Gastrointestinal health composition

Abstract

The present invention relates generally to compositions for improving gastrointestinal health, nutritional supplements and / or dietary supplements formulated for improving gastrointestinal health, and the use of such compositions and / or supplements for gastrointestinal reprogramming.

Description

Technical Field

[0001] The present invention generally relates to compositions for improving gastrointestinal health, nutritional supplements formulated for such use, and / or functional foods, and the use of such compositions and / or supplements for gastrointestinal reprogramming.

Background Art

[0002] Gastrointestinal diseases Gastrointestinal diseases and disorders such as irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD) are caused by the interaction of several different factors including microbial, immune, genetic, and environmental factors (Bernstein CN, 2010). Enterotoxemia of gastrointestinal bacteria has downstream effects such as mucosal inflammation, immune disruption, and mucosal damage (Bernstein CN, 2010). Gastrointestinal disorders of IBD and IBS share common disease characteristics such as changes in the microbiome, intestinal dysfunction, and mucosal damage. The factor that differentiates these two diseases is the level of inflammation observed. In contrast to IBD, which is characterized by chronic inflammation, IBS is mild inflammation. Current treatment approaches for IBD aim to suppress inflammation for the induction and maintenance of remission. In contrast, gastroenterologists treating IBS mainly treat symptoms such as abdominal pain, abdominal distension, and alternating episodes of constipation and diarrhea. The current treatment approaches employed for treating both IBD and IBS have a high treatment failure rate.

[0003] Therefore, there is still an unmet medical need in the art for products that can successfully treat, and / or alleviate, or eliminate these conditions and their symptoms from patients with IBD and IBS without suffering from the currently observed high treatment failure rate.

[0004] Thus, it is an object of the present invention to provide a gastrointestinal functional food that can at least help to address this unmet need and / or provide at least a useful option to the public.

[0005] In this specification, which refers to a patent specification, other external documents, or other information sources, this is generally intended to provide a context for discussing the features of the present invention. Specifically, unless otherwise indicated, such references to external documents should not be construed as an admission that such documents, or such information sources, are prior art in any jurisdiction or form part of the common general knowledge in the art.

Summary of the Invention

[0006] In one aspect, the present invention relates to a composition comprising the following components. (a) Bromelain, (b) One or more SCFAs or esters thereof, (c) L-Threonine, and (d) Vitamin D or a metabolite thereof In another aspect, the present invention relates to an oral dosage form comprising from about 1 to about 10 g of the composition of the present invention.

[0007] In another aspect, the present invention relates to an oral dosage form consisting essentially of the composition of the present invention, wherein at least two components of the composition are included in a delayed-release formulation. In another aspect, the present invention relates to a method of preventing, treating, or managing IBD or IBS, comprising administering a therapeutically effective amount of the composition or oral dosage form of the present invention to a subject in need thereof.

[0008] In another aspect, the present invention relates to a method of at least partially reducing at least one symptom of IBD or IBS, comprising administering a therapeutically effective amount of the composition or oral dosage form of the present invention to a subject in need thereof.

[0009] In another aspect, the present invention relates to a method of improving at least one parameter associated with IBD or IBS, comprising administering a therapeutically effective amount of the composition or oral dosage form of the present invention to a subject in need thereof.

[0010] In another aspect, the present invention relates to a method for reducing the adhesion of bacteria to cells of the gastrointestinal tract of a subject in need thereof, comprising contacting the bacteria with an effective amount of the composition or oral dosage form of the present invention. In another aspect, the present invention relates to a method for reducing the invasion of bacteria into cells of the gastrointestinal tract of a subject in need thereof, comprising contacting the bacteria with an effective amount of the composition of the present invention, or an oral dosage form.

[0011] In another aspect, the present invention relates to a method for increasing the gastrointestinal mucosal layer of a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the composition of the present invention, or an oral dosage form. In another aspect, the present invention relates to a method for improving gastrointestinal barrier integrity in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the composition of the present invention.

[0012] In another aspect, the present invention relates to a method for restoring the homeostasis of the microbiota of a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the composition of the present invention. In another aspect, the present invention relates to a method for treating gastrointestinal inflammation in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the composition of the present invention.

[0013] Other aspects of the present invention are merely illustrative and may become apparent from the following description given with reference to the accompanying drawings.

Brief Description of the Drawings

[0014] The present invention will now be described by way of example with reference to the following drawings.

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Mode for Carrying Out the Invention

[0015] Definitions The following definitions are presented for reference by those skilled in the art in the practice of the present invention to better define the present invention.

[0016] Unless otherwise specified, all technical and scientific terms used herein should be understood to have the same meaning as commonly understood by one of ordinary skill in the relevant technical fields to which this disclosure pertains.

[0017] The practice of the present invention can also be considered to be carried out using standard cell biology, microbiology, molecular biology, pharmacology, and biochemistry protocols and procedures known in the art and described in numerous references commonly available in the art relevant to the fields to which this disclosure pertains.

[0018] The term "food" means any substance, whether processed, semi-processed or raw, intended for consumption by animals including humans, and includes, but is not limited to, solid and semi-solid foods, alcoholic beverages, beverages, and chewing gums. The food composition of the present invention includes the composition described herein in combination with food.

[0019] The term "dietary supplement" means a product intended to supplement the diet and containing one or more of the following ingredients: enzymes, vitamins, minerals, herbs, metabolites, extracts, etc. Dietary supplements are not usually intended to be used as a single item of diet and can be taken independently of food. The dietary supplement of the present invention includes the composition described herein in combination with one or more dietary supplements.

[0020] The term "medical food" means a food that is prescribed to be taken enterally (e.g., via a feeding tube or nasogastric tube) or administered under the supervision of a physician. Medical foods are intended for the dietary management of conditions with specific nutritional requirements. Examples of medical foods include, but are not limited to, nutritional products from a single source, oral rehydration solutions, and products intended for use in the dietary management of metabolic disorders. The medical foods contemplated herein include the composition described herein that is prescribed as a medical food.

[0021] One or more ingestible excipients may be present in the dietary supplement formulation, food composition, dietary supplement, or medical food described herein. Such ingestible excipients may include, as will be apparent to those skilled in the art, carriers, diluents, binders, adhesives, lubricants, plasticizers, disintegrants, colorants, extenders, flavoring agents, sweeteners, buffering agents, absorbents, etc.

[0022] Examples of binders that may be used in the compositions and oral dosages described herein include cellulose derivatives such as ethyl cellulose, hydroxypropyl cellulose, or hydroxypropyl methyl cellulose, povidone, crospovidone, pharmaceutical glazes, rubbers, milk derivatives such as whey and casein, starch, and starch derivatives. Sweeteners may include dipeptide sweeteners, water-soluble natural sweeteners, and artificial sweeteners. Flavoring agents may include synthetic and natural flavor oils including peppermint oil, spearmint oil, cinnamon oil, wintergreen oil; natural and artificial fruit flavors such as citrus oils including lemon, lime, grapefruit, and orange; and fruit essences including cherry, pineapple, strawberry, raspberry, apple, and the like. Bulking substances may include glucose, sucrose, lactose, starch, gelatin, and silicon dioxide. The binders, sweeteners, flavoring agents, and bulking substances described above are all illustrative and not limiting thereto.

[0023] As used herein, the term "different types of pills, tablets, minitabs, capsules, granules, particles, or microparticles" and grammatical variations thereof mean that a given pill, tablet, minitab, capsule, granule, particle, or microparticle described herein as a "type" (including type A, type B, and type C described herein) contains different components, is formulated to contain, or is the main component of, and is delivered to different parts of the intestine and is active in different parts of the intestine. In one embodiment, the different parts of the intestine are the duodenum and the colon.

[0024] As used herein, the terms "treat", "treating", and "treatment" mean a treatment means that reduces, alleviates, improves, manages, prevents, suppresses, stops, or reverses symptoms caused by, or associated with, IBD and / or IBS, including symptoms associated with, or related to, IBD and / or IBS. As is known in the art, a subject may exhibit an observable or measurable (statistically significant) decrease in one or more symptoms associated with, or related to, IBD and / or IBS as an indication of improvement.

[0025] As used herein, the term "effective amount" means an amount effective to prevent, delay, reduce, stabilize, improve, or treat at least one symptom of IBD and / or IBS known in the art and / or described herein. In some embodiments, an effective amount is an amount sufficient to achieve a statistically different result compared to an untreated control or a placebo-treated control.

[0026] A "therapeutically effective amount" of the compositions described herein is an amount sufficient to achieve at least a reduction of at least one symptom associated with IBD and / or IBS during or in anticipation of treatment, or sufficient to achieve a change in that symptom having a therapeutically beneficial effect. Examples of symptom changes contemplated herein include, but are not limited to, a reduction in gastrointestinal inflammation, an increase in the thickness of the gastrointestinal mucosal layer, and / or an improvement in gastrointestinal barrier integrity.

[0027] As used herein, the terms "manage", "managing", and "management" in the context of treatment application to a subject mean a beneficial effect that a subject derives from treatment, but does not result in a cure of the condition. For example, managing a condition includes preventing the condition from worsening.

[0028] As used herein, the terms "prevent", "preventing", and "prevention" in the context of therapeutic application to a subject mean preventing or inhibiting the recurrence, onset, or manifestation of a condition due to the therapeutic application.

[0029] The term "about" when used in connection with a reference numerical indication means a reference numerical indication that varies by up to 10% of that reference numerical indication. For example, "about 100" means from 90 to 110, and "about 6" means from 5.4 to 6.6.

[0030] The term "comprising" as used herein means "consisting at least in part of". When interpreting a description in this specification that includes that term, all of the features presupposed by that term in each description must be present, but other features can also be present. Related terms such as "comprise" and "comprised" are interpreted similarly.

[0031] The term "consisting essentially of" as used herein means a particular material or step and those that do not substantially affect the basic and novel characteristics of the claimed invention.

[0032] The term "consisting of" as used herein means the materials or steps specifically recited in the claimed invention, excluding elements, steps, or components not specifically recited in the claims.

[0033] References to numerical ranges (e.g., 1 to 10) disclosed herein include all rational numbers within that range (e.g., 1, 1.1, 2, 3, 3.9, 4, 5, 6, 6.5, 7, 8, 9, and 10), and also include references to any range of rational numbers within that range (e.g., 2 to 8, 1.5 to 5.5, and 3.1 to 4.7). Thus, all sub-ranges of all ranges explicitly disclosed herein are intended to be explicitly disclosed by this specification. These are merely examples of what is specifically intended, and all possible combinations of numbers between the recited minimum and maximum values should likewise be considered to be explicitly recited in this application.

[0034] Description Gastrointestinal diseases and disorders such as irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD) are caused by several different factors in individuals, including microbial, immune, genetic, and environmental factors.

[0035] The following three factors contribute to the onset and chronic persistence of IBD, IBS, and related symptoms such as changes in bowel habits (i.e., diarrhea, constipation, or a mixture thereof), abdominal pain, and abdominal distension.

[0036] - Loss of gastrointestinal microbial homeostasis - Mucosal damage - Inflammation.

[0037] Currently, there are few market - available products, including dietary supplements, that can successfully re - establish the homeostasis of the gastrointestinal microbiota, reduce inflammation, and promote mucosal healing. Re - establishing microbiota homeostasis is thought to be beneficial in reducing the effects of diseases associated with diarrhea, inflammation, obesity, diabetes, and cardiovascular disease (Vamanu, 2018).

[0038] However, the inventors have discovered a composition of agents that act together to act on all three of these causative factors, which provides effective management of chronic intestinal conditions such as IBD and IBS.

[0039] Disclosed herein is a Gastrointestinal Reprogramming Product (GaRP). GaRP is an all-natural dietary product specifically developed by the inventors to support and maintain gastrointestinal health. The GaRP described herein may be formulated as, or included in, a nutritional supplement, medical food, or pharmaceutical for addressing the major underlying factors associated with gastrointestinal disorders. Without wishing to be bound by theory, the inventors believe that GaRP acts by supporting the individual gastrointestinal microbiota and restoring normal microbial intestinal function. Further, the formulation components reduce inflammation, which is important for restoring normal gastrointestinal tract function.

[0040] Thus, in one aspect, the present invention relates to a composition comprising the following components: (a) Bromelain, (b) one or more SCFAs or esters thereof, (c) one or more amino acids selected from the group consisting of L-threonine, L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof, and (d) vitamin D, or a metabolite thereof.

[0041] In one embodiment, the composition consists essentially of the following components: (a) Bromelain, (b) one or more SCFAs or esters thereof, (c) one or more amino acids selected from the group consisting of L-threonine, L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof, and (d) vitamin D, or a metabolite thereof.

[0042] In one embodiment, (c) comprises, or consists essentially of, one or more amino acids selected from L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof.

[0043] In one embodiment, (c) comprises, or consists essentially of, L-tryptophan, L-arginine, L-glutamine, and L-methionine. In one embodiment, (c) comprises, or consists essentially of, L-arginine, L-glutamine, and L-methionine. In one embodiment, (c) comprises, or consists essentially of, L-tryptophan, L-glutamine, and L-methionine. In one embodiment, (c) comprises, or consists essentially of, L-tryptophan, L-arginine, and L-methionine. In one embodiment, (c) comprises, or consists essentially of, L-tryptophan, L-arginine, and L-glutamine.

[0044] In one embodiment, (c) comprises, or consists essentially of, L-tryptophan and L-arginine. In one embodiment, (c) comprises, or consists essentially of, L-tryptophan and L-glutamine. In one embodiment, (c) comprises, or consists essentially of, L-tryptophan and L-methionine. In one embodiment, (c) comprises, or consists essentially of, L-arginine and L-glutamine. In one embodiment, (c) comprises, or consists essentially of, L-arginine and L-methionine. In one embodiment, (c) comprises, or consists essentially of, L-glutamine and L-methionine.

[0045] In one embodiment, (c) comprises, or consists essentially of, L-tryptophan, L-arginine, L-glutamine, or L-methionine. In another aspect, the present invention relates to a composition comprising the following components: (a) Bromelain, (b) one or more SCFAs or esters thereof, (c) L-threonine, and (d) vitamin D, or a metabolite thereof.

[0046] In one embodiment, the composition consists essentially of the following components: (a) Bromelain, (b) one or more SCFAs or their esters, (c) L - threonine, and (d) vitamin D, or its metabolite.

[0047] In one embodiment, the L - threonine of (c) is used in place of one or more different amino acids selected from the group consisting of L - tryptophan, L - arginine, L - glutamine, and L - methionine. In one embodiment, the composition comprises at least one of the following additional amino acid components: L - tryptophan, L - arginine, L - glutamine, L - methionine, or any combination thereof.

[0048] In one embodiment, the composition consists essentially of the following components: bromelain, SCFA, L - threonine, and vitamin D, or their metabolites. In one embodiment, the one or more SCFAs or their esters are butyric acid, propionic acid, acetic acid, or their physiologically acceptable salts. The SCFA is preferably sodium butyrate, calcium butyrate, sodium propionate, or sodium acetate, more preferably sodium butyrate.

[0049] In one embodiment, the composition consists essentially of the following components: bromelain, butyrate, preferably sodium butyrate, L - threonine, and vitamin D, or their metabolites.

[0050] In one embodiment, the composition contains menthol as a component. In one embodiment, the composition is a dietary supplement, a nutritional supplement, or a food. In one embodiment, the food is a medical food.

[0051] In one embodiment, the composition comprises one or more additional components that are ingestible excipients. Component The inventor has surprisingly confirmed that the various components of the composition of the present invention act synergistically to restore normal gastrointestinal function and reduce the symptoms of IBD and IBS, particularly diarrhea.

[0052] Bromelain is a general term for proteolytic extracts obtained from the stem of pineapple (Ananas comosus). The term "bromelain" is also used to refer to the two main proteolytic enzymes present in pineapple, stem bromelain protease (CAS 37189-34-7 / EC 3.4.22.32) and fruit bromelain protease (EC 3.4.22.33). Although the names represent the main locations in the plant, both enzymes are present in the extract of stem bromelain. Bromelain may contain proteases such as ananain, cosmosin, and canain, as well as small amounts of other compounds, in addition to stem bromelain protease and fruit bromelain protease.

[0053] Bromelain extract is prepared by mixing the skin, stem, leaves, and other non-edible parts of pineapple, filtering the mixture to obtain a liquid containing soluble bromelain enzyme, and then it can be purified, fractionated, concentrated, and / or dried. Bromelain can be fractionated using ion exchange chromatography. The fractions corresponding to the main absorbance peaks can be subjected to SDS-PAGE to isolate and identify the individual enzymes.

[0054] As used herein, the term "bromelain" refers to an extract of proteolytic enzymes obtained from the stem of pineapple and includes crude extracts, purified extracts, or any fraction, or combinations of those fractions.

[0055] Bromelain is a well-known dietary supplement that has not been approved by the US Food and Drug Administration for the treatment of gastrointestinal disorders but has been suggested to be useful as a digestive aid and a general anti-inflammatory agent. It is also suggested to be useful in supporting the body's response to stress, assisting in protein digestion, reducing swelling in the nose and sinuses, removing dead and damaged tissue after burns, relaxing muscles, preventing pulmonary edema, stimulating muscle contraction, and shortening labor, as well as in the treatment of joint pain, sinus infections, allergies, asthma, cancer, varicose veins, hemorrhoids, urinary tract infections, bronchitis, gout, fever, and carpal tunnel syndrome.

[0056] The composition of the present invention combines bromelain with a specific agent that reduces three causative factors of IBD and IBS, which are inflammatory gastrointestinal diseases. In one embodiment, the composition is a unit dosage form containing from about 50 mg to about 2500 mg, preferably from about 250 mg to about 2250 mg, from about 500 mg to about 2000 mg, from about 750 mg to about 1750 mg, from about 1000 mg to about 1600 mg, from about 1300 mg to about 1550 mg, from about 1400 mg to about 1500 mg, from about 1420 mg to about 1460 mg, more preferably about 1440 mg of bromelain. In one embodiment, the composition contains about 1440 mg, preferably about 1030 mg, more preferably about 720 mg of bromelain.

[0057] In one embodiment, the composition is a unit dosage form containing from about 460 mg to about 500 mg of bromelain, preferably about 480 mg of bromelain. In one embodiment, the composition is a unit dosage form containing from 460 mg to 500 mg of bromelain, preferably 480 mg of bromelain.

[0058] In one embodiment, the composition is in a unit dosage form containing 50 mg to 2500 mg, preferably 250 to 2250 mg, 500 to 2000 mg, 750 to 1750 mg, 1000 to 1600 mg, 1300 to 1550 mg, 1400 to 1500 mg, 1420 to 1460 mg, more preferably 1440 mg, 1030 mg, and even more preferably 720 mg of bromelain. In one embodiment, the composition contains 1440 mg, preferably 1030 mg, and more preferably 720 mg of bromelain.

[0059] Short-chain fatty acids (SCFAs) are the end products of the microbial fermentation of major nutrients such as plant polysaccharides that cannot be digested in humans lacking the required glycoside hydrolases and polysaccharide lyases. The microbiome supplies the missing enzymes and produces SCFAs that can promote intestinal epithelial barrier integrity. SCFAs such as butyric acid can also help restore the homeostasis of gut microbiota.

[0060] When the concentration of SCFA increases, the regrowth of beneficial bacteria that reduce inflammation through the inhibition of NF-κB and the decrease of pro-inflammatory cytokines is promoted. The terms "short-chain fatty acid" and "SCFA" refer to weak acids containing 2 to 7 carbon atoms, including the anions of specific basic acids and the salt forms. Regarding the anions, SCFAs include acetate, butyrate, propionate, valerate, caproate, isobutyrate, 2-methyl-isobutyrate, and isovalerate. SCFA salts for use in the compositions of the present invention include alkali metals (e.g., Na and K), and alkaline earth metals (e.g., Mg and Ca).

[0061] The term "its SCFA ester" refers to an ester of SCFA. SCFA esters for use in the compositions of the present invention include methyl, ethyl, etc. The SCFAs and selected amino acids present in the compositions of the present invention promote the regeneration of the intestinal mucosa, which restores the integrity of the gastrointestinal tract and gut microbiota.

