carrier. In another embodiment, the composition may include sweeteners, preservatives, flavoring agents, thickeners, or combinations thereof. In yet another embodiment, the composition may further include coatings, isotonic agents, absorption retarders, binders, adhesives, lubricants, disintegrants, colorants, flavoring agents, sweeteners, absorbents, surfactants, emulsifiers, antioxidants, vitamins, minerals, proteins, lipids, carbohydrates, or combinations thereof. In some embodiments, the formulation may include a polymer that sustainably releases a particular compound. Almost any number or type of component necessary to manufacture a specifically requested composition or formulation can be utilized.

[0072] In one embodiment, the composition may be in the form of an orally administered formulation. In one embodiment, the orally administered formulation may be a capsule, tablet, powder, beverage, syrup, gum, wafer, confectionery, suspension, or food. In another embodiment, the orally administered formulation may be a capsule, tablet, softgel, lozenge, sachet, powder, beverage, syrup, suspension, or food. In another embodiment, the orally administered formulation may be formulated into a food or beverage, for example, as a snack bar, cereal, beverage, gum, or other easily ingestible form. In one embodiment, the orally administered formulation may be incorporated into a liquid beverage such as water, milk, juice, or soda. In another embodiment, the orally administered formulation may be formulated into a nutritional beverage. The nutritional beverage may be a premixed formulation or a powder mixture that can be added to a beverage. In another embodiment, the powder mixture may be in the form of granules. In another embodiment, the composition may be a powder that can be sprinkled on food.

[0073] In one embodiment, the oral dosage form may be designed to be administered once daily to a subject who requires it. In another embodiment, the oral dosage form may be designed to be administered to the subject in the morning. In yet another embodiment, the oral dosage form may be administered in the afternoon or evening. In yet another embodiment, the dosage form may be designed to be administered intermittently and temporarily. For example, the dosage form may consist of 2 days of administration followed by 1 day of rest, 3 days of administration followed by 2 days of rest, 3 days of administration followed by 4 days of rest, 4 days of administration followed by 3 days of rest, 5 days of administration followed by 2 days of rest, or 6 days of administration followed by 1 day of rest, and each of these dosage methods may be repeated continuously for a certain period. In yet another embodiment, the dosage method may alternate between administration and rest on a daily basis. The duration of administration can also be varied. For example, the dosage form may be designed to be administered for 2 weeks, 3 weeks, 1 month, 6 weeks, 2 months, 3 months, 4 months, 5 months, 6 months, 1 year, 1.5 years, or indefinitely.

[0074] The oral dosage form may include all of the compositions specified herein. In one example, the oral dosage form has a component selected from the group consisting of black rice component, blueberry component, blackcurrant component, crowberry component, bilberry component, black chokeberry component, or a combination thereof.

[0075] In one embodiment, the oral administration form may contain the black rice component, and the amount of the black rice component may be in the range of approximately 500 mg to approximately 800 mg of the oral administration form. In another embodiment, the black rice component may have a standardized anthocyanin content of approximately 15 wt% to approximately 30 wt%.

[0076] In one embodiment, the oral administration form may contain a blueberry component, and the amount of the blueberry component may range from approximately 100 mg to approximately 3,000 mg of the oral administration form. In another embodiment, the amount of the blueberry component may range from approximately 50 mg to approximately 500 mg. In yet another embodiment, the amount of the blueberry component may range from approximately 2,000 mg to approximately 3,000 mg. In yet another embodiment, the blueberry component may have a standardized anthocyanin content of approximately 0.5 wt% to approximately 25 wt%.

[0077] In one embodiment, the oral administration form may contain a blackcurrant component, and the amount of the blackcurrant component may range from approximately 200 mg to approximately 3,000 mg of the oral administration form. In another embodiment, the amount of the blackcurrant component may range from approximately 50 mg to approximately 500 mg. In yet another embodiment, the amount of the blackcurrant component may range from approximately 2,000 mg to approximately 3,000 mg. In yet another embodiment, the blackcurrant component may have a standardized anthocyanin content of approximately 2.5 wt% to approximately 30 wt%.

[0078] In one embodiment, the oral administration form may contain a crowberry component, and the amount of the crowberry component may range from about 100 mg to about 1,000 mg of the oral administration form. In one embodiment, the crowberry component has a standardized anthocyanin content of about 1 wt% to about 50 wt%. In another embodiment, the crowberry component has a standardized anthocyanin content of about 1 wt% to about 30 wt%. In yet another embodiment, the crowberry component has a standardized anthocyanin content of about 40 wt% to about 50 wt%.

[0079] In another embodiment, the oral administration form may contain a bilberry component, and the amount of the bilberry component may range from about 100 mg to about 700 mg of the oral administration form. In one embodiment, the bilberry component may have a standardized anthocyanin content of about 30 wt% to about 40 wt%.

[0080] In further embodiments, the oral administration form may contain the black chokeberry component, which may be in the range of approximately 50 mg to approximately 700 mg. In another embodiment, the black chokeberry component may be in the range of approximately 100 mg to approximately 600 mg. In yet another embodiment, the black chokeberry component may be in the range of approximately 200 mg to approximately 500 mg. In one embodiment, the black chokeberry component may have a standardized anthocyanin content of approximately 1 wt% to approximately 35 wt%.

[0081] In one embodiment, the oral administration form may further include the prebiotics or prebiotic mixture. The prebiotics or prebiotic mixture may include inulin, fructooligosaccharides, or a combination thereof. In one embodiment, the inulin may be present in the oral administration form in an amount of about 1 to 2 grams. In another embodiment, the inulin may provide about 1 to 2 grams of fiber in the oral administration form.

[0082] In one embodiment, the oral administration form may include fructooligosaccharide (FOS). In one embodiment, the FOS in the oral administration form may be in the range of about 1 gram to about 1.5 grams. In another embodiment, the FOS in the oral administration form may be in the range of about 3 grams to about 4 grams.

[0083] The oral administration form can provide various amounts of anthocyanins. In one embodiment, the oral administration form can provide a total of about 200 mg to about 300 mg of anthocyanins. In another embodiment, the oral administration form can provide about 50 mg to about 100 mg of anthocyanins. In one embodiment, the oral administration form can provide about 80 mg of anthocyanins. In another embodiment, the oral administration form can provide about 215 g of anthocyanins.

[0084] The oral administration form can also provide various amounts of fiber. In one embodiment, the oral administration form can provide a total of about 1.5 grams to about 3 grams of fiber. In one embodiment, the oral form can provide about 2.6 g, 2.7 g, or 2.9 g of fiber.

[0085] A method for preparing the composition is also included in the present invention. In one embodiment, the method may include the steps of mixing the components (whether raw materials or extracts) and processing the combined components into a desired form of the composition. In one embodiment, the desired form of the composition may be a tablet, capsule, powder, food, or beverage. Procedures that can be used to mix the components are well known to those skilled in the art.

[0086] Furthermore, this specification presents a method for treating intestinal hyperpermeability in a subject. In one embodiment, the method may include administering to the subject a composition that promotes intestinal health. In one embodiment, the composition that promotes intestinal health may be as described herein.

[0087] In another embodiment, the intestinal health-promoting composition may be administered to the subject daily. In one embodiment, the administration may be carried out in the morning. In yet another embodiment, the administration may be carried out over a long period of time, such as daily for at least 3 weeks, at least 4 weeks, at least 6 weeks, at least 8 weeks, at least 12 weeks, at least 6 months, at least 9 months, at least 1 year, at least 2 years, or indefinitely.

[0088] Furthermore, methods for treating conditions or diseases related to the gastrointestinal health of subjects are presented. In one embodiment, the method may include a step of maximizing the tight junction integrity of the subject's gastrointestinal epithelial cells. In another embodiment, the method may include a step of suppressing the subject's intestinal hyperpermeability.

[0089] In one embodiment, the method can improve the gastrointestinal health of the subject. The improvement in gastrointestinal health may vary.

[0090] In one embodiment, the improvement may include an improvement in the subject's bowel habits compared to the subject's bowel habits before the method was performed. In another embodiment, the improvement in the subject's bowel habits may include a reduction in straining during and after bowel movements. In yet another embodiment, the improvement may be a reduction in abdominal distension, discomfort, flatulence, or a combination thereof. In yet another embodiment, the improvement may be a reduction in the subject's intestinal permeability. In yet another embodiment, the improvement may be a reduction in symptoms / onset of leaky gut syndrome.

[0091] In further embodiments, the improvement in gastrointestinal health may be due to the suppression of intestinal dysbiosis. In one embodiment, the improvement can be demonstrated by a decrease in fecal calprotectin levels. In yet another embodiment, the improvement may be due to an increase in single-chain fatty acid levels.

[0092] In one embodiment of the method, the condition or disease may include inflammation, inflammatory bowel disease, irritable bowel syndrome, chronic bowel disease, celiac disease, Crohn's disease, ulcerative colitis, food intolerance, dyspepsia, low-level chronic enteritis, gastrointestinal infection, or a combination thereof.

