WO2003099306A1

WO2003099306A1 - Process for preparing apigenin-7-o-¥â-d-glucuronide from clerodendron trichotomum folium

Info

Publication number: WO2003099306A1
Application number: PCT/KR2003/001037
Authority: WO
Inventors: Uy-Dong Sohn; Wan-Kyunn Whang; In-Hye Ham; Young-Sil Min; Ki-Lyong Bae; Sung-Hyuk Yim; Yul-Pyo Lee
Original assignee: Uy-Dong Sohn; Wan-Kyunn Whang; In-Hye Ham; Young-Sil Min; Ki-Lyong Bae; Sung-Hyuk Yim; Yul-Pyo Lee
Priority date: 2002-05-27
Filing date: 2003-05-27
Publication date: 2003-12-04
Also published as: KR20030091403A; KR100489520B1; AU2003234352A1

Abstract

The present invention relates to a process for preparing apigenin-7-O-b-D-glucuronide(AGC) isolated from Clerodendron trichotomum Folium and a pharmaceutical composition comprising above compound for the prevention and treatment of gastritis, and reflux esophagitis comprising above compound. The present invention provide a process for preparing apigenin-7-O-b-D-glucuronide (AGC) isolated from Clerodendron trichotomum Folium comprising the steps of; obtaining the crude extract Clerodendron trichotomum Folium; defatting the crude extract by using non-polar solvent; removing sugar and mineral salts from above defatted extract and obtaining flavonoid fraction by using non-ionic exchange column chromatography; and purifying the fraction by recrystallizing to obtain above compound.

Description

PROCESS FOR PREPARING APIGENIN-7-O-β-D-GLUCURONIDE FROM CLERODENDRON TRICHOTOMUM FOLIUM

Technical Field

The present invention relates to novel process for preparing apigenin-7-O-β-D- glucuronide isolated from Clerodendron trichotomum Folium and pharmaceutical composition comprising the same for the prevention and the treatment of inflammation of gastrointestinal tract, gastritis and reflux esophagitis.

Background Art

As Korean intakes the hot and salty food and the alcoholic beverage in excess, gastritis diseases have been occurred frequently. However, there has not been yet developed therapeutically effective treatment to protect gastritis diseases in Korea. Gastric acid has been widely known as the cause of disease such as gastritis diseases and reflux esophagitis. Recently, various scientific studies on the astritis diseases and pharmacological treatments of the disease have been reported so that the secret pharmacological effects are paid attention to modern scientific pproaches (Nakamura K, et al; Jpn. J. Pharmacol. 32(3), pp445-56, 1982 ; Okabe S, et al; Jpn. J. Pharmacol. 69(4), pp317-23, 1995)

Clerodendron trichotomum Thunb belonging to Nerbenaceae family grows naturally at domestic central and southern districts of Korea. It is reported that the plant contains lectin clerodendrin, alkaloid, meso-inositol, glucose, acacetin-7- glucurono (l->2)-glucuronide), and etc. (Kitagaki-Ogawa, H. et al., Eur. J. Biochem., 161, p779, 1986), arid haves physiological activities such as depressor, ataralgesia and Clerodendron inerme shows anti-inflammatory activity, and hypertension treating activity, apoplexy and dermatitis (Lu. G. W. et al, J. Ethanopharmacol, 42, p77, 1994).

However, there have been no disclosure or suggestion about therapeutic activity of apigenin-7-O-β-D-glucuronide against gastritis and reflux esophatitis. Disclosure of the invention

Accordingly, it is an object of the present invention to provide a process for preparing apigenin-7-O-β-D-glucuronide of the following general formula (1) from Clerodendron trichotomum Folium.

[Chemical Formula 1]

Glucuronic acid-0

It is another object of the present invention to provide a pharmaceutical composition comprising a pharmaceutically effective amount of apigenin-7-O-β-D- glucuronide (AGC) compound with a pharmaceutically acceptable carrier, exupient and diluents for the prevention and treatment of gastritis and reflux esophagitis.

It is another object of the present invention to provide use of AGC compound for preparation of medicament for preventing or treating gastritis and reflux esophagitis.

It is another object of the present invention to provide a health food comprising above quercetin-3-0-β -D-glucuronide for the prevention of gastritis diseases and reflux esophagitis and sitologically acceptable additive.

The AGC compound of the present invention may be prepared in accordance with the following preferred embodiment:

In preferred embodiment of the present invention, there provided a process for preparing AGC compound from Clerodendron trichotomum Folium comprising the steps of: obtaining the crude extract from Clerodendron trichotomum Folium; defatting the crude extract by using non-polar solvent; removing sugar and mineral salts from above defatted extract and obtaining flavonoid fraction by using non-ionic exchange column chromatography; and purifying the fraction by recrystallizing to obtain above compound.

