WO2005068485A1

WO2005068485A1 - Pharmaceutical composition comprising flavonoid compounds isolated from the extract of sedum sarmentosum bunge for preventing and treating hypertension

Info

Publication number: WO2005068485A1
Application number: PCT/KR2004/002315
Authority: WO
Inventors: Ho Sub Lee; Dae Gill Kang; Hyuncheol Oh; Tae Oh Kwon; Ji Wung Kwon; Seung Yeob Lee; Duck Bae Lee
Original assignee: Ho Sub Lee; Dae Gill Kang; Hyuncheol Oh; Tae Oh Kwon; Ji Wung Kwon; Seung Yeob Lee; Duck Bae Lee
Priority date: 2004-01-14
Filing date: 2004-09-10
Publication date: 2005-07-28
Also published as: KR20050074753A; KR100642151B1

Abstract

The present invention relates to the pharmaceutical composition and health care food comprising flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge for preventing and treating hypertension. The flavonoid compounds of the present invention inhibit effectively the increase of blood pressure caused by the action of angiotensin converting enzyme by inhibiting angiotensin converting enzyme. Therefore, it can be used as therapeutic agents or health care food for treating and preventing hypertension with safe.

Description

PHARMACEUTICAL COMPOSITION COMPRISING FLAVONOID COMPOUNDS ISOLATED FROM THE EXTRACT OF SEDUM SARMENTOSUM BUNGE FOR PREVENTING AND TREATING HYPERTENSION

BACKGROUM) OF THE INVENTION

Technical Field

The present invention relates to a pharmaceutical composition comprising flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge for preventing and treating hypertension.

B ackground Art

The term, blood pressure, denotes a pressure of blood stream flowing through blood vessels, i.e., arterial pressure. Hypertension induces an increase of resistance on the internal wall of blood vessel thus resulting in the maximum blood pressure (systolic blood pressure or highest blood pressure) of 150-160 mmHg and the minimum blood pressure (diastolic blood pressure or lowest blood pressure) of 90 mmHg or more. The former is called as a systolic hypertension and the latter is called as a diastolic hypertension. Although both of them may arise separately, it is general that they occur simultaneously. Moreover, hypertension may occur in the thirties in men, but it mainly occurs more frequently during the pre- and post-climacteric period in men rather than women.

According to the etiological origin, hypertension can be classified into essential hypertension and symptomatic hypertension. Although the cause of essential hypertension is not fully clarified as yet, essential hypertension occurs in many cases of fatty persons having a potent genetic predisposition. However, essential hypertension may also be observed in many cases of the other persons. This essential hypertension is closely correlated with arteriosclerosis. In this context, it appears that hypertension may result in arteriosclerosis. In case of essential hypertension, blood pressure is fluctuated at the early stage and then is maintained at a high level when arterial wall got hardened. Symptomatic hypertension arises from other causes and is mainly induced by renal diseases, particularly acute nephritis, toxemia of pregnancy, climacteric disorders, etc.

In many cases, hypertension at the early stage does not show any symptom and is accidentally revealed through the measurement of blood pressure. As hypertension becomes progressed, various subjective symptoms such as headache, anxiety, fatigue, vertigo, palpitation, excitation and constipation are developed and blood pressure is greatly fluctuated. In a chronic state of the disease, hypertrophic extension of left ventricle, coronary artery disorders, heart failure, etc. are progressed to cause congestive heart failure or cardiac infarction which may lead to death, or to give rise to the change in a renal artery and nephrosclerosis showing typical syndromes of renal failure such as polyuria, night polyuria, proteinuria, etc., which leads to fatal death because of uremia. Besides the syndromes, the other complications such as arteriosclerosis in eyeground, aneurysm etc., may also be developed (K m, K.H., A lecture of pharmacology of Lee W.J., 4^th, pp447-465, 2001).

Although the cause of hypertension is very diverse, hypertension generally stems from the hypergasia of sympathetic nerve, abnormality of hormone secretion, abnormality of renal artery, etc. Thus, spontaneous hypertension is concerned with at least three or more genes, upon which environmental factors such as excess ingestion of salts or stress, act to induce hypertension. In the early stage of disease, the neurological factors play an important role and the disease is progressed by factors including abnormalities of metabolism and the structure of blood vessels. If hypertension has been progressed, the contraction of artery is fixed, plasma norepinephrine is reduced, and collagen and non-collagen metabolism in cardiovascular system is accelerated, thereby fixing the state of hypertension. A change in the internal layer of blood vessel to support the elevated pressure thereof affects on the blood flow towarding heart and kidney thus increasing the mortality due to diseases in circulatory system such as arteriosclerosis, cardiac infarction, cerebral stroke, etc.