[0093] In another embodiment, the condition or disease may be defined as inflammation, and the inflammation in the subject's digestive tract may be reduced compared to the subject's gastrointestinal inflammation before performing the method. In yet another embodiment, the reduction may be approximately 50% of the inflammation. In yet another embodiment, the reduction may be approximately 60% of the inflammation. In yet another embodiment, the reduction may be approximately 70% of the inflammation. In one embodiment, the reduction may be up to 73% of the inflammation. In yet another embodiment, inflammatory biomarkers may be moderately reduced with 3 weeks of supplementation.

[0094] In further embodiments, the condition or disease may include nutrient malsorption, endotoxemia, intestinal hyperpermeability, or a combination thereof.

[0095] In another embodiment, the condition or disease may include obesity, obesity-related symptoms, allergies, cardiovascular conditions, type 1 diabetes, type 2 diabetes, rheumatoid arthritis, insulin resistance, cancer, metabolic syndromes, asthma, neurodegenerative diseases, or a combination thereof.

[0096] In one embodiment, the condition or disease may be a cardiovascular condition or disease. In one embodiment, the cardiovascular condition or disease may be an increase in high-density lipoprotein cholesterol in a subject not taking cardiovascular medication. In another embodiment, the cardiovascular condition or disease may be a decrease in HbA1c levels. In yet another embodiment, the decrease in HbA1c levels, measured as a decrease from 6-6.4% to less than 6%, may be from a prediabetic level to a normal level. In yet another embodiment, the decrease in HbA1c levels may be from a diabetic level to a prediabetic level. This is a decrease from a level above 6.5% to a level between 6% and 6.4%.

[0097] In further embodiments, the cardiovascular condition or disease may be associated with elevated plasma zonulin levels. In one embodiment, performing the method may result in a decrease in plasma zonulin levels compared to the plasma zonulin levels before performing the method. Elevated plasma zonulin may indicate increased intestinal permeability. Zonulin can regulate intestinal permeability by breaking down tight junctions and allowing larger molecules such as lactose to pass through. By administering the compositions herein, the integrity of tight junctions can be maximized, thereby minimizing the amount of zonulin that passes through the inner lining of the intestine.

[0098] In another embodiment, the condition may be peripheral insulin resistance, and the method can suppress peripheral insulin resistance. In yet another embodiment, the condition or disease may be type 1 diabetes or type 2 diabetes.

[0099] In yet another embodiment, the condition or disease may be a nitric oxide-related disorder, iNOS expression, COX-2 expression, NADPH oxidase, or a combination thereof. In one embodiment, the condition or disease may be caused by pathogens, antigens, and pro-inflammatory factors that can pass through the tight junction.

[0100] In one embodiment, the method may include steps to maximize tight junction integrity or to suppress intestinal hyperpermeability. The step to maximize tight junction integrity may include protecting the subject's gastrointestinal tract from increased permeability of the epithelial cell monolayer induced by TNFα. In another embodiment, the amount of the protective factor may be a concentration dependent on the amount of cyanidin and delphinidin in the subject's gastrointestinal tract. In one embodiment, the amount of the protective factor is dose-dependent.

[0101] In yet another embodiment, the method may further include a step of increasing the transepithelial electrical resistance of the epithelial cells. In yet another embodiment, the method may include a step of increasing FITC-dextran paracellular transport.

[0102] In one embodiment, the condition or disease may be caused by a pro-inflammatory factor. In one embodiment, the pro-inflammatory factor may be an advanced glycation end product. In another embodiment, the pro-inflammatory factor may be a lipopolysaccharide. In a further embodiment, the pro-inflammatory factor may include the cytokine tumor necrosis factor α (TNF-α), IL-6, or a combination thereof.

[0103] In one embodiment of the method described above, the condition or disease may be associated with a signaling pathway involving NF-κB, ERK1 / 2, or a combination thereof.

[0104] In one embodiment, a process to maximize the tight junction integrity may reduce the increased intestinal permeability induced by high fat content.

[0105] In another embodiment, the epithelial cells may include a Caco-2 cell monolayer.

[0106] In another embodiment, the method may include a step of optimizing the balance of the gut microbiota of the gastrointestinal tract. In one embodiment, the step of optimizing the balance of the gut microbiota may include increasing the level of symbiotic bacteria in the gastrointestinal tract by comparing it to the level of symbiotic bacteria in the gastrointestinal tract before the administration of the method. In another embodiment, the symbiotic bacteria may belong to the genus Bifidobacterium. In one embodiment, the symbiotic bacteria may belong to the phylum Bacteroidetes. In another embodiment, the symbiotic bacteria may be Bacterdies caccae, Bacteriodes uniformis, or a combination thereof. In yet another embodiment, the increase in the symbiotic bacteria after the subject has been given the method daily for eight weeks may be at least 20%. In another embodiment, the increase in the symbiotic bacteria after the subject has been given the method daily for eight weeks may be about 5%, about 10%, about 15%, or about 25%.

[0107] In one embodiment of the method, the step of optimizing the balance of the gut microbiota may include a step of increasing bacterial diversity by comparing the number of bacteria present in the gut before the method was performed with other positive bacteria. In one embodiment, the bacterial diversity may include at least 200 species. In another embodiment, the method may include a step of reducing harmful enteric bacteria by comparing the level of harmful enteric bacteria present in the gut before the method was performed with other positive bacteria. In one embodiment, the harmful enteric bacteria may include the phylum Firmicutes. Some studies have shown that the phylum Firmicutes constitutes a high proportion of the gut microbiota of obese individuals. In yet another embodiment, the reduction in the phylum Firmicutes after the subject has been given the method daily for 8 weeks may be a reduction of 15% or more in the level of the phylum Firmicutes. In yet another embodiment, the reduction in the phylum Firmicutes after the subject has been given the method daily for 8 weeks may be a reduction of 5% or more, a reduction of about 10%, a reduction of about 12%, a reduction of about 15%, or a reduction of about 20%.

[0108] In one embodiment, the method may include a step of optimizing the gut microbiota by changing the Firmicutes:Bacteroides ratio. In one embodiment, the Firmicutes:Bacteroides ratio may decrease by approximately 3% after the subject has been given the method daily for 8 weeks. The Firmicutes:Bacteroides ratio may be a contributing factor to obesity. Individuals with a high body mass index show differences in their gut microbiota at the phylum level, with potentially higher Firmicutes concentrations and lower Bacteroides concentrations. The changes in the microbiota were not associated with calorie intake but were found to be associated with body mass index. Therefore, by changing this ratio, it may be possible to reduce the weight of obese individuals when administered for a certain period of time.

[0109] In another embodiment of the method for reducing harmful enteric bacteria, the harmful enteric bacteria may include the phylum Actinomycetes. In one embodiment, the reduction of the Actinomycetes after the subject has been given the method daily for eight weeks may be at least about 5%.

[0110] In yet another embodiment of the method for reducing harmful intestinal bacteria, the harmful intestinal bacteria may include Helicobacter pylori, which may be associated with ulcers and heartburn. In yet another embodiment of the method, the harmful intestinal bacteria may include Clostridium. In yet another embodiment, the harmful intestinal bacteria may include Crab Sierra.

[0111] In one embodiment, the method may further include a step of providing a fuel source to symbiotic bacteria of the gut microbiota. In this step, the method can form a larger proportion of symbiotic bacteria that ultimately contribute to systemic health benefits.

[0112] In another embodiment, a method is provided for maximizing the tight junction integrity of the gastrointestinal epithelial cells of the subject. In one embodiment of the method, the improvement of tight junction integrity may include a step of restoring tight junction integrity. In another embodiment of the method, the improvement may include a step of maintaining tight junction integrity. In a further embodiment, the method may provide systemic health benefits. These health benefits may include improvements in conditions or diseases such as celiac disease, IBS, Crohn's disease, ulcerative colitis, food intolerance, allergies, dyspepsia, mild chronic inflammation, obesity, type 1 diabetes, type 2 diabetes, rheumatoid arthritis, insulin resistance, metabolic syndromes, asthma, atopic dermatitis, leaky intestinal barrier, tight junction barrier dysfunction, decreased plasma glucose levels, plasma free fatty acid levels, high-density lipoprotein levels, fatty liver, intestinal filmictes:bacteroides levels, abdominal distension, abdominal gas, abdominal pain, bowel function, proliferation of beneficial intestinal bacteria, single-chain fatty acid production, plasma zonulin levels, HbA1c levels, diabetes, prediabetes, nutrient absorption, and combinations thereof.

[0113] The benefits of the various methods described above can be achieved by administering the aforementioned gut health-promoting composition to subjects. In one embodiment, the composition may be administered daily for an extended period. In another embodiment, the composition may be administered intermittently and sporadically based on the administration method. For example, the administration method may involve administering the drug for two days followed by a one-day break. In another embodiment, the administration method may involve administering the drug for five days followed by a two-day break. In yet another embodiment, the administration method may involve administering the drug for three days followed by a one, two, three, or four-day break. In yet another embodiment, the administration method may involve administering the drug for four days followed by a one, two, three, or four-day break. In some embodiments, the extended period may vary. In one embodiment, the extended period may be approximately four weeks. In another embodiment, the extended period may be approximately six weeks, eight weeks, twelve weeks, sixteen weeks, twenty weeks, six months, nine months, one year, or more than one year. In some embodiments, the benefits of implementing the method for a certain period may increase with longer administration periods.

