KR101961641B1 - Anti-Helicobacter pylori composition comprising 2-alkoxy-6-acetyl-7-methyljuglone as effective component - Google Patents

Abstract

The present invention relates to an anti-Helicobacter pylori composition comprising 2-alkoxy-6-acetyl-7-methyl juglone of formula (1) as an active ingredient, wherein the 2-alkoxy- It can be used not only for prevention, improvement or treatment of infectious diseases caused by Helicobacter pylori but also for antibiotic resistance since it does not cause side effects due to natural products and does not cause antibiotic resistance, Is expected to be effectively used as a food or pharmaceutical composition.

Description

The present invention relates to an anti-Helicobacter pylori composition comprising 2-alkoxy-6-acetyl-7-methyljuglone as an effective component and a 2-alkoxy-

The present invention relates to an anti-Helicobacter pylori composition containing 2-alkoxy-6-acetyl-7-methyl juglone as an active ingredient.

Helicobacter pylori is a pathogenic bacterium parasitic to the gastric mucosa of humans, gram-negative bacteria, spiral, microaerophilic, and bivalve. Helicobacter pylori produces urease, which decomposes urea to form ammonia and carbon dioxide, and this ammonia neutralizes the acidic environment. These bacteria produce VacA (vacuolating toxin) toxin, and the products made by the CagA (cytotoxin-associated gene) gene have been reported to cause toxicity. Helicobacter pylori was found by Warren and Marshall in 1983 in the gastric pylorus of patients with chronic gastritis and is known to cause gastrointestinal diseases such as gastritis, gastrointestinal ulcer, and gastric cancer. In addition, recent studies have shown that Helicobacter pylori is associated with other diseases such as chronic hepatitis C and Parkinson's disease.

Helicobacter pylori infection treatments include proton pump inhibitors, triple therapy to prescribe clarithromycin or metronidazole to amoxicillin, sequential therapies to prescribe them in turn, sequential therapy with bismuth carassylate (bismuth subsalicylate-based sequential therapy), which is based on bismuth subsalicylate and triple therapy, but side effects such as relapse, recurrence, vomiting and diarrhea are still present do. Therefore, unlike the previously reported antimicrobial substances, there are no side effects, resistance and stability are not raised, and there is a growing demand for development of a Helicobacter pylori antimicrobial composition which is easy to take. Particularly, there is a desperate need for research on a substance having antimicrobial activity against Helicobacter pylori centered on natural products which can be used as an adjuvant while having few side effects.

Polygoni cuspidatum is a tall, perennial herb that belongs to Polygonaceae. The stem of this plant grows straight and hollow, and the nodes have red or purple spots. This plant is widely distributed in Asia or North America. The roots are known to contain mainly secondary metabolites emodin, polydatin, resveratrol, physcion and anthraglycoside B. Until recently, over 67 compounds such as quinones, stilbenes, flavonoids, coumarins and lignans have been isolated and reported. On the other hand, more than one hundred kinds of prescriptions including Hojangun have been used for inflammation, jaundice or skin rash. Recently, anti-inflammatory, antioxidant, anti-cancer, fat control, anti-bacterial, antiviral and antifungal activity have been reported.

In the present invention, the structure of the physiologically active ingredient was isolated from the 70% ethanol extract of Root muscle, which had strong antihericobacterial effect, and the anti-Helicobacter pylori activity of these substances was evaluated.

Korean Patent No. 1032066 discloses a pharmaceutical composition for the prevention and treatment of colds comprising the extracts, fractions thereof or stilbene compounds thereof, and Korean Patent Publication No. 1077202 discloses a composition for preventing and treating Helicobacter pylori Quot; antimicrobial extracts for use in the present invention, " but the anti-Helicobacter pylori composition containing the 2-alkoxy-6-acetyl-7-methyl juglone of the present invention as an active ingredient has not been disclosed.