[0062] In one embodiment, the composition is a unit dosage form comprising about 45 to 1800 mg of SCFA, preferably about 75 to about 1650 mg, about 250 to about 1500 mg, about 500 to about 1400 mg, about 750 to about 1500 mg, about 1000 to about 1400 mg, about 1100 to about 1300 mg, more preferably about 1200 mg of SCFA. In one embodiment, the composition comprises about 1200 mg, preferably about 600 mg, more preferably about 300 mg of SCFA.

[0063] In one embodiment, the composition is a unit dosage form comprising 45 to 1800 mg of SCFA, preferably 75 to 1650 mg, 250 to 1500, 500 to 1400, 750 to 1500, 1000 to 1400, 1100 to 1300 mg, more preferably 1200 mg of SCFA. In one embodiment, the composition comprises 1200 mg, preferably 600 mg, more preferably 300 mg of SCFA.

[0064] In one embodiment, the composition is a unit dosage form comprising about 45 to 1800 mg of sodium butyrate, sodium propionate or sodium acetate, preferably about 75 to about 1650 mg, about 250 to about 1500 mg, about 500 to about 1400 mg, about 750 to about 1500 mg, about 1000 to about 1400 mg, about 1100 to about 1300 mg, more preferably about 1200 mg of sodium butyrate. In one embodiment, the composition comprises about 1200 mg, preferably about 600 mg, more preferably about 300 mg of sodium butyrate, sodium propionate or sodium acetate, preferably sodium butyrate.

[0065] In one embodiment, the composition is a unit dosage form comprising 45 to 1800 mg of sodium butyrate, sodium propionate or sodium acetate, preferably 75 to 1650 mg, 250 to 1500, 500 to 1400, 750 to 1500, 1000 to 1400, 1100 to 1300 mg, more preferably 1200 mg of sodium butyrate. In one embodiment, the composition comprises 1200 mg, preferably 600 mg, more preferably 300 mg of sodium butyrate, sodium propionate or sodium acetate, preferably sodium butyrate.

[0066] In one embodiment, the composition is a unit dosage form comprising about 600 mg of sodium butyrate. In one embodiment, the composition is a unit dosage form comprising 600 mg of sodium butyrate.

[0067] L-Threonine is included as a component of the compositions described herein. Threonine is an essential amino acid used in protein biosynthesis, which can produce collagen and contribute to immune health.

[0068] Additional amino acids may also be included in the compositions described herein. In some embodiments, L-threonine may be replaced in the composition by another amino acid selected from the group consisting of L-tryptophan, L-glutamine, L-arginine, and L-methionine. Without wishing to be bound by theory, the inventors believe that each amino acid selected from L-threonine, L-tryptophan, L-glutamine, L-arginine, and L-methionine, and any combination thereof in the composition, may reduce inflammation and / or promote mucosal healing. The preferred amino acid is L-threonine. Together with bromelain, these amino acids also act to reduce pro-inflammatory cytokines, which may promote the restoration of the microbiome and reduce inflammation.

[0069] In one embodiment, the composition is a unit dosage form comprising from about 67.5 to about 2700 mg, preferably from about 112.5 to about 2250 mg, from about 250 to about 2000 mg, from about 500 to about 1900 mg, from about 900 to about 1800 mg, more preferably about 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. In one embodiment, the composition comprises about 1800 mg, preferably about 900 mg, more preferably about 450 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof.

[0070] In one embodiment, the composition is a unit dosage form comprising from about 67.5 to about 2700 mg, preferably from about 112.5 to about 2250 mg, from about 250 to about 2000 mg, from about 500 to about 1900 mg, from about 900 to about 1800 mg, more preferably about 1800 mg of L-threonine. In one embodiment, the composition comprises about 1800 mg, preferably about 900 mg, more preferably about 450 mg of L-threonine.

[0071] In one embodiment, the composition is a unit dosage form comprising 67.5 to 2700 mg, preferably 112.5 to 2250 mg, 250 to 2000 mg, 500 to 1900 mg, 900 to 1800 mg, more preferably 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. In one embodiment, the composition comprises 1800 mg, preferably 900 mg, more preferably 450 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof.

[0072] In one embodiment, the composition is a unit dosage form containing 67.5 to 2700 mg, preferably 112.5 to 2250 mg, 250 to 2000 mg, 500 to 1900 mg, 900 to 1800 mg, more preferably 1800 mg of L-threonine. In one embodiment, the composition contains 1800 mg, preferably 900 mg, more preferably 450 mg of L-threonine.

[0073] Vitamin D is included as a component of the composition described herein. Vitamin D has an anti-inflammatory effect and helps to restore the microbiome to homeostasis, similar to tightening gap junctions between L cells in the gastrointestinal tract and reducing intestinal permeability.

[0074] The term "vitamin D" refers to a group of fat-soluble secosteroids including vitamin D2 (ergocalciferol) and 1,25-dihydroxyvitamin D3 (cholecalciferol), which is vitamin D3. Vitamin D without a subscript refers to D2, D3, or both. Both forms of vitamin D are collectively known as calciferol. The active metabolite of vitamin D, known as alfacalcidol, does not require the second hydroxylation step in the kidney.

[0075] In one embodiment, the composition is a unit dosage form containing about 4 to about 150 μg, about 6.5 to about 140 μg, about 13 to about 130 μg, about 25 to about 120 μg, about 35 to about 110 μg, about 50 to about 105 μg, preferably about 100 μg, more preferably about 50 μg, still more preferably about 25 μg, still more preferably about 12.5 μg of vitamin D.

[0076] In one embodiment, the composition is a unit dosage form containing about 4 to 150 μg, 6.5 to 140 μg, 13 to 130 μg, 25 to 120 μg, 35 to 110 μg, 50 to 105 μg, preferably 100 μg, more preferably 50 μg, still more preferably 25 μg, still more preferably 12.5 μg of vitamin D.

[0077] In one embodiment, the vitamin D is cholecalciferol (vitamin D3). Menthol can be included as a component of the compositions described herein, and can be an artificially produced or organic compound obtained from mint oils such as peppermint. Both peppermint leaves and peppermint essential oil have been used for health purposes.

[0078] In one embodiment, the composition is a unit dosage form containing about 3 to about 150 mg, preferably about 6 to about 145 mg, about 12 to about 140 mg, about 24 to about 135 mg, 48 to about 130 mg, about 60 to about 125 mg, about 80 to about 120 mg, about 90 to about 110 mg, about 100 mg, more preferably about 96 mg of menthol. In one embodiment, the composition contains about 100, preferably about 96, more preferably about 50 mg, still more preferably about 48 mg of menthol.

[0079] In one embodiment, the composition is a unit dosage form containing 3 to 200 mg, preferably 6 to 150 mg, 12 to 145 mg, 24 to 140 mg, 48 to 135 mg, 60 to 130 mg, 80 to 120 mg, 90 to 110 mg, 100 mg, more preferably 96 mg of menthol. In one embodiment, the composition contains 100, preferably 96, more preferably 50 mg, still more preferably 48 mg of menthol.

[0080] In one embodiment, the menthol is L-menthol. In one embodiment, the menthol is D-menthol. The compositions disclosed herein may be presented in unit dosage form and can be prepared by any of the methods well known in the fields of pharmacy and pharmaceutical formulations. The term "unit dosage form" means a single dose that is available in a single container or package, and a single dose in which all the active agents and inactive ingredients are combined and administered to a patient in a suitable system that contains the entire "unit dosage form". A "unit dosage form" does not include the mixing of components from two or more containers or packages.

[0081] Typical examples of unit dosage forms include pills, tablets, minitablets, capsules, granules, particles, and / or microparticles (i.e., oral dosage forms) for oral administration of the compositions described herein, or transdermal patches (i.e., topical dosage forms) containing unit doses of the compositions described herein. These examples of unit dosage forms are not intended to be limiting in any way and are merely intended to represent typical examples in the pharmacy art of unit dosage forms.

[0082] Thus, in another aspect, the present invention relates to unit dosage forms for the treatment or prevention of IBD or IBS, comprising from about 1 to about 10 g of the composition of the present invention. In one embodiment, the unit dosage form is an oral dosage form.

[0083] In one embodiment, the oral dosage form comprises from about 1 to about 8 g, from about 1 to about 7 g, from about 1 to about 6 g, from about 1 to about 5 g, from about 1 to about 4 g, from about 1 to 3 g of the composition. In one embodiment, the oral dosage form comprises 1 - 8 g, 1 - 7 g, 1 - 6 g, 1 - 5 g, 1 - 4 g, 1 - 3 g of the composition.

[0084] In one embodiment, the oral dosage form comprises a composition from about 4 g to about 4.5 g, preferably about 4.22 g of the composition. In one embodiment, the oral dosage form comprises a composition of 4 g - 4.5 g, preferably 4.22 g of the composition.

[0085] In one embodiment, the oral dosage form comprises a composition from about 2 g to about 2.5 g, preferably about 2.21 g of the composition. In one embodiment, the oral dosage form comprises a composition of 2 g - 2.5 g, preferably 2.21 g of the composition.

[0086] In one embodiment, the oral dosage form consists essentially of from about 1 to about 8 g, from about 1 to about 7 g, from about 1 to about 6 g, from about 1 to about 5 g, from about 1 to about 4 g, from about 1 to 3 g of the composition. In one embodiment, the oral dosage form consists essentially of 1 - 8 g, 1 - 7 g, 1 - 6 g, 1 - 5 g, 1 - 4 g, 1 - 3 g of the composition.

[0087] In one embodiment, the oral dosage form consists essentially of a composition of about 1 to about 10 g, about 1.25 to about 8 g, preferably about 1.5 to about 6 g. In one embodiment, the oral dosage form consists essentially of a composition of 1 to 10 g, 1 to 8 g, preferably 1.5 to 6 g.

[0088] In one embodiment, the oral dosage form consists essentially of a composition of about 4 g to about 4.5 g, preferably about 4.22 g. In one embodiment, the oral dosage form consists essentially of a composition of 4 to 4.5 g, preferably 4.22 g.

[0089] In one embodiment, the oral dosage form consists essentially of a composition of about 2 g to about 2.5 g, preferably about 2.21 g. In one embodiment, the oral dosage form consists essentially of a composition of 2 to 2.5 g, preferably 2.21 g.

[0090] In one embodiment, the oral dosage form contains 50 - 2500 mg of bromelain, preferably 720 mg of bromelain, 45 - 1800 mg of sodium butyrate, preferably 300 mg of sodium butyrate, 67.5 - 2700 mg of L-threonine, preferably 450 mg of L-threonine, 4 - 150 μg of vitamin D, preferably 25 μg of vitamin D, and optionally 3 - 150 mg of menthol, preferably 48 mg of menthol.

[0091] In one embodiment, the oral dosage form contains about 460 mg to about 500 mg of bromelain, preferably about 480 mg of bromelain, about 550 mg to about 650 mg of sodium butyrate, preferably about 600 mg of sodium butyrate, about 850 mg to about 950 mg of L-threonine, preferably about 900 mg of L-threonine, about 10 μg to about 15 μg of vitamin D, preferably about 12.5 μg of vitamin D, and optionally about 65 mg to about 75 mg of menthol, preferably about 70 mg of menthol.

[0092] In one embodiment, the oral dosage form comprises 460 mg to 500 mg of bromelain, preferably 480 mg of bromelain, 550 mg to 650 mg of sodium butyrate, preferably 600 mg of sodium butyrate, 850 mg to 950 mg of L-threonine, preferably 900 mg of L-threonine, 10 μg to 15 μg of vitamin D, preferably 12.5 μg of vitamin D, and optionally 65 mg to 75 mg of menthol, preferably about 70 mg of menthol.

[0093] In one embodiment, the oral dosage form consists essentially of about 460 mg to about 500 mg of bromelain, preferably about 480 mg of bromelain, about 550 mg to about 650 mg of sodium butyrate, preferably about 600 mg of sodium butyrate, about 850 mg to about 950 mg of L-threonine, preferably about 900 mg of L-threonine, about 10 μg to about 15 μg of vitamin D, preferably about 12.5 μg of vitamin D, and optionally about 65 mg to about 75 mg of menthol, preferably about 70 mg of menthol.

[0094] In one embodiment, the oral dosage form consists essentially of 460 mg to 500 mg of bromelain, preferably 480 mg of bromelain, 550 mg to 650 mg of sodium butyrate, preferably 600 mg of sodium butyrate, 850 mg to 950 mg of L-threonine, preferably 900 mg of L-threonine, 10 μg to 15 μg of vitamin D, preferably 12.5 μg of vitamin D, and optionally 65 mg to 75 mg of menthol, preferably about 70 mg of menthol.

[0095] In one embodiment, the oral dosage form comprises about 700 mg to about 740 mg of bromelain, preferably about 720 mg of bromelain, about 550 mg to about 650 mg of sodium butyrate, preferably about 600 mg of sodium butyrate, about 850 mg to about 950 mg of L-threonine, preferably about 900 mg of L-threonine, about 10 μg to about 15 μg of vitamin D, preferably about 12.5 μg of vitamin D, and optionally about 100 mg to about 110 mg of menthol, preferably about 105 mg of menthol.

[0096] In one embodiment, the oral dosage form comprises 700 mg to 740 mg of bromelain, preferably 720 mg of bromelain, 550 mg to 650 mg of sodium butyrate, preferably 600 mg of sodium butyrate, 850 mg to 950 mg of L-threonine, preferably 900 mg of L-threonine, 10 μg to 15 μg of vitamin D, preferably 12.5 μg of vitamin D, and optionally 100 mg to 110 mg of menthol, preferably 105 mg of menthol.

[0097] In one embodiment, the oral dosage form consists essentially of about 700 mg to about 740 mg of bromelain, preferably about 720 mg of bromelain, about 550 mg to about 650 mg of sodium butyrate, preferably about 600 mg of sodium butyrate, about 850 mg to about 950 mg of L-threonine, preferably about 900 mg of L-threonine, about 10 μg to about 15 μg of vitamin D, preferably about 12.5 μg of vitamin D, and optionally about 100 mg to about 110 mg of menthol, preferably about 105 mg of menthol.

[0098] In one embodiment, the oral dosage form consists essentially of 700 mg to 740 mg of bromelain, preferably 720 mg of bromelain, 550 mg to 650 mg of sodium butyrate, preferably 600 mg of sodium butyrate, 850 mg to 950 mg of L-threonine, preferably 900 mg of L-threonine, 10 μg to 15 μg of vitamin D, preferably 12.5 μg of vitamin D, and optionally 100 mg to 110 mg of menthol, preferably 105 mg of menthol.

[0099] In one embodiment, the oral dosage form is formulated to administer 4.22 g of the dosage form twice daily per subject. In this embodiment, the oral dosage form provides a total daily dose of 140 mg of menthol, 960 mg of bromelain, 1800 mg of threonine, 1200 mg of sodium butyrate, and 25 μg of colecalciferol.

[0100] In another embodiment, the oral dosage form is formulated to administer 2.21 g twice daily per subject. In this embodiment, the oral dosage form provides a total daily amount of 70 mg of menthol, 480 mg of bromelain, 900 mg of threonine, 600 mg of sodium butyrate, and 12.5 μg of cholecalciferol.

[0101] In one embodiment, the oral dosage form comprises at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing 1440 mg of bromelain targeting the small intestine and 288 mg of menthol, and at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing a colon-targeted dose of 1200 mg of butyric acid (sodium salt), 1800 mg of L-threonine, and 0.025 mg of vitamin D3.

[0102] In one embodiment, the oral dosage form comprises at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing approximately 480 mg of bromelain targeting the small intestine and approximately 70 mg of menthol, and at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing a colon-targeted dose of approximately 600 mg of butyric acid (sodium salt), approximately 900 mg of L-threonine, and approximately 0.0125 mg of vitamin D3.

[0103] In one embodiment, the oral dosage form comprises at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing 480 mg of bromelain targeting the small intestine and 70 mg of menthol, and at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing a dosage targeting the colon of 600 mg of butyric acid (sodium salt), 900 mg of L-threonine, and 0.0125 mg of vitamin D3.

[0104] In one embodiment, the oral dosage form consists essentially of at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing approximately 480 mg of bromelain targeting the small intestine and approximately 70 mg of menthol, and at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing a dosage targeting the colon of approximately 600 mg of butyric acid (sodium salt), approximately 900 mg of L-threonine, and approximately 0.0125 mg of vitamin D3.

[0105] In one embodiment, the oral dosage form consists essentially of at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing 480 mg of bromelain targeting the small intestine and 70 mg of menthol, and at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing a dosage targeting the colon of 600 mg of butyric acid (sodium salt), 900 mg of L-threonine, and 0.0125 mg of vitamin D3.

[0106] In one embodiment, the oral dosage form comprises at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or microparticle, or any combination thereof, containing about 960 mg of bromelain targeting the small intestine and about 140 mg of menthol, and at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or microparticle, or any combination thereof, containing a dosage targeting the colon of about 1200 mg of butyric acid (sodium salt), about 1800 mg of L-threonine, and about 0.025 mg of vitamin D3.

[0107] In one embodiment, the oral dosage form comprises at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or microparticle, or any combination thereof, containing 960 mg of bromelain targeting the small intestine and 140 mg of menthol, and at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or microparticle, or any combination thereof, containing a dosage targeting the colon of 1200 mg of butyric acid (sodium salt), 1800 mg of L-threonine, and 0.025 mg of vitamin D3.

[0108] In one embodiment, the oral dosage form consists essentially of at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or microparticle, or any combination thereof, containing about 960 mg of bromelain targeting the small intestine and about 140 mg of menthol, and at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or microparticle, or any combination thereof, containing a dosage targeting the colon of about 1200 mg of butyric acid (sodium salt), about 1800 mg of L-threonine, and about 0.025 mg of vitamin D3.

[0109] In one embodiment, the oral dosage form consists essentially of at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing 960 mg of bromelain targeting the small intestine and 140 mg of menthol, and at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing a colon-targeted dosage of 1200 mg of butyric acid (sodium salt), 1800 mg of L-threonine, and 0.025 mg of vitamin D3.

[0110] In one embodiment, the oral dosage form provides the concentration of components per enterocyte and bacterium contained within the gut microbiota, as shown in Table 1. In one embodiment, the oral dosage form includes one or more pills, tablets, minitablets, capsules, granules, particles, or microparticles, or any combination thereof.

[0111] In one embodiment, the oral dosage form includes, or consists essentially of, a composition comprising two different types of pills, tablets, minitablets, capsules, granules, particles, or microparticles, wherein each different type is formulated to release its contents in different parts of the gastrointestinal tract. In one embodiment, the different parts of the gastrointestinal tract are the duodenum (also referred to herein as the "upper intestine") and the colon (also referred to herein as the "lower intestine").

[0112] The Applicant has surprisingly found that the effectiveness of the composition of the present invention increases when the components of the composition are formulated into two different types of pills, tablets, minitablets, capsules, granules, particles, or microparticles (referred to herein as Type A and Type B). Herein, each type is formulated to deliver the components contained therein to a specific part of the intestine.

[0113] In one example, the components of the composition work together to reduce the pain of inflammation in the upper intestine (type A) and restore the function of tight junctions and mucin production in the lower intestine (type B), acting simultaneously throughout the upper and lower intestines (type A), thereby reducing the symptoms of IBD and / or IBS. The unexpected benefit of this approach devised by the applicant can be clearly seen in the endoscopic results obtained in mice (Examples 11 and 12) and the improvement in its histopathology (Example 14).

[0114] In one embodiment, the two different types of pills, tablets, mini-tabs, capsules, granules, particles, or microparticles are type A and type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles, where the type A and type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles each contain at least one component, preferably at least two components, of the composition described herein.

[0115] In one embodiment, the type A and type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles each contain at least one different component, preferably at least two different components of the composition, from each other.