[0114] Embodiment One embodiment presented herein contains a composition that promotes intestinal health, comprising cyanidin and delphinidin in an amount sufficient to treat intestinal hyperpermeability.

[0115] In one embodiment of the composition, the cyanidin and delphinidin are present collectively in amounts that maintain intestinal permeability.

[0116] In one embodiment of the above composition, the cyanidin and delphinidin are present collectively in amounts that suppress intestinal hyperpermeability.

[0117] In one embodiment of the composition, the composition may include at least one raw material of cyanidin and delphinidin derived from black rice, blueberry, blackcurrant, crowberry, bilberry, black chokeberry, or a combination thereof.

[0118] In one embodiment of the above composition, the raw materials for cyanidin and delphinidin are derived from black rice components, blueberry components, and blackcurrant components.

[0119] In one embodiment of the composition, the composition contains a black rice component, the black rice component is derived from a component selected from the group consisting of black rice grains, black rice concentrate, black rice extract, black rice powder, or a combination thereof.

[0120] In one embodiment of the above composition, the black rice component is a black rice extract.

[0121] In one embodiment of the above composition, the black rice component is derived from black rice grains.

[0122] In one embodiment of the composition, the black rice component constitutes approximately 2.5 wt% to approximately 20 wt% of the active fraction of the composition.

[0123] In one embodiment of the composition, the black rice component constitutes about 10 wt% to about 15 wt% of the active fraction of the composition.

[0124] In one embodiment of the composition, the black rice component constitutes approximately 2.5 wt% to approximately 7.5 wt% of the active fraction of the composition.

[0125] In one embodiment of the composition, the black rice component has a standardized anthocyanin content of about 10 wt% to about 30 wt%.

[0126] In one embodiment of the composition, the black rice component has a standardized anthocyanin content of about 20 wt%.

[0127] In one embodiment of the composition, the black rice component has a standardized anthocyanin content of about 25 wt%.

[0128] In one embodiment of the above composition, the black rice component is derived from Oryza sativa L.

[0129] In one embodiment of the above composition, the composition has the blueberry component, and the blueberry component is an ingredient selected from the group consisting of blueberry fruit, blueberry extract, blueberry concentrate, blueberry juice, blueberry powder, or a combination thereof.

[0130] In one embodiment of the above composition, the blueberry component is blueberry powder.

[0131] In one embodiment of the above composition, the blueberry component is blueberry juice.

[0132] In one embodiment of the composition, the blueberry component constitutes about 1 wt% to about 30 wt% of the active fraction of the composition.

[0133] In one embodiment of the composition, the blueberry component constitutes about 1 wt% to about 10 wt% of the active fraction of the composition.

[0134] In one embodiment of the composition, the blueberry component constitutes about 25 wt% to about 30 wt% of the active fraction of the composition.

[0135] In one embodiment of the composition, the blueberry component has a standardized anthocyanin content of about 0.5 wt% to about 30 wt%.

[0136] In one embodiment of the composition, the blueberry component has a standardized anthocyanin content of about 0.5 wt% to about 5 wt%.

[0137] In one embodiment of the composition, the blueberry component has a standardized anthocyanin content of about 20 wt% to about 30 wt%.

[0138] In one embodiment of the above composition, the blueberry component is derived from Vaccinium uliginosum L.

[0139] In one embodiment of the composition, the composition contains a blackcurrant component, the blackcurrant component comprising a component selected from the group consisting of blackcurrant fruit, blackcurrant extract, blackcurrant concentrate, blackcurrant juice, blackcurrant powder, or a combination thereof.

[0140] In one embodiment of the above composition, the blackcurrant component is a blackcurrant extract.

[0141] In one embodiment of the composition, the blackcurrant component constitutes about 0.5 wt% to about 15 wt% of the active fraction.

[0142] In one embodiment of the composition, the blackcurrant component constitutes about 1 wt% to about 5 wt% of the active fraction.

[0143] In one embodiment of the composition, the blackcurrant component has a standardized anthocyanin content of about 20 wt% to about 40 wt%.

[0144] In one embodiment of the above composition, the blackcurrant component has a standardized anthocyanin content of about 30 wt%.

[0145] In one embodiment of the above composition, the blackcurrant component is derived from Ribes nigrum.

[0146] In one embodiment of the composition, the composition has the crowberry component, the crowberry component is an ingredient selected from the group consisting of crowberry fruit, crowberry extract, crowberry concentrate, crowberry juice, crowberry powder, or a combination thereof.

[0147] In one embodiment of the above composition, the crowberry component is crowberry fruit.

[0148] In one embodiment of the above composition, the crowberry component is a crowberry extract.

[0149] In one embodiment of the composition, the crowberry component constitutes about 1 wt% to about 30 wt% of the active fraction of the composition.

[0150] In one embodiment of the composition, the crowberry component is present in an amount of about 5 wt% to about 25 wt% of the composition.

[0151] In one embodiment of the composition, the crowberry component has a standard anthocyanin content of about 40 wt% to about 50 wt%.

[0152] In one embodiment of the composition, the crowberry component has a standardized anthocyanin content of about 46.7 wt%.

[0153] In one embodiment of the above composition, the crowberry component is derived from Empetrum nigrum.

[0154] In one embodiment of the composition, the composition contains the bilberry component, the bilberry component comprising a component selected from the group consisting of bilberry fruit, bilberry extract, bilberry concentrate, bilberry juice, bilberry powder, or a combination thereof.

[0155] In one embodiment of the above composition, the bilberry component is a bilberry extract.

[0156] In one embodiment of the composition, the bilberry component is in the range of about 0.5 wt% to about 30 wt% of the active fraction of the composition.

[0157] In one embodiment of the composition, the bilberry component is in the range of about 2 wt% to about 20 wt% of the composition.

[0158] In one embodiment of the composition, the bilberry component has a standardized anthocyanin content of about 1 wt% to about 30 wt%.

[0159] In one embodiment of the composition, the bilberry component has a standardized anthocyanin content of about 5 wt% to about 15 wt%.

[0160] In one embodiment of the above composition, the bilberry component contains 36 wt% anthocyanins when measured by HPLC, or 25 wt% anthocyanins when measured by UV.

[0161] In one embodiment of the above composition, the bilberry component is derived from Vaccinium myrtillus.

[0162] In one embodiment of the composition, at least one of the cyanidin and delphinidin raw materials is derived from black rice components, blueberry components, and blackcurrant components.

[0163] In one embodiment of the composition, the black rice component, the blueberry component, and the blackcurrant component are in a ratio of approximately 1:1:1.

[0164] In one embodiment of the composition, the black rice component, the blueberry component, and the blackcurrant component are in a ratio of approximately 1:1.4:4.3.

[0165] In one embodiment of the composition, the composition further comprises a prebiotic mixture.

[0166] In one embodiment of the above composition, the prebiotic mixture comprises inulin.

[0167] In one embodiment of the above composition, the inulin is chicory inulin having oligosaccharides and polysaccharides having fructose units linked by β(2-1) bonds.

[0168] In one embodiment of the composition, the raw material for the inulin is derived from banana, onion, wheat flour, garlic, asparagus, wheat, rye, leeks, chicory root, sugar beet, or a combination thereof.

[0169] In one embodiment of the composition, the inulin is present in an amount of about 15 wt% to about 60 wt% of the composition.

[0170] In one embodiment of the composition, the inulin is present in an amount of about 15 wt% to about 25 wt% of the composition.

[0171] In one embodiment of the composition, the inulin constitutes about 40 wt% to about 60 wt% of the composition.

[0172] In one embodiment of the above composition, the prebiotic mixture comprises fructooligosaccharides.

[0173] In one embodiment of the above composition, the fructooligosaccharide is a short-chain fructooligosaccharide (DP ≤ 5).

[0174] In one embodiment of the above composition, the short-chain fructooligosaccharide is derived from sucrose.

[0175] In one embodiment of the above composition, the short-chain fructooligosaccharide is derived from sugarcane.

[0176] In one embodiment of the composition, the fructooligosaccharide constitutes about 10 wt% to about 40 wt% of the active fraction of the composition.

[0177] In one embodiment of the composition, the fructooligosaccharide constitutes about 10 wt% to about 20 wt% of the active fraction of the composition.

[0178] In one embodiment of the composition, the fructooligosaccharide constitutes about 25 wt% to about 40 wt% of the active fraction of the composition.

[0179] In one embodiment of the above composition, the fructooligosaccharide comprises a galactooligosaccharide.

[0180] In one embodiment of the above composition, the composition further comprises a prebiotic mixture of inulin and fructooligosaccharides.