The study of hypertension has been extensively and continuously performed with a medical concern in an effort to develop several therapeutic agents till now. However, several problems occur in continuous medication such that, if one side is inhibited by the treating agent, the other side may be promoted in controlling the blood pressure, which induce an unbalance of hypertension. Therefore, more satisfactory therapeutic agent for treating hypertension has been needed till now. Angiotensin converting enzyme (ACE) is a zinc-dipeptidyl dipeptidase converting angiotensin I to angiotensin π physiologically and a potent vasoconstrictor. Angiotensin II is also known to stimulate both synthesis and release of aldosteron from the adrenal cortex, and this event increases blood pressure via sodium retention (Lacaille-Dubois, M.A., et al., Phytomedicine, 8, pp47-52, 2001). Therefore, the inhibition of ACE has been considered as one of the effective therapeutic approach for the treatment of hypertension. The conventional drugs, for example, Captopril , Enalepril^®, has been prescribed to the patients suffering from hypertension or kidney trouble up to now.

The plant belonged to Sedum genus (Crassulaceae) is mainly distributed over East- European regions and lots of the species have been used as medicines by dint of their various pharmacological activities (Mulinacci, N., et al., Phytochemistry, 38, pp531- 533, 1995). Sedum sarmentosum Bunge, in particular, has been used for the treatment of chronic viral hepatitis in Asian countries (Kang, T.H., et al., J. of Ethnopharmacology, 70, ppl77-182, 2000). The plant of Sedum species has been known to contain alkaloids, flavonoids, tannins, and cyanogenic compounds together with few metabolites especially in Sedum sarmentosum (Kim, J.H., et al., Phytochemistry, 41_, ppl319-1324, 1996; He, A, et al., Phytochemistry, 49, pp2609- 2610, 1998).

However, there has been not reported or suggested that the flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge has inhibitory effect on ACE and the compounds are useful for preventing and treating hypertension.

Therefore, the present inventors have endeavored to study the inhibitory effect on ACE, and finally found that flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge has inhibitory effect on ACE by experimenting in vitro ACE inhibition test.

SUMMARY OF THE INVENTION According to one aspect, the present invention provides a novel compounds isolated from the extract of Sedum sarmentosum Bunge for preventing and treating hypertension. The present invention also provides a pharmaceutical compositions comprising flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge or the pharmaceutically acceptable salt as an active ingredient in an amount effective for preventing and treating hypertension.

The present invention still provides a use of the compounds for the preparation of pharmaceutical composition for treating and preventing hypertension.

Disclosure of the invention

Thus, the present invention provides a novel quercetin-3-O-α-(6'"- caffeoylglucosyl-/J-l,2-rhamnoside) represented by the following chemical formula (la), the pharmaceutically acceptable salt or the isomer thereof:

The present invention also provides a pharmaceutical composition comprising the compounds of following general formula (I) or the pharmaceutically acceptable salt as an active ingredient in an amount effective for prevention and treatment of hypertension:

wherein Ri and R₂ are a hydrogen atom, OR' or NR', independently; R' is a hydrogen atom or a lower alkyl group having 1 to 3 carbon atoms.

The preferred ones of general formula (I) compounds comprises quercetin-3-O-cc- (6"'-caffeoylglucosyl-/i-l,2-rhamnoside) wherein both of Ri and R₂ are hydroxyl group and quercetin-3-O-α-(6'"- ?-coumaroylglucosyl-/?-l,2-rhamnoside) wherein Ri is a hydrogen atom, R₂ is a hydroxyl group respectively.

The present invention also provides a pharmaceutical composition comprising the compounds of following general formula (II) or the pharmaceutically acceptable salt as an active ingredient in an amount effective for prevention and treatment of hypertension:

wherein Ri and R₂ are a hydrogen atom or a lower alkyl group having 1 to 3 carbon atoms, independently.

The preferred ones of general formula (II) compounds comprises isorhamnetin-3- ?- glucopyranoside wherein Ri is a hydroxyl group, R₂ is a methyl group respectively and quercetin-3-/J-glucopyranoside wherein Ri is a hydroxyl group, R₂ is a hydrogen atom respectively.

The present invention also provides a pharmaceutical composition comprising the compounds of following general formula (III) or the pharmaceutically acceptable salt as an active ingredient in an amount effective for prevention and treatment of hypertension:

(πD wherein Ri and R₂ are a hydrogen atom, OR' or NR', independently; R' is a hydrogen atom or a lower alkyl group having 1 to 3 carbon atoms. The preferred ones of general formula (III) compounds comprises kaempferol-3-o arbinopyranoside wherein Ri is a hydroxyl group and R₂ is a hydrogen atom respectively. The inventive flavonoid compounds represented by general formula (I), (II) and (III) can be prepared by various method. For example, it is prepared by the simple chemical synthetic method or the extraction method from sedum genus plant.

