KR100665628B1 - Compositions for Inhibiting the Activity of ???-? and Lowering Blood Glucose Level - Google Patents

Abstract

The present invention relates to a composition for inhibiting the activity of dipeptidyl peptidase-IV (DPP-IV) and a hypoglycemic composition comprising at least one plant extract selected from the group consisting of essential, bitter peppermint and nadomus root as an active ingredient will be. The composition of the present invention effectively inhibits the activity of DPP-IV to reduce abnormally elevated blood glucose levels, and has an effect as a therapeutic agent for diabetes, especially type 2 diabetes. The composition of the present invention has the advantage that there is no harm to the human body because the plant extract used in the prior art as an active ingredient as an active ingredient.

Dipeptidyl Peptidase-IV, Blood Sugar, Diabetes, Prevalence, Bitter Mint, Nado

Description

Compositions for Inhibiting the Activity of DPI-IV and Lowering Blood Glucose Level             

1 is a graph showing that the essential and bitter peppermint extracts of the present invention inhibit the activity of dipeptidyl peptidase-IV (DPP-IV).

2 is a graph showing that the essential extract of the present invention exhibits the effect of lowering blood glucose levels.

3 is a graph showing that the bitter peppermint extract of the present invention exhibits an effect of lowering blood glucose levels.

Figure 4 is a graph showing that the Nado root extract of the present invention exhibits the effect of lowering blood glucose levels.

The present invention relates to a composition for inhibiting the activity of dipeptidyl peptidase-IV (DPP-IV) and lowering blood sugar.

Incretin hormone is a hormone that promotes insulin secretion in response to nutrient digestion of the intestines. There are two types of hormones: Glucose-dependent insulinotropic polypeptide (GIP) and Glucagon like peptide-1 (GLP-1).

GLP-1 is composed of a total of 30 amino acids and is secreted in response to nutrient digestion, resulting in insulin secretion associated with glucose metabolism, biosynthesis of insulin, inhibition of glucagon secretion, and food intake (inhibiting digestion and exercise of digestive organs, especially Inhibits gastrointestinal motility).

Both GIP and GLP-1 are essential hormones to maintain normal glucose metabolism, but especially in type 2 diabetic patients, blood levels of GIP remain normal or near normal, but GLP-1 secretion is reported to be reduced. GLP-1 is one of the most potent insulin secreting substances and has an EC ₅₀ (half-maximal effective concentration) of about 10 pmol / L.

It was first reported in 1992 that it was effective in diabetes using GLP-1. The report showed that blood glucose can be normalized with little need for insulin during the instillation of GLP-1 using artificial pancreas. Diabetes patients with secondary failure to oral hypoglycemic agents were instilled with GLP-1 for 4 hours to fully normalize blood glucose, demonstrating the potential of GLP-1 as a therapeutic for diabetes. The mean fasting blood glucose of these patients was 13 mmol / L and HbA1c was also high, but GLP-1 was effective.

Subsequent studies showed that GLP-1 lowered blood glucose levels in all patients with type 2 diabetes, regardless of the severity or duration of diabetes, the type of past treatment, and the presence of complications. In addition, GLP-1 was administered intravenously until 1 week in patients with type 2 diabetes to maintain almost normal blood glucose levels even during regular meals.

GLP-1 is synthesized in visceral endocrine cells, with two major forms being "7-36 amide" and "7-37 amide". The resulting GLP-1 is "GLP-1 (9-36) amide" as the two amino acids at the N-terminus are cleaved by an enzyme called dipeptidyl peptidase IV (DPP-IV) in the body. It also disappears.

DPP-IV recognizes and cleaves the proline and alanine at the N-terminus of GLP-1, and simply changing these positions results in resistance to DPP-IV and prolongs action time compared to living GLP-1.

Background of the development of DPP-IV inhibitors is that inhibiting the action of DPP-IV increases the concentration of GLP-1 and decreases the blood glucose level in patients with type 2 diabetes. IV-Deficient Fischer Rats.Biochem Biophys Res Commun. 8; 284 (2): 501-6 (2001); and Marguet D, et al., Enhanced insulin secretion and improved glucose tolerance in mice lacking CD26.Proc Natl Acad Sci USA 97 (12): 6874-6879 (2000). In other words, if there is a substance capable of selectively inhibiting DPP-IV, it is possible to prevent the degradation of exogenous or endogenous GLP-1, thereby enhancing the effect of promoting insulin secretion and eventually reducing blood glucose.

