Composition for Anti-Diabetes Using an Extract of Persicaria longiseta

Abstract

The present invention discloses an antidiabetic activity using a Polygonum hydropiper extract that inhibits α-glucosidase inhibitory activity and suppresses STZ-induced toxicity in rat insulinoma-derived pancreatic beta cells RIN-m5F.

Description

Technology Field

[0001] The present invention is a knotweed ( Persicaria longiseta This relates to an antidiabetic composition using an extract. Background Technology

[0003] Diabetes mellitus is a disease in which insulin, a glucose-regulating hormone secreted by the beta cells of the pancreas, is insufficient or does not function properly, causing blood sugar to accumulate in the blood without being used as energy, leading to hyperglycemia and the detection of sugar in the urine.

[0004] Diabetes is classified into insulin-dependent type 1 diabetes, which occurs when pancreatic beta cells are destroyed by an autoimmune response and fail to secrete a sufficient amount of insulin, and insulin-independent type 2 diabetes, which occurs when pancreatic beta cells initially increase to compensate for insulin resistance but eventually decrease and function declines (A balanced overview. Diabetes Care 1992, 15: 318-368; Diabetes Res Clin Pract 2010, 87(1): 4-14).

[0005] Type 1 diabetes is an insulin-dependent type and is divided into immune-mediated type 1 diabetes, in which pancreatic beta cells are destroyed by immunological mechanisms leading to impaired insulin production, and idiopathic type 1 diabetes, the cause of which has not yet been identified (Lancet, 2006, 367:847-858). Type 2 diabetes is a disease involving both insulin resistance and impaired insulin secretion, and is divided into insulin-resistant type 2 diabetes, in which insulin resistance plays a major role, and insulin-deficient type 2 diabetes, in which impaired insulin secretion plays a major role (BMJ, 2000, 321:405-412).

[0006] Currently, 90% of diabetes cases are Type 2 diabetes, and since Type 2 diabetes causes persistent hyperglycemia due to insulin resistance, reduced insulin action, and inefficient insulin production by the pancreas (Int J Appl Res Nat Prod, 2016, 9 : 33-38), if it becomes chronic, it is accompanied by very serious complications such as diabetic retinopathy, renal failure, hypertension, and arteriosclerosis. Therefore, type 2 diabetes is emerging as an urgent issue that needs to be addressed both medically and socioeconomically.

[0007] In particular, in the case of type 2 diabetes, chronic high glucose, high fat, and various inflammatory mediators secreted by adipose tissue destroy beta cells, preventing them from secreting insulin (Nature 2006, 444(7121):840-846). Therefore, since beta cells are damaged in both type 1 and type 2 diabetes, preventing the destruction of beta cells is known to be one of the important treatment strategies for diabetes.

[0008] Currently known treatments for type 2 diabetes include biguanides, which transport glucose to muscle cells and inhibit glucose synthesis in the liver; thiazolidinediones, which activate PPARγ involved in adipocyte differentiation; sulfonylureas, which indirectly induce insulin secretion; and α-glucosidase inhibitors, which inhibit the enzyme that produces glucose in the small intestine. However, diabetes treatment using these drugs is accompanied by many side effects (Diabetes, 2008, 57:737-745), so the World Health Organization (WHO) actively recommends the use of natural products with fewer side effects for diabetes (Journal of Ethnopharmacology, 2000, 73:461-470).

[0009] Meanwhile, streptozotocin (STZ) is known to be toxic to pancreatic beta cells, which are cells that synthesize insulin to maintain blood glucose homeostasis, so substances that inhibit STZ-induced toxicity to pancreatic beta cells could be candidate substances for the treatment of type 1 and type 2 diabetes (Molecular Medicine REPORTS 22: 1831-1838, 2020; Oxid Med Cell Longev. 2017; 2017: 7054272).

[0010] The present invention discloses an antidiabetic activity using a Polygonum hydropiper extract that inhibits α-glucosidase inhibitory activity and suppresses STZ-induced toxicity in rat insulinoma-derived pancreatic beta cells RIN-m5F. The problem to be solved

[0012] The objective of the present invention is to provide an antidiabetic composition using an extract of Polygonum hydropiper.

[0013] Other objects or specific objectives of the present invention will be presented below. means of solving the problem

[0015] The present invention was completed by confirming that, as confirmed in the following examples and experimental examples, the extract of Polygonum hydropiper has inhibitory activity against α-glucosidase and also has activity that inhibits the toxicity of STZ against rat insulinoma-derived pancreatic beta cells RIN-m5F.