[0116] In one embodiment, the type A pills, tablets, mini-tabs, capsules, granules, particles, or microparticles contain bromelain or consist essentially of bromelain. In one embodiment, the type A pills, tablets, mini-tabs, capsules, granules, particles, or microparticles contain menthol.

[0117] In one embodiment, the type A pills, tablets, mini-tabs, capsules, granules, particles, or microparticles contain an amount of bromelain that provides a dosage of about 10 - 40 fg / bacteria, about 15 - 35 fg / bacteria, about 20 - 30 fg / bacteria, preferably about 28 fg / bacteria, more preferably about 25 fg / bacteria, and even more preferably about 20 fg / bacteria in the ileum / small intestine of the subject. In one embodiment, the dosage is an effective amount.

[0118] In one embodiment, a type A pill, tablet, minitablet, capsule, granule, particle, or microparticle contains an amount of bromelain that provides a dosage of 10 - 40 fg / bacterium, 15 - 35 fg / bacterium, 20 - 30 fg / bacterium, preferably 28 fg / bacterium, more preferably 25 fg / bacterium, even more preferably 20 fg / bacterium of bromelain in the ileum / small intestine of a subject. In one embodiment, the dosage is an effective amount.

[0119] In one embodiment, the effective amount is about 10 - 40 fg / bacterium, about 15 - 35 fg / bacterium, about 20 - 30 fg / bacterium, preferably about 28 fg / bacterium, more preferably about 25 fg / bacterium, even more preferably about 20 fg / bromelain in the ileum / small intestine of a subject.

[0120] In one embodiment, the effective amount is 10 - 40 fg / bacterium, 15 - 35 fg / bacterium, 20 - 30 fg / bacterium, preferably 28 fg / bacterium, more preferably 25 fg / bacterium, even more preferably 20 fg / bromelain in the ileum / small intestine of a subject.

[0121] In one embodiment, a type A pill, tablet, minitablet, granule, capsule, particle, or microparticle contains an amount of bromelain that provides a dosage of about 25 - 125 pg / ileal enterocyte (pg / ie), about 50 - 100 pg / ie, about 60 - about 90 pg / ie, about 70 - 80 pg / ie, preferably about 75 pg / ie of bromelain in the ileum / small intestine of a subject. In one embodiment, the dosage is an effective amount.

[0122] In one embodiment, a type A pill, tablet, minitablet, granule, capsule, particle, or microparticle contains an amount of bromelain that provides a dosage of 25 - 125 pg / ileal enterocyte (pg / ie), 50 - 100 pg / ie, 60 - 90 pg / ie, 70 - 80 pg / ie, preferably 75 pg / ie of bromelain in the ileum / small intestine of a subject. In one embodiment, the dosage is an effective amount.

[0123] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain, or consist essentially of, about 400 mg to 1100 mg of bromelain, about 600 mg to 1000 mg, about 700 mg to 900 mg, about 750 mg to 850 mg, preferably about 800 mg of bromelain.

[0124] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain, or consist essentially of, 400 mg to 1100 mg of bromelain, 600 mg to 1000 mg, 700 mg to 900 mg, 750 mg to 850 mg, preferably 800 mg of bromelain.

[0125] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain, or consist essentially of, about 460 mg to about 500 mg of bromelain, about 470 mg to about 490 mg, about 475 mg to about 485 mg, preferably about 480 mg of bromelain.

[0126] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain, or consist essentially of, 460 mg to 500 mg of bromelain, 470 mg to 490 mg, 475 mg to 485 mg, preferably 480 mg of bromelain.

[0127] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain, or consist essentially of, about 700 mg to about 740 mg of bromelain, about 710 mg to about 730 mg, about 715 mg to about 725 mg, preferably about 720 mg of bromelain.

[0128] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain, or consist essentially of, 700 mg to 740 mg of bromelain, 710 mg to 730 mg, 715 mg to 725 mg, preferably 720 mg of bromelain.

[0129] In one embodiment, the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles contain menthol. In one embodiment, the type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles contain L-threonine, L-tryptophan, L-glutamine, L-arginine, L-methionine or any combination thereof, sodium butyrate, and vitamin D.

[0130] In one embodiment, the type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles contain L-threonine, sodium butyrate, and vitamin D. In one embodiment, the type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles consist essentially of L-threonine, L-tryptophan, L-glutamine, L-arginine, L-methionine or any combination thereof, sodium butyrate, and vitamin D.

[0131] In one embodiment, the type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles consist essentially of L-threonine, sodium butyrate, and vitamin D. In one embodiment, vitamin D is vitamin D3.

[0132] In one embodiment, vitamin D is vitamin D3. In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle comprises, or consists essentially of, from about 67.5 to about 2700 mg, preferably from about 112.5 to about 2250 mg, from about 250 to about 2000 mg, from about 500 to about 1900 mg, from about 900 to about 1800 mg, more preferably about 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle comprises, or consists essentially of, about 1800 mg, preferably about 900 mg, more preferably about 450 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof.

[0133] In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle comprises, or consists essentially of, from about 67.5 to about 2700 mg, preferably from about 112.5 to about 2250 mg, from about 250 to about 2000 mg, from about 500 to about 1900 mg, from about 900 to about 1800 mg, more preferably about 1800 mg of L-threonine. In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle comprises, or consists essentially of, about 1800 mg, preferably about 900 mg, more preferably about 450 mg of L-threonine.

[0134] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle contains, or consists essentially of, 67.5 to 2700 mg, preferably 112.5 to 2250 mg, 250 to 2000 mg, 500 to 1900 mg, 900 to 1800 mg, more preferably 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle contains, or consists essentially of, 1800 mg, preferably 900 mg, more preferably 450 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof.

[0135] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle contains, or consists essentially of, 67.5 to 2700 mg, preferably 112.5 to 2250 mg, 250 to 2000 mg, 500 to 1900 mg, 900 to 1800 mg, more preferably 1800 mg of L-threonine. In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle contains, or consists essentially of, 1800 mg, preferably 900 mg, more preferably 450 mg of L-threonine.

[0136] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle contains, or consists essentially of, about 45 to 1800 mg of sodium butyrate, preferably about 75 to about 1650 mg, about 250 to about 1500 mg, about 500 to about 1400 mg, about 750 to about 1500 mg, about 1000 to about 1400 mg, about 1100 to about 1300 mg, more preferably about 1200 mg of sodium butyrate. In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle contains about 1200 mg, preferably about 600 mg, more preferably about 300 mg of sodium butyrate.

[0137] In one embodiment, the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles comprise, or consist essentially of, 45 to 1800 mg of sodium butyrate, preferably 75 to 1650 mg, 250 to 1500, 500 to 1400, 750 to 1500, 1000 to 1400, 1100 to 1300 mg, more preferably 1200 mg of sodium butyrate. In one embodiment, the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles comprise, or consist essentially of, 1200 mg, preferably 600 mg, more preferably 300 mg of sodium butyrate.

[0138] In one embodiment, the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles comprise, or consist essentially of, about 4 to about 150 μg, about 6.5 to about 140 μg, about 13 to about 130 μg, about 25 to about 120 μg, about 35 to about 110 μg, about 50 to about 105 μg, preferably about 100 μg of vitamin D.

[0139] In one embodiment, the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles comprise, or consist essentially of, 4 to 150 μg, 6.5 to 140 μg, 13 to 130 μg, 25 to 120 μg, 35 to 110 μg, 50 to 105 μg, preferably 100 μg of vitamin D.

[0140] In one embodiment, the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles comprise, or consist essentially of, about 1 to about 50 μg of vitamin D, preferably about 10 to about 40 μg, about 20 to about 30 μg, more preferably about 25 μg of vitamin D.

[0141] In one embodiment, the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles comprise, or consist essentially of, 1 to 50 μg of vitamin D, preferably 10 to 40 μg, 20 to 30 μg, more preferably 25 μg of vitamin D.

[0142] In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle contains, or consists essentially of, about 5 to about 30 μg of vitamin D, preferably about 10 to about 25 μg, about 12 to about 13 μg, more preferably about 12.5 μg of vitamin D.

[0143] In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle contains, or consists essentially of, 5 to 30 μg of vitamin D, preferably 10 to 25 μg, 12 to 13 μg, more preferably 12.5 μg of vitamin D.

[0144] In one embodiment, the vitamin D is cholecalciferol (vitamin D3). In one embodiment, the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle can include subtypes of pills, tablets, minitablets, granules, capsules, particles, or microparticles such as type B1 and B2.

[0145] In one embodiment, the type B1 pill, tablet, minitablet, granule, capsule, particle, or microparticle contains the L-threonine and sodium butyrate components contained in the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle, and the type B2 pill, tablet, minitablet, granule, capsule, particle, or microparticle contains the vitamin D component contained in the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle.

[0146] An additional embodiment of the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle that includes the subtypes B1 and B2 pills, tablets, minitablets, granules, capsules, particles, or microparticles is specifically contemplated herein as an option for all of the specific embodiments described herein and is presented as the type B pill, tablet, minitablet, granule, capsule, particle, or microparticle.

[0147] In some embodiments, the type A, and / or type B pills, tablets, minitabs, capsules, granules, particles, or microparticles can be modified for delayed release of one or more components, including sustained release and delayed release. One of ordinary skill in the art can design an oral dosage form according to any of the sustained release dosage forms known and described in the art (see, e.g., the disclosure of U.S. Patent No. 7,108,865, which is incorporated herein by reference).

[0148] In some embodiments, the pills, tablets, minitabs, capsules, granules, particles, or microparticles include a water-soluble, water-insoluble, or enteric coating. Enteric and other pH-sensitive polymers that are relatively insoluble and impermeable at gastric pH but more soluble and permeable at the pH of the small intestine and colon include polyacrylamide; acid phthalates of carbohydrates, amylose acetate phthalate, cellulose acetate phthalate, other cellulose phthalate esters, cellulose phthalate ethers, hydroxypropylcellulose phthalate, hydroxypropyl ethylcellulose phthalate, hydroxypropyl methylcellulose phthalate, methylcellulose phthalate, polyvinyl acetate phthalate, hydrogen phthalate polyvinyl acetate, sodium cellulose acetate phthalate, starch phthalate, styrene-maleic acid dibutyl phthalate copolymer, styrene-maleic acid polyvinyl acetate phthalate copolymer, and other phthalic acid derivatives; styrene, and maleic acid copolymers; polyacrylic acid, and acrylic acid ester copolymers, polymethacrylic acid, and its esters, polyacrylic acid methacrylic acid copolymers, and other polyacrylic acid derivatives; shellac; and vinyl acetate, and crotonic acid copolymers. Preferred pH-sensitive polymers include shellac; phthalic acid derivatives, particularly cellulose acetate phthalate, polyvinyl acetate phthalate, and hydroxypropyl methylcellulose phthalate; polyacrylic acid derivatives, particularly polymethyl methacrylate mixed with acrylic acid, and acrylic acid ester copolymers; and vinyl acetate and crotonic acid copolymers.

[0149] In one embodiment, the type A pills, tablets, mini-tabs, capsules, granules, particles, or microparticles include an enteric coating. In one embodiment, the enteric coating forms an outer layer on the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles. In some embodiments, the enteric coating is contained within an inner layer within the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles.

[0150] In one embodiment, the type A pills, tablets, mini-tabs, capsules, granules, particles, or microparticles are formulated with an excipient that releases the components contained therein in the upper part of the small intestine. In one embodiment, the excipient includes one or more diluents, binders, lubricants, and disintegrants.

[0151] In one embodiment, the diluent is microcrystalline cellulose, dicalcium phosphate dihydrate, or a combination thereof. In one embodiment, the disintegrant is crospovidone, croscarmellose sodium, or a combination thereof. In one embodiment, the polymer is hydroxypropyl methylcellulose, hydroxypropyl methylcellulose phthalate HP55, or Eudragit L100-55. In one embodiment, the lubricant is magnesium stearate, talc, or a combination thereof.

[0152] In one embodiment, the type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles are formulated with an excipient that releases the components contained therein in the lower part of the small intestine. In one embodiment, the excipient includes one or more diluents, binders, lubricants, and disintegrants.

[0153] In one embodiment, the diluent is microcrystalline cellulose, dicalcium phosphate dihydrate, or a combination thereof. In one embodiment, the disintegrant is crospovidone, croscarmellose sodium, or a combination thereof. In one embodiment, the polymer is hydroxypropylmethylcellulose, ethylcellulose, Eudragit S100, shellac, or Eudraguard biotec. In one embodiment, the lubricant is magnesium stearate, talc, or a combination thereof.

[0154] In one embodiment, the type B pills, tablets, mini-tabs, capsules, granules, particles, or microparticles include an enteric coating. In one embodiment, the enteric coating forms an outer layer on the type B pills, tablets, mini-tabs, granules, capsules, particles, or microparticles. In some embodiments, the enteric coating is contained within an inner layer within the type B pills, tablets, mini-tabs, granules, capsules, particles, or microparticles.

[0155] It is important that the enteric coating is designed to dissolve at the optimal location identified by the inventors for releasing the components of the composition contained therein from the pills, tablets, mini-tabs, capsules, granules, particles, or microparticles into the gastrointestinal (GI) tract of the subject in the region of the GI tract.

[0156] In one embodiment, the enteric coating dissolves from the type A pills, tablets, mini-tabs, granules, capsules, particles, or microparticles at about pH 6.0. In one embodiment, the pH is less than 6.0.

[0157] In one embodiment, the enteric coating dissolves from the type A microparticles at a predetermined pH. In one embodiment, the predetermined pH is about pH 6.0. In one embodiment, the predetermined pH is greater than 6.0.

[0158] In one embodiment, the enteric coating is formulated to release bromelain and menthol from type A pills, tablets, minitabs, granules, capsules, particles, or microparticles in the ileum / small intestine of a subject.

[0159] In one embodiment, the enteric coating dissolves from type A pills, tablets, minitabs, granules, capsules, particles, or microparticles while releasing bromelain and menthol within the ileum / small intestine of a subject.

[0160] In one embodiment, the enteric coating dissolves from type B pills, tablets, minitabs, granules, capsules, particles, or microparticles at neutral pH. In one embodiment, the pH is greater than 6.5. In one embodiment, the pH is about 7.0.

[0161] In one embodiment, the enteric coating dissolves from type B microparticles at a predetermined pH. In one embodiment, the predetermined pH is neutral pH. In one embodiment, the pH is greater than 6.5. In one embodiment, the pH is about 7.0.

[0162] In one embodiment, the enteric coating is formulated to release L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, sodium butyrate, and vitamin D from pills, tablets, minitabs, capsules, granules, particles, or microparticles near and / or within the colon of a subject.

[0163] In one embodiment, the enteric coating is formulated to release L-threonine, sodium butyrate, and vitamin D from pills, tablets, minitabs, capsules, granules, particles, or microparticles near and / or within the colon of a subject.

[0164] In one embodiment, the enteric coating dissolves from type B pills, tablets, minitabs, capsules, granules, particles, or microparticles while releasing L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, sodium butyrate, and vitamin D in the vicinity of and / or within the colon of the subject.

[0165] In one embodiment, the enteric coating dissolves from type B pills, tablets, minitabs, capsules, granules, particles, or microparticles while releasing L-threonine, sodium butyrate, and vitamin D in the vicinity of and / or within the colon of the subject.

[0166] In some embodiments, the enteric coating is designed to deliver the components of the composition to a predetermined region of the GI tract. In some embodiments, the enteric coating delivers the components of the composition to a predetermined region of the GI tract.

[0167] In some embodiments, the enteric coating is designed to dissolve from pills, tablets, minitabs, capsules, granules, particles, or microparticles at a predetermined pH within the GI tract of the subject and release the components of the composition contained therein. In some embodiments, the enteric coating dissolves at a predetermined pH within the GI tract of the subject.

[0168] In some embodiments, type A and / or type B pills, tablets, minitabs, capsules, granules, particles or microparticles are formulated to deliver an effective amount of at least one, preferably at least two, more preferably at least three, even more preferably at least four, and even more preferably five components of the composition to bacteria within the GI tract of the subject.

[0169] In some embodiments, different types of pills, tablets, minitabs, capsules, granules, particles, or microparticles deliver an effective amount of at least 1, preferably at least 2, more preferably at least 3, even more preferably at least 4, and even more preferably 5 components of the composition to bacteria in the subject's GI tract when the enteric coating dissolves.

[0170] In some embodiments, delivery of the effective amount includes delivery to the ileum / small intestine of the subject. In some embodiments, the effective amount is bromelain at 10 - 40 fg / bacterium, 15 - 35 fg / bacterium, 20 - 30 fg / bacterium, preferably 28 fg / bacterium, more preferably 25 fg / bacterium in the ileum / small intestine of the subject.

[0171] In one embodiment, the effective amount is bromelain at 10 - 40 fg / bacterium, 15 - 35 fg / bacterium, 20 - 30 fg / bacterium, preferably 28 fg / bacterium, more preferably 25 fg / bacterium, and even more preferably 20 fg / bacterium in the ileum / small intestine of the subject.

[0172] In some embodiments, delivery of the effective amount includes delivery to the colon of the subject. In some embodiments, the effective amount is bromelain at 25 - 125 pg / ileal enterocyte (pg / ie), 50 - 100 pg / ie, 60 - 90 pg / ie, 70 - 80 pg / ie, preferably 75 pg / ie in the ileum / small intestine of the subject.

[0173] In one embodiment, at least a first one of the pills, tablets, minitabs, capsules, granules, particles or microparticles contains an amount of bromelain sufficient to provide an effective amount of 25 - 125 pg / ileal enterocyte (pg / ie), 50 - 100 pg / ie, 60 - 90 pg / ie, 70 - 80 pg / ie, preferably 75 pg / ie in the ileum / small intestine of the subject.

[0174] As will be understood by those skilled in the art, several different types of enteric coatings that dissolve at a predetermined location within the GI tract can be prepared by those skilled in pharmaceutical technology by following the guidelines for enteric coatings according to conventional formulation practices, such as (Allen) and (Encyclopaedia of Pharmaceutical Technology), which are incorporated herein by reference. The preparation of such enteric coatings is considered to be within the scope of the art. For example, the compositions described herein are formulated to facilitate the targeted delivery of bromelain to the colon. Oral drug delivery systems targeting the colon are well understood in the art (Amidon, 2016).

[0175] This application provides a substantial disclosure regarding where the components of the compositions of the invention are released in the GI tract of the subject. Thus, those skilled in the art, in combination with common general knowledge, can formulate an appropriate enteric coating and release bromelain and menthol components to the ileum / small intestine of the subject according to the instructions provided in the disclosure of this application.

[0176] For example, the components of the composition can be dispersed in any pharmaceutically acceptable carrier, which can be an immediate-release or sustained-release carrier. Carriers may include microcrystalline cellulose (MCC), dextran, corn starch, wheat flour, talc, sucrose, mannitol, lactose, calcium carbonate, polyvinylpyrrolidone (PVP), polyethylene oxide, hydroxypropylmethylcellulose (HPMC), hydroxypropylcellulose (HPC), polyvinyl alcohol (PVA), etc., known in the art.

[0177] In one non-limiting example, the carrier can be compressed into a solid or semi-solid and then coated with a polymer coating to alter the release profile of the components. The coating can include a water-soluble polymer such as polyvinylpyrrolidone (PVP), polyvinylpolypyrrolidone (crospovidone), or polyethylene glycol, or a water-insoluble polymer including ethers of cellulose, esters of cellulose, cellulose acetate, ethyl cellulose, polyvinyl acetate, neutral copolymers based on ethyl acrylate, and methyl methacrylate, copolymers of acrylic acid having a quaternary ammonium group, and methacrylic acid esters, pH-insensitive ammoniomethacrylic acid copolymers, and mixtures thereof. The coating can also include methylcellulose, ethylcellulose, hydroxyethylcellulose, hydroxypropylmethylcellulose (HPMC), hydroxypropylcellulose (HPC), or combinations of such natural polymers. For example, the use of methylcellulose in combination with hydroxypropylmethylcellulose (HPMC) is well known.