The representative DPP-IV target compound, LAF237, recently completed a 12-week phase II clinical trial in Novartis in patients with type 2 diabetes, and showed excellent hypoglycemic effects when administered alone or in combination with metformin. After 1 week of treatment with metformin with LAF237 or a placebo, HbA1c differences after 52 weeks in both groups were 1.1 W 0.2% (p <0.0001).

However, there are no studies on natural products that can inhibit the activity of DPP-IV.

Throughout this specification, numerous citations and patent documents are referenced and their citations are indicated. The disclosures of cited documents and patents are incorporated herein by reference in their entirety, so that the level of the technical field to which the present invention belongs and the contents of the present invention are more clearly explained.

The present inventors have diligently researched to develop a substance capable of inhibiting the activity of dipeptidyl peptidase-IV (DPP-IV) and ultimately lowering abnormal hyperglycemia. As a result, the essential and bitter peppermint extracts, which are conventionally used as herbal medicines, are described in detail. The discovery of one efficacy has led to the completion of the present invention.

Accordingly, an object of the present invention is to provide a composition for inhibiting the activity of dipeptidyl peptidase-IV (DPP-IV).

Another object of the present invention to provide a food composition for lowering blood sugar.

Another object of the present invention to provide a pharmaceutical composition for lowering blood sugar.

Other objects and advantages of the present invention will become apparent from the following detailed description, claims and drawings.

According to one aspect of the present invention, the present invention is for inhibiting the activity of dipeptidyl peptidase-IV (DPP-IV) comprising at least one plant extract selected from the group consisting of essential, bitter peppermint and nadomus root as an active ingredient To provide a composition.

According to another aspect of the present invention, the present invention provides a food composition for lowering blood sugar comprising at least one plant extract selected from the group consisting of essential, bitter peppermint and nado root water as an active ingredient.

According to another aspect of the present invention, the present invention provides a pharmaceutical composition comprising (a) a pharmaceutically effective amount of at least one plant extract selected from the group consisting of essential, bitter peppermint and nadomus; And (b) provides a pharmaceutical composition for lowering blood sugar comprising a pharmaceutically acceptable carrier.

The present inventors screened the ability to inhibit dipeptidyl peptidase-IV (DPP-IV) activity against various plant extracts, and finally, extracts of the essential and bitter peppermint inhibit DPP-IV activity, finally exhibiting hypoglycemic effect Found.

Primer used as an active ingredient in the composition of the present invention has been used in India for thousands of years to treat acid, swelling, wags, cold, heart pain and blood pressure (Indian Drugs, June, 384-388, 1984), these mainly It is cultivated in Korea, imported from Korea, and used as a herbal medicine under the name of 'Pil'. Considerable studies have been made on the physiological activity of the essential tree, medically anti-allergic effects (Indian J. Physiol. Pharmacol., 43: 486-490, 1999), anti-inflammatory effects (Indian J. Exp. Biol., 32: 633-636, 1994) and hepatoprotective effects (Planta Med., 59: 413-417, 19 93).

As another active ingredient of the present invention, bitter mint (Horehound; Marrubium vulgare ) is used to treat wounds, and is also used to treat anorexia and indigestion.

Oryzae Radix, which is used as an active ingredient of the present invention, refers to dried root stems or roots of perennial herbaceous rice belonging to the family Obese. This herb is commonly prescribed for patients with chronic hepatitis, sweating or pulmonary tuberculosis in the afternoon due to mild fever and cold sweating.

The extract of the present invention is an extractant such as (a) water, (b) anhydrous or hydrous lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, propanol, butanol, normal-propanol, iso-propanol and normal-butanol, etc.), (c) a mixed solvent of the lower alcohol with water, (d) acetone, (e) ethyl acetate, (f) chloroform, (g) 1,3-butylene glycol, (h) hexane (i) diethyl ether As an extraction solvent, it was obtained from the above-mentioned essential, bitter peppermint and nado root.

In addition, the extract of the present invention includes not only the extract by the above-described extraction solvent, but also the extract that has undergone a conventional purification process. Obtained by various additional purification methods, such as, for example, separation using ultrafiltration membranes having a constant molecular weight cut-off value, separation by various chromatography (manufactured for separation according to size, charge, hydrophobicity or affinity). The fraction is also included in the plant extract of the present invention.

