Composition containing cucumis melo leaf extract for preventing, alleviating or treating diabetes

Abstract

The present invention relates to an anti-diabetic effect of a Cucumis melo leaf extract, particularly an alcohol extract of Cucumis melo leaves. The Cucumis melo leaf extract of the present invention can provide an anti-diabetic effect through a blood glucose-lowering effect. In addition, the Cucumis melo leaf extract of the present invention has a high yield of about 21%, and thus can be high feasibility as an excellent anti-diabetic material.

Description

Composition for the prevention, improvement, or treatment of diabetes containing Korean melon leaf extract

[0001] The present application claims priority to Korean Patent Application No. 10-2024-0141227 filed on October 16, 2024, and the entire specification is a reference to the present application.

[0002]

[0003] The present invention relates to the antidiabetic effect of Korean melon leaf extract, particularly Korean melon leaf ethanol extract.

[0004]

[0005] The present invention was completed with the support of the Ministry of Science and ICT of the Republic of Korea by Project No. RS-2023-00274576 (2710003021) and Project No. 2021R1A6A1A03039211 (RS-2021-NR060118, 2340006719).

[0006]

[0007] The Korean melon (*Cucumis melo var. makuwa*) is a plant belonging to the Cucurbitaceae family. Its origin is believed to be India, and depending on the region of its spread, it developed into Asian varieties and Western varieties. Currently, it is mainly cultivated in Korea, China, Japan, and other regions. It is estimated that Korean melons have been cultivated in Korea since before the Silla Dynasty, and various native varieties were grown in different regions. As a fruit and vegetable with a sweet taste and unique aroma, its refreshing flavor suits Korean palates, and it has been widely consumed as a summer fruit since ancient times.

[0008] However, the leaves generated after harvesting Korean melons at farms are difficult to dispose of, and disposal incurs costs. Therefore, research is needed to utilize the leaves produced during cultivation as a high-value-added material.

[0009] Diabetes mellitus refers to a condition in which insulin, a glucose-regulating hormone secreted by the β-cells of the pancreas, is insufficient or does not function properly, causing blood sugar to accumulate in the blood without being utilized as energy, leading to hyperglycemia and the detection of sugar in the urine.

[0010] Diabetes can be classified into Type 1 diabetes, which is insulin-dependent diabetes in which no insulin is produced at all, and Type 2 diabetes, which is insulin-independent diabetes in which insulin is produced but relatively insufficient, depending on whether insulin is produced. In Korea, it is known that more than 90% of diabetes patients are Type 2 diabetes patients.

[0011] Type 1 diabetes develops because the beta cells of the pancreas are destroyed due to congenital factors, viral infection, or severe damage to the pancreas, resulting in the body producing little to no insulin. It is also called juvenile diabetes because it typically develops acutely during childhood, and it is treated by controlling blood sugar levels through the administration of insulin.

[0012] Type 2 diabetes primarily affects adults and is caused by insulin resistance resulting from impaired insulin secretion in response to baseline conditions and stimuli, increased endogenous glucose production in the liver, and reduced glucose utilization in peripheral tissues. In addition to genetic factors, causes include high-calorie and high-fat diets resulting from the Westernization of eating habits, lack of exercise, stress, obesity, and medication use. Treatment involves taking blood glucose-lowering agents along with dietary control and exercise.

[0013] There are three main types of hypoglycemic agents used to treat type 2 diabetes. One type stimulates pancreatic beta cells to promote insulin secretion and includes sulfonylurea and meglitinide drugs; another type improves insulin resistance and includes biguanide and thiazolidinedion drugs; and yet another type is an alpha-glucosidase inhibitor that inhibits the breakdown of sugar and inhibits the absorption of sugar in the intestines, such as acarbose and miglitol.

[0014] However, since the aforementioned drugs are chemically manufactured, there are issues with side effects such as impaired kidney or liver function, cardiovascular abnormalities, abdominal distension, and diarrhea, as well as patient tolerance. Therefore, there is a continuous demand for research on natural-derived diabetes treatments that have fewer side effects and superior efficacy.

[0015]

[0016] [Prior Art Literature]

[0017] [Patent Literature]

[0018] (Patent Document 1) KR 10-2014-0109742 (2014-08-22)

[0019]

[0020] As a result of diligent efforts to provide a material among naturally derived materials capable of providing excellent anti-diabetic effects, the inventors confirmed that an extract of Korean melon leaves can provide excellent anti-diabetic effects, such as a blood sugar-lowering effect, and completed the present invention.

[0021] Therefore, the objective of the present invention is to provide an antidiabetic effect of Korean melon leaf extract, particularly an alcoholic extract of Korean melon leaves.

[0022]

[0023] The present invention provides a pharmaceutical composition for the prevention or treatment of diabetes comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0024] According to a preferred embodiment of the present invention, the Korean melon leaf extract is,

[0025] i) Step of extracting Korean melon leaf powder;

[0026] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0027] iii) A step of volatilizing the solvent after the above re-extraction;

[0028] It was manufactured through.

[0029] According to a preferred embodiment of the present invention, the extract is an alcohol extract of Korean melon leaves.

[0030] According to a preferred embodiment of the present invention, the alcohol is 1 to 100% ethanol.

[0031] According to a preferred embodiment of the present invention, the composition reduces blood sugar.

[0032] In addition, the present invention provides a health functional food composition for preventing or improving diabetes comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0033] In addition, the present invention provides an antidiabetic feed additive comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0034] In addition, the present invention provides a blood glucose-lowering composition comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0035] In addition, the present invention comprises: i) a step of extracting Korean melon (Cucumis melo var. makuwa) leaf powder;

[0036] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0037] iii) A step of volatilizing the solvent after the above re-extraction;

[0038] A method for preparing a composition for preventing, improving, or treating diabetes comprising a Korean melon (Cucumis melo var. makuwa) leaf extract is provided.