Hereinafter, the present invention shall be described in detail. An inventive flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge can be prepared by following procedures. The inventive crude extract of Sedum sarmentosum Bunge can be prepared by follows, for example, Sedum sarmentosum Bunge is dried, cut, crushed and mixed with 1 to 10-fold, preferably, about 1 to 5 -fold volume of distilled water, lower alcohols having 1 to 4 carbon atoms such as methanol, ethanol, butanol and the like, or the mixtures thereof, preferably, the mixture of approximately ratio of 1:0.1 to 1:10, and then is subjected to the extraction, reflux extraction, or ultra-sonication extraction at room temperature for the period ranging from 1 day to 5 weeks, preferably 1 to 5 days, and repeated 2 to 5 times, preferably 3 times, consecutively. The residue is filtered to obtain the supernatant to be concentrated with rotary evaporator, and then concentrated under reduced pressure to obtain crude extract of Sedum sarmentosum Bunge.

Also, non-polar solvent soluble extract of present invention can be prepared by following procedure, for example, the crude extract prepared by above step, is suspended in water, and then is mixed with 1 to 100-fold, preferably, 1 to 5-fold volume of non polar solvent such as ethyl acetate, chloroform, hexane and the like and fractionated 1 to 10 times, preferably 2 to 5 times, the non-polar solvent soluble layer is collected to obtain non-polar solvent soluble extract of the present invention. Also, above described procedures may be modified or subjected to further step to fractionate or isolate more potent fractions or compounds by conventional procedure well-known in the art, for example, the procedure disclosed in the literature (Harborne J. B. Phytochemical methods: A guide to modern techniques of plant analysis, 3^rd Ed. pp6-7, 1998).

More particularly, above crude extract of S. sarmentosum, preferably methanol- soluble extract of S. sarmentosum is suspended in distillated water, and sequentially partitioned with n-hexane, EtOAc, n-BuOH in the manner to increase the polarity of the solvent. The EtOAc-soluble fraction showing potent pharmacological activity is subjected to silica gel vacuum flash column chromatography. The column is eluted with a stepwise application of solvent mixture consisting of linear gradient, i.e., starting with 20% hexane in CH₂C1₂, followed by MeOH in CH₂C1₂ (0% ~ 50%) to give 15 sub-fractions. Then, the 10^th fraction eluted with 10% MeOH in CH₂C1₂ is subjected to LH 20 column chromatography with a stepwise application of solvent mixture consisting of linear gradient, i.e., in the polarity order of CH Cl₂-acetone (1:1), CH₂Cl₂-acetone (1:4), acetone, acetone-MeOH (4:1), and MeOH. Fractions showing similar component spectra determined by TLC analysis are pooled and subjected to further purification to afford pharmacologically active compounds. For example, isorhamnetin-3-/?- glucopyranoside (compound 3) and quercetin-3- ?-glucopyrarιoside (compound 4) are isolated in fractions 31 to 34, and 35, respectively. And, for purify the fractions 26 to 30, reversed-phase HPLC with increasing polarity gradient from 20 to 60% acetonitrile is performed to obtain isorharnnetin-3- ?- glucopyranoside (compound 3) and kaempferol-3-α-arbinopyranoside (compound 5). Also, the fraction eluted 20% MeOH in CH₂C1₂ is subjected to LH 20 column chromatography with a stepwise application of solvent mixture containing linear gradient, i.e., H₂O (140 mi), MeOH-H₂O (1:9, 160 mi), MeOH-H₂O (1 :4, 60 m ), MeOH-H₂O (1:1, 140 mi), and MeOH (60 mi) to collect 20 mi of respective fractions. Fractions having similar component spectra between each other determined by TLC analysis are pooled. For purify fractions 21 to 23, reversed-phase HPLC is performed to obtain quercetin-3-O-α-(6"'-caffeoylglucosyl- ?-l,2- rhamnoside(compound 1) and quercetin-3-O-α-(6"'- -coumaroylglucosyl-/?-l,2- rhamnoside(compound 2). The inventive flavonoid compounds of the present invention may be also synthesized by the conventional synthetic method in accordance with a using method well known in the art (Herbert O. House, Modern Synthetic Reactions, 2ⁿ Ed., The Benjamin/Cummings Publishing Co., 1972).

The inventive compounds represented by general formula (I), (II) and (III) can be transformed into their pharmaceutically acceptable salt and solvates by the conventional method well known in the art. For the salts, acid-addition salt thereof formed by a pharmaceutically acceptable free acid thereof is useful and can be prepared by the conventional method. For example, after dissolving the compound in the excess amount of acid solution, the salts are precipitated by the water-miscible organic solvent such as methanol, ethanol, acetone or acetonitrile to prepare acid addition salt thereof and further the mixture of equivalent amount of compound and diluted acid with water or alcohol such as glycol monomethylether, can be heated and subsequently dried by evaporation or filtrated under reduced pressure to obtain dried salt form thereof. As a free acid of above-described method, organic acid or inorganic acid can be used. For example, organic acid such as methansulfonic acid, j^-toluensulfonic acid, acetic acid, trifluoroacetic acid, citric acid, maleic acid, succinic acid, oxalic acid, benzoic acid, lactic acid, glycolic acid, gluconic acid, galacturonic acid, glutamic acid, gmtaric acid, glucuronic acid, aspartic acid, ascorbic acid, carbonylic acid, vanillic acid, hydroiodic acid and the like, and inorganic acid such as hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid and the like can be used herein.