[0178] Standard components and preparation methods for various oral dosage forms are described in "Remington: The Science and Practice of Pharmacy", which is incorporated herein by reference. Additional excipients include, but are not limited to, lubricants, disintegrants, and the like.

[0179] In one embodiment, the composition of the present invention can include in a single container a unit dosage form as described herein, preferably an oral dosage form as described herein, comprising a mixture of pills, tablets, minitablets, capsules, granules, particles, or microparticles of type A and type B as described herein.

[0180] In one embodiment, the composition of the present invention can include in a single container a unit dosage form as described herein, preferably an oral dosage form as described herein, comprising pills, tablets, minitablets, capsules, granules, particles, or microparticles of type A or type B as described herein.

[0181] In one embodiment, the container is a sachet. In one embodiment, the mixture is suspended in a carrier. In one embodiment, the carrier is an ingestible aqueous liquid selected from the group consisting of water, juice, artificial juice, artificially flavored beverages, animal milk, whey, whey-based beverages, vegetable milk, and nut milk. In one embodiment, the carrier is an ingestible gel. In one embodiment, the ingestible gel is yogurt.

[0182] In one embodiment, the oral dosage form comprises particles, or microparticles, of type A and type B, preferably microparticles. In one embodiment, the oral dosage form comprises particles, or microparticles, of type A or type B, preferably microparticles.

[0183] In one embodiment, the oral dosage comprises suspended microparticles. In one embodiment, the suspended microparticles are suspended in a liquid or a gel. In one embodiment, the oral dosage comprises dispersed microparticles. In one embodiment, the dispersed microparticles are dispersed in a powder.

[0184] In one embodiment, the oral dosage comprises a mixture of type A and type B microparticles in a single container. In one embodiment, the mixture is suspended in a carrier. In one embodiment, the carrier is an ingestible liquid selected from the group consisting of water, juice, artificial juice, artificially flavored beverages, animal milk, vegetable milk, nut milk, whey, and whey-based beverages. In one embodiment, the carrier is an ingestible gel. In one embodiment, the ingestible gel is yogurt.

[0185] In one embodiment, the subject is a mammal. In one embodiment, the subject is a human. The subject can be any subject, including male, female, adult, elderly, or pediatric subjects. In some embodiments, the subject has received a clinical diagnosis of IBD, particularly ulcerative colitis or Crohn's disease. Veterinary uses in mammals are also contemplated herein. Thus, in some embodiments, the subject is a non-human mammal. In one embodiment, the non-human mammal is a cat or a dog.

[0186] In another aspect, the invention relates to an oral dosage form consisting essentially of the composition of the invention, wherein at least two components of the composition are included in a delayed release formulation. Specifically contemplated herein as embodiments of the oral dosage form of the invention consisting essentially of the composition of the invention, wherein at least two components of the composition are included in a delayed release formulation, are all embodiments contemplated within the aspects of the invention disclosed herein, which are oral dosage forms comprising a composition including (a)-(d) and about 1 to about 6 g of the composition of the invention.

[0187] In another aspect, the invention relates to an oral dosage form that is a unit dosage form, preferably in an approximate amount, more preferably in that amount, shown as total dose / 80 kg of human (mg / day), and that contains or consists essentially of the components listed in Table 1, wherein the components are included in either of the indicated types, type A or type B, and type A and type B are formulated for targeting the small intestine and the large intestine, respectively, or for the release of the components indicated in the small intestine and the large intestine, respectively.

[0188]

Table 1

[0189] In another aspect, the invention relates to a unit dosage form that includes as separate parts type A and type B pills, tablets, minitabs, granules, capsules, particles, or microparticles, wherein The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type A contain bromelain and are formulated to release bromelain into the small intestine of a subject, where the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type A optionally contain menthol, The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type B consist essentially of sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, and vitamin D, and the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type B are formulated for release in the lower intestine of a subject.

[0190] In another aspect, the present invention relates to a unit dosage form containing the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type A and Type B as separate parts, where The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type A consist essentially of 720 mg of bromelain and are formulated to release bromelain into the small intestine of a subject, where the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type A optionally contain 144 mg of menthol, The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type B consist essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.0125 mg of vitamin D, and sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D are formulated for release into the lower intestine of a subject.

[0191] In another aspect, the present invention relates to a unit dosage form containing the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of Type A and Type B as separate parts, where The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A consist essentially of 720 mg of bromelain and are formulated to release bromelain into the small intestine of the subject, where the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A optionally contain 105 mg of menthol, The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type B consist essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.0125 mg of vitamin D, and are formulated to release sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D into the lower intestine of the subject.

[0192] In another aspect, the invention relates to a unit dosage form comprising the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A and type B as separate parts, where The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A consist essentially of 480 mg of bromelain and are formulated to release bromelain into the small intestine of the subject, where the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A optionally contain 70 mg of menthol, The pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type B consist essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.0125 mg of vitamin D, and are formulated to release sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D into the lower intestine of the subject.

[0193] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, and 0.0125 mg of vitamin D.

[0194] In another aspect, the present invention relates to a unit dosage form comprising the type A and type B pills, tablets, mini-tabs, granules, capsules, particles, or microparticles as separate portions, wherein the type A pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of 1440 mg of bromelain and is formulated to release bromelain into the small intestine of a subject, wherein the type A pill, tablet, mini-tab, granule, capsule, particle, or microparticle optionally contains 288 mg of menthol, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of 1200 mg of sodium butyrate, 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.025 mg of vitamin D, and the sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D are formulated to be released into the lower intestine of a subject.

[0195] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of 1200 mg of sodium butyrate, 1800 mg of L-threonine, and 0.025 mg of vitamin D.

[0196] In one embodiment, the type A and type B pills, tablets, mini-tabs, granules, capsules, particles, or microparticles are of similar size and characteristics to each other and may be mixed in any useful ratio depending on the stage of the condition being treated.

[0197] In one embodiment, the type A and type B pills, tablets, minitabs, granules, capsules, particles, or microparticles are mixed in a ratio of 1.94 (part A) to 2.28 (part B) (w / w in mg), providing a component ratio of 70 (menthol):480 (bromelain):900 (threonine):600 (sodium butyrate):0.0125 (colecalciferol).

[0198] In another embodiment, the type A and type B pills, tablets, minitabs, granules, capsules, particles, or microparticles are mixed in a ratio of 2.91 (part A) to 2.28 (part B) (w / w in mg), providing a component ratio of 105 (menthol):720 (bromelain):900 (threonine):600 (sodium butyrate):0.0125 (colecalciferol).

[0199] The inventors have also surprisingly found that a composition containing sodium butyrate, L-threonine, and vitamin D is surprisingly effective in treating the symptoms of IBD / IBS by promoting the healing of lesions associated with gastrointestinal dysfunction. When these components are used together as described herein, sodium butyrate, L-threonine, and vitamin D act synergistically to increase the expression of several mucin genes involved in intestinal integrity, maintain the diversity of the gut microbiota, and prevent inflammation. This effect can be seen in the results shown in Example 10. These same genes were not affected when the components were administered individually. Further evidence that the combination of sodium butyrate, threonine, and vitamin D3 is effective in the treatment of colitis in a mouse model is found in Example 11. In the mice in the described experiment, a significant 135% reduction in the lesion score was shown when measured 7 days after induction with TNBS. The results obtained from Example 12 further show that the combination of sodium butyrate, threonine, and vitamin D3 has similar efficacy to 2 mg / kg of prednisolone in the treatment of lesions induced by TNBS.

[0200] Accordingly, in another aspect, the present invention relates to a composition comprising sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine or L-methionine, and vitamin D.

[0201] In one embodiment, the composition consists essentially of sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine or L-methionine, and vitamin D.

[0202] In one embodiment, the composition is formulated to release sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D into the lower intestine of a subject.

[0203] In one embodiment, when administered, the composition administers sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D to the lower intestine of a subject.

[0204] In one embodiment, the composition comprises sodium butyrate, L-threonine, and vitamin D. In one embodiment, the composition consists essentially of sodium butyrate, L-threonine, and vitamin D.

[0205] In another aspect, the present invention relates to a composition comprising 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.0125 mg of vitamin D.

[0206] In one embodiment, the composition comprises 600 mg of sodium butyrate, 900 mg of L-threonine, and 0.0125 mg of vitamin D. In one embodiment, the composition consists essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, and 0.0125 mg of vitamin D.

[0207] In one embodiment, the composition is formulated to release sodium butyrate, L-threonine, and vitamin D into the lower intestine of the subject. In one embodiment, when administered, the composition administers sodium butyrate, L-threonine, and vitamin D to the lower intestine of the subject.

[0208] In another aspect, the present invention relates to a composition comprising 1200 mg of sodium butyrate, 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.025 mg of vitamin D.

[0209] In one embodiment, the composition comprises 1200 mg of sodium butyrate, 1800 mg of L-threonine, and 0.025 mg of vitamin D. In one embodiment, the composition consists essentially of 1200 mg of sodium butyrate, 1800 mg of L-threonine, and 0.025 mg of vitamin D.

[0210] In one embodiment, the composition is formulated to release sodium butyrate, L-threonine, and vitamin D into the lower intestine of the subject. In one embodiment, when administered, the composition administers sodium butyrate, L-threonine, and vitamin D to the lower intestine of the subject.

[0211] Specifically contemplated herein as embodiments of each aspect of the present invention with respect to compositions comprising and / or consisting essentially of sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D, are all of the embodiments described herein with respect to unit dosage forms and oral dosage forms, including their determination, formulation, and administration.

[0212] In another aspect, the present invention relates to a unit dosage form comprising a type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle, wherein the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle comprises sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, and vitamin D.

[0213] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle or microparticle consists essentially of sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, and vitamin D.

[0214] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle or microparticle is formulated to release sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, and vitamin D into the lower intestine of a subject.

[0215] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle or microparticle is formulated to release sodium butyrate, one or more of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, and vitamin D into the lower intestine of a subject.

[0216] In one embodiment, the unit dosage form, when administered, delivers the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle to the lower intestine of a subject. In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle comprises sodium butyrate, L-threonine, and vitamin D.

[0217] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of sodium butyrate, L-threonine, and vitamin D. In one embodiment, the unit dosage form is an oral dosage form as described herein.

[0218] In another aspect, the present invention relates to a unit dosage form comprising a type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle, wherein the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.0125 mg of vitamin D.

[0219] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle comprises 600 mg of sodium butyrate, 900 mg of L-threonine, and 0.0125 mg of vitamin D.

[0220] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle consists essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, and 0.0125 mg of vitamin D.

[0221] In one embodiment, the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle is formulated to release sodium butyrate, L-threonine, and vitamin D into the lower intestine of the subject.

[0222] In one embodiment, when administered, the unit dosage form administers the type B pill, tablet, mini-tab, granule, capsule, particle, or microparticle to the lower intestine of the subject. In one embodiment, the unit dosage form is an oral dosage form as described herein.

[0223] In another aspect, the present invention relates to a unit dosage form comprising a Type B pill, tablet, minitablet, granule, capsule, particle, or microparticle, wherein the Type B pill, tablet, minitablet, granule, capsule, particle, or microparticle consists essentially of 1200 mg of sodium butyrate, 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.025 mg of vitamin D.

[0224] In one embodiment, the Type B pill, tablet, minitablet, granule, capsule, particle, or microparticle comprises 1200 mg of sodium butyrate, 1800 mg of L-threonine, and 0.025 mg of vitamin D.

[0225] In one embodiment, the Type B pill, tablet, minitablet, granule, capsule, particle, or microparticle consists essentially of 1200 mg of sodium butyrate, 1800 mg of L-threonine, and 0.025 mg of vitamin D.

[0226] In one embodiment, the Type B pill, tablet, minitablet, granule, capsule, particle, or microparticle is formulated to release sodium butyrate, L-threonine, and vitamin D into the lower intestine of the subject.

[0227] In one embodiment, the unit dosage form, when administered, delivers the Type B pill, tablet, minitablet, granule, capsule, particle, or microparticle to the lower intestine of the subject. Specifically contemplated herein as embodiments of the above aspects of the present invention with respect to compositions and Type B pills, tablets, minitablets, granules, capsules, particles, or microparticles that contain and / or consist essentially of sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D, are all embodiments explicitly set forth herein with respect to unit dosage forms and oral dosage forms, including the dosages, formulations, and administrations of such forms described herein.

[0228] Method In another aspect, the present invention relates to a method for reducing the adhesion of bacteria to cells of the gastrointestinal tract of a subject, comprising contacting the bacteria with an effective amount of a composition, unit dosage form, or oral dosage form of the present invention.

[0229] In another aspect, the present invention relates to a method for reducing the invasion of bacteria into cells of the gastrointestinal tract of a subject, comprising contacting the bacteria with an effective amount of a composition, unit dosage form, or oral dosage form of the present invention.

[0230] In another aspect, the present invention relates to a method for improving gastrointestinal barrier integrity in a subject, comprising administering to the subject a therapeutically effective amount of a composition, unit dosage form, or oral dosage form.

[0231] In one aspect, the present invention relates to a method for preventing, treating, or managing IBD or IBS, comprising administering to a subject in need thereof a therapeutically effective amount of a composition, unit dosage form, or oral dosage form of the present invention.

[0232] In another aspect, the present invention relates to a method for at least partially reducing at least one symptom of IBD or IBS, comprising administering to a subject in need thereof a therapeutically effective amount of a composition, unit dosage form, or oral dosage form.

[0233] In one embodiment, the at least one symptom is selected from the group comprising diarrhea, abdominal pain, abdominal cramps, fever, fatigue, nausea, flatulence, abdominal distension, weight loss, anorexia, and bloody stools.

[0234] In another aspect, the present invention relates to a method for improving at least one parameter associated with IBD or IBS, comprising administering to a subject in need thereof a therapeutically effective amount of a composition, unit dosage form, or oral dosage form.

[0235] In one embodiment, the at least one parameter associated with IBD or IBS is selected from the group comprising the following. Circulating serum levels of inflammatory markers such as IL-6, IL-8, IL-17A, IL-17F, IL-18, IL-21, IL-22, TL1A, TGF-β, ICAM-1, IFN-gamma, MADCAM-1, and E-selectin ·Fecal markers of inflammation such as calprotectin, lactoferrin, and lysozyme ·Circulating levels of serum oxidative stress markers such as oxidized LDL, serum oxidizability, and F2-isoprostanes ·Biological pathogens in fecal samples such as bacteria belonging to Escherichia coli of the Enterobacteriaceae family of the Proteobacteria phylum, Klebsiella spp. of the Enterobacteriaceae family of the Proteobacteria phylum, Pseudomonas spp. of the Pseudomonadaceae family of the Proteobacteria phylum, Salmonella spp. of the Enterobacteriaceae family of the Proteobacteria phylum, Bilophilia wadsworthia of the Desulfovibrionaceae family of the Proteobacteria phylum, Desulfovibrio spp. of the Desulfovibrionaceae family of the Proteobacteria phylum, Desulfuromonas spp. of the Desulfuromonadaceae family of the Proteobacteria phylum, Campylobacter concisus of the Campylobacteraceae family of the Proteobacteria phylum, Campylobacter jejuni of the Campylobacteraceae family of the Proteobacteria phylum, Bacteroides fragilis of the Bacteroidaceae family of the Bacteroidetes phylum, Atopobium parvulum of the Coriobacteriaceae family of the Actinobacteria phylum, and Clostridium difficile of the Clostridiaceae family of the Firmicutes phylum

[0236] In the above aspects: In one embodiment, the subject has ulcerative colitis. In one embodiment, the subject has Crohn's disease. In one embodiment, the subject has IBS. In one embodiment, the administration is oral administration.

[0237] In one embodiment, the administration is about 20 to 90 minutes, preferably about 30 to 60 minutes, more preferably about 45 minutes before a meal. In one embodiment, the administration is 20 to 90 minutes before a meal, preferably 30 to 60 minutes, more preferably 45 minutes.

[0238] In one embodiment, the administration is 1 to 5 times a day before a meal, preferably 3 times a day before a meal, more preferably 2 times a day before a meal. In one embodiment, the administration is before breakfast and lunch. In one embodiment, the administration is before breakfast and dinner.

[0239] In one embodiment, the administration is about 20 to 90 minutes, preferably about 30 to 60 minutes, more preferably about 45 minutes after a meal. In one embodiment, the administration is 20 to 90 minutes after a meal, preferably 30 to 60 minutes, more preferably 45 minutes.

[0240] In one embodiment, the administration is 1 to 5 times a day after a meal, preferably 3 times a day after a meal, more preferably 2 times a day after a meal. In one embodiment, the administration is after breakfast and lunch. In one embodiment, the administration is after breakfast and dinner.

[0241] Subjects in need of prevention, treatment, or management of IBD or IBS are subjects diagnosed with such a condition, having a predisposition and / or risk for such a condition, or having recovered from or being in remission from such a condition. The subject may have a predisposition and / or risk for the condition due to genetic and / or environmental factors.

[0242] The amount of the compositions, unit dosage forms, or oral dosage forms described herein that are effective for the prevention, treatment, or management of the foregoing conditions will vary depending on factors specific to the subject, such as age, weight, gender, past medical history, as well as the nature and severity of the condition and the route by which the extract is administered.

[0243] Generally, the compositions or dosage forms used for the treatment of acute conditions will contain a greater amount of the bromelain composition, unit dosage, or oral dosage of the present invention than those used for the treatment of chronic conditions. Formulations of specific dosage forms for addressing either acute or chronic conditions are believed to be achievable by those skilled in the art of pharmacy in combination with the disclosure of the present invention.

[0244] In some embodiments, the subject is a non-human mammal. In one embodiment, administration to a non-human mammal comprises administration of a composition or unit dosage form described herein that comprises or consists essentially of the oral dosage form described herein.

[0245] In one embodiment, the oral dosage form administered to a non-human mammal comprises from about 1 to about 50 mg of bromelain per kg of the subject, preferably about 5 to about 25 mg, about 7 to about 20 mg, about 9 to about 18 mg, about 10 to about 15 mg, about 13 mg of bromelain.

[0246] In one embodiment, the oral dosage form administered to a non-human mammal comprises about 9 mg / kg, about 13 mg or about 18 mg of bromelain per kg of the subject. In one embodiment, the oral dosage form administered to a non-human mammal comprises from 1 to 50 mg of bromelain per kg of the subject, preferably 5 to 25 mg, 7 to 20 mg, 9 to 18 mg, 10 to 15 mg, or 13 mg of bromelain.

[0247] In one embodiment, the oral dosage form administered to a non-human mammal comprises about 9 mg / kg, about 13 mg or about 18 mg of bromelain per kg of the subject. In one embodiment, the oral dosage form is an initial dosage determined based on an acceptable conversion in the art from the human dosage contemplated herein using the industry guidance "Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutic Drugs in Adult Healthy Volunteers" issued by the Center for Drug Evaluation and Research (CDER) of the U.S. Food and Drug Administration in July 2005, which is hereby incorporated by reference in its entirety.

[0248] In one embodiment, the human dosage in mg / kg is multiplied by 1.8 to provide the dog dosage in mg / kg. In one embodiment, the non-human mammal is a household pet. In one embodiment, the household pet is selected from the group consisting of dogs, cats, rats, mice, hamsters, guinea pigs, ferrets, and gerbils. In one embodiment, the household pet is a dog. In one embodiment, the household pet is a cat.

[0249] In this specification, which refers to patent specifications, other external documents, or other information sources, this is generally for the purpose of providing a context for discussing the features of the present invention. Unless specifically indicated otherwise, such references to external documents should not be construed as an admission that such documents, or such information sources, are prior art in any jurisdiction or form part of the common general knowledge in the art.