SUMMARY OF THE INVENTION The present invention has been made in view of the above-mentioned needs, and it is an object of the present invention to identify the structure of nine physiologically active ingredients from a 70% ethanol extract of Hojanggok, which has a strong effect on anti-Helicobacter pylori. 2-methoxy-6-acetyl-7-methyl juglone and 2-ethoxy-6-acetyl-7-methyl juglone have excellent anti-helicobacter pylori activity.

In order to solve the above problems, the present invention provides an antimicrobial composition comprising 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.

The present invention also provides a health food composition for preventing or ameliorating infectious diseases caused by Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.

The present invention also provides a pharmaceutical composition for preventing or treating infectious diseases caused by Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.

The 2-alkoxy-6-acetyl-7-methyl juglone of the present invention shows excellent antimicrobial activity against Helicobacter pylori, so that it can be effectively used for preventing, improving or treating infectious diseases caused by Helicobacter pylori. It is expected that it can be effectively used as a functional food or a pharmaceutical composition because it does not cause side effects and does not cause a problem of antibiotic resistance.

Fig. 1 shows the chemical structures of Compounds 1 to 9 isolated from callus root.
Fig. 2 shows the anti-helicobacter activity of the compounds 1 to 9 isolated from callus root.

In order to achieve the object of the present invention, the present invention provides an antimicrobial composition comprising 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.

The 2-alkoxy-6-acetyl-7-methyl juglone of the present invention may be 2-methoxy-6-acetyl-7-methyl juglone or 2-ethoxy- But is not limited thereto.

The term " antimicrobial " means the ability to reduce, prevent, inhibit, or eliminate the growth or survival of a microorganism at any concentration, and may preferably mean anti-bacterial, May be Helicobacter, and more preferably Helicobacter pylori, but is not limited thereto.

In one embodiment of the present invention, 2-alkoxy-6-acetyl-7-methyl juglone is characterized in that it has an antibacterial effect against Helicobacter pylori.

The 2-alkoxy-6-acetyl-7-methyl juglone of the present invention can be isolated, purified or chemically synthesized from Hojokgang extract and preferably isolated from Hojanggang extract. However, the present invention is not limited thereto. The chemical synthesis of the compound may be performed by a general method known in the art.

The callus root extract according to the present invention can be prepared by extracting various plant tissues with an organic solvent. But it is not limited thereto. As the organic solvent for extraction in the present invention, solvents such as n -hexane, ethyl acetate, acetone, butanol, and ethanol may be used alone. In the present invention, a fractionation process commonly used in the art may also be performed. Preferably, the extract may be a fraction obtained by adding ethyl acetate to the ethanol extract obtained by adding ethanol to the root muscle, but the present invention is not limited thereto.

The compound can be prepared by a method comprising fractionating an organic solvent extract of phytophthora inflorescences by column chromatography, and isolating and purifying the extract. Examples of the usable column include a silica gel column, a reversed phase silica gel column or a sephadex column. The eluent may be an organic solvent such as hexane, ethyl acetate, chloroform, methylene chloride, acetone and lower alcohols, Preferably n -hexane, ethyl acetate, acetone, methanol, but most preferably ethyl acetate.

In one embodiment of the present invention, nine kinds of active compounds having anti-helicobacter activity were identified from the ethyl acetate fraction of acanthin gum, and 2-methoxy-6-acetyl-7-methyl juglone And the 2-alkoxy-6-acetyl-7-methyl juglone compound of 2-ethoxy-6-acetyl-7-methyl juglone had the best anti-helicobacter activity. This was the first report by the inventors of the activity of 2-alkoxy-6-acetyl-7-methyl juglone.

The present invention also provides a health food composition for preventing or ameliorating infectious diseases caused by Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.