Further, the pharmaceutically acceptable metal salt form of inventive compounds may be prepared by using base. The alkali metal or alkali-earth metal salt thereof can be prepared by the conventional method, for example, after dissolving the compound in the excess amount of alkali metal hydroxide or alkali-earth metal hydroxide solution, the insoluble salts are filtered and remaining filtrate is subjected to evaporation and drying to obtain the metal salt thereof. As a metal salt of the present invention, sodium, potassium or calcium salt are pharmaceutically suitable and the corresponding silver salt can be prepared by reacting alkali metal salt or alkali-earth metal salt with suitable silver salt such as silver nitrate.

The pharmaceutically acceptable salt of the compound represented by general formula (I), (II) and (III) comprise all the acidic or basic salt which may be present at the compounds, if it does not indicated specifically herein. For example, the pharmaceutically acceptable salt of the present invention comprise the salt of hydroxyl group such as the sodium, calcium and potassium salt thereof; the salt of amino group such as the hydrogen bromide salt, sulfuric acid salt, hydrogen sulfuric acid salt, phosphate salt, hydrogen phosphate salt, dihydrophosphate salt, acetate salt, succinate salt, citrate salt, tartarate salt, lactate salt, mandelate salt, methanesulfonate(mesylate) salt and j^-toluenesulfonate (tosylate) salt etc, which can be prepared by the conventional method well known in the art. There may exist in the form of optically different diastereomers since the compounds represented by chemical formula (la) have chiral centers(*), accordingly, the compounds of the present invention comprise all the optically active isomers, R or S stereoisomers and the mixtures thereof. Present invention also comprises all the uses of racemic mixture, more than one optically active isomer or the mixtures thereof as well as all the preparation or isolation method of the diastereomer well known in the art.

In order to confirm the inhibitory effects on angiotensin converting enzyme of the flavonoid compounds represented by general formula (I), (II) and (III) obtained in the above preparation method, the ACE inhibition experiment was performed. The results of this experiment demonstrate that the inventive flavonoid compounds represented by general formula (I), (II) and (III) exhibit potent inhibitory effects on angiotensin converting enzyme (ACE) activity.

The inventive pharmaceutical composition for treating and preventing hypertension comprises above compounds as 0.1 ~ 90 % by weight based on the total weight of the composition. The inventive composition may additionally comprise conventional carrier, adjuvants or diluents in accordance with a using method well known in the art. It is preferable that said carrier is used as appropriate substance according to the usage and application method, but it is not limited. Appropriate diluents are listed in the written text of Remington's Pharmaceutical Science (Mack Publishing co, Easton PA).

Hereinafter, the following formulation methods and excipients are merely exemplary and in no way limit the invention. Pharmaceutical formulations containing present composition may be prepared in any form, such as oral dosage form (powder, tablet, capsule, soft capsule, aqueous medicine, syrup, elixirs pill, powder, sachet, granule), or topical preparation (cream, ointment, lotion, gel, balm, patch, paste, spray solution, aerosol and the like), or injectable preparation (solution, suspension, emulsion). The composition according to the present invention can be provided as a pharmaceutical composition containing pharmaceutically acceptable carriers, adjuvants or diluents, e.g., lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starches, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, polyvinyl pyrrolidone, water, methylhydroxy benzoate, propylhydroxy benzoate, talc, magnesium stearate and mineral oil. The formulations may additionally include fillers, anti-agglutinating agents, lubricating agents, wetting agents, flavoring agents, emulsifiers, preservatives and the like. Solid dosage forms for oral administration can include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the compounds of this invention are ordinarily combined with one or more adjuvants appropriate to the indicated route of administration. If administered per os, the compounds can be admixed with lactose, sucrose, starch powder, cellulose esters of alkanoic acids, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric and sulfuric acids, gelatin, acacia gum, sodium alginate, polyvinylpyrrolidone, and/or polyvinyl alcohol, and then tableted or encapsulated for convenient administration. Liquid dosage forms for oral administration can include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs containing inert diluents commonly used in the art, such as water. Such compositions can also comprise adjuvants, such as wetting agents, emulsifying and suspending agents, and sweetening, flavoring, and perfuming agents. For therapeutic purposes, formulations for parenteral administration can be in the form of aqueous or non-aqueous isotonic sterile injection solutions or suspensions. These solutions and suspensions can be prepared from sterile powders or granules having one or more of the carriers or diluents mentioned for use in the formulations for oral administration. The compounds can be dissolved in water, polyethylene glycol, propylene glycol, ethanol, corn oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, and/or various buffers. Other adjuvants and modes of administration are well and widely known in the pharmaceutical art.