[0250] Here, the present invention will be described in a non-limiting manner by reference to the following examples. 1. Examples General Methodology Culture of macrophage cells: Human monocytic THP-1 cells were cultured in RPMI 1640 medium modified to contain 2-mM L-glutamine, 10 mM HEPES, 1 mM sodium pyruvate, 4500 mg / L glucose, and 1500 mg / L sodium bicarbonate, with the addition of 10% FBS, 100 U / ml penicillin, 100 mg / ml streptomycin, and 5 mM β-mercaptoethanol. The cell density was 1×106 does not exceed / ml. To induce differentiation, THP-1 cells were seeded at 5 - 10×10 5 cells / cm 2 in a 24-well plate and treated with 200 nM PMA for 48 hours. After 48 hours, before starting the experiment, the macrophages were washed and grown in a medium without PMA and antibiotics for 24 hours. The cell cultures were incubated at 37°C in a humidified 5% CO2 incubator. All experiments were performed with cells within passage numbers 10 - 30.

[0251] Culture of Caco-2 cells: The human colorectal adenocarcinoma cell line Caco-2-BBe1 was grown and maintained in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% v / v fetal bovine serum (FBS), 4 mM L-glutamine, 100 U / ml penicillin, and 100 mg / ml streptomycin. All experiments were performed with Caco-2-BBe1 cells within passage number 12 after thawing. The cell cultures were incubated at 37°C in a humidified 5% CO2 incubator.

[0252] Caco-2:HT-29-MTX (75:25) co-culture: This co-culture model was used to simulate the human colonic epithelium (Hilgendorh C, 2000). Caco-2 cells (passage number 8) and HT-29-MTX cells (passage number 9) were combined at a ratio of 75:25 in DMEM containing 10% FBS. The combined cells (6.4×10 4 cells / ml) were seeded into 25 cm 2 at a total of 5 ml per flask. The co-cultures were incubated at 37°C in a 5% CO2 atmosphere while replacing the medium every 48 hours. On the 15th day after seeding, the monolayer of the co-culture was washed three times with DMEM, the medium was replaced with 5 ml of pre-warmed test solution, and incubated at room temperature for 24 hours.

[0253] Bacterial strains and growth conditions: AIEC strains HM95, HM605, HM615, and HM484 were cultured in LB medium. After growing all the bacteria overnight at 37°C in air, they were diluted 1:100 in fresh medium and then cultured for an additional 2 hours. The bacteria were Ab 600nmIt was used in the experiment in the mid-logarithmic phase confirmed by measuring 0.8 at

[0254] Transepithelial electrical resistance (TER) measurement: The integrity of the Caco-2 monolayer was determined by measuring the TER value. Tight junctions function as a barrier to paracellular diffusion, where TER indicates the level of integrity between epithelial cells (Mynott TL, 1999). Before measuring TER, the cell culture was equilibrated at room temperature for 30 minutes. TER was measured using a Millicell-ERS device (Millipore, Eschborn, Germany).

[0255] Quantification of cytokines and TNF-α: Commercially available ELISA kits were used to determine the concentrations of IL-8, IL-6, and TNF-α. At the end of the culture, the apical and basolateral media were collected, centrifuged at 400 gav for 5 minutes, and the supernatant was frozen at -80 °C until analysis. The frozen supernatant was used without repeated thawing. All ELISAs were performed according to the manufacturer's instructions.

[0256] Inflammatory model of the Caco-2 / THP-1 macrophage co-culture system: Caco-2-BBe1 cells were seeded at 5×10 5 cells / well in 24-well Transwell insert plates (4.67 cm 2 , pore size 0.4 μm). The cell culture medium was changed every 3 days until the cells were fully differentiated (TER value > 1200 / cm 2 ). In another 24-well plate, THP-1 monocyte cells were seeded at 1×10 6 cells / well, differentiated with 200 nM PMA for 48 hours, and then cultured in normal medium for 24 hours before the experiment. After replacing all media with RPMI1640, all Transwell inserts containing the differentiated Caco-2 monolayer were filled into the wells containing the differentiated THP-1 cells. In experiments to evaluate the anti-inflammatory effects of various bromelain preparations, 0.25 ml of the test sample was applied to the apical side for 24 hours, and simultaneously 50 ng / ml of LPS was added to the basolateral side of this model. After 24 hours of culture, the culture supernatant from the apical side was collected for IL-8 measurement.

[0257] Inflammatory mediator-stimulated Caco-2 polarized monolayer inflammation model with LPS, TNF-α, and IFN-β: The polarized Caco-2 model was generated as described above. When the TER baseline of cell culture reached >1000 / cm 2 of the monolayer was stimulated by adding 0.6 ml containing 50 ng / ml IFN-β, 10 μg / ml LPS, and 5 ng / ml TNF-α to the basolateral compartment. Immediately after the addition of the stimulation cocktail, 0.25 ml of GaRP ± 0 - 80 μg / ml bromelain was applied to the apical compartment. After 24 hours of culture, the culture supernatant was collected from the apical side for quantification of IL-8.

[0258] Gentamicin protection assay - Measurement of bacterial uptake, survival, and replication: THP-1 cells were infected at a multiplicity of infection (MOI) of 30 bacteria per macrophage. After 3 hours of culture, the medium was removed and fresh cell culture medium containing 100 μg / ml gentamicin was added to kill extracellular bacteria. To determine the number of intracellular bacteria, at 1 hour and 3 hours, the medium was removed and 0.5 ml of 1% Triton-100 (in deionized water) was added to each well for 5 minutes to lyse the THP-1 cells. This concentration of Triton-100 did not affect the survival rate of bacteria for at least 30 minutes. The samples were mixed, diluted, and plated on LB agar plates to measure the number of colony-forming units (cfu) recovered from the lysed monolayer. The number of intracellular bacteria at 1 hour was compared to the RPMI untreated control and these were considered as adherent bacteria. The number of bacteria recovered 1 hour after gentamicin treatment was defined as 100% and the average percentage of bacteria recovered 4 hours after infection was represented as the replication of bacteria in comparison to this.

[0259] Quantitative polymerase chain reaction (qPCR) analysis of gene expression Before RNA extraction, cells were harvested by centrifugation at 5000 g for 5 minutes at 4°C, the cell pellet was washed 3 times with ice-cold PBS, and then resuspended in 500 μl of ice-cold PBS. Using the PureLink RNA Mini Kit, 2 × 10 6Total RNA of the cells was extracted, and 1.5 μg of RNA (A260 / A280 ratio between 2.00 and 2.20) was reverse-transcribed into cDNA using a high-capacity cDNA reverse transcription kit. The expression of the target gene was quantified using PowerUp SYBR Green Master Mix. Briefly, using a Biomek-4000 automated liquid handler (Beckman Coulter, Brea, California, USA), three sets of qPCR reactions were set up in a MicroAmp optical 384-well reaction plate using 20 ng of cDNA and 500 nM primers. Amplification was performed on a ViiA7 real-time PCR system (Thermo Fisher Scientific) under the following conditions: 20 seconds at 95°C, followed by 40 cycles of 1 second at 95°C and 20 seconds at 60°C, and a melting curve stage: 15 seconds at 95°C; 60 seconds at 60°C. The results of qPCR were analyzed by the 2-ΔΔCT method as described above using QuantStudio software V1.3 (Applied Biosystems). Relative gene expression values were normalized by using GAPDH and 18S as multiple endogenous reference genes.

[0260] Statistical analysis Results were generated from three independent experiments (n = 3) using three technical replicates unless otherwise specified. Data analysis was performed in Microsoft Excel. Results were illustrated using SigmaPlot v11, and the variation between results was represented as the standard deviation (SD). Data were statistically analyzed by one-way ANOVA and post hoc Dunnett's test using SigmaPlot v11 unless otherwise specified. Values with p ≤ 0.05 were considered statistically significant. No differences were seen graphically for p-values between 0.05 and 0.001.

[0261] Example 1: Modeling of Components and Dosages The components of the compositions of the present invention described herein, including their sources, are presented in Table 2 below. In some embodiments, the compositions were prepared by mixing the amounts of bromelain shown in Table 2a or 2b and the remaining components.

[0262]

Table 2a

[0263]

Table 2b

[0264] Modeling of the potential doses of the components of the present invention that reach the ileum and colon after administration of GaRP Dose justification modeling To determine the dose range of bromelain tested in a proof-of-concept in vitro study, the following gastrointestinal distribution model was developed.

[0265] Assumptions: · Oral bioavailability of less than 1% · Bromelain tablets dissolve rapidly and uniformly in ileal fluid · Bromelain fully maintains its biological activity when entering the colon · Mixing of bromelain in the colon is uniform and does not occur as gradient mixing

[0266]

Table 2c

[0267] Ileal enterocytes: To calculate the surface area of the ileum, a cylinder with a length of 3 m and a diameter of 2 cm was considered (Sender, 2016). Next, considering the small intestinal mucosal folds, the total surface area increases threefold (Mudie DM, 2014). Cattaneo and Baratta (Borley, 2005) estimated 1000 ± 3.4 cells per villus. With 13 ± 1 villi / mm 2 the total number of cells covering them is estimated to be 1.35×10 10 . The total number of cells covering the crypts represents 12 / 50 of the crypts covering each villus (Sender, 2016)(Teodori, 1987). If the estimated numbers of crypts and villi are considered to be approximately the same (Sender, 2016), 3.24 ± 0.14×109 It is. Therefore, the total number of intestinal cells was calculated to be 1×10 per 3 m of ileum. 10 as calculated.

[0268] Colonic intestinal cells: To calculate the surface area of the colon, a cylinder with a length of 1.5 m and a diameter of 7.5 cm was considered (Cattaneo L, 1989), which is 1 / 30 of the small intestine due to the absence of villi (Weiss L, 1981). Therefore, the total number of intestinal cells per 1.5 m of colon is 3.3×10 8 is.

[0269] Concentration of bromelain in the ileum: When 1.5 g of ileum / colon-targeted bromelain was orally administered to a 60-kg person together with a cup of water, bromelain was distributed at 94±24 ml (Weiss L, 1981), or 16 mg / ml. At this concentration, assuming that the bromelain tablets are completely dissolved in the ileum, the maximum exposure per bacterium is 170 fg / bacterium.

[0270] Concentration of bromelain in the colon: When 1.5 g of ileum / colon-targeted bromelain was orally administered to a 60-kg person, bromelain was distributed at 400 ml of colonic fluid, or 40 μg / ml. At this concentration, assuming that bromelain is completely mixed with the colonic fluid, the maximum exposure per bacterium is 38 fg / bacterium.

[0271] Concentration of bromelain per ileal intestinal cell: When 1.5 g of ileum / colon-targeted bromelain was orally administered to a 60-kg person together with a cup of water, bromelain was distributed at 94±24 ml (Weiss L, 1981), or 16 mg / ml. At this concentration, assuming that the bromelain tablets are completely dissolved in the ileum, the maximum exposure per intestinal cell is 150 pg / intestinal cell.

[0272] Concentration of bromelain per colonic enterocyte: When 1.5 g of ileum / colon-targeted bromelain was orally administered to a 60 kg person, bromelain was distributed at 400 ml of colonic fluid, or 40 μg / ml. At this concentration, assuming that bromelain is completely mixed with the colonic fluid, the maximum exposure per bacterium is 4.5 ng / enterocyte.

[0273] Using the above model, when an 80 kg person was administered a total daily dose of 1440 mg of bromelain targeted to the small intestine, and 288 mg of menthol enteric-coated, and a dose of 1200 mg of butyric acid (sodium salt), 1800 mg of L-threonine, and 0.025 mg of vitamin D3 targeted to the colon, the following concentrations per enterocyte and bacterium contained in the intestinal microbiota were achieved (Table 3).

[0274]

Table 3

[0275] The observed therapeutically effective amount was determined in the in vitro experiments described herein. Table 4 summarizes the observed therapeutically effective amounts for each GaRP formulation component.

[0276]

Table 4

[0277] Note: No relevance means that a particular formulation component did not exert an anti-adhesion or anti-invasive effect against AIEC obtained from patients with IBD and IBS. Without wishing to be bound by theory, the inventors recognize that * was not tested in vitro, but published literature suggests that 190 - 240 mg / day (35% of 540 mg / day of enteric-coated peppermint oil) is the achieved concentration in the ileum and colon described above.

[0278] In conclusion, the inventors believe that the GaRP formulations described herein successfully achieve therapeutic dosages in both the ileum and the colon, except for the menthol dosage in the modeling that needs to be achieved in the ileum.

[0279] Example 2: Evaluation of the effect of bromelain pretreatment on the invasion and intracellular proliferation of adherent-invasive Escherichia coli (AIEC) in THP-1 macrophages In these studies, the bacteria were pretreated with bromelain.

[0280] Macrophage culture Human monocyte THP-1 cells (ATCC TIB-202) were modified to contain 2-mM L-glutamine, 10 mM HEPES, 1 mM sodium pyruvate, 4500 mg / L glucose, and 1500 mg / L sodium bicarbonate, and cultured in RPMI 1640 medium supplemented with 10% FBS, 100 U / ml penicillin, 100 mg / ml streptomycin, and 5 mM β-mercaptoethanol. The cell density did not exceed 1×10 6 / ml. To induce differentiation, THP-1 cells were seeded at 1 - 10×10 5 cells / cm 2 in 24-well plates and treated with 200 nM PMA for 48 hours. After 48 hours, before starting the experiment, the macrophages were washed and grown for 24 hours in medium without PMA and antibiotics.

[0281] Strains and growth conditions AIECHM605 (from CD patients), AIECHM615 (from CD patients), and Escherichia coli HM484 (isolated from IBS patients) were grown overnight on nutrient agar, suspended in sterile saline, and left at room temperature for 24 hours. This is because it has been found to maximize the expression of fimbriae (HM Martin, 2004). On the day of the experiment, the bacteria were centrifuged at 300 g for 15 minutes at room temperature and resuspended in treatment medium or control medium at 1.4×10 7 bacteria / ml. The treatments are summarized in Table 5.

[0282]

Table 5

[0283] The bacteria were cultured at 37 °C for 2 hours with constant shaking. At the end of the culture, the bacteria were washed three times with RPMI and resuspended in THP-1 medium to a final concentration of 1.4×10 7 bacteria / ml. Assays for bacterial uptake, survival, and replication were measured by the gentamicin protection assay THP-1 cells were infected at a multiplicity of infection (MOI) of 30 bacteria per macrophage. After 3 hours of culture, the medium was removed and fresh cell culture medium containing 100 μg / ml gentamicin was added to kill extracellular bacteria. To determine the number of intracellular bacteria, at 1 hour and 3 hours, the medium was removed and 0.5 ml of 1% Triton-100 (in deionized water) was added to each well for 5 minutes to lyse the THP-1 cells. This concentration of Triton-100 had no effect on bacterial viability for at least 30 minutes. Samples were mixed, diluted, and plated on LB agar plates to measure the number of colony-forming units (cfu) recovered from the lysed monolayer. The number of intracellular bacteria at 1 hour was compared to the RPMI untreated control and these were considered adherent bacteria. The number of bacteria recovered 1 hour after gentamicin treatment was defined as 100% and bacterial replication was represented as the mean percentage of bacteria recovered 4 hours after infection compared to this.

[0284] Results / Discussion: In the case of HM605 and HM484 bacteria, at bromelain concentrations of 20 fg / bacterium or higher, invasion of bacteria into THP-1 macrophages could be inhibited (Table 6). Pretreatment of bacteria with bromelain was less effective in inhibiting intracellular growth.

[0285]

Table 6

[0286] The data in Table 5 were expressed as intestinal cells / cm 2、and by inserting into the gastrointestinal distribution model described herein based on bacteria / intestinal cells, the inventors believe that in order to achieve an effective dose of 20 fg / bacterium, humans would require bromelain targeting the ileum at 867 mg / 80 kg (targeting the small intestine?). In one embodiment, the composition described herein provides bromelain targeting the ileum at a total daily dose of 1440 mg.

[0287] Example 3: Pretreatment of AIEC with bromelain and its effect on the invasion and intracellular growth of AIEC in Caco-2 intestinal cells Cell culture Caco-2-BBe1 intestinal cells were seeded at 2×10 5 cells / cm 2 in 24-well plates and grown until confluent. Before starting the experiment, the cells were grown in antibiotic-free medium for 24 hours. On the day of the experiment, HM605 and HM484 Escherichia coli were centrifuged at 300 gav for 15 minutes at room temperature, and the bacteria resuspended in Caco-2 medium ± 20 - 640 μg / ml bromelain at 1.4×10 7 bacteria / ml were cultured at 37°C for 2 hours with constant shaking. At the end of the culture period, the bacteria were washed three times with DMEM and resuspended in Caco-2 medium to a final concentration of 1.4×10 7 bacteria / ml. Next, the gentamicin protection assay was performed as described above. To simulate the intestinal environment, a series of experiments were carried out in which the bacteria were resuspended in fasting-state simulated intestinal fluid (Rieder F, 2012) containing 20 - 640 μg / ml bromelain.

[0288] Results / Discussion: In the case of HM605 and HM484 bacteria, at bromelain concentrations of 20 fg / bacterium or higher, the invasion of bacteria into intestinal cells and the intracellular growth of bacteria could be inhibited (Table 7).

[0289] Pretreatment of the bacteria in simulated intestinal fluid increased the concentration of bromelain required to achieve the same level of suppression of bacterial invasion and intracellular proliferation (from 20 fg to 28 fg). Thus, when using the gastrointestinal exposure model described herein, to achieve 20 fg of bromelain per bacterium, the appropriate patient dosage is 800 mg / 60 kg / day.

[0290]

Table 7

[0291] Inserting the data in Table 7 into the gastrointestinal distribution model based on the intestinal cells / cm 2 and bacteria / intestinal cells described herein, the inventor believes that to achieve an effective dose of 28 fg / bacterium, the appropriate patient dosage is bromelain targeting the small intestine at 1067 mg / 80 kg / day.

[0292] Example 4 - GaRP containing bromelain is effective in inhibiting the release of TNF-α from LPS-stimulated THP-1 macrophages Macrophage culture Human monocyte THP-1 cells (ATCC TIB-202) were modified to contain 2-mM L-glutamine, 10 mM HEPES, 1 mM sodium pyruvate, 4500 mg / L glucose, and 1500 mg / L sodium bicarbonate, and cultured in RPMI 1640 medium supplemented with 10% FBS, 100 U / ml penicillin, 100 mg / ml streptomycin, and 5 mM β-mercaptoethanol. The cell density does not exceed 1×10 6 / ml. To induce differentiation, THP-1 cells were seeded at 5×10 5 cells / cm 2 in a 96-well plate and treated with 200 nM PMA for 48 hours. After 48 hours, before starting the experiment, the macrophages were washed and grown for 24 hours in medium without PMA and antibiotics. The transwell is shown in Figure 1. The treatments are shown in Table 8.

[0293]

Table 8

[0294] As a test compound and as a control, unstimulated cultures were used as controls, and cells in which macrophages were stimulated with 50 ng / ml LPS were applied. After 8 hours, the medium was removed, and TNF-α was quantified using ELISA.

[0295] Results / Discussion: As shown in Figure 2, bromelain was able to inhibit the release of TNF-α from LPS-stimulated THP-1 macrophages. When compared with the RPMI control, at bromelain concentrations higher than 13 pg / macrophage, significant inhibition was seen in the release of TNF-α (p < 0.001, Mann-Whitney rank sum test).

[0296] Conclusion: Although not wishing to be bound by theory, the inventors believe that their modeling using the gastrointestinal distribution model indicates that when humans are administered 800 mg of enteric-coated bromelain, the exposure is 75 pg / cell in ileal enterocytes and 4500 pg / cell in the colon.

[0297] Example 5 - GaRP containing bromelain inhibits the release of IL-8 from polarized Caco-2 monolayers co-cultured with LPS-stimulated THP-1 macrophages Using the industry standard model most commonly used for the evaluation of foods and pharmaceuticals, the effect of bromelain on the release of IL-8 was studied. The apical compartment represents the lumen (the fluid and mucosa of the GIT). Co-culture cell culture Caco-2 cells were seeded at 1×10 6 cells / well in a Transwell insert plate (1.12 cm 2 , pore size 0.4 μm). The cell culture medium was changed every 3 days until the cells were fully differentiated (TER value > 1200 / cm 2 ), and the cells were used within passage 12 after thawing.