[0039] In addition, the present invention provides a method for treating diabetes comprising the step of administering a Korean melon (Cucumis melo var. makuwa) leaf extract to an individual in need.

[0040] According to a preferred embodiment of the present invention, the extract is an alcohol extract of Korean melon leaves.

[0041] According to a preferred embodiment of the present invention, the alcohol is 1 to 100% ethanol.

[0042] According to a preferred embodiment of the present invention, the extract reduces blood sugar.

[0043]

[0044] The present invention will be described in more detail below.

[0045]

[0046] The term "prevention" in the present invention refers to any act that causes diabetes or diabetes-related diseases to be suppressed or their onset to be delayed due to the Korean melon leaf alcohol extract of the present invention.

[0047] The term "improvement" or "treatment" of the present invention refers to any act that causes parameters related to diabetes or diabetes-related diseases, such as the severity of symptoms, to improve or become beneficial due to the Korean melon leaf ethanol extract of the present invention.

[0048]

[0049] The present invention relates to the preventive, improving, or therapeutic effects of a Korean melon leaf alcohol extract on diabetes.

[0050] The inventors confirmed the effect of the Korean melon leaf ethanol extract on blood glucose reduction in C57BL / 6J mice with obesity induced by a high-fat diet. It was confirmed that the Korean melon leaf ethanol extract of the present invention exhibited an antidiabetic effect by reducing blood glucose in obese mice induced by a high-fat diet without reducing food intake. This meant that the Korean melon leaf ethanol extract exhibited an antidiabetic effect through blood glucose reduction.

[0051]

[0052] Accordingly, the present invention can provide a pharmaceutical composition for the prevention or treatment of diabetes comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0053] According to a preferred embodiment of the present invention, the Korean melon leaf extract is,

[0054] i) Step of extracting Korean melon leaf powder;

[0055] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0056] iii) A step of volatilizing the solvent after the above re-extraction;

[0057] It may be manufactured through.

[0058] In step i) above, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:10 to 1:20, and preferably, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:12 to 1:17.

[0059] In step ii) above, the weight ratio of the residue to the extraction solvent may be 1:5 to 1:15, and preferably, the weight ratio of the residue to the extraction solvent may be 1:7 to 1:12.

[0060] According to a preferred embodiment of the present invention, the extract may be an alcohol extract of Korean melon leaves.

[0061] According to a preferred embodiment of the present invention, the alcohol may be 1 to 100% ethanol. Preferably, the alcohol may be 50 to 80% ethanol.

[0062] According to a preferred embodiment of the present invention, the composition may reduce blood sugar.

[0063] The above diabetes may be type 2 diabetes.

[0064] The pharmaceutical composition of the present invention may be in various oral or parenteral formulations. When formulating the composition, it may be prepared using one or more buffers (e.g., saline solution or PBS), antioxidants, bacteriostatic agents, chelating agents (e.g., EDTA or glutathione), fillers, extenders, binders, adjuvants (e.g., aluminum hydroxide), suspending agents, thickening agents, wetting agents, disintegrants or surfactants, diluents or excipients.

[0065] Solid dosage forms for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid dosage forms are prepared by mixing at least one excipient with one or more compounds, for example, starch (including corn starch, wheat starch, rice starch, potato starch, etc.), calcium carbonate, sucrose, lactose, dextrose, sorbitol, mannitol, xylitol, erythritol, maltitol, cellulose, methyl cellulose, sodium carboxymethylcellulose and hydroxypropylmethylcellulose, or gelatin, etc. For example, tablets or sugar-coated tablets can be obtained by combining an active ingredient with a solid excipient, grinding the mixture, adding a suitable auxiliary agent, and processing it into a granular mixture.

[0066] In addition, lubricants such as magnesium stearate and talc are also used in addition to simple excipients. Liquid formulations for oral administration include suspensions, liquid formulations, emulsions, or syrups, and may contain various excipients, such as humectants, sweeteners, flavorings, or preservatives, in addition to commonly used simple diluents like water and liquid paraffin. Additionally, cross-linked polyvinylpyrrolidone, agar, alginic acid, or sodium alginate may be added as disintegrants, and may additionally include anticoagulants, flavorings, emulsifiers, solubilizers, dispersants, flavorings, antioxidants, packaging agents, pigments, and preservatives.

[0067] Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, or suppositories. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspensions. Witepsol, macrogol, Tween 61, cocoa paste, laurin paste, glycerol, gelatin, etc. may be used as bases for suppositories.

[0068] The composition of the present invention may be administered orally or parenterally, and when administered parenterally, it may be formulated into the abdominal cavity, rectum, vein, muscle, or subcutaneous according to methods known in the art.

[0069] The above-mentioned injectable must be sterilized and protected from contamination by microorganisms such as bacteria and fungi. Suitable carriers for the injectable may include, but are not limited to, water, ethanol, polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), mixtures thereof, and / or vegetable oils, as solvents or dispersion media. More preferably, suitable carriers may include Hanks' solution, Ringer's solution, PBS (phosphate buffered saline) containing triethanolamine, sterile water for injection, isotonic solutions such as 10% ethanol, 40% propylene glycol, and 5% dextrose. To protect the injectable from microbial contamination, various antimicrobial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, and thimerosal may be additionally included. Additionally, the injectable may, in most cases, further include isotonic agents such as sugars or sodium chloride.

[0070] The composition of the present invention is administered in a pharmaceutically effective amount. A pharmaceutically effective amount refers to an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level may be determined based on factors including the type and severity of the patient's disease, drug activity, sensitivity to the drug, time of administration, route of administration and elimination rate, duration of treatment, concurrently used drugs, and other factors well known in the medical field. The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered as a single or multiple doses. That is, the total effective amount of the composition of the present invention may be administered to the patient as a single dose, or administered via a fractionated treatment protocol involving long-term administration of multiple doses. It is important to administer an amount that obtains maximum effect with a minimum amount without side effects by considering all the aforementioned factors, and this can be easily determined by a person skilled in the art.