Above described AGC compound can be prepared by following steps:

1. 1^st step: the preparation of crude extract

Specially, at the 1^st step, Clerodendron trichotomum Folium is sliced and dried in the shade. Then, dried plant is mixed with one to twenty times, preferably, three to ten times volumes of water or ~ C₅ lower alcohol, or the mixture thereof; and is extracted at a temperature ranging from 60 - 90 °C, preferably 80 °C, for a peroid ranging from 1 to 24 hours, preferably 5 hours, with 3 to 10 times, preferably 7 times, to obtain an aqueous crude extract. The extract is centrifuged, filtered, concentrated under a reduced pressure and the crude eatract is sealed and kept in refrigerator until use. Additionally, above concentreate can be lyophilized to obtain an extract powder.

2. 2^nd step: defatting step

The crude eatract obtained from 1^st step, is suspended in distilled water, separated to water-soluble layer and precipitates; mixed with non-polar organic solvent, such as chloroform, ether, methylene chloride hexane, ethyl-acetate, and acetone to remove fatty component therefrom.

3. 3^rd step: obtaining step of flavonoid containing fraction

The defatted extract prepared in 2" step, is subjected to removement of non polar organic solvent, adsorption to column chromatography non ionic resin and elution with water in order to eliminate sugar and mineral components, and consecutively, elution with methanol, 10 to 50 % ethyl alcohol, or Ci ~ C₅ lower alcohol or obtain flavonoid containing fraction.

4. 4^th step: separation of AGC The flavonoid containing fraction obtained in 3^rd step, is subjected to sephadex column chromatography such as sephadex LH 20, and recrystallized to separate and obtain pure AGC. The present invention also provides a pharmaceutical composition comprising apigenin-7-O-β-D-glucuronide (AGC) prepared by above-mentioned process as an active ingredient for the prevention and treatment of gastritis and reflux esophagitis.

The Pharmaceutical composition of the present invention for the treatment of gastritis and reflux esophagitis contains between 0.5 to 50 % by weight of AGC based on the total weight of the composition.

The inventive composition comprising AGC may additionally comprise conventional carrier, excipients or diluents in accordance with a using method. 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.

The pharmaceutical composition of the present invention in pharmaceutically acceptable carriers, excipient and diluents may be contains 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, emulsifϊers, preservatives and the like. The composition of the invention may be formulated so as to provide quick, sustained or delayed release of the active ingredient after their administration to a patient by employing any of the procedures well known in the art.

For example, the composition of the present invention can be dissolved in oils, propylene glycol or other solvents which are commonly used to produce an injection.

Suitable examples of the carriers include physiological saline, polyethylene glycol, ethanol, vegetable oils, isopropyl myristate, etc., but are not limited to them. For topical administration, the compounds of the present invention can be formulated in the form of ointments and creams.

Pharmaceutical formulations containing present composition may be prepared in any form, such as oral dosage form (powder, tablet, capsule, soft capsule, suspension, emulsion, syrup, aerosol, elixirs pill, 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 crude drug composition of the present 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 AGC compound of the present invention may varies depending on the condition of the age, sex, and body weight of the individual patient. However, the dose may be administered in single or divided into several times as the dosage between 0.1 to 500 mg/kg by weight per day. The amount of administration may increase and decrease according to route of administration, condition of disease, the sex, the weight, the age, etc. Therefore, the above dose should not be intended to further illustrate the present invention without limiting its scope. The pharmaceutical composition comprising AGC compound of present invention can be using diversely to medicine for treatment and prevention, food, and beverage and the like.

Examples of addable food comprising AGC compound of the present invention are e.g., various food, beverage, gum, vitamin complex, health improving food and the like.

Above described AGC compound therein can be added to food or beverage for prevention of gastritis and reflux esophagitis. For the purpose of preventing gastritis and reflux esophagitis, wherein, the amount of above described AGC compound of the present invention in food or beverage may generally range from about 0.1 to 15 w/w %, preferably 1 to 10 w/w % of total weight of food for the health food composition and 1 to 30 g, preferably 3 to 10 g on the ratio of 100 ml of the health beverage composition. Providing that the health beverage composition of present invention contains above of described AGC compound of the present invention 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 syntlietic 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 πu2 of present beverage composition.

The other components than aforementioned composition are various nutrients, a vitamin, a mineral or an electrolyte, synthetic or natural 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 crude drug composition therein are various food, beverage, gum, vitamin complex, health improving food and the like.