[0298] In another 12-well plate, THP-1 monocytes were seeded at 8.5×10 5 cells / well, differentiated with 200 nM PMA for 48 hours, and then cultured in normal medium for 24 hours before the experiment. After replacing all the medium with RPMI 1640, a transwell insert with Caco-2 cells on top was filled into the wells containing THP-1 cells. When the cell culture reached the TER baseline of >1000 / cm 2 the monolayer was stimulated by adding 0.6 ml containing 50 ng / ml IFN-β, 10 μg / ml LPS, and 5 ng / ml TNF-α to the basolateral compartment. Immediately after the addition of the stimulation cocktail, 0.25 ml of GaRP ± 0 - 80 μg / ml bromelain (0 - 20 pg / enterocyte) was applied to the apical compartment. After 24 hours of culture, the culture supernatants from the apical side and the basolateral side were collected for IL-8 measurement.

[0299] Results / Discussion: As shown in Figure 3, bromelain can inhibit the release of IL-8 from Caco-2 cells co-cultured with LPS-stimulated THP-1 macrophages. GaRPv1 without bromelain was able to inhibit the release of IL-8 by only 18 ± 6.

[0300] Example 6 - Evaluation of the effect of GaRPv1 containing bromelain on the recovery of intestinal integrity measured by transepithelial electrical resistance (TER) In preclinical trials, vitamin D3 and sodium butyrate have been shown to be effective in regulating tight junction (Kong J, 2008) (Ma X, 2012), which was a discovery that played a role in the selection as part of the GaRP formulation. To determine whether GaRPv1 restores intestinal integrity, the M cell / Caco-2 transwell model was infected with AIECHM605 ± GaRP, and TER was monitored for 2 days after infection.

[0301] M cell / Caco-2 model Caco2-cl1 cells were grown and maintained as described above. Human Burkitt lymphoma cell line RajiB was maintained in RPMI 1640 medium supplemented with 10% FBS, 8 mM L-glutamine, 100 U / ml penicillin, and 100 mg / ml streptomycin. RajiB cells were seeded at 3×10 5 / ml and the cell suspension was allowed to settle every 3 days. Two-thirds of the medium was replaced with fresh medium. Every 9 days, the cells were split 1:3. All cells were maintained at 37 °C with 5% CO2 in a humidified atmosphere.

[0302] M cells were generated by seeding 5×10 5 Caco-2 cells on the apical surface of 12-mm Transwell cell culture inserts pre-coated with Matrigel basement membrane matrix and placing DMEM medium in the basolateral compartment. The cells were maintained for 14 - 18 days until the TER exceeded 600 Ω (equivalent to 300 Ω·cm 2 tissue monolayer). On the day of co-culture, RajiB cells were harvested, resuspended in complete DMEM, and 5×10 5 cells were added to the basal Transwell compartment. Importantly, both the apical and basal media were exchanged daily while taking care not to disturb the RajiB cells after co-culture. The co-culture was maintained for 4 - 6 days until M cells were generated. In parallel, a single culture of Caco2-cl1 (without RajiB cells in the basal compartment) was also generated on Transwell inserts and maintained by M cells. TER was measured throughout to monitor successful monolayer formation.

[0303] Infection of the M cell model with AIECHM605 It has been previously reported (Bailey JR, 2011) that AIECHM615 causes disruption of tight cell junctions and induces cell death in Caco-2 and T84 monolayers. To reproduce this effect, when an M cell model was generated (days 20 - 24 depending on the time of the accompanying decrease in TER), the medium was changed to antibiotic-free, and bacteria were added to the apical well of the transwell insert at an MOI of 30 for 4 hours. After 4 hours, the cells were washed and cultured in the presence of 100 μg / ml gentamicin to kill the remaining bacteria. At that point, GaRP without bromelain was placed on the cells, and the TER of all monolayers was measured every 2 hours for up to 12 hours, and the bacteria in the basal well were counted every hour for up to 4 hours.

[0304] Results / Discussion: Recovery of the gastrointestinal cell layer after AIEC infection When AIECHM605 was suspended in normal medium (untreated control) or GaRP (without bromelain) and applied to the M cell model, the GaRP formulation provided a protective effect on the integrity of the cell layer (Figure 4).

[0305] Example 7 - Comparison of anti-inflammatory activities in various gastrointestinal reprogramming compositions without bromelain The human colorectal adenocarcinoma cell line Caco-2-BBe1 was grown and maintained in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% v / v fetal bovine serum (FBS), 4 mM L-glutamine, 100 U / ml penicillin, and 100 mg / ml streptomycin. All experiments were performed with C2BBe1 cells within 12 passages after thawing. Caco-2 cells were seeded at 5×10 5 cells / well in a transwell insert plate (0.33 cm 2 , pore size 0.4 μm). The cell culture medium was changed every 3 days until the cells were fully differentiated (starting average TER of 1121 ± 196 cm 2 ; mean ± SD), and the cells were used within 12 passages after thawing.

[0306] In an experiment to evaluate the anti-inflammatory effects of bromelain or various GaRP formulations, a 0.6 ml cocktail containing 50 ng / ml IFN + 10 μg / ml LPS + 5 ng / ml TNF was applied to the basolateral side in this model, and at the same time, 0.25 ml of the sample was applied to the apical side for 24 hours. After 24 hours of culture, the culture supernatant was collected from the apical side, and IL-8 was quantified by ELISA.

[0307] Table 9 shows an overview of various substitutions and combinations of formulation components that were tested for their ability to inhibit the release of IL-8 from polarized Caco-2 monolayers stimulated with LPS, TNF-α, and IFN-β, as previously described herein.

[0308]

Table 9

[0309] Results / Discussion: The data shown in Table 10 confirm that GaRP without bromelain can significantly reduce the release of IL-8 compared to the standard growth medium (DMEM) (p < 0.001). When all GaRP formulations were compared to the original GaRPv1, there was no statistically significant difference, but when ranking the anti-inflammatory activities from highest to lowest, generally clear observations as shown in Table 11 could be made.

[0310]

Table 10

[0311]

Table 11

[0312] When all GaRP formulations are ranked from highest to lowest for anti-inflammatory activity, the inventors believe that several clear observations can be made (Table 11). That is, sodium butyrate, L-threonine, and cholecalciferol are all correlated with a better anti-inflammatory effect, and generally, when formulations containing L-glutamine and L-tryptophan are used, a higher anti-inflammatory effect is observed.

[0313] In the examples provided herein, it is confirmed that bromelain is an important anti-inflammatory component of the GaRP dietary supplements described herein, and as the sole agent, bromelain inhibited the secretion of IL-8 by 70±7 (mean±SEM). Without wishing to be bound by theory, the inventors believe that the additional components (sodium butyrate, amino acids, and vitamin D3) of the GaRP formulations described herein act as sources of cellular energy and mucin synthesis precursors that promote the anti-inflammatory activity of bromelain. These experiments confirm previously published studies that each formulation component alone exhibits low levels of anti-inflammatory activity. In one embodiment, the GaRP formulations described herein comprise, or consist essentially of, 80 μg / ml bromelain, 400 μg / ml sodium butyrate, 400 μg / ml L-threonine, and 0.48 μg / ml vitamin D3.

[0314] Example 8 - Comparison of the anti-inflammatory activities of various gastrointestinal reprogramming compositions containing 80 μg / ml (equivalent to 40 pg / cell) of bromelain To determine whether any of the GaRP formulations exerted an additive or synergistic effect on the anti-inflammatory activity of bromelain, each formulation was retested after adding 80 μg / ml bromelain (equivalent to 20 pg / cell).

[0315] Methodology The human colorectal adenocarcinoma cell line Caco-2-BBe1 was grown and maintained in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% v / v fetal bovine serum (FBS), 4 mM L-glutamine, 100 U / ml penicillin, and 100 mg / ml streptomycin. All experiments were performed with C2BBe1 cells within 12 passages after thawing. Caco-2 cells were seeded at 5×10 5 cells / well in a transwell insert plate (0.33 cm 2 , pore size 0.4 μm). The cell culture medium was changed every 3 days until the cells were fully differentiated (starting average TER of 1121 ± 196 cm 2 ; mean ± SD), and the cells were used within 12 passages after thawing.

[0316] In an experiment to evaluate the anti-inflammatory effects of bromelain or various GaRP formulations, 0.6 ml of a cocktail containing 50 ng / ml IFN + 10 μg / ml LPS + 5 ng / ml TNF was applied to the basolateral side in this model, and simultaneously, 0.25 ml of the sample was applied to the apical side for 24 hours. After 24 hours of culture, the culture supernatant was collected from the apical side, and IL-8 was quantified by ELISA.

[0317] Results / Discussion: Table 12 shows the results of adding 80 μg / ml of bromelain (equivalent to 40 pg / cell) to the formulations tested in Example 7. The added bromelain increased the ability to suppress IL-8 secretion by 30 - 70%.

[0318] [Table 12-1]

[0319] [Table 12-2]

[0320] As shown in Table 13, the highest anti-inflammatory effect was observed when the formulations containing bromelain were ranked.

[0321]

Table 13

[0322] Example 9 - Quantification of the effects of GaRP and its individual formulation components on mucin - like receptors and the expression of inflammatory - inducing genes The 75:25 Caco - 2 / HT - 29 co - culture was established as previously described herein, and the effects of 24 - hour exposure to bromelain, threonine, sodium butyrate, vitamin D3, and two different GaRP formulations on the expression of the following MUC genes were studied using qPCR. At the end of the 24 - hour treatment, the supernatant in the 25 cm 2 flask was collected, and the co - culture monolayer was trypsinized as previously described herein.

[0323]

Table 14

[0324] When interpreting the gene expression data, the MUC genes and their associated mucins were classified as follows: · Secreted mucins · Membrane - bound mucins · Non - gel - forming mucins Secreted mucins MUC2, MUC5AC, MUC5B, MUC6, and MUC19 act as secreted extracellular viscous secretions. For an overview of the effects of various concentrations of bromelain dissolved in GaRP formulation components targeting the colon of sodium butyrate, L - threonine, and vitamin D3, see Figure 5.

[0325] Table 14 summarizes the association of each mucin with IBS or IBD.

[0326]

Table 15

[0327] The expression of the MUC2 gene and protein has been found to be significantly decreased in IBD and IBS and is considered a very important mucin for maintaining the mucosal barrier. The glycan content bound to mucin MUC2 functions not only as a nutrient for bacteria but also as a binding site, inducing the selection of specific microbial species essential for integrity, homeostasis, and maintenance of intestinal function. As seen in Figure 6, bromelain at 20 and 40 μg / ml in GaRP was able to increase gene expression 5 - 25-fold.

[0328] MUC6 is decreased in IBD and IBS. At bromelain concentrations of 10 μg / ml and above, the GaRP formulation component appeared to act additively, if not synergistically, with bromelain (GaRPv1), significantly increasing the expression of MUC6 up to 30-fold.

[0329] The expression of MUC19 is not characterized in IBS or IBD. At bromelain concentrations of 10 μg / ml and above, the components of the GaRPv1 formulation appeared to have an additive effect on the activity of bromelain.

[0330] Membrane-bound mucin MUC1 - 4, MUC12, MUC15 - 17, and MUC20 are considered membrane-bound mucins within the glycocalyx. See Figure 6 for an overview of the effects of various concentrations of bromelain dissolved in the GaRP formulation components targeting the colon with sodium butyrate, L-threonine, and vitamin D3.

[0331] Table 15 summarizes the association of each mucin with IBS or IBD.

[0332]

Table 16

[0333] Except for the expression of the MUC1 gene, it appears that at bromelain concentrations of 10 μg / ml or higher, the GaRP / bromelain formulation significantly increases the expression of MUC3, MUC4, and MUC15. Considering that these mucins are decreased in IBD and sometimes in IBS, these data suggest that GaRP products may be able to stimulate mucin production to re - establish intestinal homeostasis. Generally, the GaRPv1 formulation targeting the colon shows better performance than GaRPv2, and protein research and mucin quantification studies are currently underway to confirm this observation.

[0334] The expression of MUC13, MUC16, and MUC17 increases in IBD. At bromelain concentrations of 20 μg / ml or higher, the GaRPv2 / bromelain formulation was able to significantly decrease gene expression.

[0335] Non - gel - forming mucin MUC7 and MUC8 are non - gel - forming mucins that are not well - characterized in IBD or IBS. Referring to Figure 7, at bromelain concentrations of 10 μg / ml or higher, gene expression could be significantly increased.

[0336] Although not wishing to be bound by theory, the inventors believe that the MUC gene expression data provided herein indicate that the concentrations of bromelain and colon - targeting formulation components used in GaRP dietary supplements are sufficient, in turn, to stimulate the expression of mucin genes that are expected to promote mucosal healing.

[0337] Example 10 - Mucosal healing ability of GaRP products: showing a synergistic relationship between L - threonine and vitamin D3 Establishment of cell line and Caco - 2:HT - 29 - MTX(75:25) co - culture (simulation of the colon) The human colorectal adenocarcinoma cell line Caco-2-BBe1 (obtained from Professor Barry Campbell of the University of Liverpool) was grown and maintained in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% v / v fetal bovine serum (FBS), 4 mM L-glutamine, 100 U / ml penicillin, and 100 mg / ml streptomycin. All experiments were performed using Caco-2-BBe1 cells within 12 passages after thawing. The cell cultures were incubated at 37°C in a humidified 5% CO2 incubator. The cells were maintained in a 5% CO2 atmosphere in a 37°C incubator, and the medium was changed every 48 hours. At 80% confluence, the cells were detached with 10 ml of 1% trypsin-EDTA (Sigma-Aldrich) for 5 minutes, and then 40 ml of DMEM containing 10% FBS was added to terminate the trypsinization. The cell suspension was pelleted by centrifugation (200 g, 5 minutes), resuspended in fresh DMEM containing 10% FBS, and subcultured into new flasks at a 1:4 dilution.

[0338] This co-culture model was used to simulate human colonic epithelium. Caco-2 cells (passage 8) and HT-29-MTX cells (passage 9) were combined at a ratio of 75:25 in DMEM containing 10% FBS. The combined cells (6.4×10 4 cells / ml) were seeded into 25 cm 2 flasks at a total of 5 ml per flask. The co-cultures were incubated at 37°C in a 5% CO2 atmosphere while changing the medium every 48 hours. On the 15th day after seeding, the monolayer of the co-culture was washed three times with DMEM, the medium was replaced with 5 ml of pre-warmed test solution, and incubated for 24 hours.

[0339] Treatment of Caco-2:HT-29-MTX (75:25) co-culture Test solutions of cholecalciferol (VD3) and L-threonine (L-Thr) (Sigma-Aldrich) in the culture medium were prepared 1 day before treatment. On the 15th day after seeding, the co-culture monolayer was washed three times with the culture medium at 1-hour intervals, the medium was replaced with 5 ml of pre-warmed test solution, and cultured for 24 hours.

[0340] Quantitative polymerase chain reaction (qPCR) analysis of gene expression At the end of 24 h of treatment, the supernatant in the 25 cm 2 flask was collected, and the co-culture monolayer was trypsinized as previously described. Cells were harvested by centrifugation at 5000 g for 5 min at 4 °C. After washing three times with ice-cold PBS, the cells were resuspended in 500 μl of ice-cold PBS. Total RNA of the cells was extracted using the PureLink RNA Mini Kit (Invitrogen) at 2 × 10 6 per sample. For real-time PCR analysis, 1.5 μg of RNA (A260 / A280 ratio between 2.00 and 2.20) was reverse transcribed into cDNA using the High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems, Foster City, CA, USA).

[0341] The expression of human MUC2 and MUC7 genes in Caco-2:HT-29-MTX (75:25) co-culture was quantified using PowerUp SYBR Green Master Mix (Applied Biosystems). Primers for detecting the expression of MUC2 (forward, 5’-CGAAACCACGGCCACAACGT-3’ (SEQ ID NO: 3); reverse, 5’-GACCACGGCCCCGTTAAGCA-3’ (SEQ ID NO: 4)), and primers for detecting the expression of MUC7 (forward, 5’-GCAACTACCCTAGACCCATCA-3’ (SEQ ID NO: 15); reverse, 5’-TAGGTGTGGTAATTGGGGCAG-3’ (SEQ ID NO: 16)) were purchased from Sigma-Aldrich. A 10 μl qPCR reaction was set up on a MicroAmp Optical 384-well reaction plate (Applied Biosystems) using a Biomek-4000 automated liquid handler (Beckman Coulter, Brea, California, USA), with 20 ng of cDNA and 500 nM primers in triplicate. Amplification was performed on a ViiA7 real-time PCR system (Thermo Fisher Scientific) under the following conditions: 20 s at 95 °C, followed by 40 cycles of 1 s at 95 °C and 20 s at 60 °C, and a melt curve stage: 15 s at 95 °C; 60 s at 60 °C. The qPCR results were analyzed by the 2-ΔΔCT method as described above using QuantStudio software V1.3 (Applied Biosystems). Relative gene expression values were normalized by using GAPDH and 18S as multiple endogenous reference genes.

[0342] Results When interpreting gene expression data, MUC genes and their associated mucins were classified into three classes: · Secreted mucins · Membrane-bound mucins · Non-gel-forming mucins Secretory mucin MUC2, MUC5AC, MUC5B, MUC6, and MUC19 act as secreted extracellular viscous secretions. Table 16 summarizes the association of each mucin with IBS or IBD.

[0343]

Table 17

[0344] Membrane-bound mucin MUC1-4, MUC12, MUC15-17, and MUC20 are regarded as membrane-bound mucins within the glycocalyx. Table 17 summarizes the association of each mucin with IBS or IBD.

[0345]

Table 18

[0346] Non-gel-forming mucin MUC7 and MUC8 are non-gel-forming mucins that are not well characterized in IBD or IBS. See Figure 8 below, which shows the synergistic effect of the combination of L-threonine and vitamin D3 on the above MUC genes.

[0347] L-threonine (400 μg / ml) as the sole agent did not statistically significantly affect the expression of any of the MUC genes tested. The effects of i) vitamin D3 and ii) the combination of 0.48 μg / ml vitamin D3 and 400 μg / ml L-threonine on MUC gene expression are summarized in Table 18.

[0348]

Table 19-1

[0349]

Table 19-2

[0350] The synergistic effect is highlighted in boldface. Evidence that L-threonine and vitamin D3 exerted an unexpected synergistic effect by increasing the expression of MUC15, MUC2, MUC6, MUC19, MUC7, and MUC8 was further investigated by measuring the potencies of L-threonine and vitamin D3. The expression of MUC2 and MUC7 was used as typical mucin genes.

[0351] As seen in FIGS. 9A and 9B, individually, neither L-threonine nor vitamin D3 had a significant effect on the expression of MUC2 (a gel-forming mucin mainly secreted in the gastrointestinal tract) or MUC7 (a non-gel-forming mucin).

[0352] In both IBS and IBD, it has been shown that changes in intestinal integrity and the resulting enterotoxemia microbiota, factors leading to systemic inflammation, decrease the expression of the MUC2 gene, protein, and mucin.

[0353] When titrating 0 to 0.48 μg / ml vitamin D3 in combination with 0 to 800 μg / ml L-threonine, it was revealed that at vitamin D3 of 0.053 μg / ml or more, the concentration of L-threonine was the main factor affecting the increase in MUC2 and MUC7 gene expression (FIGS. 10A and 10B).

[0354] Example 11 - Bromelain and GaRP can reduce the endoscopic disease in the TNBS colitis model Evaluation of GaRP in the TNBS mouse model 2,4,6-Trinitrobenzenesulfonic acid solution (TNBS) colitis was induced in 5-week-old C56 / BL / 6 mice by rectal introduction of TNBS (5 w / v%) in 50% ethanol. The animals were randomly divided into treatment groups and treated as outlined in FIG. 11.

[0355] Eight treatment groups (n = 4 in the healthy control group, 8 - 9 animals / TNBS group) were completed. A detailed description of the treatment groups is shown in Table 19.