[0071] The dosage of the pharmaceutical composition of the present invention may vary depending on the patient's body weight, age, gender, health condition, diet, time of administration, method of administration, excretion rate, and severity of the disease.

[0072] The composition of the present invention can be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy, and biological response modifiers.

[0073]

[0074] In addition, the present invention may provide a health functional food composition for preventing or improving diabetes comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0075] According to a preferred embodiment of the present invention, the Korean melon leaf extract is,

[0076] i) Step of extracting Korean melon leaf powder;

[0077] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0078] iii) A step of volatilizing the solvent after the above re-extraction;

[0079] It may be manufactured through

[0080] In step i) above, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:10 to 1:20, and preferably, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:12 to 1:17.

[0081] In step ii) above, the weight ratio of the residue to the extraction solvent may be 1:5 to 1:15, and preferably, the weight ratio of the residue to the extraction solvent may be 1:7 to 1:12.

[0082] According to a preferred embodiment of the present invention, the extract may be an alcohol extract of Korean melon leaves.

[0083] According to a preferred embodiment of the present invention, the alcohol may be 1 to 100% ethanol. Preferably, the alcohol may be 50 to 80% ethanol.

[0084] According to a preferred embodiment of the present invention, the composition may reduce blood sugar.

[0085] The above diabetes may be type 2 diabetes.

[0086] The food composition according to the present invention can be prepared in various forms according to conventional methods known in the art. General foods may be prepared by adding the Korean melon leaf alcohol extract of the present invention to beverages (including alcoholic beverages), fruits and their processed foods (e.g., canned fruit, bottled fruit, jam, marmalade, etc.), fish, meat and its processed foods (e.g., ham, sausage, corned beef, etc.), breads and noodles (e.g., udon, buckwheat noodles, ramen, spaghetti, macaroni, etc.), fruit juices, various drinks, cookies, malt syrup, dairy products (e.g., butter, cheese, etc.), edible vegetable oils, margarine, vegetable proteins, retort foods, frozen foods, and various seasonings (e.g., soybean paste, soy sauce, sauces, etc.), although not limited thereto. Additionally, nutritional supplements may be prepared by adding the Korean melon leaf alcohol extract of the present invention to capsules, tablets, pills, etc. In addition, as a health functional food, it is not limited thereto, but for example, the Korean melon leaf alcohol extract of the present invention itself can be consumed by liquefying, granulating, encapsulating, and powdering it so that it can be prepared in the form of tea, juice, and drink for consumption (health beverage). In addition, to use the Korean melon leaf alcohol extract of the present invention as a food additive, it can be prepared and used in the form of a powder or concentrate. Furthermore, the Korean melon leaf alcohol extract of the present invention can be prepared in the form of a composition by mixing it with a known active ingredient known to have an effect of preventing or improving diabetes.

[0087] When the Korean melon leaf ethanol extract of the present invention is used as a health drink, the health drink composition may contain various flavoring agents or natural carbohydrates as additional ingredients, as in conventional beverages. The aforementioned natural carbohydrates may be monosaccharides such as glucose and fructose; disaccharides such as maltose and sucrose; polysaccharides such as dextrin and cyclodextrin; or sugar alcohols such as xylitol, sorbitol, and erythritol. As sweeteners, natural sweeteners such as thaumatin and stevia extract; or synthetic sweeteners such as saccharin and aspartame may be used. The proportion of the natural carbohydrates is generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 mL of the composition of the present invention.

[0088] In addition, the Korean melon leaf ethanol extract of the present invention may be included as an active ingredient in a health functional food composition for the prevention or improvement of diabetes. The amount is not specifically limited to an amount effective for achieving the prevention or improvement of diabetes, but it is preferable to be 0.01 to 100 weight% with respect to the total weight of the composition. The health functional food composition of the present invention may be prepared by mixing the Korean melon leaf ethanol extract with other active ingredients known to be effective against diabetes.

[0089] In addition to the above, the health functional food of the present invention may contain various nutritional supplements, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, or carbonating agents. Furthermore, the health functional food of the present invention may contain fruit pulp for the production of natural fruit juice, fruit juice beverages, or vegetable beverages. These ingredients may be used independently or in combination.

[0090]

[0091] In addition, the present invention can provide an antidiabetic feed additive comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0092] According to a preferred embodiment of the present invention, the Korean melon leaf extract is,

[0093] i) Step of extracting Korean melon leaf powder;

[0094] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0095] iii) A step of volatilizing the solvent after the above re-extraction;

[0096] It may be manufactured through.

[0097] In step i) above, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:10 to 1:20, and preferably, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:12 to 1:17.

[0098] In step ii) above, the weight ratio of the residue to the extraction solvent may be 1:5 to 1:15, and preferably, the weight ratio of the residue to the extraction solvent may be 1:7 to 1:12.

[0099] According to a preferred embodiment of the present invention, the extract may be an alcohol extract of Korean melon leaves.

[0100] According to a preferred embodiment of the present invention, the alcohol may be 1 to 100% ethanol. Preferably, the alcohol may be 50 to 80% ethanol.

[0101] According to a preferred embodiment of the present invention, the composition may reduce blood sugar.

[0102] The above diabetes may be type 2 diabetes.

[0103] The feed additive of the present invention may further include auxiliary components such as amino acids, inorganic salts, vitamins, antibiotics, antimicrobial substances, antioxidants, antifungal enzymes, and other microbial preparations in the form of live bacteria; grains, for example, ground or crushed wheat, oats, barley, corn, and rice; plant protein feeds, for example, those with rapeseed, soybeans, and sunflowers as main components; animal protein feeds, for example, blood meal, meat meal, bone meal, and fish meal; dry components consisting of sugars and dairy products, for example, various milk powders and whey powders; main components such as lipids, for example, animal fats and vegetable fats optionally liquefied by heating; and additives such as nutritional supplements, digestion and absorption enhancers, growth promoters, and disease preventive agents.

