WO2008048044A1

WO2008048044A1 - Extract of agastache rugosa for treating and preventing gastrointestinal diseases and a process for preparing the same

Info

Publication number: WO2008048044A1
Application number: PCT/KR2007/005081
Authority: WO
Inventors: Sung Sick Woo; Ji Min Cha; Dong Seon Kim; Seon Gil Do; Young Chul Lee; Jeong Bum Nam; Tae Woo Kim
Original assignee: Unigen, Inc.
Priority date: 2006-10-17
Filing date: 2007-10-17
Publication date: 2008-04-24
Also published as: KR100807645B1

Abstract

The present invention relates to a composition comprising Agastache rugosa extract for the treatment and prevention of gastrointestinal disease, and a process for preparing the same. The Agastache rugosa extract of the present invention which is extracted from natural plant, also provides a composition for the treatment and prevention of gastrointestinal disease having good in vivo stability.

Description

EXTRACT OF AGASTACHE RUGOSA FOR TREATING AND PREVENTING GASTROINTESTINAL DISEASES AND A PROCESS FOR PREPARING THE SAME.

TECHNICAL FIELD

The present invention relates to a composition comprising Agastache rugosa extract for the treatment and prevention of gastrointestinal diseases, and a process for preparing the same.

10 BACKGROUND ART

Gastrointestinal disease is one of the widespread digestive system diseases, and indicates that ulcer is formed in the parietal mucosa by secretion of gastric acid. About 10% of the population experiences gastrointestinal disease more than once in their lives.

The stomach wall consists of mucosa, submucosa, muscularis externa, and serosa.

15 The symptoms of gastrointestinal disease are varied according to the damaged region.

The pathogenesis of gastrointestinal disease is not completely understood. It is generally known that imbalance between attack factors and defense factors, i.e., increase of attack factors or decrease of defense factors, can develop gastrointestinal disease. Attack factors are increased by increase of acids or pepsin secretion, and the like. Defense factors are weakened by defect of the structure or form of gastric mucosa, or decrease of secretion of mucus or bicarbonate ion, inhibition of production of prostaglandin, and the like. Recently, there are some reports that Helicobacter pylori is associated with gastrointestinal disease.

Among these factors, the main cause of development of gastrointestinal disease is excessive secretion of gastric acid. Gastric acid is a high concentration of hydrochloric acid (HCl) secreted in the stomach, and activates pepsinogen into pepsin, thereby breaking the bonds linking amino acids, which is a process known as proteolysis. Gastric acid also critically affects damaged gastroparietal layer.

Gastric acid is secreted from parietal cells in the stomach, and three (3) receptors are associated with the secretion of gastric acid. These three receptors are stimulated by histamine, gastrin, or acetylcholine, to secrete gastric acid.

As therapeutic agents for gastrointestinal disease, drugs to enhance defense factors are used, with including antiacid that can neutralize gastric acid secreted but has no effect to a total amount of gastric acid secreted; drug to inhibit secretion of gastric acid; accelerator of secretion of prostaglandin; coating agent for stomach wall, and the like.

However, gastrointestinal disease is caused by complex factors including psychological stress, and the cause has not been completely understood. Thus, there is no absolute method to cure gastrointestinal disease.

Early 19^th century, Beaumont reported that string liquor can cause erythema and pathological change in gastric mucosa of human (Beaumont W: Experiments and observations on the gastric juice and the physiology of digestion. Plattsburgh, NY; EP Allen, 237-9, 1833). Ever since, there have been reports about the effect of alcohol to the gastric mucosa in various species including human. These reports correspond with a report that mucous and submucous bleeding, and necrosis edema can be developed in an alcohol having more than 35% of concentration.

Despite extensive studies in human and animal, etiology of gastric mucosa injury by alcohol is not completely understood yet. Palmer reported that the development of acute gastrointestinal disease is mediated by the change of blood stream upon drinking a large amount of alcohol. Wallace et al. reported that a material having cell-protective property can suppress deep mucosa bleeding caused by alcohol. Gute et al. reported that 100% of alcohol can completely block the blood stream in an injured mucosa region. In short, alcohol can induce the obstruction of gastric blood stream to cause bleeding quickly.