[0016] Considering the foregoing, the present invention can be understood as an antidiabetic composition comprising Polygonum hydropiper extract as an active ingredient.

[0017] In this specification, "Polygonum hydropiper extract" refers to an extract obtained by leaching the stem, branch, leaf, root, whole plant, or mixture thereof of Polygonum hydropiper that is the subject of extraction using water, a lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, butanol, etc.), methylene chloride, ethylene, acetone, hexane, ether, chloroform, ethyl acetate, butyl acetate, N,N-dimethylformamide (DMF), dimethyl sulfoxide (DMSO), 1,3-butylene glycol, propylene glycol, or a mixture of these solvents (i.e., an extract soluble in the said extraction solvent), an extract obtained using a supercritical extraction solvent such as carbon dioxide or pentane, or a fraction obtained by fractionating the extract. The extraction method may be any method such as cold maceration, reflux, heating, ultrasonic radiation, or supercritical extraction, taking into account the polarity of the active substance, the degree of extraction, and the degree of preservation. The term "fractionated extract" includes fractions obtained by suspending an extract in a specific solvent and then mixing and settling it with a solvent of different polarity, and fractions obtained by adsorbing the crude extract onto a column packed with a polar or non-polar solid phase, such as silica gel, and then using a hydrophobic solvent, a hydrophilic solvent, or a mixture thereof as the mobile phase. Furthermore, the term "extract" includes concentrated liquid extracts or solid extracts from which the extraction solvent has been removed by methods such as freeze-drying, vacuum drying, hot air drying, or spray drying. Preferably, it refers to an extract obtained using water, ethanol, or a mixture thereof as the extraction solvent, and more preferably, an extract obtained using a mixture of water and ethanol (50% to 99%) as the extraction solvent.

[0018] Furthermore, in this specification, "anti-diabetic" means blood glucose lowering and the prevention, treatment, improvement, or delay of onset of diabetes. Specifically, it means the prevention, treatment, improvement, or delay of onset of pathological symptoms, such as hyperglycemia caused by the failure to produce insulin or insulin deficiency, or the detection of sugar in the urine.

[0019] Furthermore, in this specification, the term "diabetes" includes the insulin-dependent type 1 diabetes and insulin-independent type 2 diabetes as described above. Type 1 diabetes refers to diabetes that occurs as a result of damage to pancreatic beta cells due to other diseases, such as hyperthyroidism, hyperadrenocorticism, diabetes caused by hypergrowth hormone or hypercatecholaminism, and gestational diabetes.

[0020] In addition, in this specification, "active ingredient" means an ingredient that exhibits the intended activity alone or can exhibit activity together with a carrier that is inactive itself.

[0021] In the composition of the present invention, the active ingredient may be included in any amount (effective amount) depending on the use, formulation, etc., as long as it can exhibit an antidiabetic effect, etc., and the typical effective amount will be determined within the range of 0.001 weight % to 15 weight % based on the total weight of the composition. Here, "effective amount" refers to the amount of the active ingredient included in the composition of the present invention that can exhibit intended medical and pharmacological effects, such as an antidiabetic effect, when the composition of the present invention is administered to mammals, preferably humans, the subjects of application, for a period of administration as recommended by medical professionals, etc. Such effective amount may be determined experimentally within the ordinary capacity of a person skilled in the art.

[0022] In addition to the active ingredient, the composition of the present invention may additionally include any compound or natural extract whose safety has already been verified and whose activity has been confirmed in the art, for the purpose of enhancing or reinforcing the anti-diabetic effect, or for the convenience of taking or consuming by adding similar activities such as reducing body fat, improving blood cholesterol, or improving blood triglycerides.