The composition of the present invention in pharmaceutical dosage forms may be used in the form of their pharmaceutically acceptable salts, and also may be used alone or in appropriate association, as well as in combination with other pharmaceutically active compounds. The desirable dose of the inventive compounds varies depending on the condition and the weight of the subject, severity, drug form, route and period of administration, and may be chosen by those skilled in the art. However, in order to obtain desirable effects, it is generally recommended to administer at the amount ranging 0.0001 to 100 mg/kg, preferably, 0.01 to 10 mg/kg by weight/day of the inventive compounds of the present invention. The dose may be administered in single or divided into several times per day. However, the range of present invention is not limited to above amount of administration.

The pharmaceutical composition of present invention can be administered to a subject animal such as mammals (rat, mouse, domestic animals or human) via various routes. All modes of administration are contemplated, for example, administration can be made orally, rectally or by intravenous, intramuscular, subcutaneous, intracutaneous, intrathecal, epidural or intracerebro ventricular injection.

The present invention provide a health care food comprising above compound represented by general formula (I), (II) or (III) and sitologically acceptable additive to prevent and improve hypertension. To develop for health food, examples of addable food comprising above compounds of the present invention are various food, beverage, gum, vitamin complex, health improving food and the like. Also, above described the compounds can be added to food or beverage for prevention and improvement of hypertension. The amount of above described compound in food or beverage may generally range from about 0.001 to 15 w/w % of total weight of food for the health food composition and 0.02 to 5 g, preferably 0.3 to 1 g on the ratio of 100 ml of the health beverage composition.

The term "sitologically acceptable additive" comprise all the conventional additives well known in the art, which shall be explained in detail as follows. Providing that the health beverage composition of present invention contains above described compound as an essential component in the indicated ratio, there is no particular limitation on the other liquid component, wherein the other component can be various deodorant or natural carbohydrate etc such as conventional beverage. Examples of aforementioned natural carbohydrate are monosaccharide such as glucose, fructose etc; disaccharide such as maltose, sucrose etc; conventional sugar such as dextrin, cyclodextrin; and sugar alcohol such as xylitol, and erythritol etc. As the other deodorant than aforementioned ones, natural deodorant such as taumatin, stevia extract such as levaudioside A, glycyrrhizin et al., and synthetic deodorant such as saccharin, aspartam et al., may be useful favorably. The amount of above described natural carbohydrate is generally ranges from about 1 to 20 g, preferably 5 to 12 g in the ratio of 100 mi of present beverage composition.

The other components than aforementioned composition are various nutrients, a vitamin, a mineral or an electrolyte, synthetic flavoring agent, a coloring agent and improving agent in case of cheese, chocolate et al., pectic acid and the salt thereof, alginic acid and the salt thereof, organic acid, protective colloidal adhesive, pH controlling agent, stabilizer, a preservative, glycerin, alcohol, carbonizing agent used in carbonate beverage et al. The other component than aforementioned ones may be fruit juice for preparing natural fruit juice, fruit juice beverage and vegetable beverage, wherein the component can be used independently or in combination. The ratio of the components is not so important but is generally range from about 0 to 20 w/w % per 100 w/w % present composition. Examples of addable food comprising aforementioned extract therein are various food, beverage, gum, vitamin complex, health improving food and the like. It will be apparent to those skilled in the art that various modifications and variations can be made in the compositions, use and preparations of the present invention without departing from the spirit or scope of the invention. Brief Description of the Drawings

The above and other objects, features and other advantages of the present invention will more clearly understood from the following detailed description taken in conjunction with the accompanying drawing, in which;

Fig. la to le represent the measurement of inhibitory effects on angiotensin converting enzyme (ACE) of flavonoid compounds of the present invention, Fig. la represents the inhibitory effects on angiotensin converting enzyme (ACE) in a dose dependent manner of quercetin-3-O-α-(6'"-caffeoylglucosyl-/?-l,2-rhamnoside) (compound 1) isolated from the extract of Sedum sarmentosum Bunge, Fig. lb represents the inhibitory effects on angiotensin converting enzyme (ACE) in a dose dependent manner of quercetin-3-O-α-(6"'-p-coumaroylglucosyl-/ ,2- rhamnoside) (compound 2) isolated from the extract of Sedum sarmentosum Bunge, Fig. lc represents the inhibitory effects on angiotensin converting enzyme (ACE) in a dose dependent manner of isorhamnetin-3- ?-glucopyranoside(compound 3) isolated from the extract of Sedum sarmentosum Bunge, Fig. Id represents the inhibitory effects on angiotensin converting enzyme (ACE) in a dose dependent manner of quercetin-3-y#-glucopyranoside(compound 4) isolated from the extract of Sedum sarmentosum Bunge, Fig. le represents the inhibitory effects on angiotensin converting enzyme (ACE) in a dose dependent manner of kaempferol-3-α-arbinopyranoside(compound 5) isolated from the extract of Sedum sarmentosum Bunge.