The plant extract of the present invention may be prepared in powder form by additional processes such as distillation under reduced pressure and freeze drying or spray drying.

Inhibition of the activity of dipeptidyl peptidase-IV (DPP-IV), the potency of the essential, bitter peppermint and nadomus root extracts of the present invention, more specifically, the effect achieved by inhibiting DPP-IV binding to normal in vivo substrates. to be. In other words, the essential, bitter peppermint and nadomus root extracts act as competitive inhibitors of DPP-IV to inhibit DPP-IV activity.

Blood glucose drop, the ultimate potency of the essential, bitter peppermint and nadomus root extract of the present invention, means an action of reducing abnormally increased blood glucose levels.

Pharmaceutically acceptable carriers included in the pharmaceutical compositions of the present invention are those commonly used in the preparation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, Calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like It doesn't happen. In addition to the above components, the pharmaceutical composition of the present invention may further include a lubricant, a humectant, a sweetener, a flavoring agent, an emulsifier, a suspending agent, a preservative, and the like.

The pharmaceutical composition of the present invention may be administered orally or parenterally, and in the case of parenteral administration, it may be administered by intravenous injection, subcutaneous injection, intramuscular injection, or the like.

Suitable dosages of the pharmaceutical compositions of the invention vary depending on factors such as the formulation method, mode of administration, age, weight, sex, morbidity, condition of food, time of administration, route of administration, rate of excretion and response to reaction, Usually a skilled practitioner can easily determine and prescribe a dosage effective for the desired treatment or prophylaxis. According to a preferred embodiment of the present invention, the daily dose of the pharmaceutical composition of the present invention is 0.001-100 mg / kg.

The pharmaceutical compositions of the present invention may be prepared in unit dosage form by formulating with a pharmaceutically acceptable carrier and / or excipient according to methods which can be easily carried out by those skilled in the art. Or may be prepared by incorporation into a multi-dose container. In this case, the formulation may be in the form of a solution, suspension or emulsion in an oil or an aqueous medium, or may be in the form of extracts, powders, granules, tablets or capsules, and may further include a dispersant or stabilizer.

The food containing the plant extract of the present invention may include ingredients that are commonly added during food production. For example, when the beverage is prepared, in addition to the plant extract of the present invention, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid and / or juice may be further included.

The composition of the present invention acts as a competitive inhibitor of DPP-IV to inhibit the activity of DPP-IV, which increases the activity of Gluca-1 like Gluca-1 in vivo, which promotes the secretion of insulin Do it. As a result, it effectively reduces abnormally high blood sugar levels and acts as a treatment for diabetes, especially type 2 diabetes. In addition, the composition of the present invention has the advantage that there is no harm to the human body because the plant extract used in the prior art as an active ingredient as an active ingredient.

On the other hand, the composition of the present invention is administered in combination with existing drugs (e.g. sulfonylurea, metformin) developed as a type 2 diabetes treatment for synergistic hypoglycemic effect, diabetes, in particular the treatment efficacy of type 2 diabetes Can be exercised.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it is to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. Will be self-evident.

Example 1 Activity Inhibition Assay of Dipeptidyl Peptidase-IV

In the following examples, the ability to inhibit the activity of dipeptidyl peptidase-IV (DPP-IV) was analyzed as follows.

First, in order to form a thin film of DPP-IV, functional groups were substituted using SPDP (functional substitution medium, 3- (2-Pyridyldithio) propionic acid N-hydroxysuccinimide ester). 1 mM DPP-IV (Sigma, USA) and 1 mM SPDP (Sigma, USA) were mixed in a volume ratio of 1: 1 followed by gel filtration. In the mixing process, the amine group of DPP-IV forms a crosslink structure with a portion of 2-DPridyldthio-propionate from which the succinimide group is removed. The disulfide group was reduced to a thiol group by mixing the crosslinked DPP-IV mixture in a solution state of 1 mM DTT (dithiothreitol). The DPP-IV solution substituted with the functional group was impregnated with a gold substrate to finally prepare a thin film having DPP-IV bound to the surface.

