KR101964054B1 - Pharmaceutical composition comprising extract of Lonicera japonica for prevention and treatment of Crohn's disease - Google Patents

Abstract

The present invention relates to an extract for the treatment or prophylaxis of Crohn's disease or ulcerative colitis containing Lonicera japonica Thunberg organic solvent extract. The gold-silver fraction according to the present invention can be effectively used for the treatment, prevention or improvement of Crohn's disease or ulcerative colitis without side effects.

Description

[0001] The present invention relates to a pharmaceutical composition for the treatment or prevention of Crohn's disease,

The present invention relates to a composition for treating or preventing Crohn's disease or ulcerative colitis, which contains, as an active ingredient, a gingival fraction having a therapeutic effect on Crohn's disease or ulcerative colitis damage or a compound isolated therefrom, (3,5-di-O-caffeoylquinic acid), 4 (4-di-O-caffeoylquinic acid) which has excellent therapeutic effect on Crohn's disease or ulcerative colitis and does not show cytotoxicity, , 5-di-O-caffeoylquinic acid as an active ingredient in the treatment or prevention of Crohn's disease or ulcerative colitis. And a health functional food composition.

Ulcerative colitis and Crohn's disease are bowel diseases that have not yet been clearly identified. Ulcerative colitis is a disease in which erosions (erosions) and ulcers are continuously formed in the mucous membranes of the large intestine. Bleeding, dyspepsia, diarrhea and abdominal pain occur in the colon. In severe cases, systemic symptoms such as fever, weight loss and anemia appear .

In addition, Crohn's disease is a disease in which lesions such as ulcers are generated discontinuously in any part of the digestive tract from the mouth to the anus. In addition to abdominal pain, diarrhea and stool, severe cases include fever, hemorrhage, weight loss, , And anemia. Both ulcerative colitis and Crohn's disease are chronic refractory diseases in which the symptoms recur temporarily and recurrence recurs, collectively referred to as inflammatory bowel disease. Although there is an indication that environmental factors such as genetic factors, immune disorders, and diet are involved in the pathogenesis of these diseases, the cause is still unclear.

In the past, the incidence of ulcerative colitis and Crohn's disease was known to be high in westerners, but recently, the number of patients has increased rapidly in Korea and other countries due to changes in lifestyle such as eating habits. However, there is a reason why the cause is unclear, and fundamental treatment is not established. For this reason, it is not the goal of complete treatment, but rather the use of medicines that alleviate symptoms (remission, temporary symptom improvement) and maintain these conditions for as long as possible. Aminosalicylic acid preparations, adrenocorticosteroids, immunosuppressants and the like are mainly used as medicines for such symptomatic therapy, but various side effects have been reported. For example, salazosulfapyridine, which is frequently used as an aminosalicylic acid preparation, has been reported to have side effects such as nausea, vomiting, anorexia, rash, headache, liver damage, leukocytosis, abnormal red blood cells, proteinuria and diarrhea. Adrenocortical steroids are generally used for oral administration of prednisolone, enema, suppository, intravenous injection, etc. However, side effects such as gastric ulcer necrosis caused by gastric ulcer or long-term use are strong. However, since discontinuation of the medication recurs, the medicines must be continuously used.

Therefore, there is a demand for the development of therapeutic agents for intestinal diseases such as ulcerative colitis and Crohn's disease which are excellent in efficacy, safe, and do not cause side effects.

Lonicerae Flos is a flower of Lonicera japonica Thunb. Belonging to the genus Caprifoliaceae. It is used for diuretic, arthritis, purulent dermatitis and bronchitis in a herbal medicine or a civilian. Tannin, inositol, sterol, chlorogenic acid, isochlogenic acid, and the like have been reported as the components of gold silver. Examples of flavonoid components include luteolin luteolin, apigenin, luteolin-7-O-rhamnoglucoside, quercetin and the like are known. The flavonoid component of this gilt-silver was reported to be anti-inflammatory (Lee, SJ; Arch. Pharm. Res. 16, 25, 1993) and antimutagenic action.