[0181] In one embodiment of the composition, the inulin and fructooligosaccharides together constitute about 55 wt% to about 95 wt% of the composition.

[0182] In one embodiment of the composition, the mixed raw material of cyanidin and delphinidin constitutes about 5 wt% to about 50 wt% of the composition.

[0183] In one embodiment of the composition, the composition further comprises a pharmaceutically acceptable carrier.

[0184] In one embodiment of the above composition, the composition further comprises a sweetener, a preservative, a flavoring agent, a thickener, or a combination thereof.

[0185] In one embodiment of the above composition, the composition is in an orally administered form.

[0186] In one embodiment of the composition, the oral administration form is a capsule, tablet, powder, beverage, syrup, gum, wafer, confectionery, suspension, or food.

[0187] In one embodiment of the composition, the oral administration form is a powder.

[0188] In one embodiment of the composition, the oral administration form is designed to be administered once daily to a subject who requires it.

[0189] In one embodiment of the composition, the oral administration form is designed to be administered to the subject in the morning.

[0190] In one embodiment of the composition, the oral administration form contains a component selected from the group consisting of black rice component, blueberry component, blackcurrant component, crowberry component, bilberry component, black chokeberry component, or a combination thereof.

[0191] In one embodiment of the composition, the oral administration form contains a black rice component, and the amount of the black rice component is in the range of approximately 500 mg to approximately 800 mg of the oral administration form.

[0192] In one embodiment of the above composition, the black rice component has a standardized anthocyanin content of about 15 wt% to about 30 wt%.

[0193] In one embodiment of the above composition, the oral administration form contains a blueberry component, and the amount of the blueberry component in the oral administration form is approximately 100 mg to approximately 3,000 mg.

[0194] In one embodiment of the composition, the blueberry component has a standardized anthocyanin content of about 0.5 wt% to about 25 wt%.

[0195] In one embodiment of the composition, the oral administration form contains a crossglycum component, and the amount of the crossglycum component in the oral administration form is approximately 200 mg to approximately 3,000 mg.

[0196] In one embodiment of the composition, the blackcurrant component has a standardized anthocyanin content of about 2.5 wt% to about 30 wt%.

[0197] In one embodiment of the composition, the oral administration form contains a crowberry component, and the crowberry component is present in an amount of approximately 100 mg to approximately 1,000 mg of the oral administration form.

[0198] In one embodiment of the composition, the crowberry component has a standardized anthocyanin content of about 1 wt% to about 30 wt%.

[0199] In one embodiment of the composition, the oral administration form contains a bilberry component, and the amount of the bilberry component is in the range of approximately 100 mg to approximately 700 mg of the oral administration form.

[0200] In one embodiment of the composition, the bilberry component has a standardized anthocyanin content of about 30 wt% to about 40 wt%.

[0201] In one embodiment of the composition, the oral administration form contains a black chokeberry component, and the amount of the black chokeberry component is in the range of approximately 50 mg to approximately 700 mg of the oral administration form.

[0202] In one embodiment of the composition, the black chokeberry component has a standardized anthocyanin content of about 1 wt% to about 35 wt%.

[0203] In one embodiment of the composition, the oral administration form comprises a prebiotic mixture.

[0204] In one embodiment of the composition, the prebiotic mixture is present in an amount of about 1 to 2 grams in the oral administration form.

[0205] In one embodiment of the composition, the prebiotic mixture provides about 1 gram to about 2 grams of fiber in the oral administration form.

[0206] In one embodiment of the above composition, the oral administration form further comprises fructooligosaccharides.

[0207] In one embodiment of the composition, the fructooligosaccharide is present in an oral administration form of approximately 1 gram to approximately 1.5 grams.

[0208] In one embodiment of the composition, the fructooligosaccharide is present in an oral dosage form of approximately 3 to 4 grams.

[0209] In one embodiment of the above composition, the oral administration form contains approximately 200 mg to approximately 300 mg of anthocyanins.

[0210] Furthermore, one embodiment of this specification presents a method for treating intestinal hyperpermeability.

[0211] In one embodiment of the method, the method may include the step of administering a composition that promotes intestinal health to a subject.

[0212] In one embodiment of this method, the composition that promotes intestinal health comprises cyanidin and delphinidin in an amount sufficient to treat intestinal hyperpermeability.

[0213] In one embodiment of the above method, the composition that promotes intestinal health further comprises a prebiotic mixture.

[0214] In one embodiment of the above method, the composition that promotes intestinal health further comprises fructooligosaccharides.

[0215] In another embodiment of this method, the composition that promotes intestinal health can be administered daily.

[0216] In one embodiment of this method, the administration can be carried out in the morning.

[0217] In one embodiment of this method, the administration can be carried out over at least three weeks.

[0218] In one embodiment of this specification, a method is presented for treating a condition or disease related to the gastrointestinal health of a subject, comprising maximizing the tight junction integrity of the subject's gastrointestinal epithelial cells.

[0219] In one embodiment of this method, the gastrointestinal health of the subject is improved compared to the gastrointestinal health of the subject before the method was performed.

[0220] In one embodiment of the method, the improved intestinal health of the subject is compared to the intestinal habits of the subject before the method was performed, and the method includes a step of improving the intestinal habits of the subject.

[0221] In one embodiment of the method, the improved gastrointestinal health of the subject includes a step of reducing the occurrence of abdominal distension, discomfort, flatulence, or a combination thereof, compared to the occurrence of abdominal distension, discomfort, flatulence, or a combination thereof, before the method was performed.

[0222] In one embodiment of the above method, the improved gastrointestinal health of the subject includes a step of reducing intestinal hyperpermeability.

[0223] One embodiment of the method includes a step of improving the intestinal dysbiosis of the subject, comparing the improved gastrointestinal health of the subject with the level of intestinal dysbiosis before the method was performed.

[0224] In one embodiment of the method described above, the improved gastrointestinal health of the subject is characterized by a decrease in fecal calprotectin levels.

[0225] In one embodiment of the method described above, the improved gastrointestinal health of the subject is characterized by an increase in short-chain fatty acid levels.

[0226] In one embodiment of the method, the condition or disease is inflammation, inflammatory bowel disease, irritable bowel syndrome, chronic bowel disease, celiac disease, Crohn's disease, ulcerative colitis, food intolerance, dyspepsia, low-level chronic enteritis, gastrointestinal infection, or a combination thereof.

[0227] In one embodiment of the above method, the inflammation is reduced overall.

[0228] In one embodiment of the above method, the inflammation is reduced by up to 73%.

[0229] In one embodiment of the above method, inflammatory biomarkers are gradually reduced by supplementing with a composition that promotes gut health for three weeks.

[0230] In one embodiment of the above method, the condition or disease involves insufficient nutrient absorption, endotoxemia, intestinal hyperpermeability, or a combination thereof.

[0231] In one embodiment of the method described above, the condition or disease is obesity, obesity-related symptoms, allergies, cardiovascular conditions, type 1 diabetes, type 2 diabetes, rheumatoid arthritis, insulin resistance, cancer, metabolic syndromes, asthma, neurodegenerative diseases, or a combination thereof.

[0232] In one embodiment of the above method, the condition or disease is a cardiovascular condition.

[0233] In one embodiment of the above method, the cardiovascular state is an increase in high-density lipoprotein cholesterol in subjects who are not taking cardiovascular therapeutic drugs.

[0234] In one embodiment of the above method, the cardiovascular state is characterized by a decrease in HbA1c levels.

[0235] In one embodiment of the above method, the decrease in the HbA1c value is from a pre-diabetic level to a normal level.

[0236] In one embodiment of the above method, the cardiovascular state is characterized by a decrease in plasma zonulin levels.

[0237] In one embodiment of the above method, the condition is peripheral insulin resistance, and the method reduces peripheral insulin resistance.

[0238] In one embodiment of the above method, the condition or disease is type 1 diabetes or type 2 diabetes.

[0239] In one embodiment of the method, the condition or disease comprises nitric oxide-related disease, iNOS expression, COX-2 expression, NADPH oxidase, or a combination thereof.

[0240] In one embodiment of the method, the condition or disease is caused by pathogens, antigens, and pro-inflammatory factors passing through the tight junctions of the epithelial cells of the gastrointestinal tract.

[0241] In one embodiment of the above method, the step of maximizing the tight junction integrity includes a step of protecting the subject's gastrointestinal tract from increased permeability of the epithelial cell monolayer induced by TNFα.

[0242] In one embodiment of the above method, the amount of the protective factor is a concentration that depends on the amount of cyanidin and delphinidin in the subject's gastrointestinal tract.

[0243] In one embodiment of the above method, the method further comprises the step of increasing the transepithelial electrical resistance of the epithelial cells.

[0244] In one embodiment of the above method, the method further comprises a step of enhancing the paracellular transport of FITC dextran.

[0245] In one embodiment of the above method, the state is derived from an inflammatory factor, and the inflammatory factor has advanced glycation end products.