The DPP-IV thin film was immersed in a solution containing a test substance to be analyzed (1 mM) and a green fluorescent protein (Clontech, USA; 1 mM) and reacted at room temperature for 24 hours. If there is NH in the test substance, DPP-IV acts to recognize NH, and DPP-IV (+) and GFP (-) act through their respective charges. DPP-IV, which interacts with a test substance with NH, loses its inherent charge due to ionic transfer between molecules, and weakly bonds with GFP to DPP-IV. Comparing results in differences in fluorescence intensity. The GFP emission wavelength was measured at 510 nm, which is an intrinsic wavelength band.

Example 2: Screening of Inhibitors of Dipeptidyl Peptidase-IV Activity

Plant extracts that inhibit the activity of dipeptidyl peptidase-IV (DPP-IV) were screened.

First of all, poppy, celandine, nado root as well as essential and bitter mint were used as screening targets.

Extracts were obtained from the plants to be screened in the following manner: by measuring electronic scales to 40 g of bite and bitter peppermint, respectively, and then pulverizing to pieces or powder with a grinder. Each comminuted raw material was divided again by 20 g using an electronic balance. The divided 20 g of the raw material was extracted with water and ethanol. Extraction was first stirred for 2 hours using a hot plate stirrer. After stirring, fine extraction was performed for 2 hours using ultrasonic waves. After the extraction was completed, the solution was filtered by using a 0.45 μm filter to remove foreign substances.

DPP-IV inhibitory activity was investigated as in Example 1 using the obtained extract, the experimental results are shown in FIG. As shown in Figure 1, in the case of pure DPP-IV (a), the fluorescence intensity is higher than the other two cases because the charge coupling with the GFP was made because no other variable occurs. On the other hand, since the essential extract (b) and bitter peppermint extract (c) interact with DPP-IV, the (+) charge of DPP-IV is lost, so that the binding through the charge of DPP-IV and GFP is weak. The fluorescence intensity was low.

Therefore, it can be seen that the essential extract and the bitter peppermint extract function as a competitive inhibitor of DPP-IV, thereby inhibiting the activity of DPP-IV.

Example 3: Animal Testing

Male Wistar rats (Charles river, Japan) were bred at 25 ° C and 30-70% humidity, fasted for 24 hours, and purified 1 hour before the experiment. The dry extract, bitter peppermint extract, and nano-root extract obtained in Example 2 were dried to prepare a powder, and the powder was liquefied for oral use by adding water. Four hours before oral glucose (2 g / kg), each extract was orally administered 10 mg / kg and 20 mg / kg. Blood was collected from the tail and venous blood was collected at -4, -0.5, 0, 0.5, 1 and 2 hours. Blood glucose analysis was performed using standard glucose oxidase method.

As can be seen in Figures 2, 3 and 4, in the rats not taking the extract of the present invention, the blood glucose level reached a maximum value of 150 mg / dl after 1 hour of glucose administration, but the essential extract, bitter peppermint extract and myocardium extract Intake of 10 mg and 20 mg resulted in reduced blood glucose levels of 60-80 mg / dl.

Therefore, it can be seen that the essential extract, bitter peppermint extract and nadomus root extract of the present invention can effectively reduce the increased blood sugar level.

Incorporating the results of Examples 2 and 3, the essential extract and bitter peppermint extract of the present invention functions as a competitive inhibitor of DPP-IV, inhibiting the activity of DPP-IV, which is in vivo GLP-1 (Glucagon like peptide) -1) increases the activity, which acts to promote the secretion of insulin. Eventually reduce blood sugar levels that are abnormally high.

As described in detail above, the present invention provides a composition for inhibiting activity of dipeptidyl peptidase-IV (DPP-IV) and a composition for lowering blood glucose. The composition of the present invention effectively inhibits the activity of DPP-IV to reduce abnormally elevated blood glucose levels, and has an effect as a therapeutic agent for diabetes, especially type 2 diabetes. The composition of the present invention has the advantage that there is no harm to the human body because the plant extract used in the prior art as an active ingredient as an active ingredient.

Claims (3)

delete A blood glucose-lowering food composition comprising an active extract of dipeptidyl peptidase-IV (DPP-IV) as an active ingredient, an essential extract, a nadomus root extract or a mixed extract of the essential and nadomus root. (a) a pharmaceutically effective amount of an active extract of dipeptidyl peptidase-IV (DPP-IV), the extract of which is essential extract, nadomus root extract, or a mixture of the essential and nadomus root; And (b) a pharmaceutical composition for hypoglycemic comprising a pharmaceutically acceptable carrier.

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