To investigate the protective effect of inventive composition on the reflux esophagitis, we induced reflux esophagitis to the mouse model and confirmed that inventive compound significantly and dose-dependently prevented the development of reflux esophagitis, and gastritis and the lipid peroxidation of cellular membrane. 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.

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. 1 shows the process for preparing AGC isolated from Clerodendron trichotomum

Folium

Fig. 2 shows the effects of AGC and apigenin on the reflux esophagitis induced surgically in rats,

Fig. 3 shows the effect of AGC and apigenin on the indomethacin-induced gastritis,

Fig. 4 shows the effect of AGC and apigenin on the gastric volume,

Fig. 5 shows the effect of AGC and apigenin on the gastric pH,

Fig. 6 shows the effect of AGC and apigenin on the gastric acid output, Fig. 7 shows the effect of AGC and apigenin on the lipid peroxidation.

Best Mode for Carrying Out the Invention

The following Example and Experimental Examples are intended to further illustrate the present invention without limiting its scope.

Example 1. Preparation of the AGC (1)

1 kg of dried Clerodendron trichotomum Folium (purchased in Kyung-dong Market) was mixed with 3 L of 70 % of methanol with 3 times and was subjected to reflux-extraction for 3 hrs at 40 ~ 60 ^°C with 2 times to obtain a crude extract thereof.

After evaporating the solvent with rotary vacuum evaporator (Eyela, FD-5N,

Japan) , obtained 100 g of concentration extract, was dissolved, suspended with 300 itljg of distilled water, filtered with filter paper (Whatman, USA) and then divided into water soluble layer and water insoluble layer.

The water insoluble layer was removed and water soluble layer was defatted with 3 L of chloroform. After water soluble layer was concentrated under reduced pressure to eliminate remaining chloroform, it was subjected to adsorption process with non-ionic resin (Amberite XAD-2) for column chromatography, and followed by eluting with distilled water to eliminate sugar and mineral salts therefrom, eluted with 30 % methanol, and obtained 3.2 g of flavonoid containing fraction. The above fraction was subjected to sephadex column chromatography (30 % ethyl alcohol, 6 ml/min, 5 cm x 1cm) with sephadex LH 20 (Pharmacia co., sweden), and finally separate 100 mg of pure and light yellow AGC which was identified and confirmed as apigenin-7-O-β-D-glucuronide having molecular weight of 632 by the result of instrumental analysis as followings (Fig .1);

mp: 216 ^°C

(-)FAB MS(m/z) : 445[M-H]^_ , 269[M-(GlcUA=H)]^" ¹H- MR(300MHz, DMSO-J₆) ;

7.94(24, d, J=8.7Hz, H-2\ 6 ), 6.93(2H, d, J=8.7Hz, H-3\ 5") 6.84(1H, s, H-8), 6.84(1H, s, H-3), 6.45(1H, d, J=l .5Hz, H-6)

5.25(1H, d, J=6.6Hz, GluUA anorneric H)

¹³C-NMR(75MHz, OMSO-d₆)

164.7(C-2), 103.4(C-3), 182.4(C-4), 161.5(C-5), 99.4(C-6), 162.8(C-7), 84.9(C-8), 157.3(C-9), 105.7(C-10), 121.3(C-r),

128.9(0-2'), 116.3(C-3'), 161.7(C-4^λ), 116.3(C-5'), 128.9(C-6'), 99.6(GluUA: C-l), 72.9(GluUA: C-2), 75.8(GluUA: C-3), 71.4(GluUA: C-4), 75.5(GluUA: C-5), 170.5(GluUA: C-6)

Example 2. Preparation of the AGC (2)

1 kg of dried Clerodendron trichotomum Folium(purchased in Kyung-dong Market) was mixed with 3 L of 70 % ethanol with 3 times and the crude extract was prepared, processed and purified according to the identical method of Example 1 except using other non-ionic resin (Amberite XAD-2) to separate inventive compound, AGC.

Finally, apigenin-7-O-β-D-glucuronide of molecular weight 632 was obtained and identified by the result of instrumental analysis as shown in Example 1.

Experimental Example 1. The inhibitory effects of AGC on reflux esophagitis and gastritis To confirm protective effect of apigenin-7-O-D-glucuropyranoside (AGC) on the reflux esophagitis and gastritis , we adopted the evaluation methods to check the effect on reflux esophagitis disclosed in the literature (Nakamura K, et al; Jpn. J. Pharamcol. 32(3), pp445-56, 1982).