[0356]

Table 20

[0357] The following endoscopic scoring system was used: i) Score 5 - 7 = mild disease, ii) Score 8 - 11 = moderate disease, iii) Score 12 - 15 = severe disease. The severity of the disease at the population baseline was as follows.

[0358] · Mild: 34% (21 / 62 animals) · Moderate: 62% (38 / 62 animals) · Severe: 4% (3 / 62 animals) 8.1: Safety Of the original 72 animals randomized in the study, 10 died from anesthesia complications or lost more than 20% of their body weight before the start of treatment and were excluded from the analysis. Study-related deaths are summarized in Table 20.

[0359]

Table 21

[0360] Animals euthanized due to weight loss looked good and were functioning normally, but were euthanized due to ethical guidelines. See Figure 12 comparing the change in body weight from Day 0 of treatment to the experimental endpoint (Day 7).

[0361] The only group that showed a significant difference in body weight at the experimental endpoint was as follows. · 75 mg / kg bromelain / 15 mg / kg menthol in TNBS - oral vehicle: 75 mg / kg bromelain / 15 mg / kg menthol in oral vehicle decreased the weight at the endpoint by 9% compared to 75 mg / kg bromelain in water or oral vehicle (p = 0.024).

[0362] · 75 mg / kg bromelain / 15 mg / kg menthol in TNBS - oral vehicle, and GaRP in rectal vehicle: When compared to 75 mg / kg bromelain in water, oral or rectal vehicle, 75 mg / kg bromelain / 15 mg / kg menthol / rectal GaRP reduced the weight at the endpoint by 5 - 12% (p = 0.001).

[0363] When these groups were compared over the treatment period, as shown in Figure 13, the animals maintained their original body weight. In published studies, administration of 10 mg / kg L - menthol to mice was found to increase core temperature, dissociate the expression of protein 1 in brown adipose tissue, and the heat - generating activity without shivering and insulin sensitivity led to attenuation of weight gain. In the GaRP product, mice ingest 15 mg / kg daily, which may explain the observed lack of weight gain. No toxicity was observed.

[0364] Efficacy Refer to Figure 14 comparing the change in endoscopic scores from day 0 (before treatment) to day 7 (experimental endpoint). Statistically significant differences were observed in the following treatment groups.

[0365] · Components targeting the colon of the GaRP product (sodium butyrate / L - threonine / vitamin D3 administered rectally): Colitis was significantly reduced as quantified by the endoscopic score. When compared to the rectal vehicle control, GaRP improved the endoscopic score by 135% (p = 0.018, n = 8).

[0366] · Orally administered bromelain / peppermint oil combined with components targeting the colon of the GaRP product (sodium butyrate / L - threonine / vitamin D3 administered rectally): When compared to oral and rectal vehicle controls, this formulation improved the endoscopic score by 148% (p = 0.012, n = 9).

[0367] · Bromelain in water administered orally: When compared to the oral vehicle control, bromelain improved the endoscopic score by 65% (p = 0.038, n = 8). When comparing individual parameters such as stool hardness, area involved, granularity of the mucosal surface, changes in the vascular pattern, and thickening of the colonic wall, there was no significant difference between the treatment groups.

[0368] See Figure 15 comparing the effects of various treatments and control vehicles on colon length, colon weight, and the ratio of colon weight / length. No significant difference was observed between the treatment groups. Conclusion: The colonic target components of the GaRP product without bromelain (sodium butyrate / L-threonine / vitamin D3 administered rectally) significantly improved experimental colitis. The addition of orally administered bromelain / menthol exerted a slight additive effect in the colitis model, but this was not significantly superior to GaRP alone. It was noted that bromelain in the oral vehicle did not function as well as bromelain in water, and the inventors hypothesized and suggested that some components of the oral vehicle may interfere with the release of bromelain in both the small and large intestines. Specific formulations that provide the release of bromelain and / or any of the components of the compositions and formulations described herein are considered to be within the scope of the art of pharmaceutical formulations for delivery to the gastrointestinal system, as described elsewhere herein.

[0369] Example 12 - Bromelain, and GaRP can reduce the endoscopic disease in the TNBS colitis model without negating the negative drug interaction with prednisolone Prednisolone is known to have 55 major, 390 moderate, and 39 minor drug interactions. Among the known adverse effects are interactions with some common ingredients such as buffers that can impair the absorption of prednisolone. Herbal medicines are a popular form of treatment and are being used, usually by an increasing number of patients whom clinicians do not normally recommend for combination use. Therefore, physicians and clinicians are aware that known or potential herb-drug interactions exist and should be screened for. This concept can, of course, be extended to non-herb-based nutritional agents. Of particular relevance to irritable bowel syndrome and inflammatory bowel disease is the common use of corticosteroids to control inflammation. According to Miller [Miller LG "Herbal medicines: selected clinical considerations focused on known or potential herb-drug interactions", Arch Intern Med, 158 (1998): 2200-11], the theoretical concern underlying these herb-drug interactions is that immunostimulatory herbs counteract or minimize the immunosuppressive effects of corticosteroids. In light of these concerns, the inventors investigated the effects of prednisolone in the presence and absence of the formulations described herein.

[0370] Mice were induced with colitis by administering TNBS in the same manner as in Example 11. Three days later, the lesions were recorded by endoscopy and the animals were treated with i) 150 mg / kg bromelain in water, ii) 150 mg of bromelain and rectally administered threonine / sodium butyrate / vitamin D3, iii) 2 mg / kg oral prednisolone, or iv) 150 mg of bromelain and rectally administered threonine / sodium butyrate / vitamin D3 and 2 mg / kg oral prednisolone. The endoscopic score was measured on day 11 and the change in endoscopic score from day 3 to day 11 was calculated (Figure 16). Table 21 shows the change in endoscopic score of the above groups and the statistical significance compared to placebo.

[0371]

Table 22

[0372] The groups were: i) placebo, ii) bromelain in 150 mg in water orally, iii) 150 mg bromelain and 15 mg / kg menthol orally, and rectally GaRP, iv) 150 mg bromelain and 15 mg / kg menthol and rectally GaRP and 2 mg / kg prednisolone, v) 2 mg / kg prednisolone rectally, and vi) GaRP rectally. GaRP is 185 mg / kg sodium butyrate, 280 mg / kg threonine, 3.84 μg / ml vitamin D3. The human equivalent doses are: bromelain 960 mg / human 80 kg; menthol 100 mg / human 80 kg; threonine 1800 mg / human 80 kg; sodium butyrate 1200 mg / human 80 kg; and vitamin D3 25 μg / human 80 kg. Threonine, sodium butyrate, and vitamin D3 were formulated and administered as a rectal forced administration in the upper colon to simulate a formulation that is released in the colon, i.e., to achieve administration to the lower intestine. Since the pH of the mouse colon is lower and the transit time is shorter than that of humans, the human formulation of the compositions described herein could not be used. This means that the formulation moves quickly through the mouse GI tract and the pH required to lose the protective coating necessary for release is not achieved, so the targeted human formulation does not release the contents in animal studies.

[0373] From the results presented herein, the inventors believe that formulations containing sodium butyrate, L-threonine, and vitamin D do not interfere with the effectiveness of prednisolone, and that 1) bromelain alone, 2) threonine / butyric acid / vitamin D3 alone, and 3) the combination of oral bromelain / menthol and rectal threonine / butyric acid / vitamin D3 are as effective as 2 mg / kg prednisolone in this experiment.

[0374] Example 13 - Formulation of Dosage Forms Part A Tablet / minitablet / capsule / Ingredients: The objective was to develop a stable enteric-coated mini-tablet formulation containing bromelain and menthol at a ratio of 6.8:1 (±30%, i.e., in the range of 4.75:1 to 8.85:1). The developed formulation was prepared by the direct compression method.

[0375]

Table 23

[0376] Manufacturing procedure: 1. Menthol premix: Menthol was melted under heating at 70°C. Fucicel was added to the melted menthol and heated with stirring. Stirring was continued until a free-flowing powder was formed.

[0377] 2. Aerosil 200, Fucicolin, and the menthol premix need to be mixed in a 10 L octagonal blender at 15 rpm for 15 minutes. 3. Bromelain, Avicel SMCC90, Crucell EXF, and crospovidone were sieved through #40 ASTM attached to a mechanical sieve and mixed with the materials from step 2 in a 30 L octagonal blender at 15 rpm for 15 minutes.

[0378] 4. Magnesium stearate was sieved through #60 ASTM. 5. Lubrication: The material from step 4 was added to the material from step 3 and mixed in a 30 L octagonal blender at 15 rpm for 5 minutes.

[0379] 6. Compression: The lubricated mixture was compressed into mini-tablets using a 2.50 × 2.50 mm round biconvex-shaped multi-tip punch. 7. Seal coating: The required amount of purified water was added to Eudraguard Control. Methocel E5 Premium LV and talc were added with stirring. Stirring was continuously carried out during the coating operation. The suspension was sprayed onto the core tablet bed of an automatic tablet coater.

[0380] 8. Enteric Coating: Glycerin was added to Eudraguard Control while stirring. The required amount of purified water was added to the Eudraguard Control and glycerin mixture while stirring. Sodium alginate and talc were placed in a plastic bag and mixed, and then slowly added to the above mixture without forming lumps. The required amount of lake dye was homogenized with a small amount of water and added to the suspension while stirring. Stirring was continued for 3 hours before the coating operation.

[0381]

Table 24

[0382] Based on the stability profile of bromelain, a direct compression strategy was selected. Alternative methods such as dissolving menthol in miglyol, embedding it in wax, or incorporating menthol micelles into vitamin E were investigated, but due to the tendency of menthol to sublime at low temperatures, an adsorption technology was adopted.

[0383] The minitablets were subjected to a dissolution test according to the standard USP method using a Type II apparatus USP29NF24<711>. The minitablets were stirred in 0.1N hydrochloric acid for 1 hour and then adjusted to pH 6.8 for 5 hours. Samples were taken at the time of pH adjustment (0), 30 minutes and 60 minutes before pH adjustment, 30 minutes and 60 minutes after adjustment, and then every 1 hour thereafter. As shown in Figure 17, bromelain was analyzed and the titer was plotted against time.

[0384] Part B Tablet / minitablet / capsule / Ingredients: The pharmaceutical was designed as a minitablet for delivering drugs to the lower part of the small intestine and colon regions. This product is physically and chemically stable and is designed to exert its efficacy during the shelf life. Formulations and methods were developed to ensure a consistent product that meets all quality criteria. The ratio of threonine to sodium butyrate is 1.5:1 ± 30% (1.05:1 to 1.95:1).

[0385]

Table 25-1

[0386]

Table 25-2

[0387] Manufacturing method: 1. Sieving: Sodium butyrate, L-threonine, microcrystalline cellulose, dicalcium phosphate anhydrous (Tab A), crospovidone (Kollidon CL) were passed through a #24 ASTM mesh.

[0388] 2. Binder solution: Butylated hydroxytoluene, vitamin D3, and povidone K90F were added to isopropyl alcohol and stirring was continued until a clear solution was formed. 3. The above materials were combined by wet granulation, the product was dried in a fluid bed dryer, and then pulverized and sieved with magnesium stearate.

[0389] 4. Compression: The mixture was compressed into minitablets using a 2.50 × 2.50 mm round biconvex multi-tip punch. 5. Seal coating: Hydroxypropyl methylcellulose in isopropyl alcohol / methylene chloride seal coat was applied.

[0390] 6. Eudraguard biotec dispersion: Eudraguard biotec, water, and PlasACRYLT 20 dispersion were sprayed onto the sealed minitablets. 7. Eudraguard Control dispersion: Glycerin, Eudraguard Control, sodium alginate, and talc were mixed to form a dispersion, which was coated onto the minitablets.

[0391]

Table 26

[0392] The mini-tablets were subjected to a dissolution test according to the standard USP method using a Type II apparatus USP29NF24<711>. The mini-tablets were stirred in 0.1 N hydrochloric acid for 1 hour, then adjusted to pH 6.2 for 1 hour and finally to pH 7.4 for 4 hours. Samples were taken at the time of pH adjustment (0), 60 minutes before pH adjustment, +60 minutes after adjustment (before the second pH adjustment), +30 minutes and +60 minutes after the second adjustment, and then every 1 hour thereafter. As shown in Figure 18, butyric acid was analyzed and the titer was plotted against time.

[0393] Finished formulation: The mini-tabs of Part A and Part B are of similar size and characteristics and may be mixed in any useful ratio depending on the stage of the condition being treated. In current clinical trials, they were mixed in a ratio of 1.94:2.28 w / w, providing a component ratio of 70 (menthol):480 (bromelain):900 (threonine):600 (sodium butyrate):0.0125 (colecalciferol). An alternative ratio of 2.91 w / w (Part A) to 2.28 w / w (Part B) provides a component ratio of 105 (menthol):720 (bromelain):900 (threonine):600 (sodium butyrate):0.0125 (colecalciferol), and a ratio.

[0394] The doses provided to patients in current clinical trials are 4.22 g twice a day, providing a total daily dose of 140 mg of menthol, 960 mg of bromelain, 1800 mg of threonine, 1200 mg of sodium butyrate, and 25 μg of colecalciferol, or 2.21 g twice a day, providing a total daily dose of 70 mg of menthol, 480 mg of bromelain, 900 mg of threonine, 600 mg of sodium butyrate, and 12.5 μg of colecalciferol. The filling tolerance of the dose is ±15%.

[0395] Histological scoring of the treated sections in Examples 14 - Examples 11, and 12 Evaluation of GaRP in the TNBS mouse model Examples 11 and 12 evaluated aspects of GaRP performance in TNBS-induced colitis in a mouse model. The endoscopic scores on day 11 compared to day 3, and the effects of various treatment groups on the weight, length, and weight-to-length ratio of the excised colon of euthanized animals were reported.

[0396] In this example, before evaluation by microscopy in a blinded manner, the colon was fixed, stained, and sectioned. The colon was examined for evidence of both active disease and recovery from the disease and scored according to the following criteria. Scoring was performed three times with a maximum score of 3 in each of six categories and the scorers were blinded.

[0397] Histological scoring of colitis Activity of colitis (0 - 3 based on severity) Hypervascularity (opening of blood vessels) Presence of mononuclear cells (structure also lost) Epithelial hyperplasia (elongated crypts and villi) Epithelial damage (loss of epithelial continuity) Presence of neutrophils Lymphoid aggregates (score 0 - 2) 0, 1 - 2, >2 Data were collated and analyzed by Student's t-test for significance against the placebo control. Summary statistics are reported in the following table, and the mean of the healthy controls subtracted from the mean data of each group (x￣ = 5.1, σ = 1.7) is shown in Figure 19.

[0398]

Table 27

[0399] Data indicate that prednisone treatment tended to improve healing, and that the complete formulations at both concentrations of GaRP (threonine / butyric acid / vitamin D), and bromelain showed a statistically significant improvement in histological scores regardless of the presence or absence of prednisone.