[0246] In one embodiment of the above method, the state is derived from an inflammatory factor, and the inflammatory factor contains lipopolysaccharides.

[0247] In one embodiment of the above method, the state is derived from a pro-inflammatory factor, and the pro-inflammatory factor is a cytokine such as tumor necrosis factor α (TNF-α), IL-6, or a combination thereof.

[0248] In one embodiment of the method, the condition or disease is associated with a condition or disease related to NF-κB, ERK1 / 2, or a combination thereof, of a signaling pathway.

[0249] In one embodiment of the above method, the step of maximizing the tight junction integrity reduces the increased intestinal permeability induced by high fat content.

[0250] In one embodiment of the above method, the epithelial cells have a monolayer of Caco-2 cells.

[0251] In one embodiment of the method, the method includes the step of optimizing the balance of the gut microbiota in the gastrointestinal tract.

[0252] In one embodiment of the method, the step of optimizing the balance of the gut microbiota includes the step of increasing the level of commensal bacteria in the gastrointestinal tract to be higher than the commensal bacteria value in the gastrointestinal tract before the implementation of the method.

[0253] In one embodiment of the method, the commensal bacteria belong to the genus Bifidobacterium.

[0254] In one embodiment of the method, the commensal bacteria belong to the phylum Bacteroidetes.

[0255] In one embodiment of the method, the commensal bacteria have bacterdies caccae, bacteriodes uniformis, or a combination thereof.

[0256] In one embodiment of the method, after the subject performed the method daily for 8 weeks, the increase in the commensal bacteria was at least 20%.

[0257] In one embodiment of the method, the optimization of the balance of the gut microbiota includes the step of increasing the bacterial diversity.

[0258] In one embodiment of the method, the bacterial diversity has at least 200 species.

[0259] In one embodiment of the method, the method includes the step of reducing the level of harmful gut bacteria compared to the level of harmful gut bacteria before the execution of the method.

[0260] In one embodiment of the method, the harmful gut bacteria have the phylum Firmicutes.

[0261] In one embodiment of the method, after the subject performed the method daily for 8 weeks, the decrease in the phylum Firmicutes was a decrease of 15% or more.

[0262] In one embodiment of the method, the Firmicutes:Bacteroides ratio decreased by approximately 3% after the subject performed the method daily for 8 weeks.

[0263] In one embodiment of the above method, the harmful enterobacteria have the phylum Actinomycetes.

[0264] In one embodiment of the above method, the reduction in the Actinomycetes after the subject performed the method daily for 8 weeks was at least 5%.

[0265] In one embodiment of the above method, the harmful intestinal bacteria include Helicobacter pylori.

[0266] In one embodiment of the above method, the harmful enteric bacteria include the genus Clostridium.

[0267] In one embodiment of the above method, the harmful enteric bacteria include the genus Crab Sierra.

[0268] In one embodiment of the above method, the method includes the step of providing a fuel source for symbiotic bacteria. [Examples]

[0269] Tabletop study on the effects of anthocyanins (AC) on tumor necrosis factor-α-induced loss of CACO-2 cell barrier integrity. The performance of seven extracts containing various types of anthocyanins (ACs) and high AC content was measured to determine their ability to suppress tumor necrosis factor-alpha (TNFα)-induced increased permeability of the Caco-2 cell monolayer. The ACs themselves were also examined, and the relationship between their chemical structure and stereochemistry was determined. The degree of AC content was then determined in relation to the degree of protective effect of the ACs in suppressing TNFα-induced increased permeability of the Caco-2 cell monolayer.

[0270] material Caco-2 cells were obtained from the American Type Culture Collection (Rockville, Massachusetts, USA).

[0271] The cell culture media and reagents were from Invitrogen / Life Technologies (Grand Island, New York, USA).

[0272] HBSS 1X (21-022-CV) was obtained from Corning, Manassas, Virginia, USA.

[0273] Millicell cell culture inserts in 12 mm and 30 mm sizes (polyester membrane with 0.4 μm pore size) (PIHP01250 and PIHP03050, respectively) were obtained from EMD Millipore (Hayward, California, USA).

[0274] Fluorescein isothiocyanate (FITC)-dextran (46944-100MG-F) and human tumor necrosis factor α (TNFα) (T6674-10UG) were obtained from Sigma Chem.Co. (St. Louis, Missouri, USA).

[0275] Human interferon-gamma (IFN-γ) (#8901SC) was obtained from Cell Signaling Technology (Danvers, Massachusetts, USA).

[0276] Pure anthocyanins: delphinidin 3-O-glucoside (myrtillin) (0938), cyanidin 3-O-glucoside (kuromanin) chloride (0915S), and malvidin-3-O-glucoside (oenin) (0911S) were obtained from Extrasynthese (Genay Cedex, France).

[0277] The powder extract with a high anthocyanin content was provided by Pharmanex Research (Nu Skin Enterprises) and included black chokeberry extract powder (total AC 35%), black rice extract (total AC 20%), wild blueberry extract (total AC 5%), bilberry extract (total AC 36%), blackcurrant extract powder (total AC 30%), blueberry extract (total AC 25%), and red grape extract (total AC minimum 10%).

[0278] Method The resolution of the chemical structure was performed by molecular mechanics (MM2) according to Allinger using a predetermined method available in ChemBio3D Ultra 11.0.1 (Cambridge Scientific Computing, Inc.). The structures of the anthocyanins are shown in FIGS. 5 to 9.

[0279] Caco-2 cells were cultured in minimum essential medium (MEM) without phenol red at 37°C in a 5% (v / v) CO2 atmosphere. 10% (v / v) fetal bovine serum, antibiotics (50 μg / ml penicillin and 50 μg / ml streptomycin), 1% non-essential amino acids (NEAA), and 1% sodium pyruvate were added to the MEM medium. The cells were cultured for 21 days until they reached confluence and differentiated into intestinal epithelial cells. The medium was changed every 3 days during the 21 days.

[0280] After 21 days, the Caco-2 cells were differentiated into a polarized monolayer using Millicell cell inserts (30 mm, polyester membrane with a pore size of 0.4 μm) and seeded in a 6-well plate. The apical chamber was composed of 1.5 ml of medium. After initially adding 15 μl of a 1 mg / ml extract solution dissolved in 20% (v / v) ethanol to the apical chamber, the cells were incubated at 37°C and 5% (v / v) CO2.

[0281] Sample collection was performed at 0, 1, and 3 hours. At the first time, 15 μl of culture medium was removed from the apical chamber and the same amount of extract was added. The plate was gently agitated, and 200 μl of sample was immediately removed from the upper chamber. Samples were collected from the apical and lateral bottom chambers at 1 and 3 hours. All samples were immediately acidified with 2.5 μL of 12M HCl and left to stand in a -80°C freezer until analysis.

[0282] Polyphenolic metabolites after interaction with the aforementioned cell layer were identified in each extract by high-performance liquid chromatography (HPLC)-mass spectrometry (MS) / MS. Liquid chromatography was performed using an Agilent Series 1200 instrument (Agilent Technologies, Santa Clara, California, USA) coupled with a diode array detector (DAD), and the cells were monitored at wavelengths of 280 nm and 520 nm. A Phenomenex Kinetex F5 pentafluorophenyl HPLC column (2.6 μM, 100 × 4.6 mm) and a SecurityGuard® cartridge (PFP, 4.0 × 2.0 mm) were used at 0.70 mL min. -1The separation was performed at a flow rate of 37°C and a temperature of 37°C. The injector temperature was set to 4°C for a 7 μL injection. A binary gradient consisting of a 1.0% formic acid (v / v) aqueous solution (mobile phase A) and a 1.0% formic acid (v / v) acetonitrile solution (mobile phase B) was used (Fisher Scientific, Fair Lawn, New Jersey, USA). The gradient was set to 1% B at 0 min, 7.5% B at 7 min, 7.6% B at 14 min, 10% B at 17 min, 12% B at 18.5 min, 30% B at 24 min, 90% B at 25 min, and 1% B from 26 to 30 min. Mass spectrum data were measured using an Agilent 6430 triple quadrupole mass spectrometer (Agilent Technologies, Santa Clara, California, USA) with electrospray injection, and multiple reaction monitoring (MRM) was selected as the acquisition mode. The optical MS / MS source parameters were set to nebulizer 40 psi, capillary voltage +4000V (or -3500V), gas temperature 325°C, and flow rate 5 L-min. The sheath gas temperature was 250°C and the rease flow rate was 11 L-min.

[0283] The anthocyanin standards consisted of malvidin-3-O-glucoside, cyanidin-3-O-glucoside, cyanidin-3-O-galactoside, delphinidin-3-O-glucoside, pelargonidin-3-O-glucoside, and peonidin-3-O-glucoside (Extrasynthese, Genay Cedex, France). Anthocyanins detected for which no standard was available were quantified as equivalent to malvidin-3-O-glucoside. The phenolic acid standards included syringic acid, vanillic acid, protocatechuic acid, 4-hydroxybenzoic acid, gallic acid (Sigma-Aldrich, St. Louis, Missouri, USA), and 3-O-methylgallic acid (Extrasynthese, Genay Cedex, France). Phloroglucinol aldehyde was supplied by Sigma-Aldrich (St. Louis, Missouri, USA).