Best Mode for Carrying Out the Invention

The present invention is more specifically explained by the following examples. However, it should be understood that the present invention is not limited to these examples in any manner.

Reference Example 1: Experimental Apparatus For structural analysis of compounds obtained from the following Example 1, the optical rotation was recorded on Optical Activity AA-10 Automatic Polarimeter. FAB- MS data was obtained on a JEOL JMS HX-100 spectrometer using 3-nitrobenzyl alcohol as a matrix and ESLMS data was obtained on a Macro Mass Quatro LC with electrospray ionization. NMR spectra (ID & 2D) was recorded in acetone-^ or MeOH- ₄ using a JEOL Ecliρse-500 MHz spectrometer (500 MHz for 1H and 150 MHz for ¹³C), and chemical shifts were referenced relative to the corresponding residual solvents signals. HMQC and HMBC data were optimized for 'Jar* 140 Hz and ⁿJc_H ⁼8 Hz, respectively.

Example 1. Isolation of flavonoid compounds from the extracts of a sedum sarmentosum 1-1. Preparation of methanol-soluble extract of a £ sarmentosum Fresh S. sarmentosum was collected in the Botanical Garden of Wonkang University, Dcsan, Korea, in July 2000. The plant was identified and authenticated, and a specimen (NO. DH-67) was deposited in the Herbarium of the Professional Graduate School of Oriental Medicine, Wonkwang University. 4.0 kg of dried S. sarmentosum was extracted with 6L of methanol at 25 °C for 3 days at three times. The resulting extracts were filtrated with vacuum filtration, combined, concentrated under reduced pressure with evaporator and dried to afford

284.4g of dried methanol-soluble extract of S. sarmentosum, which was used as a test sample in further experiment.

1-2. Preparation of various fractions of a S. sarmentosum 284.4g of the methanol-soluble extract of S. sarmentosum in Example 1-1 was suspended in 2L of distillated water, and sequentially partitioned with n-hexane, EtOAc, n-BuOH and water. At first, the extract in Example 1-1 was suspended in 2L of distilled water and 2L of n-hexane was added thereto. The solution was subjected to fractionation at three times, concentrated in vacuum to obtain 52.6g of n-hexane fraction. 2L of EtOAc was added to remaining water fraction and the solution was subjected to fractionation at three times, concentrated in vacuo to obtain 61.2g of EtOAc fraction. 2L of BuOH was added to the remaining water fraction to obtain 70.5g of BuOH fraction. Finally, remained water fraction was concentrated under reduced pressure to obtain 49.6g of water fraction.

1-3. Purification of compounds from the EtOAc fraction 1-3-1. Isolation of sub-fractions 6.6g of bioactive EtOAc fraction in Example 1-2 was subjected to silica gel vacuum flash column chromatography (4.5x10 cm; 15-40μm of particle size). The column was eluted with a stepwise application of solvent mixture consisting of linear gradient, i.e., starting with 20%> hexane in CH₂C1₂ (900m£; collecting 300m£ volume of each fractions), followed by solvent mixture of MeOH in CH₂C1₂ (0% 600ml, 5% 600ml, 7% 300m£, 10% 600ml, 20% 600ntf, 30% 900m£ and 50% 600m£, respectively; made to 300 fractions) to give 15 sub-fractions. 1-3-2. Isolation of Isorhamnetin-3- 3- glucopyranoside and quercetin-3-<3- glucopyranoside 399.3mg of fraction 10 obtained in Example 1-3-1 was subjected to LH-20 column chromatography with a stepwise application of solvent mixture containing linear gradient, i.e., starting with CH₂Cl₂-acetone (1:1, 30( ), CH₂Cl₂-acetone (1:4, 200m£), acetone (160ml), acetone-MeOH (4:1, 40ml), and MeOH (60ml) to collect total 38 fractions with respective volume of 20ml. Fractions having similar component spectra between determined by TLC analysis are pooled together. Among the fractions, compound 3 and compound 4 are isolated in fractions 31 to 34, and 35, respectively.

And, the isolated compounds prepared from above Examples were identified by comparing their spectral data with those of same compounds disclosed in the literature (Markham, K.R., et al., Phytochemistry, 31, pp 1009-11, 1992; Markham, K.R., et al, Tetrahedron, 34, pp 1389-97, 1978; Markham, K.R., et al, Chapman and Hall, pp 19- 134, 1982), compound 3 and 4 were identified as isorhamnetin-3-/?-glucopyranoside and quercetin-3-yø-glucopyranoside, respectively.