[0023] These compounds or extracts include compounds or extracts listed in pharmacopoeias of each country (in Korea, the “Korean Pharmacopoeia”), health functional food codes of each country (in Korea, the “Standards and Specifications for Health Functional Foods” notified by the Ministry of Food and Drug Safety), compounds or extracts for which product approval has been obtained in accordance with the laws of each country regulating the manufacture and sale of pharmaceuticals (in Korea, the “Pharmaceutical Affairs Act”), and compounds or extracts for which functionality has been individually recognized in accordance with the laws of each country regulating the manufacture and sale of health functional foods (in Korea, the “Act on Health Functional Foods”).For example, as individually recognized ingredients under the Korean Code of Health Functional Foods or the Korean "Act on Health Functional Foods," ingredients with blood sugar regulating functional properties such as L-arabinose, nopal extract, cinnamon extract powder, guava leaf extract, indigestible maltodextrin, freeze-dried silkworm powder, hemp ethanol extract, banaba leaf extract, mulberry leaf extract, Seomoktae (mouse-eye bean) peptide complex, pine needle distillate concentrate, silk protein enzyme hydrolysate, albumin, ginseng hydrolysate concentrate, a mixture of Citrus aurantium and mulberry leaf extracts, fermented soybean extract, tagatose, defatted evening primrose seed extract, pinitol, complex extract of Rhodiola rosea, etc., hydroxypropylmethylcellulose, etc., or ingredients with body fat reduction functional properties such as Garcinia cambogia peel extract, conjugated linolenic acid (free fatty acid), conjugated linolenic acid (triglyceride), green tea extract, chitosan, green mate extract, green coffee bean extract, perilla leaf extract, soybean germ extract, etc. Compounds, alcohol extract powder of *Gynostemma pentaphyllum* leaves, lactoferrin (milk refined protein), lemon balm extract mixed powder, mate hot water extract, or complex extract of seaweed (Xantigen), etc., or green tea extract or spirulina, bamboo leaf extract, barley beta-glucan extract, Pu-erh tea extract, plant stanol ester, alcohol extract of *Gamtae*, flaxseed, aloe complex extract or aloe extract, etc., having blood cholesterol improvement function, or DHA concentrated oil, indigestible maltodextrin, bamboo leaf extract, vegetable oil diglyceride, refined squid oil, globin hydrolysate or refined sardine oil, ginkgo leaf extract, natto culture powder, natto culture, melon extract or cacao powder or red ginseng extract, etc., having blood circulation improvement function, would be such compounds or extracts.

[0024] One or more of these compounds or extracts may be included in the composition of the present invention together with the active ingredients.

[0025] The composition of the present invention can be understood as a food composition in specific embodiments.

[0026] The food composition of the present invention may be manufactured in any form, for example, as beverages such as tea, juice, carbonated beverages, and isotonic drinks; processed dairy products such as milk and yogurt; food products such as chewing gum, rice cakes, Korean confectionery, bread, confectionery, and noodles; and health functional food preparations such as tablets, capsules, pills, granules, liquids, powders, flakes, pastes, syrups, gels, jellies, and bars. Furthermore, the food composition of the present invention may take on any product classification in terms of legal and functional classification, as long as it complies with the regulations in effect at the time of manufacture and distribution. For example, it may be a health functional food under the Korean "Act on Health Functional Foods," or confectionery, legumes, teas, beverages, special dietary foods, etc., according to each food type in the Food Code (notified by the Ministry of Food and Drug Safety as "Standards and Specifications for Food") under the Korean "Food Sanitation Act."

[0027] The food composition of the present invention may include food additives in addition to its active ingredients. Food additives can generally be understood as substances added to, mixed with, or permeated into food during the manufacture, processing, or preservation of food; since they are consumed daily and over a long period of time along with food, their safety must be guaranteed. Food additive codes in accordance with national laws governing the manufacture and distribution of food (the "Food Sanitation Act" in Korea) restrictively define food additives with guaranteed safety in terms of composition or function. In the Korean Food Additive Code (Notification of the Ministry of Food and Drug Safety, "Standards and Specifications for Food Additives"), food additives are classified and defined in terms of composition into chemically synthesized products, natural additives, and mixed preparations; and in terms of function, these food additives are classified into sweeteners, flavorings, preservatives, emulsifiers, acidulants, thickeners, etc.

[0028] Sweeteners are used to impart a suitable sweetness to food, and both natural and synthetic sweeteners can be used in the food composition of the present invention. Preferably, natural sweeteners are used, and examples of natural sweeteners include corn syrup solids, honey, sucrose, fructose, lactose, maltose, etc.

[0029] Flavoring agents are used to improve taste or aroma, and both natural and synthetic types may be used. Preferably, natural ones are used. When natural ones are used, nutritional enhancement can be achieved in addition to flavor. Natural flavoring agents may be obtained from apples, lemons, citrus fruits, grapes, strawberries, peaches, etc., or from green tea leaves, Solomon's seal, bamboo leaves, cinnamon, chrysanthemum leaves, jasmine, etc. Additionally, those obtained from ginseng (red ginseng), bamboo shoots, aloe vera, ginkgo, etc., may be used. Natural flavoring agents may be liquid concentrates or solid extracts. In some cases, synthetic flavoring agents may be used, and synthetic flavoring agents may include esters, alcohols, aldehydes, terpenes, etc.