In case reflux esophagitis, male Sprague-Dawley rats (Hanlim, Korea) with a body weight of about 200 g were used for the experiments. The rats were starved for 24 hrs before the experiments, but were freely accessed to drinking water. All animals were kept in raised mesh-bottom cages to prevent coprophagy. Under ether anesthesia, the abdomen was incised along the midline and then both the pylorus and limiting ridge (transitional region between the forestomach and corpus) were simultaneously ligated. A longitudinal cardiomyotomy of about 1 cm length across the gastroesophageal junction was performed to enhance reflux. Six hours later, the animals were killed by cervical dislocation and then the esophagus was harvested and collected gatric juice.

AGC obtained from example 1 and apigenin (A3145, Sigma) dissolved in 0.01% dimethyl sulfoxide were administered intraduodenally (i.d.) immediately after ligation of the pylorus and limiting ridge. The volume of the drug or vehicle was 1 ml/kg of body weight. The drugs were prepared freshly each time.

In case gastritis, male Sprague-Dawley rats (Hanlim, Korea) with a body weight of about 200 g were used for the experiments. The rats were starved for 24 hrs before the experiments, but were freely accessed to drinking water. All animals were kept in raised mesh-bottom cages to prevent coprophagy. AGC and apigenin were administered per se (p.o.) one hour before indomethacin (50 mg/kg, 17378, Sigma) administration. The volume of the drug or vehicle was 2 ml/kg of body weight. The drugs were prepared freshly each time. After five hours, the animals were sacrificed, fixed with 3% formalin and then their stomachs were excised. The stomachs were opened along the greater curvature and spread out with pins on a cork board.

The area (mm²) of mucosal erosive lesions was measured under a dissecting microscope with a squared grid (XI 0; Olympus, Tokyo, Japan).

To detect lipid peroxidation, esophageal mucosa was harvested, and sonicated in 1 ml of Tris-HCl buffer (pH 7.0). After centrifugation at 600 g for 5 min at 4 °C (Micro 17TR, Hanil, Korea), 0.9 ml of trichloroacetic acid (8%) was added to 0.3 ml of supernatant. After centrifugation at 10,000 g for 5 min at 4 °C, 0.25 ml of TBA (1 %) was added to 1 ml of supernatant and the resulting solution was heated at 100 °C for 20 min. The tubes were cooled, 2 ml of n-butanol was added and each tubes was vortexed for 90 sec. After centrifugation at 3,000 g for 5 min at 4 °C, 1 ml of butanol phase was utilized for TBARS assay at 532 nm (UN-160A, Shimadzu, Japan) against malonaldehydebis(dimethylacetal) standards. Results were expressed as pmol/mg protein. Protein assay was determined according to the Bradford method.

In the experiment of reflux esophagitis, severe ulcerations were observed in the esophagus of control group. While both AGC (0.55mg/kg) and apigenin (lOmg/kg) treated groups decreased the development of reflux esophagitis significantly (Fig. 2). As shown in Fig 1, the level of ulcer index shows that AGC has more potent efficacy than apigenin, 1.7 and 2.0, respectively. AGC and apigenin also reduced a significant increase in index of gastric ulceration (4) induced by exposure of the gastric mucosa to indomethacin. (0, no visible lesions; 1, a few erosions; 2, total area of lesions 30 mm² 3, total area of lesions 30 mm² 4, perforation, Significant differences from the corresponding control, respectively, *P<0.05). As shown in Fig. 3, in gatritis, apigenin and AGC administered i.d. (0.3 mg/kg) group showed the gastric lesion of ulceration 1.7 cm² and 0.4 cat, respectively. AGC significantly reduced a increase in gastric lesion of ulceration (2.8) induced by exposure of the gastric mucosa to indomethacin. Significant differences from the corresponding control, apigenin, *P<0.05, #P<0.05, respectively. Apigenin and AGC administered i.d. (0.3 mg/kg) group decreased the gastric volume 0.9 mi and 0.8 iuE, respectively (Fig. 4). This result showed that decreased the gastric volume of indomethacin administration (1.8 ml) group. Significant differences from the corresponding control, respectively. *P<0.05). As shown in Fig. 5, about pH of the gastric content, apigenin (0.3 mg/kg) and AGC (0.3 mg/kg) administered i.d. observed the gastric pH 3.2 and 4.4, respectively. This result showed that significantly neutralized pH of indomethacin administration (2.5). Significant differences from the corresponding control denotes, respectively. *P<0.05. Administration of apigenin (3 mg/kg) and AGC (0.3 mg/kg) significantly and dose-dependently decreased acid output, the level of acidity were 51 mEq/hr and 49 mEq/hr, respectively(Fig. 6). This result showed effect reduced comparing with that of control (145 mEq/hr) adminstrated with indomethacin. From these results, it is suggested that the inhibitory effects of AGC and apigenin on the reflux esophagitis and gastritis on rats was confirmed. Significant differences from the corresponding control denotes, respectively. *P<0.05.