[0284] To measure transepithelial electrical resistance (TEER), cells were cultured in a transwell insert (12 mm, 0.4 μm pore polyester membrane) to differentiate into a polarized monolayer, which was then seeded into a 12-well plate. First, the epithelial cell monolayer was incubated with interferon-γ for 24 hours to upregulate the TNF-α receptor. Next, the monolayer in the upper chamber was pre-incubated for 30 minutes with anthocyanin-rich extracts (1-10 μg / ml) or purified compounds, or with miltilin chloride or chromatin chloride, and oenin chloride was added to the apical compartment at concentrations of 0.25, 0.5, and 1 μM. After this, TNFα (5 ng / ml) was added to the basal compartment, and the cells were incubated for a further 6 hours.

[0285] To perform TEER evaluation, the incubation medium was removed from the apical and basal compartments, the cells were washed with HBSS 1X, the same solution was added to both compartments, and the TEER was measured. TEER was measured using a Millicell-ERS Resistance System (Millipore, Bedford, Massachusetts, USA) including a dual-electrode volt-ohmmeter. See Figure 10. TEER was, TEER = (Rm - Ri) x A (I) The formula is calculated as follows: Rm: transmembrane resistance, Ri: endogenous resistance of cell-free medium, and A: membrane surface area (cm²). 2 ) was.

[0286] To determine paracellular transport, the clearance of FITC-dextran (4kDa) from the apex to the base was measured. After incubation with TNFα for 6 hours, the culture medium in both compartments was replaced with MEM without fresh serum and phenol red. Then, FITC-dextran was added to the apex compartment (final concentration 100 μM) and incubated for 3.5 hours. After this, 100 μl of culture medium from the base compartment was collected and diluted with 100 μl of HBSS 1X. Fluorescence was measured using a fluorescence plate reader at λexc: 485 nm and λem: 530 nm. Data were analyzed by one-way analysis of variance (ANOVA) using Statview 5.0 (SAS Institute Inc., Cary, North Carolina, USA). Fisher's test of minimum significance was used to examine the difference in group means. A P value < 0.05 was considered statistically significant. Data are shown as mean ± SEM.

[0287] result Table 5 shows the AC concentrations of various extracts evaluated by HPLC-MS / MS. [Table 5] The total AC content of the various extracts ranged from 8.5 to 82 μmol / g per extract. For individual ACs, the content was 0 to 30.05, 0 to 37.43, 0 to 9.70, 0 to 3.45, and 0 to 34.56 μmol / g per extract for cyanidin, delphinidin, petunidine, peonidin, and malvidin glycosides, respectively.

[0288] Figures 5-9 show the chemical and stereostructures of the non-glycosylated forms (anthocyanidins) of AC found in the examined extracts. While the A and C rings are identical in all cyanidins, the didric angle values ​​of the B ring relative to the C ring were shown to be 39, 37, 34, 39, and 43 degrees for cyanidin, delphinidin, petunidine, peonidine, and malvidin, respectively. The position of the B ring may be related to the beneficial properties of cyanidin and delphinidin. As can be seen in Figures 7-9, petunidine, peonidine, and malvidin do not incorporate the B ring in the same position.

[0289] Crowberry extract had the highest total AC content at 82 mol / g and exhibited one of the greatest diversity of individual ACs (cyanidin, delphinidin, petunidine, peonidine, and malvidin glycoside) (only bilberry AC showed greater diversity). Therefore, the crowberry extract was selected to determine the concentration-dependent performance of extracts rich in ACs in preventing TNFα-induced permeability enhancement of Caco-2 monolayers, and was evaluated by measuring TEER and FITC-dextran paracellular transport. Caco-2 monolayers were incubated in the bottom chamber (side-bottom side of the Caco-2 monolayer) with 5 ng / ml TNFα present. This resulted in a significant decrease in TEER (28%, p<0.05) and an increase in FITC-dextran paracellular transport (220%, p<0.05). These findings indicate that, under the experimental conditions, TNFα increased the permeability enhancement of Caco-2 monolayers. Adding crowberry extract (1–10 μg / ml) to the upper chamber (apical side of the Caco-2 monolayer) induced a concentration-dependent recovery of the monolayer TEER and suppression of TNFα-inducible FITC-dextran transport to the lower chamber. As can be seen in Figures 11–12, the TEER value increased with increasing extract volume, and the paracellular transport value also decreased with increasing extract volume. This showed a dose-dependent effect. Next, the relative ability of all extracts containing high levels of AC to suppress TNFα-inducible permeability enhancement of the Caco-2 cell monolayer was determined. At a concentration of 5 μg / ml, the extract had a differential effect in inhibiting TNFα-inducible changes in TEER and FITC-dextran paracellular transport. See Figures 13 and 14. [Table 6] The extracts of black chokeberry (1), black rice grains (2), and blueberry (6) were most effective in inhibiting TNFα-induced changes in TEER, while the extracts of black rice grains (2), bilberry (4), and crowberry (5) were most effective in inhibiting TNFα-induced changes in FITC-dextran paracellular transport. The preventive effect of the extracts against TNFα-induced monolayer permeability enhancement was concentration-dependent.

[0290] To evaluate whether the AC present in the extract is involved in the beneficial effects of the extract on the integrity of the Caco-2 cell barrier, the relationship between TEER and the AC content of the total and individual extracts was examined. There was no significant correlation between TEER values ​​and the total AC content or the peonidin, malvidin, and petunidine glycoside content in the extract. However, TEER values ​​were significantly associated (p<0.03) with the cyanidin (r:0.73) and delphinidin (r:0.81) glycoside content of the extract (see Figures 15 and 16), suggesting the protective effects of these specific ACs. [Table 7] [Table 8] Epicatechin and catechins were found to prevent TNFα-induced loss of gut barrier integrity. See Figures 17-20. [Examples]

[0291] Toxicity testing Toxicity studies were conducted to determine the toxicity of a composition containing cyanidin, delphinidin, and a prebiotic mixture after oral administration to Wistar rats for 90 days. The study also evaluated the dose-response relationship and the no-observed-adverse-effect level (NOAEL).

[0292] Materials-Environment Healthy Wistar rats (Rattus norvegicus) were randomly selected and included in this study. Sixty female rats and sixty male rats, aged 6-8 weeks, were assigned to six groups.

[0293] Each rat was housed individually in a standard-sized (40.5 x 24 x 18.5 cm) polycarbonate cage, with sterilized cornmeal as bedding. The bedding was replaced weekly and as needed to keep the rats clean and dry.

[0294] The room temperature was maintained at 22±3℃ and the relative humidity at 30-70%. Artificial lighting was on for 12 hours and off for 12 hours. The animal room was ventilated at least 12 times per hour.

[0295] The rats were fed pelletized rodent feed and given free access to filtered water. The water was provided in polycarbonate bottles fitted with stainless steel water inlet tubes.

[0296] method The selected rats were examined by a veterinarian and acclimatized to the study conditions for 5 days before the first dose. Rats were assigned to 6 groups before the start of the study using a computer-generated randomization table. Rat body weight variability was minimal, not exceeding ±20% of the mean body weight. [Table 9] Oral formulations were prepared on each administration day. The formulations were dissolved in aqueous solution. High doses were expected to be equivalent to human doses.

[0297] The aforementioned preparation consisted of blueberry extract (3.6%), blackcurrant extract (5.2%), black rice extract (15.6%), chicory inulin (48%), and short-chain fructooligosaccharides (27.6%). It was administered orally once daily at the same time for 90 consecutive days. The amount of the preparation administered was 10 ml / kg body weight.

[0298] Clinical signs and symptoms were observed daily in all rats. Each rat was observed individually in its home cage, then removed from the cage and observed in the hand, and again in a large open area outside the cage, with sensory responses also being observed. In the home cage, the rat's posture, respiratory rate and range, clonic involuntary movements, tonic involuntary movements, vocalization, and eyelid closure were observed. When observed in the hand, the rat's eyelid closure, lacrimation, eye and skin examination, piloerection, and salivation were observed. When observed in a large open area outside the cage, the rat's gait, mobility, arousal level, respiration, clonic movements, tonic movements, vocalization, feeding, urine pooling, fecal masses, stereotypic behavior, and paranoid behavior were observed. Sensory responses were evaluated, including click responses, touch responses, tail pinch responses, and approach responses.

[0299] The rats' body weight was measured on day 1, once a week during the experiment, and on the day of necropsy.

[0300] Urine samples were collected during the final week of administration and analyzed for physical parameters and microscopic examination. To collect urine, rats were housed for 16–18 hours in metabolic cages equipped with a graduated tube at the bottom of the cage.