The compounds of the present invention are effective for preventing or ameliorating infectious diseases caused by Helicobacter pylori. In the present invention, infectious disease caused by Helicobacter pylori is a disease caused or weakened by infection, survival or proliferation of Helicobacter pylori in a living body, and the symptom can be improved by eliminating Helicobacter pylori. Examples of such diseases include, but are not limited to, gastritis, gastric ulcer, duodenal ulcer, non-ulcer digestive syndrome, gastric MALT lymphoma, gastric hyperplastic polyps, gastric cancer, gastrointestinal cancer, pancreatitis, inflammatory bowel disease and the like.

In addition, the compound of the present invention can be added to food or beverage for the purpose of preventing or improving infection by Helicobacter pylori. The amount of the compound in the food or drink may be 0.01 to 15% by weight of the total food, and the health beverage composition may be added in a proportion of 0.02 to 5 g, preferably 0.3 to 1 g, based on 100 ml.

The health functional beverage composition of the present invention is not particularly limited to the other ingredients other than the above-mentioned compounds as essential ingredients in the indicated ratios and may contain various flavors or natural carbohydrates as additional ingredients such as ordinary beverages. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. Natural flavors (tautatin, stevia extract, for example, rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.) can be advantageously used as flavors other than those described above . The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the composition of the present invention.

In addition to the above-mentioned composition, the composition of the present invention can be used as a flavoring agent such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, coloring agents and intermediates (cheese, chocolate etc.), pectic acid and its salts, Salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated beverages and the like. In addition, the composition of the present invention may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks. These ingredients may be used independently or in combination, and although the proportion of such additives is not so critical, the compound of the present invention is generally selected in the range of 0 to about 20 parts by weight per 100 parts by weight.

The health food of the present invention is selected from the group consisting of dairy products, fermented milk, drink, meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen, gum, ice cream, soup, beverage, alcoholic beverage and vitamin complex But is not limited thereto.

The present invention also provides a pharmaceutical composition for preventing or treating infectious diseases caused by Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.

In the pharmaceutical composition of the present invention, the infectious diseases caused by Helicobacter pylori include gastritis, gastric ulcer, duodenal ulcer, non-ulcer dyspeptic syndrome, gastric MALT lymphoma, gastric hyperplastic polyps, gastric cancer, gastrointestinal cancer, pancreatitis, inflammatory bowel disease , But is not limited thereto.

The pharmaceutical composition of the present invention may contain 0.02 to 80% by weight, preferably 0.02 to 50% by weight, of the compound based on the total weight of the composition.

Compositions comprising the compounds of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of pharmaceutical compositions.

The pharmaceutical dosage forms of the compounds of the present invention may also be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds as well as in a suitable set.

The composition containing the compound according to the present invention may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, oral preparations such as syrups and aerosols, external preparations, suppositories, And can be used as formulations. Examples of carriers, excipients and diluents that can be contained in the composition including the compound include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium Various compounds or mixtures including silicates, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, oligosaccharides, sucrose, (sucrose), lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Propyleneglycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. Examples of the suppository base include witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like.

The preferred dosage of the compound of the present invention varies depending on the condition and weight of the patient, the degree of disease, the type of drug, the route of administration and the period of time, but can be appropriately selected by those skilled in the art. However, for the desired effect, the compound of the present invention is preferably administered at 0.0001 to 100 mg / kg, preferably 0.001 to 100 mg / kg per day. The administration may be carried out once a day or divided into several times. The dose is not intended to limit the scope of the invention in any way.

The compounds of the present invention may be administered to mammals such as rats, mice, livestock, humans, and the like in a variety of routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

Hereinafter, the present invention will be described in detail with reference to examples. However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

Materials and methods

1. Instruments and reagents

Nuclear magnetic resonance (NMR) spectrometer was a Bruker DRX-300 and DRX-500 spectrometer (Germany). FAB / MS and EI / MS were obtained from JEOL JMS-700 (Akishima, Japan). For the medium pressure liquid chromatography (MPLC), YMC GEL ODS-A (12 nm, S-150 M) (YMC Co. Ltd., Japan) and 25 g Biotage Isolera One system (USA) were used. TLC was performed on Silica gel 60 F _{254 (} Merck, Germany). Column chromatography was performed using silica gel 60 (0.063-0.43 mm; Merck KGaA, Germany) and Sephadex LH-20 (GE Healthcare, Sweden). For the cultivation of Helicobacter pylori, CO ₂ incubator ASTEC SCA-80DS (Japan) model was used. The quercetin used as a reference material was Sigma (USA). The other reagents were limited products.