[0030] Calcium sorbate, sodium sorbate, potassium sorbate, calcium benzoate, sodium benzoate, potassium benzoate, EDTA (ethylenediaminetetraacetic acid), etc. may be used as preservatives, and acacia gum, carboxymethylcellulose, xanthan gum, pectin, etc. may be used as emulsifiers, and acetic acid, malic acid, fumaric acid, adipic acid, phosphoric acid, gluconic acid, tartaric acid, ascorbic acid, acetic acid, phosphoric acid, etc. may be used as acidifiers. Acidifiers may be added to the food composition to achieve an appropriate acidity for the purpose of inhibiting the growth of microorganisms in addition to enhancing flavor. As thickening agents, suspending agents, precipitating agents, gel-forming agents, puffing agents, etc. may be used.

[0031] In addition to the food additives described above, the food composition of the present invention may include physiologically active substances or minerals known in the art and whose safety as food additives is guaranteed, for the purpose of supplementing and reinforcing functionality and nutritional value.

[0032] Examples of such physiologically active substances include catechins contained in green tea, vitamins such as vitamin B1, vitamin C, vitamin E, and vitamin B12, tocopherol, dibenzoylthiamine, etc., and examples of minerals include calcium preparations such as calcium citrate, magnesium preparations such as magnesium stearate, iron preparations such as iron citrate, chromium chloride, potassium iodide, selenium, germanium, vanadium, zinc, etc.

[0033] The food composition of the present invention may include the food additives described above in an appropriate amount to achieve the purpose of their addition, depending on the product type.

[0034] Regarding other food additives that may be included in the food composition of the present invention, reference may be made to the food codes or food additive codes in accordance with the laws of each country.

[0035] The composition of the present invention may be understood as a pharmaceutical composition in other specific embodiments.

[0036] The pharmaceutical composition of the present invention may be prepared as an oral or parenteral formulation according to the route of administration by conventional methods known in the art, including a pharmaceutically acceptable carrier in addition to the active ingredient. Here, "pharmaceutically acceptable" means that it does not inhibit the activity of the active ingredient and does not possess toxicity beyond an acceptable level for the subject of application (prescription).

[0037] When the pharmaceutical composition of the present invention is prepared as an oral formulation, it may be prepared in the form of powder, granules, tablets, pills, coated tablets, capsules, liquids, gels, syrups, suspensions, wafers, etc., in accordance with methods known in the art together with a suitable carrier. Examples of suitable pharmaceutically acceptable carriers include sugars such as lactose, glucose, sucrose, dextrose, sorbitol, mannitol, and xylitol; starches such as corn starch, potato starch, and wheat starch; celluloses such as cellulose, methylcellulose, ethylcellulose, sodium carboxymethylcellulose, and hydroxypropylmethylcellulose; polyvinylpyrrolidone; water; methylhydroxybenzoate, propylhydroxybenzoate, magnesium stearate; mineral oil; malt; gelatin; talc; polyols; vegetable oils, etc. In the case of formulation, the formulation may include diluents and / or excipients such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants as needed.

[0038] When the pharmaceutical composition of the present invention is prepared as a parenteral formulation, it may be formulated in the form of eye drops, injections, transdermal administrations, nasal inhalers, or suppositories according to methods known in the art with a suitable carrier. When formulated as eye drops, suitable carriers may include sterile water, saline solution, isotonic solutions such as 5% dextrose, etc., and if necessary, benzalkonium chloride, methylparaben, ethylparaben, etc., may be added for preservative purposes. When formulated as an injection, suitable carriers may include sterile water, ethanol, polyols such as glycerol or propylene glycol, or mixtures thereof; preferably, Ringer's solution, PBS (phosphate buffered saline) containing triethanolamine, sterile water for injection, or isotonic solutions such as 5% dextrose may be used. When formulated as a transdermal administration, it can be formulated in the form of ointments, creams, lotions, gels, external solutions, pastes, liniments, aerosols, etc. In the case of a nasal inhaler, it can be formulated in the form of an aerosol spray using a suitable propellant such as dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, or carbon dioxide, and when formulated as a suppository, the base may be Witepsol, Tween 61, polyethylene glycols, cocoa starch, laurin starch, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene stearates, sorbitan fatty acid esters, etc.

[0039] Specific formulations of pharmaceutical compositions are known in the art, and reference may be made to literature such as [Remington's Pharmaceutical Sciences (19th ed., 1995)]. The said literature is to be considered part of this specification.