[0301] On day 91, blood was drawn for hematological and chemical analysis, followed by necropsy. The animals were fasted overnight for 16-18 hours prior to blood collection. On day 91, animals from groups 1-4 were necropsyed after CO2 overdose and underwent physical examination. The cranial, thoracic, and abdominal cavities were opened and examined visually. Organs were decontaminated and weighed. Next, the organs and tissues were collected, and, excluding the testicles fixed in modified Davidson fixative, were preserved in 10% buffered formalin. On day 105, animals from groups 5-4 were necropsyed, physical examinations were performed, and organs were weighed.

[0302] result During the study, physical findings were mostly normal and / or consistent across all groups, and toxicity was judged to be minimal. No clonic or tonic behavior was observed. The only unusual behavior was paper chewing. Paper chewing was observed daily in all groups, including the control group. Contact and approach responses were rapid. Tail pinching was observed as stifling, and pupillary responses were normal.

[0303] No significant differences were observed in blood and clinical chemistry tests between the treatment group and the control group. No significant differences were observed in urine chemistry tests across any of the experimental groups; the results were comparable in rats from groups G1 to G4.

[0304] Average body weight was similar across the test group. This average body weight gradually increased during the study. No changes were observed in dietary intake. Slight changes in organ weight ratios were recorded but were considered toxicologically insignificant. Histopathological damage was randomly distributed in both males and females, and in the control group. Therefore, the damage was considered virtually random.

[0305] This study concluded that none of the doses administered in this study resulted in significant toxicological changes in physical, physiological, neurobehavioral, biochemical, hematological, or histopathological parameters. No toxicologically significant changes were observed with administration. [Examples]

[0306] Animal Test 1 Using a mouse model of obesity induced by a high-fat diet, we investigated the potential of anthocyanin-rich diets to prevent and / or mitigate obesity-induced intestinal inflammation, increased intestinal barrier permeability, and insulin resistance. We evaluated the effects of anthocyanin supplementation on (a) intestinal inflammation, (b) intestinal permeability, (c) gut microbiota, and (d) anthocyanin metabolism changes induced by a high-fat diet.

[0307] material Sixty healthy male C57BL / 6J mice (20-23g) were obtained from Jackson Laboratories and housed in standard stainless steel cages (4 mice / cage). A reinforced environment was provided using mouse cages and bedding. Mice were acclimatized for one week before administration began. Mice were grouped (10 mice / group / time point) and given one of the following diets: control diet, control diet + anthocyanins, high-fat diet, high-fat diet + 2% anthocyanins, high-fat diet + 20% anthocyanins, or high-fat diet + 40% anthocyanins. The components of these diets are described below.

[0308] The control diet TD.06416 was obtained from Harlen Teklad (Wisconsin, USA) and its fat content was adjusted to approximately 10% of the calories.

[0309] The high-fat diet TD.06414 was obtained from Harlen Teklad (Wisconsin, USA) and was a 60% fat diet. This diet is known to cause weight gain over time and lead to obesity. In parallel with weight gain, this diet is known to cause elevated lipid levels (accumulation of triglycerides, cholesterol, and adipocytes), elevated blood glucose levels, the development of insulin insensitivity, and, if administered for a long period, diabetes.

[0310] The anthocyanin mixture was obtained from Nu Skin Enterprises. The mixture contained black rice extract, blackcurrant extract, and blueberry extract.

[0311] method The mice were housed in stainless steel cages. Four mice were housed in each cage. Each group consisted of 10 mice. [Table 10] Each mouse group was given one of the special diets listed above. Diet intake was monitored weekly. Body weight was measured every two weeks, and fresh food was provided. Feces were collected and examined at weeks 0, 2, 4, 6, 8, 10, and 12. Urine was collected at weeks 4 and 10. Forced feeding was performed at weeks 8 and 13. Blood was drawn at week 10 and midway through weeks 12-13. Mice were euthanized at week 14.

[0312] To measure increased intestinal permeability, paracellular transport of FITC dextrin was performed by forced feeding at 8 weeks.

[0313] Gene arrays were generated at various locations in the digestive tract.

[0314] Tight junction integrity was measured as an evaluation of the expression and regulatory mechanisms of tight junction proteins.

[0315] Inflammation was determined by measuring F480+ macrophage infiltration in the intestinal mucosa, plasma CRP, TNF, MCP-1, and iNOS in the liver / intestinal mucosa.

[0316] Furthermore, ITT / GTTs were performed at weeks 10 and 11 to assess the relationship between insulin sensitivity, gut health, and microbiome.

[0317] result During the aforementioned dietary therapy, mice in each group steadily increased their body weight. The greatest weight gain occurred between weeks 0 and 8, while the least weight gain occurred between weeks 8 and 12. Mice fed a high-fat diet and a high-fat diet plus 2% AC gained the most weight overall during the study. [Table 11] Mice fed a diet in which 60% of calories came from fat sources developed obesity, insulin resistance, and increased intestinal permeability within 8 to 16 weeks of starting the diet.

[0318] Despite weight gain, mice fed a high-fat diet consumed less food than mice fed a control diet. Food intake for each group ranged from 2 to 5 grams per week during the study. Animals fed a control diet and a control-plus-anthocyanin diet typically consumed 3 1 / 4 to 4 1 / 2 grams per week. Animals fed a high-fat diet and a high-fat-plus-anthocyanin diet typically consumed 2 1 / 4 to 3 1 / 4 grams per week. Energy intake was similar across all groups. While this study strengthened the correlation between diet and total body weight, previous studies had shown that anthocyanin supplementation generally suppresses the overall tendency toward weight gain.

[0319] Furthermore, the total colon length and body weight of the mice were measured (Figures 21, 22, and 23). FTC-dextran permeability, intestinal permeability, and endotoxins were also measured (Figures 24, 25, and 26, respectively). Mice fed a high-fat diet showed the highest paracellular transport and high intestinal permeability. Even small amounts of anthocyanin addition suppressed or maintained low levels of intestinal permeability in the high-fat diet. Mice fed a high-fat diet also had higher levels of endotoxins.

[0320] Blood glucose levels and insulin concentrations from blood tests are shown in Figures 27 and 28. [Table 12] Mice fed a high-fat diet and a diet containing 20% ​​AC showed the highest blood glucose and insulin levels. Furthermore, glucose tolerance and fasting insulin levels also increased as endotoxins increased (Figures 29 and 30). Endotoxin levels increased with rising IL-6 and IL-1α levels, but IL-β levels did not show a correlation with endotoxin levels (Figures 31-33).

[0321] HOMA-IR (Hostasis Model Assessment of Insulin Resistance, a biomarker of insulin sensitivity), adiponectin, and leptin levels are also shown in Figures 34, 35, and 36, respectively. Adiponectin is an adipocyte-derived cytokine with anti-atherosclerotic, anti-inflammatory, and anti-diabetic properties, and is decreased in obesity. Leptin is another adipocyte-derived cytokine involved in regulating satiety and energy expenditure. Leptin insensitivity, like insulin insensitivity, was associated with weight gain and obesity. Ghrelin is a hormone known to stimulate appetite.

[0322] Furthermore, triglyceride and cholesterol levels were measured in plasma, liver, and stool. See Figures 37–40. As expected, a high-fat diet increased plasma triglycerides, but this increase was prevented by all three AC regimens. Figure 37.

[0323] Since it is well established that a high-fat diet causes fatty liver in these mice, liver triglyceride and cholesterol levels were also measured. Triglycerides significantly increased in the high-fat control group, but lipid accumulation in the liver was prevented in the 40% AC diet, and liver triglyceride levels were similar in the control group, control + AC group, and HF + 40% AC group. Figure 38. Cholesterol levels were lower in all diets containing anthocyanins than in the high-fat diet. Figures 39 and 40. Liver triglyceride levels were also lower in the anthocyanin-containing diets than in the high-fat diet. Figure 41. Representative images of the liver and feces of mice fed each diet can be seen in Figures 42 and 43.

[0324] Interestingly, the highest fecal triglyceride levels were observed in mice fed the high-fat diet + 40% AC diet, suggesting that the anthocyanins inhibited fat absorption. This suggests a mechanism in which the AC mixture prevented hepatic steatosis and insulin insensitivity induced by the high-fat diet. Plasma cholesterol levels increased in mice fed the high-fat diet and the high-fat diet + 2% and 20% AC groups, with only the 40% AC diet reducing plasma cholesterol levels. This may indicate that anthocyanin-containing supplements can lower total cholesterol levels in plasma. All high-fat diet groups showed elevated fecal cholesterol levels, a parameter that appears unaffected by the addition of AC to the diet. [Examples]

[0325] Human trials This study was conducted to determine the effects of compositions containing anthocyanins, inulin, and prebiotic fibers on the microbial composition of obese adults.

[0326] material The single dose of the composition shown in Table 14 below contained 1.9 g of inulin, 1.1 g of fructooligosaccharide, 144 mg of blueberry, 206 mg of blackcurrant extract, and 618 mg of black rice extract. [Table 13]

[0327] method Initial screening was conducted two weeks prior to the start of the trial. During this initial screening, prospective participants' medical histories were reviewed, all combination therapies were presented, and inclusion and exclusion criteria were identified. Resting blood pressure, heart rate, weight, height, and body mass index of prospective participants were also measured. Pregnancy tests were performed as appropriate.