2. Plant material

The roots of Hojangun are purchased at Kyungdong Market (Jejudo, Seoul) in October 2013. The feelings of plants were made by Professor An Mi Jung of the College of Pharmacy of Gyeongsang National University. The sample (No. GSC-104) is stored in the Botanical Sampling Room, College of Pharmacy, Gyeongsang National University.

3. Plant Extraction and Component Separation

The dried rhizome root (4 kg) was crushed finely and extracted with 70% ethanol at room temperature. This ethanol extract was concentrated under reduced pressure to obtain crude extract (1.12 kg). After this suspension was added distilled water to the n - hexane, ethyl acetate, n - and fractionation in order to butanol n - hexane fraction (14.6 g), ethyl acetate fraction (407 g), n - butanol fraction (174 g) and aqueous layer ( 468 g), respectively. Among them, the ethyl acetate fraction was subjected to silica gel column chromatography with hexane, methylene chloride and methanol mixed solvent (1: 1: 0 → 0: 1: 0 → MeOH) to obtain 7 fractions (fr. 7). The fractions fr.2a to fr.2c and fr.3a (respectively) were subjected to silica gel column chromatography with a mixture of hexane and ethyl acetate as a developing solvent (100: 0 to 0: 100) ~ fr.3b. Fr. Compound 1 (894 mg) was isolated from 2b and compound 2 (18 g) from fr.2b by recrystallization. Compound 7 (80 mg) was isolated from fr.3a by silica gel column chromatography using hexane and methylene chloride as eluting solvents. Compound 8 (7 mg) and Compound 9 (10 mg) were obtained from fr.3a by silica gel chromatography using hexane and methylene chloride as eluent solvents, followed by Sephadex LH-20 column chromatography using methanol as a developing solvent . The other two fr.4 and fr.5 were subjected to silica gel column chromatography using methylene chloride and methanol as developing solvents to separate the fractions fr.4a to fr.4e and fr.5a to fr.5c, respectively. Among these fractions, compounds 3 (345 mg) and 4 (42 mg) and 5 (120 mg) were obtained from the fractions fr.4c, fr.5a and fr.5b through recrystallization. The fractional fraction fr. 5c was subjected to MPLC using water and methanol as elution solvents to obtain Compound 6 (11 g).

Physcion (1) - Yellowish powder. C ₁₆ H ₁₂ O ₅ ; EI-MS ( m / z ): 284 [M] ^< ^{+ &gt};; _{^{1 H-NMR (CDCl 3,}} 300MHz): 12.34 (1H, s, 1-OH), 12.14 (1H, s, 8-OH), 7.64 (1H, d, J = 1.2 Hz, H-4), 7.38 (1H, d, J = 2.6 Hz, H-5), 7.10 (1H, d, J = 1.2 Hz, H-2), 6.70 (1H, d, J = 2.6 Hz, H-7), 3.96 (3H _{, s, 6-OCH 3)} , 2.47 (3H, s, 3-CH 3); ¹³ C-NMR (CDCl ₃ , 125 MHz): Table 2.

Emodin (2) - Orange needles, C ₁₅ H ₁₀ O ₅ ; EI-MS ( m / z ): 270 [M] ^< ^{+ &gt};; ¹ H-NMR (DMSO d ₆ , 300 MHz): 12.07 (1H, s, 1 -OH), 12.01 brd, H-2), 7.09 (1H, d, J = 2.4 Hz, H-5), 6.57 (1H, d, J = 2.4 Hz, H-7), 2.47 (3H, s, 3-CH 3) ; _{^{13 C-NMR (DMSO- d 6}} , 125MHz): Table 2.