In a study on gingkohwang, Kang Ok-hee (study on the pharmacological action of gingkohwa) reported that the methanol extract of gingkohwa had antibacterial activity against several gram-positive and gram-negative bacteria. Lee et al. Reported that the butanol fraction of gingko- In carrageenin-induced plantar edema experiments, we reported a less potent, but dose-dependent, improvement in plantar edema than prednisolone. In addition, Tae et al. Reported that the water extract of Ganoderma lucidum exhibited anti-inflammatory effects by inhibiting NFκB p65 activity and degradation of IκBα in rat liver sepsis induced by Lipopolysaccharide (LPS) (Tae, J .; Clin Chim Acta . 330 (1-2), 165-171, 2003)

The inventors of the present invention confirmed in Korean Patent Registration No. 10-0878436 that "Ganoderma lucidum extract" is effective for the treatment of peptic ulcer including gastritis and ulcer, as "a pharmaceutical composition for treating or preventing peptic ulcer including Ginkgo biloba extract" In Korean Patent Registration No. 10-1074839, it was confirmed that the gingkowort extract had a therapeutic effect on GERD with the statement that " a pharmaceutical composition for treating or preventing reflux esophagitis including Ginkgo biloba extract ".

The present inventors have found that gastritis against Gingko nuts extract. While developing a therapeutic agent for gastric ulcer and reflux esophagitis (GERD), it was confirmed that specific fractions of Gil Eul Hwa were excellent in Crohn's disease or ulcerative colitis disease. The inventors of the present invention found that the specific fractions of Gil- . Accordingly, the present inventors have found that the gold-eutectic fraction or the compound isolated therefrom is not only highly effective in treating Crohn's disease or ulcerative colitis, but also exhibits a preventive effect against Crohn's disease or ulcerative colitis, Crohn's disease or Crohn's disease or ulcerative colitis. The present invention has been accomplished on the basis of these findings.

Korean Patent Registration No. 10-0878436 (Pharmaceutical Composition for the Treatment or Prevention of Peptic Ulcer Containing Gold Euphoria Extract, 2009.01.13 Announcement) Korean Patent Registration No. 10-1074839 (Pharmaceutical Composition for the Treatment or Prevention of Reflux Esophagitis Containing Gold Euphoria Extract, October 19, 2011)

Lee, S. J .; Arch. Pharm. Res. 16, 25, 1993. Tae, J .; Clin Chim Acta. 330 (1-2), 165-171, 2003.

It is an object of the present invention to provide a pharmaceutical composition for the treatment and prevention of Crohn's disease or ulcerative colitis, which comprises a gold-eurafil fraction or a compound of the following general formulas 1 to 2 as an active ingredient.

It is another object of the present invention to provide a health functional food composition for preventing or ameliorating Crohn's disease or ulcerative colitis, which comprises a gingival fraction or the compound of Chemical Formulas 1 and 2 as an active ingredient.

In order to achieve the above object, the present invention relates to a gold-silverated fraction having antioxidant activity and treating Crohn's disease or ulcerative colitis, or a 3,5-di-O-caffeoylquinic acid (3,5-di-O -caffeoylquinic acid, hereinafter referred to as 3,5-di-CQA), 4,5-di-O-caffeoylquinic acid (4,5- di-CQA) as an active ingredient. The present invention also provides a pharmaceutical composition for treating or preventing Crohn's disease or ulcerative colitis, which comprises at least one compound selected from the group consisting of di-CQA

The present invention relates to a gold-silverated fraction having an antioxidative activity and treating Crohn's disease or ulcerative colitis, or a combination of 3,5-di-CQA of Formula 1 and 4,5-di-CQA of Formula 2 The present invention provides a health functional food composition for improving or preventing Crohn's disease or ulcerative colitis, which comprises the above compound as an active ingredient.

The gold-eutectin extract of the present invention can be used as a medicament for the treatment of Crohn's disease or ulcerative colitis in clinical trials (for example, sulfasalazine, budesonide, mesalamine, capsaicin, Saglamostim, Nitta ysannanide, prednisolone, , Adalimumab, etc.), in combination or in combination, to provide a pharmaceutical composition for the treatment and prevention of Crohn's disease or ulcerative colitis.

Hereinafter, the present invention will be described in detail.