Agastache rugosa is a perennial herb in Labiatae family, and is distributed in Korea, China, Japan, and North-East Asia. In Korea, Agastache rugosa grows wild especially in south region of Korea, or some are cultivated. In Korean traditional oriental medicine, the whole plant of Agastache rugosa is called "Agastache Herba." One of the Korean traditional oriental medicine book called Myung-ui Byulrok discloses that Agastache Herba has antiemetic effect, antipyretic effect, and elimination of bad energy. Also, leaves of Agastache rugosa have been used as flavor of cooking food, and the flowers of Agastache rugosa have been used as the sucking source of a bee.

As the elements of Agastache rugosa, there are reported essential oil [J. Essent. Oil Res. 8(2), 135-138, 1996; ibid 4(6), 585-587, 1992; Korean Journal of Food Science and Technology 23(5), 582-586, 1991; J. Agric. Food Chem. 40(8), 1362-1366, 1992], sesquiterpenes [Yakugakuzasshi 92(7), 908-909, 1972], diterpenes [Kor. Patent No. 10- 9608662; Chin. Pharm. Sci. 6(3), 115-118, 1997; Nat. Prod. Sci. 25(4), 319-327, 1994; ibid 18(2), 99-102, 1987], triterpenes [Yaozue Xuebao 26(12), 906-910, 1991; Nat. Prod. Sci. 19(2), 97-98, 1988; ibid 18(1), 50-53, 1987], flovonoids [Yaozue Xuebao 26(12), 906-910, 1991; Chem. Pharm. Bull. 29(6), 1777-1779, 1981], phenylpropanoids [Yakugaku Zasshi 106(12)1108-1111, 1986], and carotenoids [Nat. Prod. Sci. 30(4), 404-

408, 1999].

As the physiological activities of Agastache rugosa, it has been reported that

Agastache rugosa extract has antibacterial activity [Phytother. Res. 14(3), 210-212, 2000; J. Food Sci. Nutr. 4(2), 97-102, 1999], antiviral activity [Arch. Pharm. Res. 22(5), 520-

523, 1999; US Patent 5776462], monoamineoxidase inhibition activity [Yakhak hoeji

42(6), 634-638, 1998]. Essential oil in the elements of Agastache rugosa has been reported to have antibacterial activity [Zhongguo Yaozue Zazhi 35(1), 9-11, 2000;

Weishengwuxue Zazhi 18(4), 1-4, 16, 1998], and mosquitoes shirking activity [Chinese Patent No. 1044205]. Also, carotenoids are reported to have anticancer activity [Nat.

Prod. Sci. 30(4), 404-408, 1999], and diterpenes are reported to have anticancer activity [J.

Nat. Prod. 58(11) 1718-1821, 1995], and antiviral activity [Arch. Pharm. Res. 22(1), 75-

77, 1999]. Further, phenylpropanoids are reported to have antiviral activity [Arch.

Pharm. Res. 22(5), 520-523, 1999], antioxidation activity [Kor. Agri. Chem. & Biotech. 42(3), 262-266, 1999], and anticomplement activity [Nat. Prod. Sci. 27(1), 20-25, 1996;

Kor. Agri. Chem. & Biotech. 39(2), 147-152, 1996].

Despite comprehensive studies and interests in gastrointestinal disease,

Agastache rugosa extract for the treatment and prevention of gastrointestinal disease has not been known in the field. Synthetic drugs generally used for treating gastrointestinal disease have a high risk of causing other side effects when used for a long time.

The inventors of the present invention have studied wild plants to find out plants which have a useful effect for gastrointestinal disease, and confirmed that Agastache rugosa extract, and tilianine, acacetin, and agastachoside isolated from the Agastache rugosa extract show a superior effect on gastrointestinal disease, to complete the present invention.

DISCLOSURE OF THE INVENTION The objection of the present invention to provide a composition comprising

Agastache rugosa extract, or tilianine, acacetin, and agastachoside isolated from Agastache rugosa extract, for the treatment and prevention of gastrointestinal disease, and a process for preparing the same.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 is a graph showing in vivo action of Agastache rugosa extract, or tilianine, acacetin, and agastachoside isolated from Agastache rugosa extract according to the present invention in a gastrointestinal disease model. Fig. 2 is a photo of a rat's stomach showing the in vivo action of Agastache rugosa extract, or tilianine, acacetin, and agastachoside isolated from Agastache rugosa extract according to the present invention, compared with the control group and negative control group in a gastrointestinal disease model.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a composition comprising Agastache rugosa extract for the treatment and prevention of gastrointestinal disease, and a process for preparing the same. The present invention relates to a composition comprising more than one components selected from the group consisting of tilianine, acacetin, and agastacoside as effective components, for the treatment and prevention of gastrointestinal disease, and a process for preparing the same.