Anthraglycoside B (3) - Yellowish powder, C ₂₁ H ₂₀ O ₁₀ ; FAB-MS ( m / z ): 433.2 [M + H] < ⁺ >; _{^{1 H-NMR (DMSO- d 6}} , 500MHz): 13.23 (1H, brs, OH-1), 7.46 (1H, brs, H-4), 7.27 (1H, d, J = 2.4 Hz, H-5) , 7.16 (1H, br s, H-2), 6.98 (1H, d, J = 2.4 Hz, H-7), 5.05 (1H, d, J = 7.6 Hz, H-1), 3.72 ~ 3.24 (glc -H 2 ~ 6), 2.41 ( 3H, s, 3-CH 3); _{^{13 C-NMR (DMSO- d 6}} , 125MHz): Table 2.

Trans-resveratrol (trans- resveratrol) (4) - Pale white powder, C 14 H 12 O 3; EI-MS ( m / z ): 228 [M] ^< ^{+ &gt};; ¹ H-NMR (DMSO d ₆ , 500 MHz): 9.54 (4-OH), 9.18 (3,5-OH), 7.39 (2H, d, J = 8.6 Hz, H- , d, J = 16.5 Hz, Hb), 6.81 (1H, d, J = 16.5 Hz, Ha), 6.75 (2H, d, J = 8.6 Hz, H-3, 5), 6.38 (2H, d, J = 2.0 Hz, H-2, 6), 6.12 (1H, t, J = 2.0 Hz, H-4); _{^{13 C-NMR (DMSO- d 6}} , 125MHz): Table 2.

Anthraglycoside A (5) - Yellowish powder, C ₂₂ H ₂₂ O ₁₀ ; ESI-MS ( m / z ): 445.1 [MH]; _{^{1 H-NMR (DMSO- d 6}} , 500MHz): 13.10 (1-OH), 7.50 (1H, brs, H-4), 7.37 (1H, brd, H-5), 7.19 (2H, brd, H- 2,7), 5.18 (1H, d , J = 7.7 Hz, H-1), 3.51 ~ 3.19 (2 ~ 6 glc-H), 3.97 (3H, s, 6-OCH 3), 2.41 (3H, s , _3-CH 3); _{^{13 C-NMR (DMSO- d 6}} , 125MHz): Table 2.

Datin poly (Polydatin) (6) - White powder, C 20 H 22 O 8; FAB-MS (m / z): 390.1 [M] +; _{^{1 H-NMR (DMSO- d 6}} , 500MHz): 7.40 (2H, d, J = 8.6 Hz, H-2, 6), 7.03 (1H, d, J = 16.3 Hz, Hb), 6.87 (1H, d , J = 16.3 Hz, Ha) , 6.76 (2H, d, J = 8.6 Hz, H-3, 5), 6.74 (1H, br t, H-2), 6.57 (1H, br t, H-6) , 6.34 (1H, br t, H-4), 4.81 (1H, d, J = 7.6 Hz, glc H-1), 3.18 ~ 3.49 (glc H-2-6); _{^{13 C-NMR (DMSO- d 6}} , 125MHz): Table 2.

2-methoxy-6-acetyl-7-methyl-week geulron (2-Methoxy-6-acetyl -7-methyljuglone) (7) Red needles, C 14 H 12 O 5; EI-MS ( m / z ): 260 [M] ^< ^{+ &gt};; _{^{1 H-NMR (CDCl 3,}} 300MHz): 12.53 (1H, s, 5-OH), 6.13 (1H, s, H-3), 7.54 (1H, s, H-8), 3.95 (3H, s, _{2-OCH 3), 2.61 (} 3H, s, 6-COCH 3), 2.37 (3H, s, 7-CH 3); ¹³ C-NMR (CDCl ₃ , 125 MHz): Table 2.