[0400] While not desiring to be bound by theory, the data provided herein indicates to the inventors that the use of GaRP components targeting the colon and bromelain / menthol targeting the small intestine can provide unexpectedly strong therapeutic benefits for the treatment of colitis. In certain embodiments, the invention may be as follows. [Embodiment 1] A composition comprising the following components. (e) Bromelain (f) One or more SCFAs or esters thereof (g) One or more amino acids selected from the group consisting of L-threonine, L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof (h) Vitamin D or a metabolite thereof [Embodiment 2] The composition according to Embodiment 1, wherein (c) comprises one or more amino acids selected from L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof. [Embodiment 3] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan, L-arginine, L-glutamine, and L-methionine. [Embodiment 4] The composition according to Embodiment 1, wherein (c) comprises L-arginine, L-glutamine, and L-methionine. [Embodiment 5] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan, L-glutamine, and L-methionine. [Embodiment 6] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan, L-arginine, and L-methionine. [Embodiment 7] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan, L-arginine, and L-glutamine. [Embodiment 8] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan and L-arginine. [Embodiment 9] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan and L-glutamine. [Embodiment 10] The composition according to Embodiment 1, wherein (c) comprises L-tryptophan and L-methionine. [Aspect 11] The composition according to Aspect 1, wherein (c) contains L-arginine and L-glutamine. [Aspect 12] The composition according to Aspect 1, wherein (c) contains L-arginine and L-methionine. [Aspect 13] The composition according to Aspect 1, wherein (c) contains L-glutamine and L-methionine. [Aspect 14] The composition according to Aspect 1, wherein (c) contains L-tryptophan, L-arginine, L-glutamine, or L-methionine. [Aspect 15] The composition according to Aspect 1, wherein (c) consists essentially of L-tryptophan, L-arginine, L-glutamine, or L-methionine. [Aspect 16] The composition according to Aspect 1, which contains (a), (b), L-threonine, and (d). [Aspect 17] The composition according to Aspect 16, which consists essentially of bromelain, SCFA, L-threonine, and vitamin D, or their metabolites. [Aspect 18] The composition according to any one of the preceding aspects, wherein one or more SCFAs, or their esters, are butyric acid, propionic acid, or acetic acid, or their salts. [Aspect 19] The composition according to any one of the preceding aspects, wherein one or more SCFAs, or their esters, are sodium butyrate, calcium butyrate, sodium propionate, or sodium acetate, preferably sodium butyrate. [Aspect 20] The composition according to any one of the preceding aspects, wherein one or more SCFAs, or their esters, are sodium butyrate. [Aspect 21] The composition according to any one of the preceding aspects, which consists essentially of bromelain, sodium butyrate, L-threonine, and vitamin D, or their metabolites. [Aspect 22] The composition according to any one of the preceding aspects, which is a unit dosage form containing about 50 to about 2500 mg, preferably about 250 to about 2250 mg, about 500 to about 2000 mg, about 750 to about 1750 mg, about 1000 mg to about 1600 mg, about 1300 mg to about 1550 mg, about 1400 to about 1500 mg, about 1420 to about 1460 mg, more preferably about 1440 mg of bromelain. [Aspect 23] The composition according to Aspect 22, containing about 1440 mg, preferably about 1030 mg, more preferably about 720 mg, still more preferably about 480 mg of bromelain. [Aspect 24] The composition according to any one of the preceding aspects, which is a unit dosage form containing about 45 to 1800 mg of SCFA, preferably about 75 to about 1650 mg, about 250 to about 1500 mg, about 500 to about 1400 mg, about 750 to about 1500 mg, about 1000 to about 1400 mg, about 1100 to about 1300 mg, more preferably about 1200 mg of SCFA. [Aspect 25] The composition according to Aspect 24, containing about 1200 mg, preferably about 600 mg, more preferably about 300 mg of SCFA. [Aspect 26] The composition according to Aspect 24 or 25, wherein the SCFA is sodium butyrate, sodium propionate, or sodium acetate, preferably sodium butyrate. [Aspect 27] The composition according to any one of the preceding aspects, which is a unit dosage form containing about 67.5 to about 2700 mg, preferably about 112.5 to about 2250 mg, about 250 to about 2000 mg, about 500 to about 1900 mg, about 900 to about 1800 mg, more preferably about 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. [Aspect 28] The composition according to Aspect 27, containing about 67.5 to about 2700 mg, preferably about 112.5 to about 2250 mg, about 250 to about 2000 mg, about 500 to about 1900 mg, about 900 to about 1800 mg, more preferably about 1800 mg of L-threonine. [Aspect 29] The composition according to aspect 28, comprising about 1800 mg, preferably about 900 mg, more preferably about 450 mg of L-threonine. [Aspect 30] The composition according to any one of the preceding aspects, wherein the composition is in a unit dosage form containing about 4 to about 150 μg, about 6.5 to about 140 μg, about 13 to about 130 μg, about 25 to about 120 μg, about 35 to about 110 μg, about 50 to about 105 μg, more preferably about 100 μg, still more preferably about 50 μg, still more preferably about 25 μg, still more preferably about 12.5 μg of vitamin D. [Aspect 31] The composition according to aspect 30, further comprising about 3 to about 150 mg, preferably about 6 to about 145 mg, about 12 to about 140 mg, about 24 to about 135 mg, 48 to about 130 mg, about 60 to about 125 mg, about 80 to about 120 mg, about 90 to about 110 mg, about 100 mg, more preferably about 96 mg of menthol. [Aspect 32] The composition according to aspect 31, comprising about 105 mg, preferably 100 mg, more preferably about 96 mg, still more preferably about 70 mg, still more preferably about 50 mg, still more preferably about 48 mg of menthol. [Aspect 33] A unit dosage form for use in the treatment or prevention of inflammatory bowel disease (IBD) or irritable bowel syndrome (IBS), comprising the composition according to any one of aspects 1 to 32 in an amount of about 1 to about 10 g. [Aspect 34] The unit dosage form according to aspect 33, which is an oral dosage form. [Aspect 35] The oral dosage form according to aspect 34, comprising the composition according to any one of aspects 1 to 32 in an amount of about 1 to about 8 g, about 1 to about 7 g, about 1 to about 6 g, about 1 to about 5 g, about 1 to about 4, about 1 to 3 g. [Aspect 36] An oral dosage form according to Aspect 34 or 35, comprising 50 to 2500 mg of bromelain, preferably 720 mg of bromelain, 45 to 1800 mg of sodium butyrate, preferably 600 mg, more preferably 300 mg of sodium butyrate, 67.5 to 2700 mg of L-threonine, preferably 900 mg, more preferably 450 mg of L-threonine, 4 to 150 μg of vitamin D, preferably 25 μg, more preferably 12.5 μg of vitamin D, and optionally 3 to 150 mg of menthol, preferably 105 mg, more preferably 70 mg, even more preferably 48 mg of menthol. [Aspect 37] An oral dosage form according to any one of Aspects 33 to 36, comprising one or more pills, tablets, minitabs, capsules, granules, particles, or microparticles, or any combination thereof. [Aspect 38] An oral dosage form according to any one of Aspects 33 to 37, comprising two different types of pills, tablets, minitabs, capsules, granules, particles, or microparticles, wherein each different type is formulated to release its contents in different parts of the gastrointestinal tract. [Aspect 39] An oral dosage form according to any one of Aspects 33 to 38, wherein the different parts of the gastrointestinal tract are the duodenum and the colon. [Aspect 40] The two different types of pills, tablets, minitabs, capsules, granules, particles, or microparticles are type A and type B pills, tablets, minitabs, capsules, granules, particles, or microparticles, and the type A and type B pills, tablets, minitabs, capsules, granules, particles, or microparticles each contain at least one component, preferably at least two components, of the composition according to any one of Aspects 1 to 32. An oral dosage form according to Aspect 38 or 39. [Aspect 41] An oral dosage form according to Aspect 40, wherein the type A and type B pills, tablets, minitabs, capsules, granules, particles, or microparticles each contain at least one different component from each other, preferably at least two different components from each other, of the composition. [Aspect 42] An oral dosage form according to Aspect 40 or 41, wherein the type A pills, tablets, minitabs, capsules, granules, particles, or microparticles contain bromelain. [Aspect 43] The oral dosage form according to any one of Aspects 40 to 42, wherein the type A pill, tablet, mini-tab, granule, capsule, particle, or fine particle contains about 400 to 1100 mg of bromelain, about 600 to 1000 mg, about 700 to 900 mg, about 750 to 850 mg, preferably about 800 mg of bromelain. [Aspect 44] The oral dosage form according to Aspect 40 or 41, wherein the type B pill, tablet, mini-tab, capsule, granule, particle, or fine particle contains L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, sodium butyrate, and vitamin D. [Aspect 45] The oral dosage form according to Aspect 44, wherein the type B pill, tablet, mini-tab, capsule, granule, particle, or fine particle contains L-threonine, sodium butyrate, and vitamin D, preferably vitamin D3. [Aspect 46] The oral dosage form according to any one of Aspects 40 to 45, wherein the type B pill, tablet, mini-tab, granule, capsule, particle, or fine particle contains about 67.5 to about 2700 mg, preferably about 112.5 to about 2250 mg, about 250 to about 2000 mg, about 500 to about 1900 mg, about 900 to about 1800 mg, preferably about 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. [Aspect 47] The oral dosage form according to Aspect 46, wherein the type B pill, tablet, mini-tab, granule, capsule, particle, or fine particle contains about 1800 mg, preferably about 900 mg, preferably about 450 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. [Aspect 48] The oral dosage form according to any one of Aspects 40 to 47, wherein the type B pill, tablet, mini-tab, granule, capsule, particle, or fine particle contains about 45 to 1800 mg of sodium butyrate, preferably about 75 to about 1650 mg, about 250 to about 1500 mg, about 500 to about 1400 mg, about 750 to about 1500 mg, about 1000 to about 1400 mg, about 1100 to about 1300 mg, more preferably about 1200 mg, and even more preferably about 600 mg of sodium butyrate. [Aspect 49] The oral dosage form according to any one of Aspects 40 to 48, wherein the type B pill, tablet, mini-tab, granule, capsule, particle, or fine particle contains about 4 to about 150 μg, about 6.5 to about 140 μg, about 13 to about 130 μg, about 25 to about 120 μg, about 35 to about 110 μg, about 50 to about 105 μg, preferably about 105 μg, more preferably about 100 μg, and even more preferably about 70 μg of vitamin D. [Aspect 50] A unit dosage form according to Aspect 33 or 34, comprising at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or fine particle, or any combination thereof, containing 1440 mg of bromelain and 288 mg of menthol targeting the small intestine, and at least one enteric-coated pill, tablet, mini-tab, capsule, granule, particle, or fine particle, or any combination thereof, containing 1200 mg of butyric acid (sodium salt), 1800 mg of L-threonine, and 0.025 mg of vitamin D3 in a dosage targeting the colon. [Aspect 51] A unit dosage form comprising type A and type B pills, tablets, mini-tabs, granules, capsules, particles, or fine particles as separate parts, wherein the type A pill, tablet, mini-tab, granule, capsule, particle, or fine particle consists essentially of 720 mg of bromelain and is formulated to release the bromelain into the target small intestine, The round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A optionally contain 144 mg of menthol, and the round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type B consist essentially of 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.0125 mg of vitamin D, and the sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D are formulated to be released into the lower intestine of a subject, in a unit dosage form. [Aspect 52] A unit dosage form comprising round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A and type B as separate portions, The round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A consist essentially of 1440 mg of bromelain, and are formulated to release the bromelain into the small intestine of a subject, and the round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A optionally contain 288 mg of menthol, The round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type B consist essentially of 1200 mg of sodium butyrate, 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and 0.025 mg of vitamin D, and the sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D are formulated to be released into the lower intestine of a subject, in a unit dosage form. [Aspect 53] A method of preventing, treating, or managing IBD, or IBS, comprising administering to a subject in need thereof a therapeutically effective amount of a composition of any one of aspects 1 to 32, or an oral dosage form of any one of aspects 34 to 49.

[0401] References Allen, e.a. (n.d.). Remington’s Pharmaceutical Sciences, 22nd Edition, ISBN978-0-85711-062-6, Amidon, S.B. (2016). Colon-Targeted Oral Drug Delivery Systems: Design Trends and Approaches. AAPS PharmSciTech, 16, No.4, 730-741. Bernstein CN, F.M. (2010). World Gastroenterology Organization Practice Guidelines for the diagnosis and management of IBD in 2010. Inflamm Bowel Dis., 16(1), 112-24. Borley, N. (2005). Jejunum and Ileum. Gray's Anatomy: Anatomical Basis of Clinical Practice, 39, 1167-1172. Cattaneo L, B.L. (1989). Stereogrammi-di-anatomia-dell’uomo: aspetti istologici, 3, Encyclopaedia of Pharmaceutical Technology. (n.d.). Hilgendorh C, L.H. (2000). JOURNAL OF PHARMACEUTICAL SCIENCES, 89, NO.1, 63-7. HM Martin, B.C. (2004). Gastroenterology, 127, 80-93. JR Bailey, C.P. (2011). PLos ONE, 6(10). Kong J, Z.Z. (2008). Novel role of the vitamin D receptor in maintaining the integrity of the intestinal mucosal barrier. Am J Physiol Gastrointest Liver Physiol., 294, G208-16. Ma X,F.P.(2012).J.Anim.Sci.,90,266-268. Mudie DM,e.a.(2014).Mol.Pharmaceutics 2014,11,3039-3047. Mynott TL,L.A.(1999).J Immunol,163(5),2568-75. Rieder F, K.T.-H.(2012).Results of the 2nd scientific workshop of the ECCO (III):basic mechanisms of intestinal healing.J Crohns Colitis.,6,373-85. Sender,e.a.(2016).PLOS Biology.doi:10.1371 / journal.pbio.1002533 August 19,2016 Teodori,U.(1987).Trattato di Medicina Interna.Roma:Societa Editrice Universo. Vamanu,E.(2018).Current Pharmaceutical Design (Vol.24). Weiss L,G.R.(1981).Istologia.Bologna:Zanichelli.

Claims

Claim 1 The following components: (a) Bromelain, (b) One or more SCFAs or salts or esters thereof, wherein the one or more SCFAs are propionic acid or butyric acid, (c) One or more amino acids selected from the group consisting of L-threonine, L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof, and (d) Vitamin D or alfacalcidol, A composition comprising the same. Claim 2 The composition according to claim 1, wherein (c) comprises one or more amino acids selected from the group consisting of L-tryptophan, L-arginine, L-glutamine, and L-methionine, and combinations thereof. Claim 3 The composition according to claim 1, wherein (c) comprises L-tryptophan, L-arginine, L-glutamine, and L-methionine. Claim 4 The composition according to claim 1, wherein (c) comprises L-arginine, L-glutamine, and L-methionine. Claim 5 The composition according to claim 1, wherein (c) comprises L-tryptophan, L-glutamine, and L-methionine. Claim 6 The composition according to claim 1, wherein (c) comprises L-tryptophan, L-arginine, and L-methionine. Claim 7 The composition according to claim 1, wherein (c) comprises L-tryptophan, L-arginine, and L-glutamine. Claim 8 The composition according to claim 1, wherein (c) comprises L-tryptophan and L-arginine. Claim 9 The composition according to claim 1, wherein (c) comprises L-tryptophan and L-glutamine. Claim 10 The composition according to claim 1, wherein (c) comprises L-tryptophan and L-methionine. Claim 11 The composition according to claim 1, wherein (c) comprises L-arginine and L-glutamine. Claim 12 The composition according to claim 1, wherein (c) comprises L-arginine and L-methionine. Claim 13 The composition according to claim 1, wherein (c) comprises L-glutamine and L-methionine. Claim 14 The composition according to claim 1, wherein (c) comprises L-tryptophan, L-arginine, L-glutamine, or L-methionine. Claim 15 The composition according to claim 1, wherein (c) consists of L-tryptophan, L-arginine, L-glutamine, or L-methionine and a carrier. Claim 16 The composition according to claim 1, comprising the aforesaid (a), the aforesaid (b), L-threonine, and the aforesaid (d).

17. The composition according to claim 16, comprising bromelain, SCFA, L-threonine, vitamin D or alfacalcidol, and a carrier.

18. The composition according to any one of claims 1 to 17, wherein one or more SCFAs are sodium butyrate, calcium butyrate, or sodium propionate.

19. The composition according to any one of claims 1 to 18, wherein one or more SCFAs are sodium butyrate.

20. The composition according to any one of claims 1 to 19, comprising bromelain, sodium butyrate, L-threonine, vitamin D or alfacalcidol, and a carrier.

21. The composition according to any one of claims 1 to 20, wherein the composition is a single-dose formulation containing 50 - 2500 mg, 250 - 2250 mg, 500 - 2000 mg, 750 - 1750 mg, 1000 mg - 1600 mg, 1300 mg - 1550 mg, 1400 - 1500 mg, 1420 - 1460 mg, or 1440 mg of bromelain.

22. The composition according to claim 21, containing 1440 mg, 1030 mg, 720 mg, or 480 mg of bromelain.

23. The composition according to any one of claims 1 to 22, wherein the composition is a single-dose formulation containing 45 - 1800 mg of SCFA, 75 - 1650 mg, 250 - 1500 mg, 500 - 1400 mg, 750 - 1500 mg, 1000 - 1400 mg, 1100 - 1300 mg, or 1200 mg of SCFA.

24. The composition according to claim 23, containing 1200 mg, 600 mg, or 300 mg of SCFA.

25. The composition according to claim 23 or 24, wherein the SCFA is sodium butyrate, calcium butyrate, or sodium propionate.

26. The composition according to any one of claims 1 to 25, wherein the composition is a single-dose formulation containing 67.5 - 2700 mg, 112.5 - 2250 mg, 250 - 2000 mg, 500 - 1900 mg, 900 - 1800 mg, or 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof. **Claim 27**: The composition according to claim 26, comprising 67.5 - 2700 mg, 112.5 - 2250 mg, 250 - 2000 mg, 500 - 1900 mg, 900 - 1800 mg, or 1800 mg of L-threonine. **Claim 28**: The composition according to claim 27, comprising 1800 mg, 900 mg, or 450 mg of L-threonine. **Claim 29** The composition according to any one of claims 1 - 28, wherein the composition is a single-dose formulation comprising 4 - 150 μg, 6.5 - 140 μg, 13 - 130 μg, 25 - 120 μg, 35 - 110 μg, 50 - 105 μg, 100 μg, 50 μg, 25 μg, or 12.5 μg of vitamin D. **Claim 30**: The composition according to claim 29, further comprising 3 - 150 mg, 6 - 145 mg, 12 - 140 mg, 24 - 135 mg, 48 - 130 mg, 60 - 125 mg, 80 - 120 mg, 90 - 110 mg, 100 mg, or 96 mg of menthol. **Claim 31**: The composition according to claim 30, comprising 105 mg, 100 mg, 96 mg, 70 mg, 50 mg, or 48 mg of menthol. **Claim 32**: A single-dose formulation for use in the treatment or prevention of inflammatory bowel disease (IBD) or irritable bowel syndrome (IBS), comprising 1 - 10 g of the composition according to any one of claims 1 - 31. **Claim 33** The single-dose formulation according to claim 32, for oral administration. **Claim 34**: The single-dose formulation according to claim 33, comprising 1 - 8 g, 1 - 7 g, 1 - 6 g, 1 - 5 g, 1 - 4 g, 1 - 3 g of the composition according to any one of claims 1 - 31. **Claim 35** The single-dose formulation according to claim 33 or 34, comprising 50 - 2500 mg of bromelain, 720 mg of bromelain, 45 - 1800 mg of sodium butyrate, 600 mg or 300 mg of sodium butyrate, 67.5 - 2700 mg of L-threonine, 900 mg or 450 mg of L-threonine, 4 - 150 μg of vitamin D, or 25 μg or 12.5 μg of vitamin D, and optionally 3 - 150 mg of menthol, or 105 mg, 70 mg or 48 mg of menthol. **Claim 36** The single-dose formulation according to any one of claims 32 - 35, comprising one or more pills, tablets, minitabs, capsules, granules, particles, or microparticles, or any combination thereof. **Claim 37** The single-dose formulation according to claim 36, comprising two different types of pills, tablets, mini-tabs, capsules, granules, particles, or microparticles, wherein each different type is formulated to release its contents in different parts of the gastrointestinal tract.

38. The single-dose formulation according to claim 37, wherein the different parts of the gastrointestinal tract are the duodenum and the colon.

39. The single-dose formulation according to claim 37 or 38, wherein the two different types of pills, tablets, mini-tabs, capsules, granules, particles, or microparticles are each contained in a single container as separate parts.

40. The single-dose formulation according to any one of claims 37 to 39, wherein the two different types of pills, tablets, mini-tabs, capsules, granules, particles, or microparticles are pills, tablets, mini-tabs, capsules, granules, particles, or microparticles of type A and type B, and the pills, tablets, mini-tabs, capsules, granules, particles, or microparticles of type A and type B each contain at least one component, or at least two components, of the composition according to any one of claims 1 to 31.

41. The single-dose formulation according to claim 40, wherein the pills, tablets, mini-tabs, capsules, granules, particles, or microparticles of type A and type B each contain at least one different component from each other, or at least two different components from each other, of the composition.

42. The single-dose formulation according to claim 40 or 41, wherein the pills, tablets, mini-tabs, capsules, granules, particles, or microparticles of type A contain bromelain.

43. The single-dose formulation according to any one of claims 40 to 42, wherein the pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A contain 400 - 1100 mg of bromelain, 600 - 1000 mg, 700 - 900 mg, 750 - 850 mg, or 800 mg of bromelain.

44. The single-dose formulation according to claim 40 or 41, wherein the pills, tablets, mini-tabs, capsules, granules, particles, or microparticles of type B contain L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, sodium butyrate, and vitamin D.

45. The type B pill, tablet, minitablet, capsule, granule, particle, or fine particle contains L-threonine, sodium butyrate, and vitamin D, or vitamin D ３ The single-dose formulation according to claim 44, comprising

46. The single-dose formulation according to any one of claims 40 to 45, wherein the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles contain 67.5 to 2700 mg, 112.5 to 2250 mg, 250 to 2000 mg, 500 to 1900 mg, 900 to 1800 mg, or 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof.

47. The single-dose formulation according to claim 46, wherein the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles contain 1800 mg, 900 mg, or 450 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof.

48. The single-dose formulation according to any one of claims 40 to 47, wherein the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles contain 45 to 1800 mg of sodium butyrate, 75 to 1650 mg, 250 to 1500 mg, 500 to 1400 mg, 750 to 1500 mg, 1000 to 1400 mg, 1100 to 1300 mg, 1200 mg, or 600 mg of sodium butyrate.

49. The single-dose formulation according to any one of claims 40 to 48, wherein the type B pills, tablets, minitabs, granules, capsules, particles, or microparticles contain 4 to 150 μg, 6.5 to 140 μg, 13 to 130 μg, 25 to 120 μg, 35 to 110 μg, 50 to 105 μg, 105 μg, 100 μg, or 70 μg of vitamin D.

50. At least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof, containing 1440 mg of bromelain and 288 mg of menthol targeting the small intestine, and 1200 mg of butyric acid (sodium salt), 1800 mg of L-threonine, and 0.025 mg of vitamin D at a dosage targeting the colon ３ A single-dose formulation according to claim 32 or 33, comprising at least one enteric-coated pill, tablet, minitablet, capsule, granule, particle, or microparticle, or any combination thereof

51. A single-dose formulation containing type A and type B pills, tablets, minitabs, granules, capsules, particles, or microparticles as separate parts, wherein the type A pills, tablets, minitabs, granules, capsules, particles, or microparticles consist of 720 mg of bromelain and a carrier, and the bromelain is formulated to be released into the small intestine of the subject. The round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A optionally contain 144 mg of menthol, and the round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type B consist of 600 mg of sodium butyrate, 900 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, 0.0125 mg of vitamin D or alfacalcidol, and a carrier, and the sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D or alfacalcidol are formulated to be released into the lower intestine of a subject, a single-dose formulation.

52. A single-dose formulation containing round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A and type B as separate portions, wherein the round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A consist of 1440 mg of bromelain and a carrier, and are formulated to release the bromelain into the small intestine of a subject, and the round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type A optionally contain 288 mg of menthol, wherein the round pills, tablets, mini-tabs, granules, capsules, particles, or microparticles of type B consist of 1200 mg of sodium butyrate, 1800 mg of L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, 0.025 mg of vitamin D or alfacalcidol, and a carrier, and the sodium butyrate, L-threonine, L-tryptophan, L-glutamine, L-arginine, or L-methionine, or any combination thereof, and vitamin D or alfacalcidol are formulated to be released into the lower intestine of a subject, a single-dose formulation.

53. Use of a therapeutically effective amount of the composition of any one of claims 1 to 31 or the single-dose formulation of any one of claims 33 to 49 in the manufacture of a medicament for preventing, treating, or managing IBD or IBS.

54. A formulation for preventing, treating, or managing IBD or IBS, comprising the composition according to any one of claims 1 to 31 or the single-dose formulation according to any one of claims 33 to 49.

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