[0104] The feed additive of the present invention may be in the form of a powder or liquid formulation and may include an excipient for feed additives. Examples of excipients for feed additives include, but are not limited to, calcium carbonate, horse powder, zeolite, corn flour, or rice bran.

[0105] The feed additive of the present invention may be administered alone or in combination with other feed additives in an edible carrier. Additionally, the feed additive may be easily administered as a top dressing, by mixing directly into the feed, separately from the feed, as a separate oral formulation, or in combination with other ingredients. Typically, as is well known in the art, a single daily intake or a divided daily intake may be used.

[0106]

[0107] In addition, the present invention can provide a blood glucose-lowering composition comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

[0108] According to a preferred embodiment of the present invention, the Korean melon leaf extract is,

[0109] i) Step of extracting Korean melon leaf powder;

[0110] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0111] iii) A step of volatilizing the solvent after the above re-extraction;

[0112] It may be manufactured through.

[0113] In step i) above, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:10 to 1:20, and preferably, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:12 to 1:17.

[0114] In step ii) above, the weight ratio of the residue to the extraction solvent may be 1:5 to 1:15, and preferably, the weight ratio of the residue to the extraction solvent may be 1:7 to 1:12.

[0115] According to a preferred embodiment of the present invention, the extract may be an alcohol extract of Korean melon leaves.

[0116] According to a preferred embodiment of the present invention, the alcohol may be 1 to 100% ethanol. Preferably, the alcohol may be 50 to 80% ethanol.

[0117] According to a preferred embodiment of the present invention, the composition may be a pharmaceutical composition, a health functional food composition, or a quasi-drug composition.

[0118] Since the above pharmaceutical composition and health functional food composition are identical to the concept used in the pharmaceutical composition for preventing or treating diabetes or the health functional food composition for preventing or improving diabetes containing the above Korean melon (Cucumis melo var. makuwa) leaf extract, the description is replaced by the description thereof.

[0119] The term "quasi-drug" in the present invention refers to articles used for the purpose of diagnosing, treating, improving, alleviating, managing, or preventing diseases of humans or animals, and having a milder effect than pharmaceuticals. For example, according to the Pharmaceutical Affairs Act, quasi-drugs are defined as articles excluding those used for pharmaceutical purposes, and include products used for the treatment or prevention of diseases of humans or animals, and products that have a mild effect on the human body or do not act directly on it.

[0120] When the Korean melon leaf extract of the present invention is used as an additive to a quasi-drug, the composition may be added as is or used together with other quasi-drug ingredients, and may be used appropriately according to conventional methods. The mixing amount of the active ingredients may be appropriately determined according to the purpose of use (prevention, health, or therapeutic treatment).

[0121]

[0122] In addition, the present invention comprises: i) a step of extracting Korean melon (Cucumis melo var. makuwa) leaf powder;

[0123] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0124] iii) A step of volatilizing the solvent after the above re-extraction;

[0125] A method for preparing a composition for preventing, improving, or treating diabetes comprising a Korean melon (Cucumis melo var. makuwa) leaf extract containing [the ingredient] can be provided.

[0126] In step i) above, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:10 to 1:20, and preferably, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:12 to 1:17.

[0127] In step ii) above, the weight ratio of the residue to the extraction solvent may be 1:5 to 1:15, and preferably, the weight ratio of the residue to the extraction solvent may be 1:7 to 1:12.

[0128] According to a preferred embodiment of the present invention, the extract may be an alcohol extract of Korean melon leaves.

[0129] According to a preferred embodiment of the present invention, the alcohol may be 1 to 100% ethanol. Preferably, the alcohol may be 50 to 80% ethanol.

[0130] According to a preferred embodiment of the present invention, the extract may reduce blood sugar.

[0131] The above diabetes may be type 2 diabetes.

[0132]

[0133] In addition, the present invention may provide a method for treating diabetes comprising the step of administering a Korean melon (Cucumis melo var. makuwa) leaf extract to an individual in need.

[0134] According to a preferred embodiment of the present invention, the Korean melon leaf extract is,

[0135] i) Step of extracting Korean melon leaf powder;

[0136] ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and

[0137] iii) A step of volatilizing the solvent after the above re-extraction;

[0138] It may be manufactured through.

[0139] In step i) above, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:10 to 1:20, and preferably, the weight ratio of Korean melon leaf powder to extraction solvent may be 1:12 to 1:17.

[0140] In step ii) above, the weight ratio of the residue to the extraction solvent may be 1:5 to 1:15, and preferably, the weight ratio of the residue to the extraction solvent may be 1:7 to 1:12.

[0141] According to a preferred embodiment of the present invention, the extract may be an alcohol extract of Korean melon leaves.

[0142] According to a preferred embodiment of the present invention, the alcohol may be 1 to 100% ethanol. Preferably, the alcohol may be 50 to 80% ethanol.

[0143] The above individuals may include normal individuals requiring blood glucose control, improvement of insulin sensitivity, or normalization of glucose metabolism, as well as individuals who have already developed or may develop diabetes. The above individuals may refer to all mammals, including but not limited to humans, dogs, cattle, horses, rabbits, mice, rats, or chickens.

[0144] Since the above administration is the same as the administration in the above pharmaceutical composition, the description is replaced by the description thereof.

[0145] According to a preferred embodiment of the present invention, the composition may reduce blood sugar.

[0146] The above diabetes may be type 2 diabetes.