Lipid peroxidation of cellular membrane is the result of cell damage by free radicals. Malondialdehyde (MDA), the end product of lipid peroxidation, was reacted with TBA to show fluorescence and that is a sensitive marker of oxidative stress (Buege JA, Aust SD. Microsomal lipid peroxidation. Methods in Enzymol. 52, pρ302-310, 1978). AGC administration (0.3mg/kg) significantly decreased the amounts of MDA formation, which shows the inhibition of reflux esophagitis- associated lipid peroxidations comparing with apigenin administration (3mg/kg) group (Fig. 7). This result suggests that AGC has an antioxidative effect.

Finally, this study demonstrates that AGC showed potent efficacy on the development of reflux esophagitis and indomethacin-induced gastritis, by the inhibition of gastric secretion and the prevention of oxidative stress. The AGC is more protent than apigenin, one of anti-inflammatory drug. We conclude that AGC may be promising drug for the treatment of reflux esophagitis and gastritis.

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

Compound of Example 1 50mg

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

Preparation of tablet

Compound of Example 1 50mg

Corn Starch lOOmg

Lactose lOOmg

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

Preparation of capsule

Compound of Example 1 50mg

Corn Starch lOOmg Lactose lOOmg

Magnesium Stearate 2mg Tablet preparation was prepared by mixing above components and filling gelatin capsule by conventional gelatin preparation method.

Preparation of injection

Compound of Example 1 50mg

Distilled water for injection optimum amount

PH controller optimum amount Injection preparation was prepared by dissolving active component, controlling pH to about 7.5 and then filling all the components in 2ml ample and sterilizing by conventional injection preparation method.

Preparation of liquid

Compound of Example 1 0.1~80g Sugar 5-1 Og

Citric acid 0.05-0.3%

Caramel 0.005-0.02%

Vitamin C 0.1-1% Distilled water 79-94%

CO₂ gas 0.5-0.82%

Liquid preparation was prepared by dissolving active component, filling all the components and sterilizing by conventional liquid preparation method.

Preparation of health care food Compound of Example 1 lOOOmg Vitamin mixture optimum amount

Vitamin A acetate 70μg

Vitamin E l.Omg

Vitamin Bi 0.13mg

Vitamin B₂ 0.15mg

Vitamin B6 0.5mg

Vitamin B 12 0.2μg

Vitamin C lOmg

Biotin lOμg

Amide nicotinic acid 1.7mg

Folic acid 50μg

Calcium pantothenic acid 0.5mg Mineral mixture optimum amount

Ferrous sulfate 1.75mg

Zinc oxide 0.82mg

Magnesium carbonate 25.3mg

Monopotassium phosphate 15mg Dicalcium phosphate 55mg Potassium citrate 90mg Calcium carbonate lOOmg Magnesium chloride 24.8mg

The above-mentioned vitamin and mineral mixture may be varied in may ways. Such variations are not to be regarded as a departure from the spirit and scope of the present invention. Preparation of health beverage Compound of Example 1 lOOOmg Citric acid lOOOmg Oligosaccharide 100g Apricot concentration 2g Taurine lg Distilled water 900mβ

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 lOOOmβ 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

The pharmaceutical composition comprising inventive apigenin-7-O-β-D- glucuronide isolated from Clerodendron trichotomum Folium leaves may be promising drug for the prevention and treatment of gastritis and reflux esophagitis.

Claims

1. A process for preparing apigenin-7-O-β-D-glucuronide (AGC) isolated from Clerodendron trichotomum Folium comprising the steps of; obtaining the crude extract Clerodendron trichotomum Folium; defatting the crude extract by using non-polar solvent; removing sugar and mineral salts from above defatted extract and obtaining flavonoid fraction by using non-ionic exchange column chromatography; and purifying the fraction by recrystallizing to obtain above compound.

2. A pharmaceutical composition comprising a pharmaceutically effective amount of apigenin-7-O-β-D-glucuronide (AGC) compound with a pharmaceutically acceptable carrier, excipients and diluents for the prevention and treatment of gastritis and reflux esophagitis.

3. The pharmaceutical composition according to claim 2, wherein said composition is containing 0.5 to 50 weight % of the above apigenin-7-O-β-D- glucuronide (AGC) compound based on the total weight of the composition.

4. Use of apigenin-7-O-β-D-glucuronide (AGC) compound for preparation of medicament for gastritis and reflux esophagitis.