The present invention relates to an agent comprising Agastache rugosa extract for the treatment of gastrointestinal disease.

The present invention relates to a method for preparing a composition comprising Agastache rugosa extract as an effective component which comprises, 1) extracting the Agastache rugosa in water or alcohol and obtaining the extract; 2) filtrating the extract; 3) concentrating the filtrated extract under reduced pressure; and 4) further concentrating the extract by freeze-drying the concentrated extract.

In an embodiment of the invention, Agastache rugosa extract of the present invention is water or alcohol extract, preferably alcohol extract of Agastache rugosa (Labiatae). Agastache rugosa extract of the present invention can be obtained through extracting the Agastache rugosa (Labiatae) in water or alcohol, preferably alcohol. In detail, Agastache rugosa was dried in shade, and powdered. The powdered material is extracted with water or lower alcohol such as methanol and ethanol, or mixture thereof (1:0.1 to 1:10, preferably 1:0.2 to 1:5, kg/L) and the amount of the extract solvent is 5 to 20 times the powdered material, preferably about 10 to 15 times. The powdered material is extracted with preferably 20-30% of ethanol, at 80-100 ^°C, preferably 90- 100°C, for 1-10 h, preferably 3~7 hr, by solvent extract, edge hot water extraction, or ultrasonic extraction (1~5 times, preferably 2-3 times). Thus obtained extract is filtrated, concentrated under reduced pressure, and freeze-dried to obtain Agastache rugosa extract of the present invention.

In another embodiment, Agastache rugosa extract of the present invention is prepared by a method comprising the following steps, for example: 1) extracting the Agastache rugosa in water or polar solvent such as methanol and ethanol, and obtaining the extract;

2) filtrating the extract with fibrous filter under reduced pressure;

3) concentrating the obtained extract under reduced pressure to remove excess solvent; and, 4) concentrating the extract by freeze-drying the concentrated extract. hi still another embodiment of the invention, the above Agastache rugosa extract is absorbed to a silica gel whose amount is 0.5 to 3 times the extract weight (kg), preferably about 1 to 2 times, and then is applied to a column filled with silica gel. Total seven (7) column fractions are obtained from dichloromethane : methanol solvent gradient (90:10 to 0:100). A crystal from sixth fraction is washed with methanol several times, and then pure tilianine having the following formula is isolated therefrom.

Also, a crystal from first fraction is washed with methylenechloride several times, and pure acacetin having the following formula is isolated therefrom.

A crystal from fourth fraction is washed with methylenechloride several times, and pure agastachoside having the following formula is isolated therefrom.

Agastache rugosa extract, tilianine, acacetin, and agastachoside obtained or isolated by the above methods were tested for the treatment effect on gastrointestinal disease in an animal model, and confirmed that they can be applied for the treatment or prevention of gastrointestinal disease.

The present invention provides a composition comprising Agastache rugosa extract as an active component obtained by the above method for the treatment or prevention of the gastrointestinal disease.

The present invention provides a composition obtained by the above method, comprising more than one component selected from the group consisting of tilianine, acacetin, and agastachoside as effective components for the treatment and prevention of gastrointestinal disease.

The present invention provides a method for preparing a composition comprising

Agastache rugosa extract as an effective component which comprises, 1) extracting the Agastache rugosa in water or alcohol and obtaining the extract; 2) filtrating the extract; 3) concentrating the filtrated extract under reduced pressure; and 4) concentrating the extract by freeze-drying the concentrated extract.

The gastrointestinal disease in the present invention includes all kinds of gastrointestinal disorders accompanying flare, bleeding or ulcer of gastrointestine, and also includes chronic or acute gastritis, gastric ulcer, gastric cancer, or gastrorrhagia, but are not limited thereto.