The ginseng extract according to the present invention can be obtained by extracting from gold silver or its dried material, and the ginseng silver can be used without limitation such as wild and human cultivated. The gold-eutectic extract according to the present invention can be obtained by a conventional extraction method such as supercritical extraction, room temperature extraction, high-temperature extraction, high-pressure extraction, ultrasonic extraction or the like, or adsorption resin including XAD and HP- Can be used, and it is preferably soluble and can be prepared by reflux extraction or extraction at room temperature, but is not limited thereto. The number of times of extraction is preferably 1 to 5 times, more preferably 3 times, but is not limited thereto.

The extract may be extracted using water, an organic solvent or a mixed solvent thereof. The organic solvent is preferably one selected from the group consisting of C1 to C4 alcohols, ethyl acetate, methylene chloride and chloroform, or a mixed solvent thereof, more preferably C1 to C4 alcohols, more preferably methanol, It is most preferable to extract them with an aqueous solution of 50 to 100% alcohol.

For example, the gingkohye extract according to the present invention can be obtained by pulverizing the dried gingivalae to an appropriate size, placing it in an extraction container, boiling the solvent by adding an extraction solvent, refluxing the extract, allowing it to stand for a predetermined time and filtering it with a filter paper or the like. The extraction time is preferably 2 to 12 hours, more preferably 3 to 6 hours. Thereafter, a method such as concentration or freeze-drying can be added.

The ginseng fraction according to the present invention can be obtained by sequential fractionation of the ginseng extract using hexane, ethyl acetate and butanol. Further, the present invention relates to a pharmaceutical composition for treating Crohn's disease or Crohn's disease comprising as an active ingredient at least one compound selected from the group consisting of 3,5-di-CQA represented by Formula 1 and 4,5-di-CQA represented by Formula 2 There is provided a pharmaceutical composition for treating or preventing ulcerative colitis.

The compounds of Formulas 1 and 2 may be prepared by adding water, an organic solvent or a mixture thereof to silver chelate to obtain a silver sulfite extract (Step 1):

The step of suspending the Ganoderma lucidum extract obtained in the above step 1 in water and then fractionating it with ethyl acetate or butanol to obtain fractions (step 2):

Separating and purifying the compounds of Formulas 1 and 2 (Step 3) by performing silica gel chromatography on the fractions obtained in Step 2 above;

And the like.

Hereinafter, the manufacturing method according to the present invention will be described in more detail. First, step 1 according to the present invention is a step of obtaining a Ganoderma lucidum extract as an extraction solvent. The dried gold silver is pulverized to an appropriate size and placed in an extraction vessel. The extraction solvent may be an alcohol of C1 to C4, an alcohol aqueous solution of 50 to 100% of C1 to C4, ethyl acetate, methylene chloride, chloroform or a mixed solvent thereof. Of these, methanol, ethanol, or 50 to 100% It is preferable to use an aqueous alcohol solution. It is extracted with ultrasound at 60 ° C for 6 hours, and then filtered with a filter paper or the like to obtain a gingival extract according to the present invention.

Next, the step 2 is a step of obtaining the fractions by fractionating the gold and silver europium extract obtained in the step 1 with a solvent having a different polarity. As the solvent, ethylacetate or butanol can be used.

Next, the step 3 is a step of separating and purifying the compounds of the formulas (1) to (2) by performing silica gel chromatography on the fractions obtained in the step (2). The silica gel chromatography can be carried out using a column for size exclusion chromatography, and preferably a column packed with HP-20 can be used. The fraction obtained in the above step 2 is subjected to silica gel chromatography using an HP-20 column using ethanol as a mobile phase. At this time, the obtained fractions can be subjected to high performance liquid chromatography to separate the compounds of the formulas (1) to (2). The high performance liquid chromatography can be carried out using a mixed solvent of water and acetonitrile gradient from 0 vol% to 5 vol% and from 5 vol% to 10 vol% acetonitrile as a developing solvent. At this time, the flow rate of the mobile phase is 2-15 mL / min.

The gingival fraction or the compounds of Formulas 1 and 2 according to the present invention may be used for the treatment or prevention of Crohn's disease or ulcerative colitis.