1-3-3. Isolation of Isorhamnetin-3-i8-glucopyranoside and kaempferol-3- - arbinopyranoside Among the total 38 fractions obtained in Example 1-3-2, the fraction numbers 26 to

30 was pooled and subjected to reversed-phase HPLC [C₁₈ column (19x300 mm; Iμm. particle size)], using a gradient from 20 to 60% acetonitrile [in H O, over 30 min; 4 m^/min; detection at 254 nm] to obtain compound 3 (13.5 mg; t_R = 92.2 - 97.3 mi) and compound 5 (4.2 mg; t_R = 98.6 - 103.7 ml). And, the isolated compounds prepared from above Examples were identified by comparing their spectral data with those of same compounds disclosed in the literature (Markham, K.R., et al, Phytochemistry, 31, pp 1009-11, 1992; Markham, K.R., et al., Tetrahedron, 34, pp 1389-97, 1978; Markham, K.R., et al., Chapman and Hall, pp 19- 134, 1982), compound 5 was identified as kaempferol-3-α-arbinopyranoside.

1-3-4. Isolation of quercetin-3-O-α-(6'"caffeoylglucosyl-_jg-l,2-rhamnoside and quercetin-3 -O- a-{6 ' " -p-coumaroylglucosyl-/?- 1 ,,2-rhamnoside^') 397.2mg of fraction 12 obtained in Example 1-3-1 was subjected to LH-20 column chromatography with a stepwise application of solvent mixture containing linear gradient, i.e., starting with H₂O (140 ml), MeOH-H₂O (1:9, 160 m ), MeOH- H₂O (1:4, 60 m ), MeOH-H₂O (1:1, 140 me), and MeOH (60 me) to collect 20me of respective fractions. Fractions having similar component spectra between determined by TLC analysis are pooled together. Further purification of fractions 21 to 23 (70 mg) using reversed-phase HPLC [BDS C₁₈ column (10x250 mm; Sμ particle size); gradient from 15 to 30% acetonitrile in H O over 60 min; 2 m^/min; detection at 254 nm] to obtain compound 1 (16.8 mg; t_R = 41.2 - 44.0 m ) and compound 2 (6.0 mg; t_R = 49.2 -49.8 e), respectively.

And, the isolated compounds prepared from above Examples were identified by comparing their spectral data with those of same compounds disclosed in the literature (Markham, K.R., et al, Phytochemistry, 31, pp 1009-11, 1992; Markham, K.R., et al., Tetrahedron, 34, PP 1389-97, 1978; Markham, K.R., et al., Chapman and Hall, pp 19- 134, 1982), compound 2 was identified as quercetin-3-O-α-(6"'-^-coumaroylglucosyl- M,2-rhamnoside. The isolated compound 1 was identified as quercetin-3-O-<2-(6"'- caffeoylglucosyl-/?-l,2-rhamnoside by the spectral analysis described in above reference example 1. quercetin-3 -O- a-(6 " ' -caffeoylglucosyl- ?- 1 ,2-rhamnoside (compound 1) LRESLMS m z: 771 [M-H]^" LRFABMS m z: 773 [M+H]⁺ HRFABMS m/z: 773.1929(calc. for C₃₆H₃₇O₁₉, 773.1929) [a]_D ²⁵ = -147.6°(c 0.21, MeOH) ¹³C-NMR, 1H-NMR, HMBC (See Table 1)

Table 1

17

Experimental Example 1 : The effect on ACE activity of the compounds The effect on Angiotensin converting enzyme of each compounds obtained in Examples 1-3 was measured by modified procedure disclosed in the literature (Santos, R.A., Hypertension, 7, pp244-252, 1985; Kang, D-Q et al., J. ofEthnopharmacology, 81, pp 49-55, 2002).

10 of serum was transferred to 490 ≠ of assay buffer solution containing 5 mM of Hip-His-Leu in 0.4 M of sodium borate buffer, and the solution was incubated at 37 °C for 15 minutes. The reaction was stopped by adding 1.2 m ₀f 0.34 N NaOH, and then 100 βi of O-phthaldialdehyde solution dissolved in MeOH adjusted the concentration to 20 %, was transferred thereto for reproducing a fluorescent material. After 10 minutes, 200 lΛ of 3 N HC1 was added to the mixture and mixed well to stop the reaction, and centrifugation was subjected at the speed of 3000 rpm for 10 minutes at room temperature to obtain final product, i.e., His-Leu. The final product was detected by spectrofluorimeter (Hitachi, model F-2000, Tokyo, Japan) at the excitation wavelength of 365 run and emmision wavelength of 495 nm. Standard curve was drawn up using His-Leu, and negative control group was pre-treated by adding 0.34 N of NaOH prior to reacting with serum.

Consequently, it is confirmed that the compounds 1 to 5 inhibited the activity of angiotensin convertion enzyme in a dose dependent manner, and the 50% inhibitory concentration (IC₅₀) values for the compounds 1 to 5 were 158.9 + 11.1 μM, 351.6 ± 3.9 μM, 408.9 ± 4.6 μM, 708.8 ± 23.1 μM, and 392.8 ± 13.4 μM, respectively (See Fig. la to le).

Hereinafter, the formulating methods and kinds of excipients will be described, but the present invention is not limited to them. The representative preparation examples were described as follows.