The pharmaceutical composition of the present invention can include pharmaceutically acceptable carriers. The carriers include conventionally used solvent, dispersion medium, absoption retardant in this field, but lactose, dextrose, sucrose, solbitol, mannitol, xylitol, erythritol, maltitol, starch, lactose, acasia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and the like are preferable. The pharmaceutical composition of the present invention can be administered by using any route as long as it can reach target area. Accordingly, the composition of the present invention can be administered by oral, intranasal, intravenous, intramuscular, rectal, subcutaneous, intrauterine, and percutaneous method, and the like. Also, the composition of the present invention can be formulated to oral preparation including powder, granule, tablet, capsule, suspension, emulsion, syrup, and aerosol; external preparation; suppository preparation; and injection preparation.

The dose of the pharmaceutical composition of the present invention depends on the age, sex, body weight, and severity of disease of a patient, and is generally 1 to 500 mg/kg, preferably 50 to 400 mg/kg, once to three times a day, but is not limited thereto. [Advantageous Effect]

Agastache rugosa extract, tilianine, acacetin, and agastachoside obtained or isolated by the above methods are useful for the treatment of gastrointestinal disease in an animal model, and so can be applied for the treatment or prevention of gastrointestinal disease.

EXAMPLES

The present invention relates to a composition comprising Agastache rugosa extract for the treatment and prevention of gastrointestinal disease.

The present invention relates to a composition comprising more than one component selected from the group consisting of tilianine, acacetin, and agastacoside as effective component, for the treatment and prevention of gastrointestinal disease. The present invention relates to an agent comprising Agastache rugosa extract for the treatment of gastrointestinal disease.

The present invention relates to a method for preparing a composition comprising

Agastache rugosa extract as an effective component which comprises, 1) extracting the

Agastache rugosa in water or alcohol and obtaining the extract; 2) filtrating the extract; 3) concentrating the filtrated extract under reduced pressure; and 4) concentrating the extract by freeze-drying the concentrated extract.

The following examples are illustrated as the best mode by the inventor for practice of the present invention, and should not be construed to limit the scope of the present invention in any manner.

Example 1. Preparation of Agastache rugosa extract

A whole plant of Agastache rugosa (Labiatae) was collected (Kyung-san, Kyung sang book-do, Republic of Korea). This whole plant was washed with clean water, and dried in shade for 10 days.

The dried material (30 kg) was powdered and extracted with 25% of ethanol (300 mL) at 95 ^°C for 5 hr (X 2). At the end, the solvent was filtered with fibrous filter

(20OuM) under reduced pressure. The extract is then concentrated under reduced pressure to remove ethanol completely. The concentrated extract (40 L) was dried to obtain powdered Agastache rugosa extract (4.2 kg).

Example 2. Preparation of tilianine

Dried powder of Agastache rugosa extract obtained from Example 1 (100 g) is absorbed to about 100 g of silica gel, and then loaded to silica gel column (5 * 45 cm) filled with about 1 kg of silica gel. Total seven (7) fractions are obtained by dichloromethane : methanol solvent gradient (90:10 ~ 0:100). A crystal from sixth fraction was washed with methanol several times, and pure tilianine was isolated (130 mg) therefrom.

Example 3. Preparation of acacetin

Dried powder of Agastache rugosa extract obtained from Example 1 (100 g) is absorbed to about 100 g of silica gel, and then loaded to silica gel column (5 x 45 cm) filled with about 1 kg of silica gel. Total seven (7) fractions are obtained by dichloromethane : methanol solvent gradient (90:10 ~ 0:100). A crystal from first fraction was washed with methylenechloride several times, and pure acacetin was isolated (42 mg) therefrom.

Example 4. Preparation of agastachoside

Dried powder of Agastache rugosa extract obtained from Example 1 (100 g) is absorbed to about 100 g of silica gel, and then loaded to silica gel column (5 x 45 cm) filled with about 1 kg of silica gel. Total seven (7) fractions are obtained by dichloromethane : methanol solvent gradient (90:10 ~ 0:100). A crystal from fourth fraction was washed with methanol several times, and pure agastachoside was isolated (22 mg) therefrom.