The compounds of the present invention may be used alone or in combination with other drugs currently used in the treatment of Crohn's disease or ulcerative colitis (for example, sulfasalazine, budesonide, mesalamine, capsaicin, For example, in combination with, or in combination with, an anti-inflammatory agent, for example, Moss Steam, Nitasusanide, Prednisolone, Natalie Juimab, Adalimumab, and the like.

The pharmaceutical composition for treating or preventing ulcerative colitis or ulcerative colitis according to the present invention may be formulated into various oral or parenteral dosage forms as described below, It does not.

Examples of formulations for oral administration include tablets, pills, light / soft capsules, liquids, suspensions, emulsions, syrups, granules and the like. These formulations may contain fillers, extenders, wetting agents, One or more diluents or excipients such as disintegrants, lubricants, binders and surfactants may be used. As the disintegrant, agar, starch, alginic acid or its sodium salt, anhydrous calcium monohydrogenphosphate, etc. may be used. As the lubricant, silica, talc, stearic acid or its magnesium salt or calcium salt, polyethylene glycol and the like may be used And as the binder, magnesium, aluminum silicate, starch paste, gelatin, methylcellulose, sodium carboxymethylcellulose, polyvinylpyrrolidine, low-substituted hydroxypropylcellulose, glycine and the like can be used as a diluent. A boiling mixture, an absorbent, a coloring agent, a flavoring agent, a sweetening agent, and the like, may be used together.

In addition, the pharmaceutical composition for treating or preventing Crohn's disease or ulcerative colitis, which comprises the extract of Gingko nuts, fractions or the compounds of formulas (1) and (2) as an active ingredient, can be administered parenterally, and parenteral administration can be carried out by subcutaneous injection, intravenous injection, The method of injecting the injector or the injector in the chest is by the method. In order to formulate the composition for parenteral administration, the gingkoffle extract, the fractions or the compounds of Formulas 1 and 2 may be mixed with a stabilizer or a buffer in water to prepare a solution or suspension, which may be prepared into a unit dosage form of ampoule or vial have.

The composition may be sterilized or contain adjuvants such as preservatives, stabilizers, wettable or emulsifying accelerators, salts for controlling osmotic pressure, buffering agents and other therapeutically useful substances and may be prepared by conventional methods such as mixing, granulating or coating . ≪ / RTI >

When the pharmaceutical composition for treating or preventing Crohn's disease or ulcerative colitis containing the gold-silver fraction of the present invention or the compounds of the formulas (1) and (2) as an active ingredient is formulated in unit dose form, these gold-containing fractions or To 2 in a unit dose of about 0.1 to 1500 mg. The dosage depends on the physician's prescription depending on factors such as the patient's weight, age and the particular nature and severity of the disease. However, the dosage required for adult therapy is usually in the range of about 1 to 500 mg per day, depending on the frequency and intensity of administration. A total dosage of 5 to 300 mg per day, usually divided into a single dose when administered intramuscularly or intravenously to an adult, may suffice, but in some patients a higher daily dose may be desirable.

The gold-silver fraction according to the present invention or the compounds of the formulas (1) and (2) may be added to a health supplement or health functional food such as foods, beverages for the purpose of treating or preventing Crohn's disease or ulcerative colitis. In this case, when using the gingival fraction or the compounds of the formulas (1) and (2) as a food additive, it may be added in an amount of 0.01 to 30% by weight, preferably 0.1 to 10% by weight based on the raw material. The amount of the active ingredient to be mixed can be appropriately determined depending on the purpose of use. However, in the case of long-term intake intended for health and hygiene purposes or for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of stability, the active ingredient may be used in an amount exceeding the above range . The fractions or the compounds of the formulas (1) and (2) can be used together with other food or food ingredients and can be suitably used according to conventional methods.

There is no particular limitation on the kind of the food. Examples of the foods to which the fractions or the compounds of Formulas 1 and 2 can be added include dairy products including meat, sausage, bread, chocolates, candy, snack, pizza, ramen, other noodles, gums, ice cream, Beverages, drinks, alcoholic beverages, and vitamin mixtures, all of which include health foods in the conventional sense.