[0328] After a two-week trial period, 81 participants were enrolled in the main study. These participants were primarily female (73%) and Caucasian (93%). Accepted participants were men and women aged 20–60 years. Their BMI ranged from 29.9–39.9 ± 1 kg / m². 2 (29.2~40.6kg / m 2 Participants agreed to maintain their exercise level, discontinue the use of prebiotics and probiotics and / or polyphenol supplements during the study period, and discontinue the consumption of anthocyanin-containing foods (blueberries, blackberries, cherries, grapes, grape juice, pomegranates, raspberries, huckleberries, strawberries, and wine) two days prior to baseline assessment and during the study.

[0329] Participants were instructed to take one sachet of the formulation each morning at breakfast, mixed with their preferred beverage or food. If they forgot to take a dose, they were instructed to take the next dose as soon as they remembered. Participants were not allowed to take more than one sachet per day. Participants also kept a diary of concomitant therapies, adverse events, food intake, daily bowel habits, and daily abdominal discomfort. In addition, participants completed questionnaires regarding abdominal distension and flatulence, and their anthropometric measurements were assessed.

[0330] The aforementioned participants met with the research team at the initial evaluation and on days 0, 29, and 57 of the trial. Stool, urine, and blood samples were collected on days 0 and 57.

[0331] Stool samples were collected on day 0 and within 48 hours of day 57. Microbial composition and calprotectin were measured in the stool samples by a laboratory at the University of Wisconsin. Microbial composition was measured by 16s rRNA at Illumina Platform and R&D Systems (Minneapolis, Minnesota, USA).

[0332] Laboratory parameters (CBC, electrolytes (N, K, Cl, Ca), HbA1c, creatinine, AST, ALT, GGT, and bilirubin) were evaluated at the initial assessment and at the end of the study.

[0333] Urine screening was performed at the KGK Synergize Clinic. Blood parameters were measured by the LifeLabs central laboratory using standard methods. [Table 14]

[0334] Data entry and verification were performed according to KGK Synergize's standard operating procedures. Statistical analysis was performed based on the results of all participants who consumed 80% or more of the prescribed dose. The normality and log-normality of the measured parameters were examined. Non-normal parameters were analyzed using appropriate non-parametric tests. Numerical efficacy and evaluation parameters were formally tested for significance using paired Student's t-tests.

[0335] result Of the 51 participants initially enrolled in the trial, 46 completed the study. Four participants dropped out after day 0 and before the first comparative measurement on day 29. One participant dropped out between the follow-up examinations on day 29 and day 57.

[0336] Microbial diversity Participants supplemented with the aforementioned composition showed favorable changes in their microbial composition, as indicated by the change in the Firmicutes:Bacteroides ratio (Figure 44). The ratio decreased from 4.98 to 1.45 (Firmicutes decreased from 74.9% to 59%, while Bacteroides increased from 13.8% to 34.5%). Furthermore, Actinomycetes decreased from 8.5% to 3.4%. After the supplementation, a total of 8 phyla (6 Bacteria, 1 Archaea, and 1 Other) and 40 genera (7 Actinomycetes, 8 Bacteroides, 1 Euryarchaea, 2 Firmicutes, and 3 Proteobacteria) changed. [Table 15] [Table 16]

[0337] inflammation After the test, fecal calprotectin levels decreased, indicating a tendency towards suppression of gastrointestinal inflammation. Calprotectin levels are summarized in the table below and shown in Figure 45. [Table 17]

[0338] toilet habits Compared to baseline, a 5% increase in the Bristol Stool Texture Score was observed at weeks 6 and 7. While the frequency of bowel movements did not change during the study, bowel habits improved (Figure 46). Participants reported a decrease in straining before defecation (33%) and at the end of defecation (51-54%), and a decrease in incomplete bowel movements. [Table 18] [Table 19]

[0339] Abdominal distension, discomfort, and flatulence. Participants' abdominal distension decreased from week 3 to week 8 compared to baseline (41% at week 4, 52% at week 5, and 50% at week 8). Figure 47. [Table 20] Participants also experienced abdominal pain and reduced flatulence. The severity of flatulence decreased from 22% at week 5 to 11% at week 8. Figure 47. [Table 21] [Table 22]

[0340] Vital signs Participants experienced a clinically insignificant decrease in diastolic blood pressure from baseline to the end of the study. [Table 23]

[0341] conclusion Overall, this study demonstrated that the formulation can improve the gut microbiota of individuals who are mildly to moderately obese but otherwise healthy. During the study, participants showed a significant decrease in the Firmicutes phylum and a significant increase in the proportion of the Bacteroidetes phylum. The Firmicutes:Bacteroidetes ratio decreased with supplementation therapy from day 0 to day 57. The ratio changed from 4.98 to 1.45. Furthermore, actinomycete levels decreased. These three phyla constitute approximately 97% of the bacterial composition at baseline (97.2%) and after supplementation therapy (96.9%).

[0342] In this study, 30 adverse events were recorded in 18 participants. Of the 30 reported events, 11 were assessed as unlikely and 7 as unrelated. Two adverse events were judged as likely, including fecal discoloration and tooth discoloration. Ten slightly likely adverse events included abdominal discomfort, diarrhea, fecal discoloration, increased bowel movement frequency, and vomiting.

[0343] Formulations, methods for producing these formulations, and uses of said formulations have been reported. However, it will be readily apparent to those skilled in the art that various modifications and alterations can be made to the apparent properties without deviating from the spirit of the invention, and all such modifications and alterations are considered to fall within the scope of the invention as defined in the appended claims. Such modifications and alterations include, but are not limited to, the capsule, tablet, powder, lotion, food, powder, or bar manufacturing process, and initial components added to act on vitamins, flavorings, and carriers. Other such modifications and alterations include the use of herbs or other plant formulations, including combinations of the preferred embodiments described above. Many additional modifications and variations of the embodiments described herein can be made without deviating from the scope, as will be obvious to those skilled in the art. The specific embodiments described herein are provided only as examples.

Claims

1. A composition for alleviating symptoms associated with tight junction barrier dysfunction, comprising cyanidin, delphinidin, or a combination thereof in an amount exceeding 5 wt% of the composition.

2. A composition according to claim 1, wherein the raw materials for cyanidin, delphinidin, or a combination thereof are derived from black rice components, blueberry components, blackcurrant components, crowberry components, bilberry components, black chokeberry components, or a combination thereof.

3. The composition according to claim 2, wherein the raw materials for cyanidin, delphinidin, or a combination thereof are derived from black rice components, blackcurrant components, and bilberry components.

4. A composition according to claim 3, wherein the black rice component comprises 2.5 wt% to 20 wt% of the composition.

5. The composition according to claim 3, wherein the black rice component has an anthocyanin content of 10 wt% to 30 wt%.

6. A composition according to claim 3, wherein the blackcurrant component is present in an amount of 0.5 wt% to 15 wt% of the composition.

7. The composition according to claim 3, wherein the blackcurrant component has an anthocyanin content of 20 wt% to 40 wt%.

8. A composition according to claim 3, wherein the bilberry component comprises 0.5 wt% to 30 wt% of the composition.

9. The composition according to claim 3, wherein the bilberry component has an anthocyanin content of 1 wt% to 40 wt%.

10. A composition according to claim 1, further comprising a pharmaceutically acceptable carrier.

11. A composition according to claim 1, further comprising a sweetener, a preservative, a flavoring agent, a thickener, or a combination thereof.

12. A composition according to claim 2, wherein the composition is in an orally administered form.

13. The composition according to claim 12, wherein the oral administration form contains 200 mg to 300 mg of anthocyanins.

14. The composition according to claim 12, wherein the black rice component is 500 mg to 800 mg in the oral administration form.

15. The composition according to claim 12, wherein the blackcurrant component is 200 mg to 3000 mg in the oral administration form.

16. The composition according to claim 12, wherein the bilberry component is 100 mg to 700 mg in the oral administration form.

17. The composition according to claim 1, wherein the dysfunction of the tight junction barrier is induced by TNFα.

18. The composition according to claim 1, wherein the symptoms associated with the dysfunction of the tight junction barrier include inflammation, inflammatory bowel disease, irritable bowel syndrome, chronic bowel disease, celiac disease, Crohn's disease, ulcerative colitis, food intolerance, indigestion, low-level chronic enteritis, gastrointestinal infection, nutrient malabsorption, endotoxemia, obesity, allergy, cardiovascular disease, type 1 diabetes, type 2 diabetes, rheumatoid arthritis, insulin resistance, cancer, metabolic syndrome, asthma, neurodegenerative disease, or a combination thereof.

19. Use of cyanidin, delphinidin, or a combination thereof in the preparation of a composition according to any one of claims 1 to 19 for treating symptoms associated with dysfunction of the tight junction barrier.

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