Preparation of powder

Quercetin-3 -O- a-(6 " ' -caffeoylglucosyl- ?- 1 ,2-rhamnoside) 20mg

Lactose lOOmg

Talc lOmg Powder preparation was prepared by mixing above components and filling sealed package.

Preparation of tablet

Quercetin-3-O-o;-(6'"-caffeoylglucosyl-f9-l,2-rhamnoside) lOmg Corn Starch lOOmg

Lactose lOOmg

Magnesium Stearate 2mg Tablet preparation was prepared by mixing above components and entabletting.

Preparation of capsule

Quercetin-3-O- a-(6 " '-caffeoylglucosyl-/?- 1 ,2-rhamnoside) 1 Omg Crystalline cellulose 3mg

Lactose 14.8mg

Magnesium Stearate 0.2mg Tablet preparation was prepared by mixing above components and filling gelatin capsule by conventional gelatin preparation method.

Preparation of injection

Quercetin-3 -O- a-(6 " ' -caffeoylglucosyl- ?- 1 ,2-rhamnoside) 1 Omg

Mannitol 18 Omg

Distilled water for injection 2974mg Na₂HPO₄,12H₂O 26mg

Injection preparation was prepared by mixing above components's amount per 1 ampul(2 ml) by conventional injection preparation method.

Preparation of liquid Quercetin-3-O-o;-(6'"-caffeoylglucosyl-_>f5-l,2-rhamnoside) 20mg

Sugar lOg

Mannitol 5g

Distilled water optimum amount Liquid preparation was prepared by dissolving active component, adding lemon flavor and distilled water and then filling all the components in 100 ml brown bottle and sterilizing by conventional liquid preparation method.

Preparation of health beverage

Quercetin-3-O-α-(6"'-caffeoylglucosyl-/?-l ,2-rhamnoside) lOOmg Vitamin C 15g

Vitamin E (powder) lOOg

Ferrous Lactate 19.75g

Zinc oxide 3.5g

Amide nicotinic acid 1.7g Vitamin A 0.2g

Vitamin Bi 0.25g

Vitamin B₂ 0.3g Distilled water optimum amount

Health beverage preparation was prepared by dissolving active component, mixing, stirred at 85 ^°C for 1 hour, filtered and then filling all the components in 2 L ample and sterilizing by conventional health beverage preparation method.

The invention being thus described, it will be obvious that the same may be varied in many ways. Such variations are not to be regarded as a departure from the spirit and scope of the present invention, and all such modifications as would be obvious to one skilled in the art are intended to be included within the scope of the following claims.

Industrial Applicability As mentioned above, a composition comprising flavonoid compounds isolated from the extract of Sedum sarmentosum Bunge according to the present invention inhibit effectively the increase of blood pressure caused the action of angiotensin converting enzyme by inhibiting angiotensin converting enzyme. Therefore, it can be used as therapeutic agents or health care food for treating and preventing hypertension with safe.

Claims

1. A novel quercetin-3-O-α-(6'"-caffeoylglucosyl-/?-l,2-rhamnoside) represented by the following chemical formula (la), the pharmaceutically acceptable salt or the isomer thereof:

2. A pharmaceutical composition comprising the compounds of following general formula (I) or the pharmaceutically acceptable salt as an active ingredient in an amount effective for prevention and treatment of hypertension:

wherein Ri and R₂ are a hydrogen atom, OR' or NR', independently;

R' is a hydrogen atom or a lower alkyl group having 1 to 3 carbon atoms.

3. The pharmaceutical composition according to claim 2 wherein said compound represented by the general formula (I) is quercetin-3-O-α-(6"'-caffeoylglucosyl- ?-l,2- rhamnoside) or quercetin-3 -O- -(6 " ' -/?-coumaroylglucosyl- ?- 1 ,2-rhamnoside) .

4. A pharmaceutical composition comprising the compounds of following general formula (II) or the pharmaceutically acceptable salt as an active ingredient in an amount effective for prevention and treatment of hypertension:

wherein

Ri and R₂ are a hydrogen atom or a lower alkyl group having 1 to 3 carbon atoms, independently.

5. The pharmaceutical composition according to claim 4 wherein said compound represented by the general formula (II) is isorharrmetin-3-/?-glucopyranoside or quercetin-3- ?-glucopyranoside.

6. A pharmaceutical composition comprising the compounds of following general formula (III) or the pharmaceutically acceptable salt as an active ingredient in an amount effective for prevention and treatment of hypertension:

P) wherein

Ri and R₂ are a hydrogen atom, OR' or NR', independently; R' is a hydrogen atom or a lower alkyl group having 1 to 3 carbon atoms.

7. The pharmaceutical composition according to claim 6 wherein said compound represented by the general formula (in) is kaempferol-3-α-arbinopyranoside.

8. A health care food comprising the compound represented by general formula (I), (II) or (III) as set forth in claims 2, 4 and 6, and sitologically acceptable additive to prevent and improve hypertension.

9. The health care food according to claim 8, wherein said health care food is provided as beverage type.