[0147]

[0148] The Korean melon leaf extract of the present invention can provide an anti-diabetic effect through a blood sugar-lowering effect. In addition, the Korean melon leaf extract of the present invention has a high yield of about 21%, so it can be economically viable as an excellent anti-diabetic material.

[0149]

[0150] Figure 1 shows the fasting blood glucose-reducing effect of Korean melon leaf ethanol extract (CHY).

[0151] Figure 2 shows the results of a glucose tolerance test of Korean melon leaf ethanol extract (CHY).

[0152] Figure 3 shows the liver lipid metabolism improvement and antioxidant effects of Korean melon leaf ethanol extract (CHY).

[0153] Figure 4 shows the serum analysis and metabolic improvement effects following treatment with Korean melon leaf ethanol extract (CHY).

[0154] Figures 5a and 5b show the results of RNA sequencing analysis for the analysis of the molecular mechanism following treatment with Korean melon leaf ethanol extract (CHY).

[0155] Figures 6a to 6f show the results of RNA sequencing analysis following treatment with Korean melon leaf ethanol extract (CHY), and represent the results of differential expression gene analysis.

[0156] Figure 7 shows the results of RNA sequencing analysis following treatment with Korean melon leaf ethanol extract (CHY), indicating up- or down-regulated signaling pathways.

[0157] Figures 8a and 8b show the results of RNA sequencing analysis following treatment with Korean melon leaf ethanol extract (CHY), representing comprehensive network analysis results.

[0158] Figures 9a and 9b show the results of analyzing a single substance of Korean melon leaf ethanol extract (CHY).

[0159]

[0160] The present invention will be described in more detail below through examples. These examples are intended solely to illustrate the present invention, and it is obvious to those skilled in the art that the scope of the present invention is not to be interpreted as being limited by these examples.

[0161]

[0162] [Example 1]

[0163] Experiment preparation

[0164] <1-1> Preparation of Korean Melon Leaf Extract

[0165] After separating leaves from completely dried Korean melon stems, 400 g of finely ground Korean melon leaf powder was immersed in 6 L of 70% ethanol (weight ratio of leaf powder to solvent = 1:15) and extracted using a reflux extraction device at 76°C for 15 hours. After extraction, the extract and residue were separated; the extract was filtered and stored at 4°C, while the residue was immersed again in 4 L of 70% ethanol (weight ratio of residue to solvent = 1:10) and re-extracted under the same conditions, followed by filtration. The ethanol from the filtered extract was completely evaporated using a rotary vacuum evaporator, and the extract was freeze-dried for 72 hours to be used as a sample. The freeze-dried extract in powder form was stored at -20°C, yielding 84.88 g of the Korean melon leaf ethanol extract, representing a yield of approximately 21%.

[0166]

[0167] <1-2> Experimental Animals and Experimental Methods

[0168] Male C57BL / 6J mice (4 weeks old) were supplied by Hana Biotech and used in the experiment after acclimatizing them in the animal laboratory for one week. During the acclimatization and experimental period, sufficient feed and water were supplied, and the temperature was maintained at 25±2℃, humidity at 50±10%, and a 12-hour light-dark cycle. The experimental groups consisted of a standard diet group (CON), a 60% kcal high-fat diet group (HFD), a high-fat diet + 0.4% Korean melon leaf ethanol extract group (CHY 0.4%), a high-fat diet + 0.8% Korean melon leaf ethanol extract group (CHY 0.8%), and a 0.8% Garcinia extract group (GRC 0.8%, positive control), with 10 mice assigned to each group. The Korean melon leaf ethanol extract and Garcinia extract were mixed into the high-fat diet at 0.4% and 0.8%, respectively, and then fed to the mice after sterilization. Based on a daily feed intake (approx. 2.5 g), the daily intake of Korean melon leaf ethanol extract was 10 mg and 20 mg, respectively. Male C57BL / 6J mice were sacrificed using isoflurane after a total of 18 weeks of experimentation, from 4–5 weeks of age to 21–22 weeks of age. Blood, subcutaneous fat, epididymal fat, liver, small intestine, and muscle were collected upon sacrifice. The blood was centrifuged at 3000 rpm for 20 minutes to separate serum, and the blood was stored at -70°C. The subcutaneous fat, epididymal fat, liver, small intestine, and muscle were flash-frozen in liquid nitrogen and stored at -70°C.

[0169]

[0170] [Example 2]

[0171] Confirmation of fasting blood sugar improvement effect

[0172] We intended to confirm the effect of the Korean melon leaf ethanol extract (CHY) prepared in <Example 1-1> above on improving fasting blood glucose in mice by administering it to C57BL / 6J mice that had been induced to obesity by a high-fat diet.

[0173] Specifically, after fasting for 16 hours, blood was drawn through the tail vein to measure fasting blood glucose and blood glucose levels were measured.

[0174] As a result, as shown in [Figure 1], it was confirmed that the alcohol extract of Korean melon leaves reduced fasting blood glucose.

[0175]

[0176] [Example 3]

[0177] Confirmation of glucose tolerance effect

[0178] We intended to confirm the effect of improving glucose tolerance in mice by administering the Korean melon leaf ethanol extract (CHY) prepared in <Example 1-1> above to C57BL / 6J mice that had obesity induced by a high-fat diet.

[0179] Specifically, after fasting for 16 hours, blood was collected via the tail vein to measure fasting blood glucose, and subsequently, 2 g of glucose per kg of mouse body weight was orally administered. Blood glucose was measured by collecting blood via the tail vein at intervals of 15, 30, 60, 90, and 120 minutes after glucose administration.

[0180] As a result, as shown in [Figure 2], it was confirmed that the alcohol extract of Korean melon leaves reduced fasting blood glucose.