[0040] A preferred dosage of the pharmaceutical composition of the present invention may be in the range of 0.001 mg / kg to 10 g / kg per day, preferably 0.001 mg / kg to 1 g / kg, depending on the patient's condition, body weight, gender, age, severity of the patient, and route of administration. Administration may be administered once a day or divided into several doses. Such dosages shall not be construed as limiting the scope of the present invention in any aspect. Effects of the invention

[0042] As described above, according to the present invention, an antidiabetic composition using an extract of Polygonum hydropiper can be provided. The composition of the present invention can be commercialized into functional foods, such as health functional foods, or pharmaceuticals, such as natural product medicines. Brief explanation of the drawing

[0044] Figure 1 shows the results showing that the extract of Polygonum hydropiper has an inhibitory activity against STZ (streoptozotocin)-induced apoptosis in RIN-m5F cells, which are pancreatic beta cells derived from rat insulinoma. Specific details for implementing the invention

[0045] The present invention will be explained below with reference to examples and experimental examples. However, the scope of the present invention is not limited to these examples and experimental examples.

[0047] <Example> Preparation of Polygonum hydropiper extract

[0048] 500g of dried Polygonum hydropiper was cold-extracted twice with 5L of 50% ethanol for 48 hours. The extract was filtered using a filter (Advantech No. 1), and the filtrate was concentrated under reduced pressure at 40℃. The concentrated extract was freeze-dried to obtain a powdered extract.

[0049] <Experimental Example> Antidiabetic activity of Polygonum hydropiper extract

[0050] <Experimental Example 1> Evaluation of α-glucosidase inhibitory activity

[0051] α-glucosidase inhibitory activity was determined by adding 10 µl of 500 mU α-glucosidase to 75 µl of 0.1 M sodium phosphate buffer, followed by 5 µl of 10 mM ρ-nitro-phenyl-α-glucopyranoside (pNP-glycoside) as a substrate, and then adding the sample. The reaction was incubated at 37°C for 30 minutes, after which the degree of α-glucosidase inhibition was measured at 405 nm using an ELISA reader by measuring p-nitrophenol, a reaction product released from pNP-glycoside. The IC50 was calculated according to the treatment concentration of the sample. 50 The values ​​were calculated and shown in Table 1 below. Acarbose was used as the positive control.

[0052] α-glucosiase inhibitory activity Sample IC 50 (μg / ml) Polygonum hydropiper 36.80 acarbose 163.19

[0053] The results in Table 1 above are the IC50 of the Polygonum hydropiper extract. 50 This shows that it has very high α-glucosiase inhibitory activity at 36.80 ug / ml compared to the positive control, acarbose.

[0054] <Experimental Example 2> Evaluation of Cell Recovery Ability Against Apoptosis of Rin-m5F Cells Induced by STZ

[0055] Rin-m5F beta cell line cells were purchased from ATCC. Rin-m5F cells were placed in a 96-well plate at a rate of 5 x 10⁶ 4100 µl was dispensed into each well and incubated for 24 hours at 37°C under 5% CO2 conditions for stabilization. Subsequently, samples were pre-treated at different concentrations for 1 hour, followed by the addition of the samples and streptozotocin, and incubated for 24 hours at 37°C under 5% CO2 conditions. Afterward, the medium was removed, medium containing 5 mg / ml MTT solution was added, and the mixture was incubated in an incubator for 3 hours. Finally, the MTT reagent was removed, DMSO was added, and formazan was dissolved. Absorbance was then measured at 540 nm using an ELISA reader. N-Acetyl-L-cysteine ​​(NAC) was used as the positive control.

[0056] The results are shown in Figure 1. The results in Figure 1 show that the Polygonum hydropiper extract protects against STZ-induced cell damage in Rin-m5F cells in a concentration-dependent manner.

Claims

Claim 1 An antidiabetic composition comprising an extract of the whole plant of Polygonum hydropiper as an active ingredient. Claim 2 A composition according to claim 1, characterized in that the antidiabetic agent is an insulin-dependent type 1 diabetes or insulin-independent type 2 diabetes that is preventive or therapeutic active. Claim 3 A composition according to claim 1, wherein the extract is a water, ethanol, or mixed solvent extract of Polygonum hydropiper. Claim 4 A composition characterized in that, in any one of claims 1 to 3, the composition is a food composition. Claim 5 A composition characterized in that, in any one of claims 1 to 3, the composition is a pharmaceutical composition.

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