Citreorosein (8) Yellow powder, C ₁₅ H ₁₀ O ₆ ; EI-MS ( m / z ): 286 [M] ^< ^{+ &gt};; _{H-NMR (DMSO- d 6,} 300MHz): 12.10 (2H, s, 1,8-OH), 7.65 (1H, brs, H-4), 7.26 (1H, brs, H-2), 7.13 (1H , d, J = 2.4 Hz, H-5), 6.59 (1H, d, J = 2.4 Hz, H-7), 4.61 (2H, s, 3-CH 2); _{^{13 C-NMR (DMSO- d 6}} , 125MHz): Table 2.

2-Ethoxy-6-acetyl-7-methyljuglone (9)

Yellow powder C ₁₅ H ₁₄ O ₅ ; EI-MS ( m / z ): 274 [M] ^< ^{+ &gt};; _{^{1 H-NMR (CDCl 3,}} 300MHz): 12.56 (1H, s, 5-OH), 7.53 (1H, s, H-8), 6.10 (1H, s, H-3), 4.14 (1H, q, J = 7.0 Hz, 2-O C H 2 CH 3), 2.61 (3H, s, 6-COCH 3), 2.38 (3H, s, 7-CH 3), 1.56 (3H, t, J = 7.0 Hz, 2-OCH ₂ C H ₃ ); ¹³ C-NMR (CDCl ₃ , 125 MHZ): Table 2.

4. Culture of Helicobacter pylori

The Helicobacter pylori 43504 strain used in this study was purchased from H. pylori Korean Type Culture Collection, Gyeongsang National University, Korea. Helicobacter pylori is a horse serum (Gibco, USA) of 10 brucellosis (Brucella) with% were cultured in an agar medium (BD Co., US), 37 ℃, 2 ~ 3 of the 100% humidity in an incubator with 10% CO ₂ condition Day.

5. Paper disc diffusion assay

The total extract and the anti - Helicobacter pylori activity for each fraction were measured by infiltration of the extract into a paper disk. That is, each sample was dissolved in DMSO, and 20 μl of each sample was absorbed on a paper disk (8 mm diameter, 0.7 mm thickness; Advantec, Japan). The concentration of the sample was 10 ㎎ / ㎖ and the growth inhibition zone (clear zone) was measured for 24 hours. Quercetin and DMSO were used as positive control and negative control, respectively.

6. Measurement of minimum inhibitory concentration

The minimum inhibitory concentration (MIC) was measured using a liquid broth dilution method. The MIC value was measured by measuring the optical density at 600 nm while culturing the minimum concentration at which the growth of the bacteria was inhibited for 24 hours to 48 hours. MIC ₅₀ and MIC ₉₀ mean the minimum concentration inhibiting 50% and 90% growth respectively, and the respective values were obtained with GraphPad Version 5.01 (GraphPad Software, Inc., USA). All experiments were performed in 2 independent experiments and the values were obtained by repeating 3 measurements.

Example 1. Identification of anti-Helicobacter pylori activity of the extract and fractions

The anti - helicobacter pylori activity of extracts and fractions of. As a result, the total extract, hexane fraction and ethyl acetate fraction showed a larger growth inhibition region than the control quercetin, and the butanol fraction and water layer showed similar inhibitory activity to quercetin (Table 1).

Example 2 Isolation of Anti-Helicobacter pylori Active Ingredients of the Fructose Ethyl Acetate Fractions

The ethyl acetate fraction, which showed the largest inhibitory region among the extracts and fractions, was separated by silica gel column chromatography, sephadex, MPLC and recrystallization method. As a result, five anthraquinone compounds, two stilbene compounds, and one naphthoquinone compound were purely separated, and were analyzed by UV, ¹ H-NMR, ¹³ C-NMR and MS data Fiji the chemical structure of each compound was 1, the emodine (2), anthraquinone glycosides B (in emodine -8- --D- O-glucoside), (3), trans-resveratrol (4), anthraquinone glycosides A (sebum on -8- --D- O-glucoside), 5, datin poly (trans-resveratrol -3- O --D-glucoside) (6 ), 2- methoxy-6-acetyl-7- Methyl juglone ( 7 ), citrolosane ( 8 ) and 2-ethoxy-6-acetyl-7-methyl juglone ( 9 ).