Experiment 1. Measurement of the treatment effect for gastrointestinal disease in rat model

In order to measure the treatment effect of Agastache rugosa extract, tilianine, acacetin, and agastachoside obtained from Examples 1 to 4, for gastrointestinal disease, the following experiments were conducted. 5-weeks of Sprague-Dawley (SD) male rats which were pre-fed for 1 week were divided into 6 groups, five (5) rats each. Two (2) groups among the 6 groups were designated as negative control group (normal group) and control group (gastrointestinal disease group), each, and the other 4 groups were designated as experimental groups. After the above male rats were fasted for 24 hours, in order to develop gastrointestinal disease, ethanol was administered to the control group except for the negative control group and experimental groups. Agastache rugosa extract, tilianine, acacetin, and agastachoside obtained from Examples 1 to 4, were administered to the experimental groups which developed gastrointestinal disease, respectively, and then observed the treatment effect for the gastrointestinal disease.

Each animal was killed and let blood out, and the stomach was cut and fixed after cutting according to greater curvature, to observe the gastric lesion with the naked eyes. Gastric lesion index (mm) were measured for each animal by Mizui and Dodeuchi's method (Jpn. J. Pharmacol., 33, 939-945, 1983), and the group means were compared and leasured as inhibition percentage (%) to evaluate the extent of the gastrointestinal disease. The results are given in Table 1 and Fig. 1. [Table 1]

As shown in Table 1 and Fig. 1, Agastache rugosa extract, tilianine, acacetin, and agastachoside were useful for the treatment of gastrointestinal disease in the animal model, compared with the control group.

The above incised stomachs of rats were observed after the histopathological treatment, and the results are shown in Fig. 2. It was confirmed from Fig. 2 that

Agastache rugosa extract, tilianine, acacetin, and agastachoside have good effect, i.e., the extent or size of gastric flare, bleeding or ulcer was decreased, in the treatment groups, compared with the control group.

Agastache rugosa extract of the present invention can be administered as the following formulations, which however are illustrated only as examples and should not be construed to limit the scope of the present invention in any way.

Formulation 1. Preparation of Pulvis

Agastache rugosa ethanol extract 100 mg

Corn starch lOO mg lactose 100 mg talc 10 mg

The above components were mixed together. An air-tight container was filled with the mixed composition, and then the pulvis was prepared.

Formulation 2. Preparation of Tablet Agastache rugosa ethanol extract 100 mg Corn starch 100 mg lactose 100 mg

Magnesium stearate 2 mg

The above components were mixed together, and the tablet was prepared by using the conventional tableting method.

Formulation 3. Preparation of Capsule Agastache rugosa ethanol extract 100 mg Corn starch 100 mg lactose 50 mg

Magnesium stearate 1 mg The above components were mixed together, and filled in a gelatin capsule by using the conventional capsulation method.

INDUSTRIAL APPLICABILITY

The present invention provides a composition comprising Agastache rugosa extract for the treatment and prevention of gastrointestinal disease, and a process for preparing the same. The Agastache rugosa extract of the present invention is extracted from natural plant, and so can provide a composition for the treatment and prevention of gastrointestinal disease that has good in vivo stability.

Claims

1. A pharmaceutical composition for the treatment and prevention of gastrointestinal disease comprising: an Agastache rugosa extract as an active component, and pharmaceutically acceptable carriers.

2. The composition according to claim 1, wherein the Agastache rugosa extract comprises tilianine, acacetin, or agastachoside.

3. The composition according to claim 1, wherein the extract is alcohol extract.

4. A pharmaceutical composition comprising more than one component selected from the group consisting of tilianine, acacetin, and agastacoside as effective components, for the treatment and prevention of gastrointestinal disease.

5. The composition according to any one of claims 1 to 4, wherein the gastrointestinal disease is chronic or acute gastritis, gastric ulcer, gastric cancer, or gastrorrhagia.

6. An agent for the treatment of gastrointestinal disease comprising Agastache rugosa extract as an active component.

7. A method for preparing a composition comprising Agastache rugosa extract as an effective component which comprises, 1) extracting the Agastache rugosa in water or alcohol and obtaining the extract; 2) filtrating the extract; 3) concentrating the filtrated extract under reduced pressure; and 4) concentrating the extract by freeze- drying the concentrated extract.

8. The method according to claim 7, wherein in step 1), extraction of the Agastache rugosa is conducted at 90-100 ^°C for 3-7 hr.