[0181]

[0182] [Example 4]

[0183] Improvement of liver lipid metabolism and antioxidant effects of Korean melon leaf extract

[0184] <4-1> Analysis of Liver Triglycerides (TG) and Liver Total Cholesterol (TC)

[0185] Since blood glucose control is closely related to liver health, 100 mg of liver tissue was homogenized in 1 mL of DPBS for lipid analysis of liver tissue. Then, 200 μL of the homogenized solution was mixed with 800 μL of chloroform (2:1, v:v) to separate the lipids. After adding an equal volume of distilled water, the mixture was centrifuged at 1000 g at 4°C for 5 minutes to collect the lower chloroform layer, which was then dried in a fume hood for 24 hours. The dried lipids were resuspended in 100 μL of methanol (9:1, v:v), and the concentrations of TG and TC were measured using a microplate reader (AMR-100, Allsheng, Hangzhou, China) according to the usage instructions of each analysis kit.

[0186] As a result, the high-fat diet (HFD) group showed significantly increased levels of liver triglycerides (TG) and total cholesterol (TC) compared to the normal diet (CON) group, whereas these lipid indicators were significantly reduced in the CHY 0.4% and 0.8% treatment groups compared to the HFD group (Fig. 3 A, B). In particular, the CHY 0.4% treatment group showed an effect of reducing triglyceride and total cholesterol levels by 55.86% and 37.76%, respectively. This suggests that CHY can contribute to the improvement of diabetes by regulating liver lipid metabolism.

[0187]

[0188] <4-2> ROS and ONOO - measurement

[0189] ROS and ONOO generation were measured using fluorescent probes. For ROS analysis, 10 μL of homogenized tissue sample was mixed with 190 μL of 50 mM phosphate buffer (PB), and the reaction was carried out for 30 minutes in a light-blocked environment using 0.125 μM DCFDA solution. Fluorescence intensity was recorded 7 times at 5-minute intervals over 30 minutes in a 96-well plate. For ONOO analysis, 10 μL of homogenized liver sample was mixed with a mixture consisting of 175.8 μL of Rhodamine, 4 μL of DTPA, and 0.2 μL of DHR123, and measurements were taken 7 times over 30 minutes at an excitation wavelength of 485 nm and an emission wavelength of 535 nm.

[0190] As a result, in the CHY treatment group, liver ROS and ONOO were lower compared to the HFD group. - The values ​​were significantly reduced (Fig. 3 C, D). This suggests that CHY has an oxidative stress reduction effect, which may provide metabolic benefits such as improved glucose metabolism through liver cell protection.

[0191]

[0192] [Example 5]

[0193] Serum Analysis and Metabolic Improvement Effects Following Treatment with Korean Melon Leaf Extract

[0194] Serum analysis was performed using a free fatty acid (NEFA) assay kit (NEFA assay reagent, Wako Pure Chemical Industries, Osaka, Japan) and an LDL-cholesterol assay kit (LDL-Cholesterol assay reagent, Wako Pure Chemical Industries, Osaka, Japan). Kits for the analysis of the remaining serum components were purchased from Asan Pharmaceutical (Seoul, Korea) for the experiment. Additionally, alkaline phosphatase (ALP) and lipase (LIPA) levels were measured using Exdia PT10V (Precision Biosensor, Korea).

[0195] Serum analysis results showed that cholesterol levels decreased by an average of 14.72% in the CHY 0.4% treatment group and by 9.08% in the CHY 0.8% treatment group compared to the high-fat diet (HFD) group (p < 0.05, Fig. 4A). However, no significant differences were observed between the low-fat diet (CON) group and the HFD group regarding triglyceride (Fig. 4B), blood glucose (Fig. 4C), and free fatty acid (NEFA) levels (Fig. 4D), indicating that no further changes were observed in the CHY treatment group. ALP levels showed a significant difference, decreasing by 15.10% in the CHY 0.4% treatment group compared to the HFD group (p < 0.05, Fig. 4E), while LIPASE levels increased in the HFD group, decreasing by an average of 14.16% in the CHY 0.4% treatment group (Fig. 4F). These results suggest that CHY treatment is effective in reducing blood cholesterol, ALP, and LIPASE levels, and that metabolic improvement and liver health protection effects were particularly evident in the 0.4% CHY treatment group. This implies that CHY may have an effect on improving diabetes through the regulation of serum lipids and liver metabolism.

[0196]

[0197] [Example 6]

[0198] Molecular mechanism analysis and gene expression analysis

[0199] <6-1> Analysis of Molecular Mechanisms via RNA Sequencing

[0200] To investigate whether the blood glucose-improving effect of CHY treatment is associated with the reduction of metabolic disorders, fatty liver disease, and hepatoprotective effects, RNA-Seq (transcriptome analysis) was performed after treating primary hepatocytes with lipid accumulation induced by fatty acid treatment with CHY. During the RNA purification process, only RNA with a RIN value of 7.0 or higher was used for RNA library construction. Libraries were independently constructed using the Illumina TruSeq Stranded mRNA Sample Prep Kit (Illumina, Inc., San Diego, CA, USA, #RS-122-2101) with 1 μg of total RNA per sample. The constructed libraries were quantified using the KAPA Library Quantification Kit for Illumina Sequencing Platforms (KAPA BIOSYSTEMS, #KK4854), and quality control was performed using TapeStation D1000 ScreenTape (Agilent Technologies, #5067-5582). The indexed library was sequenced using the Illumina NovaSeqX platform (Illumina, Inc., San Diego, CA, USA) in a paired-end (2 × 100 bp) manner. Sequencing reads were mapped to a reference genome using HISAT2 (version 2.2.1), and then read counts were calculated using featureCounts (version subread-2.0.8).

[0201]

[0202] <6-2> Analysis of Differentially Expressed Genes

[0203] Differential expression gene analysis was performed using TMM normalization with the edgeR package (version 4.0.16). The No Replicate method described in the “Analysis of Sequence Read Count Data User's Guide” was used for TMM normalization, and a square root variance value of 0.1 was applied. Differential expression genes were filtered based on log2 fold change ≥ 1, log2 fold change ≤ -1, and FDR < 0.01, and the filtered genes were used for Gene Ontology analysis.