Example 3. Confirmation of anti-Helicobacter pylori activity of the compound of the present invention

The MIC values indicating the anti-Helicobacter pylori activity for each compound identified in Example 2 were measured by the broth dilution method. As a result, the anti-helicobacter pylori activity of all the isolated compounds was lower than that of the quercetin, which is a positive control (Fig. 2 and Table 3). Among them, compounds 2 and 7 showed strong inhibitory activity. In particular, 2-methoxy-6-acetyl-7-methyl juglone ( 7 ) showed the strongest inhibitory effect, with a MIC ₅₀ value of 0.30 μM and an MIC ₉₀ value of 0.39 μM. In the case of compound No. 9 , the MIC ₅₀ value was 0.30 μM, which showed similar inhibitory activity as the compound No. 7, but the MIC ₉₀ was 0.67 μM. Compounds 4 and 8 showed MIC ₅₀ values of 59.3 and 37.8 μM, respectively, showing the inhibitory activity following compounds 2 and 7 .

It is presumed that the activity decreases when the α-position is glycosylated in the anthraquinone structure in view of the difference in the anti-helicobacter pylori activity of emodin ( 2 ) and anthraglycoside B ( 3 ). It is presumed that inhibition activity is reduced when glycosylation occurs at position C-3 in stilbene compounds trans-resveratrol ( 4 ) and polyadin ( 6 ). In the case of fizzy on ( 1 ), the hydroxyl group at the C-6 position of emmidine ( 2 ) was replaced by methoxy group, resulting in increased liposolubility, resulting in poor activity due to low solubility in aqueous media. Compared to the years of age 8 to emodine (2) and the sheet Leo, when the α- hydroxy anthraquinones C-3 position hydroxymethyl (hydroxymethyl) group is substituted can be estimated to be lower in inhibitory activity, 1 , And 4-naphthoquinone have been reported to increase anti-helicobacter pylori activity at the methoxy group at the C-2 position and the hydroxyl group at the C-5 position. Recent studies have shown that some of the 2-hydroxy-1,4-naphthoquinone compounds, which are not cytotoxic and mitotoxic, are essential for the growth of Helicobacter pylori and the thymidylate synthase Respectively.

Although the anti-Helicobacter pylori activity of horsetail roots and Compounds 2 and 4 has been previously reported, anti-Helicobacter pylori activity against other compounds isolated from the active ingredients of this plant has been reported for the first time. Based on the above results, it is considered that the roots or isolated active ingredients of Hojangun may be useful for Helicobacter pylori infection.

Claims (7)

6. An antimicrobial composition for Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.
[Chemical Formula 1]

delete The antimicrobial composition for Helicobacter pylori according to claim 1, wherein the 2-alkoxy-6-acetyl-7-methyl juglone is isolated from the callus root extract. [Claim 4] The antimicrobial composition for Helicobacter pylori according to claim 3, wherein the callus root extract is an ethyl acetate fraction of a callus root extract. A health food composition for preventing or ameliorating infectious diseases caused by Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.
[Chemical Formula 1]

The infectious disease according to claim 5, wherein the infectious disease is any one selected from the group consisting of gastritis, gastric ulcer, duodenal ulcer, non-ulcer digestive syndrome, gastric MALT lymphoma, gastric hyperplastic polyps, gastric cancer, digestive cancer, pancreatitis and inflammatory bowel disease Which is effective for preventing or ameliorating infectious diseases caused by Helicobacter pylori. A pharmaceutical composition for preventing or treating infectious disease caused by Helicobacter pylori containing 2-alkoxy-6-acetyl-7-methyl juglone or a salt thereof as an active ingredient.
[Chemical Formula 1]

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