[0204]

[0205] <6-3> Results

[0206] A total of 17,415 genes showed statistically significant differential expression (p < 0.01), and the CHY treatment group was found to have 2,541 upregulated genes and 2,649 downregulated genes compared to the FFA alone group.

[0207] An analysis was performed using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database to identify the associated signaling pathways among the upregulated and downregulated genes. The upregulated genes were found to be primarily involved in pathways related to maintaining cell structure and tissue repair. The motor protein pathway plays a crucial role in cell migration, intracellular material transport, and maintaining structural stability, exhibiting particularly high activity. Furthermore, the upregulation of the ECM-receptor interaction and focal adhesion pathways indicates improved ECM remodeling, cell adhesion, and intracellular signaling, suggesting that signaling systems essential for tissue repair and cell survival were activated by CHY processing (Fig. 5a). Conversely, signaling systems associated with inflammation and oxidative stress were downregulated. The cytokine-cytokine receptor interaction pathway and IL-17 signaling pathway reflect reduced activation of inflammatory cytokine signaling, while the downregulation of the non-alcoholic fatty liver disease pathway indicates an improvement in overall signaling associated with fatty liver disease (Fig. 5b).

[0208] To gain a deeper understanding of functional changes, Gene Ontology (GO) analysis was performed on differentially expressed genes (DEGs) in the categories of biological processes (BP), molecular functions (MF), and cellular components (CC) (Figs. 6a–6f). The BP analysis showed significant increases in pathways related to membrane potential (GO:0042391), lipid transport (GO:0006869), ECM organization (GO:0030198), glucose homeostasis (GO:0042593), hormone metabolism (GO:0042445), and regeneration (GO:0031099) (Fig. 7A). These results suggest that CHY treatment promotes processes involved in lipid metabolism activation, energy homeostasis restoration, cell repair, and regeneration. On the other hand, pathways related to lipopolysaccharide response (GO:0032496), cytokine-mediated signaling pathway (GO:0019221), regulation of inflammatory response (GO:0050727), leukocyte migration (GO:0050900), myeloid leukocyte migration (GO:0097529), and chemotaxis (GO:0006935) were significantly inhibited (Fig. 7 B). This indicates a reduction in inflammation and immune cell tissue infiltration, reflecting the anti-inflammatory effect of CHY treatment.

[0209] CHY treatment in primary hepatocytes with induced lipid accumulation exhibited unique gene expression patterns related to lipid transport, glucose homeostasis, cytokine signaling, and the regulation of inflammatory responses. Accordingly, a comprehensive network analysis was performed to identify detailed genetic changes, and specifically, genes associated with lipid transport, inflammatory responses, and major regulatory pathways were analyzed in CHY-treated hepatocytes, as well as genes related to the improvement of metabolic disorders and fatty liver disease.

[0210] In the GO analysis, the "regulation of membrane potential" category included 21 genes, and significant changes in expression were observed in Cacna1g, Chrne, Edn1, Flna, Gabrr2, and Gja5. In the "regeneration" category, significant changes were identified in Adm, Dysf, Igf1r, and Tnc. In the "lipid transport" category (containing 19 genes), high changes in expression were observed in Abca6 and Abca9. Additionally, in the "hormone metabolic process" category, Adm, Corin, Duox2, and Igf1r showed high changes in expression, and in the "glucose homeostasis" category, significant changes in expression were observed in Csrp3, Hk1, Igf1r, Myt1, and Myh9. Finally, in the extracellular matrix organization category, prominent changes in expression were observed in Axin2, Ccn2, Crispld2, Ihh, and Loxl4 (Fig. 8a).

[0211] Regarding downregulated genes in the GO analysis, the "response to lipopolysaccharide" category showed a significant decrease in the genes Cd14, Tlr4, Il1a, Il1b, and Tnf, indicating a reduction in the inflammatory response. In the "regulation of inflammatory response" category, significant inhibition of Il1b, Il6, Tnf, Ccl2, and Cxcl2 was observed, indicating a decrease in cytokine signaling and immune activation. In the "myeloid leukocyte migration" category, decreased expression of Ccr1, Ccr2, Cxcl1, Cxcl2, and Il1b suggests reduced tissue infiltration by myeloid immune cells. Similarly, in the "leukocyte migration" category, the genes Ccl2, Ccr5, Itgam, Cd44, and Sele were inhibited, also demonstrating a decrease in the migration and tissue infiltration of various immune cells. In addition, in the "cytokine-mediated signaling pathway" category, the genes Il1b, Il6, Tnf, Ccl2, and Cxcl1 were significantly reduced, confirming that pro-inflammatory cytokine signaling was suppressed. Finally, in the "chemotaxis" category, the expression of Ccl2, Ccl5, Cxcl1, Cxcl2, and Il8 was reduced, showing that immune cell motility and inflammatory responses were decreased (Fig. 8b).

[0212] In summary, mechanistic analysis via RNA sequencing suggests that CHY treatment is closely associated with the inhibition of inflammation and oxidative stress signaling systems in hepatocytes, the activation of cellular metabolism, and the regulation of signaling pathways related to hepatocyte structural recovery. Therefore, the improvement of blood glucose indicators by CHY may be associated with the maintenance of hepatic homeostasis.

[0213]

[0214] [Example 7]

[0215] Single-substance analysis of Korean melon leaf extract

[0216] <7-1> Sample Preparation for UPLC-MS Analysis

[0217] The ethanol extract (1 g) of Korean melon leaves of Example <1-1> was precipitated by adding 10 times the amount of a solution containing 1% formic acid to methanol:water (80:20, v / v). This mixture was sonicated for 30 minutes, stored overnight at 4°C, and then centrifuged at 12,000 RPM for 10 minutes. The finally obtained supernatant (120 μL) was recovered and stored at 4°C until MS analysis.

[0218]

[0219] <7-2> Ultra-high Performance Liquid Chromatography (UPLC)

[0220] Chromatographic analysis was performed using a UPLC system (Bruker, Massachusetts, USA). Precipitated CML samples were injected into a BEH C18 column (100 mm × 2.1 mm, 1.7 μm, 1 / pk; Waters, Massachusetts, USA), and the column temperature was maintained at 40°C. The flow rate was set to 300 μL / min. The mobile phase consisted of water containing 0.1% formic acid (Phase A) and acetonitrile containing 0.1% formic acid (Phase B). The UPLC automatic sampler was maintained at 10°C, and the injection volume for each sample was 1 μL.

[0221]

[0222] <7-3> Phytochemical Analysis Using UPLC / Q-TOF Mass Spectrometry

[0223] After separation by UPLC, mass spectrometry was performed using a Q-TOF Premier (Bruker, Massachusetts, USA) with an electron spray ionization (ESI) source (Waters, Massachusetts, USA). The settings in negative mode were as follows: capillary voltage: 2 kV, cone voltage: 40 V, source temperature: 100 ℃, desolvate temperature: 250 ℃, desolvate gas flow: 600 L / h, cone gas flow: 50 L / h, and data acquisition was performed in the range of 100–1300 m / z.

[0224]

[0225] <7-4> Data Processing and Statistical Analysis

[0226] Screened data were included in the analysis after correcting for personal bias using QC and blank data. Orthogonal Partial Least Squares Discriminant Analysis (OPLS-DA), Projected Weight Variable (VIP), and coefficient analysis by VIP score were performed using EZinfo 3.0 software controlled by Progenesis QI 3.0 software (Waters, Massachusetts, USA).

[0227]

[0228] <7-5> Results

[0229] LC-MS analysis was performed to identify plant compounds contained in the CHY ethanol extract (Figs. 9a, 9b). The analysis was conducted in negative ionization mode, and a total of 438 compounds were detected. Among these, the top 20 compounds were selected based on low raw abundance values ​​and CV% values. Detailed information such as the retention time (R. Time), base m / z values, and compound names of each compound is provided in [Table 1] and [Table 2] below.

[0230]

[0231]

[0232]

[0233]

[0234]

[0235] While most of the identified compounds have not yet been clearly characterized or are under-studied, Soyacerebroside (retention time: 26.73, baseline m / z: 712.5358) is a noteworthy substance as a glycosphingolipid reported to possess anti-inflammatory properties. This compound plays a crucial role in maintaining cell membrane structure and promoting signal transduction, and it is likely to possess functions that regulate lipid metabolism and signaling pathways. Furthermore, as changes in sphingolipid metabolism are associated with insulin resistance and obesity, the possibility is raised that this compound contributes to metabolic regulation. In addition to Soyacerebroside, a tripeptid named Glu-Glu-Arg (L-α-glutamyl-L-α-glutamyl-L-arginine) was identified in the CHY ethanol extract; this compound is listed in the ChEBI database. This compound is expected to play an important role in protein signaling and metabolic pathways. The identification of Glu-Glu-Arg and its potential regulatory function are considered factors that further emphasize the diversity of bioactive compounds in the CHY extract. [Figure 9a] shows the LC-MS chromatogram, and [Figure 9b] visually presents the importance of phytochemicals by highlighting compounds that show relatively high abundance.

[0236] Taken together, these results suggest that the CHY ethanol extract contains various bioactive compounds such as Soyacerebroside and Glu-Glu-Arg, indicating that these components may contribute to lipid metabolism regulation and blood glucose control.

[0237]

[0238] The Korean melon leaf extract of the present invention can provide an anti-diabetic effect through a blood sugar-lowering effect. In addition, the Korean melon leaf extract of the present invention has a high yield of about 21%, making it economically viable as an excellent anti-diabetic material, thus having industrial applicability.

Claims

1. A pharmaceutical composition for the prevention or treatment of diabetes comprising a leaf extract of Korean melon (Cucumis melo var. makuwa).

2. In claim 1, the Korean melon leaf extract is, i) Step of extracting Korean melon leaf powder; ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and iii) A step of volatilizing the solvent after the above re-extraction; A pharmaceutical composition characterized by being manufactured through 3. A pharmaceutical composition according to claim 1, characterized in that the extract is an alcohol extract of Korean melon leaves.

4. A pharmaceutical composition according to claim 3, characterized in that the alcohol is 1 to 100% ethanol.

5. A pharmaceutical composition according to claim 1, characterized in that the composition reduces blood sugar.

6. A health functional food composition for preventing or improving diabetes containing Korean melon (Cucumis melo var. makuwa) leaf extract.

7. Antidiabetic feed additive containing Korean melon (Cucumis melo var. makuwa) leaf extract.

8. A blood glucose-lowering composition comprising Korean melon (Cucumis melo var. makuwa) leaf extract. 9.i) Step of extracting Korean melon (Cucumis melo var. makuwa) leaf powder; ii) a step of separating the extract and the residue after the above extraction and re-extracting the residue; and iii) A step of volatilizing the solvent after the above re-extraction; A method for preparing a composition for the prevention, improvement, or treatment of diabetes comprising a Korean melon (Cucumis melo var. makuwa) leaf extract.

10. A method for treating diabetes comprising the step of administering a Korean melon (Cucumis melo var. makuwa) leaf extract to an individual in need.

11. A treatment method according to claim 10, characterized in that the extract is an alcohol extract of Korean melon leaves.

12. A treatment method according to claim 10, characterized in that the alcohol is 1 to 100% ethanol.

13. A treatment method according to claim 10, wherein the extract is characterized by reducing blood sugar.

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