As the food preservative additive of the present invention, various flavors or natural carbohydrates can be used. The natural carbohydrates are sugar saccharides such as monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. Examples of sweeteners include natural sweeteners such as tau Martin and stevia extract, synthetic sweeteners such as saccharin and aspartame, and the like. The ratio of the natural carbohydrate is generally about 0.01 to 0.04 part by weight, preferably 0.02 to 0.03 part by weight per 100 parts by weight of the composition according to the present invention.

In addition to the above, the composition according to the present invention may further comprise various nutrients, vitamins, electrolytes, flavors, colorants, pect and its salts, alginic acid and its salts, organic acids, protective colloid concentrating agents, pH adjusting agents, stabilizers, preservatives, , A carbonating agent used in carbonated drinks, and the like. These components can be used independently or in combination, and the proportion of the additive is not critical, but is generally selected in the range of 0.001 to 1 part by weight per 100 parts by weight of the composition according to the present invention.

The composition comprising the gold-silver fraction according to the present invention or at least one compound selected from the group consisting of 3,5-di-CQA of Formula 1 and 4,5-di- Or ulcerative colitis disease without any adverse effects compared with the existing therapeutic agents and exhibits an excellent therapeutic effect on Crohn's disease or ulcerative colitis disease.

The extract according to the present invention or the compounds of the formulas (1) and (2) can be usefully used for the treatment and prevention of Crohn's disease or ulcerative colitis.

FIG. 1 shows the result of measuring the inflammation area of the large intestine obtained after treating the TNBS-induced inflammatory bowel disease animal with Gingko nail extract.
Fig. 2 shows the results of measuring the effect of Ganoderma lucidum extract on IL-1? Activity in an TNBS-induced inflammatory bowel disease animal model.
Fig. 3 is a photograph of the colon length and the extract obtained after treating the TNBS-induced inflammatory bowel disease animal with Ganoderma lucidum extract.

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 by the following examples.

< Example  1> Preparation of Ganoderma lucidum ethanol extract

The powder of the granule used in the present invention was dried before blossoming. The powder obtained by pulverization was pulverized to an appropriate size, and 1 kg of the powder was added to the extraction vessel. 10 L of 70% ethanol solvent was added thereto, and the mixture was stirred at 60 캜 for 6 hours And filtered through a filter paper to obtain an extract. The extraction procedure was repeated three times, and then the solvent was concentrated under reduced pressure and dried to obtain 300 g of ethanol extract.

< Example  2 > Ethyl acetate Fraction  Produce

The ethanol extract of Example 1 was suspended in 3 L of water and then extracted 3 times with 3 L of ethyl acetate. The extract was filtered under reduced pressure using a filter paper. The solvent of the obtained ethyl acetate extract was removed, and ethyl acetate fraction 30 g.

< Example  3> Butanol Fraction  Produce

The ethanol extract of Example 1 was suspended in 3 L of water and then 3 L of butanol was added 3 times. The mixture was repeatedly extracted three times. The mixture was filtered under reduced pressure using a filter paper. The solvent of the butanol extract was removed, and 75 g of butanol fraction .

Example 4: Preparation of phenolic compounds from gold-silver fraction

(Example 4-1) Preparation of 3,5-di-CQA

The ethyl acetate fraction obtained in Example 2 was subjected to chromatography using an HP-20 column with 100% ethanol as a mobile phase to obtain fractions 1 to 4. After fraction 3 of the fraction was dissolved in ethanol, high performance liquid chromatography was performed to obtain 3,5-di-CQA. At this time, a mixed solvent of water and acetonitrile was used as a mobile phase, and the concentration was gradually adjusted with polarity from 0 vol% to 20 vol% acetonitrile for 50 minutes. The flow rate was 10 mL / min, Capcell pak UG120 Mm, 5 탆) column was used.

3,5-di-CQA Compound: ¹ H-NMR (CD ₃ OD)? 2.11-2.34 (4H, m, H-2,6), 3.95 (1H, dd, J = 3.0, 7.8 Hz, ), 5.37-5.44 (2H, m, H-3,5), 6.26, 6.36 (1H each, J = 15.9Hz, H-8 ', 8 "), 6.77 (2H, d, J = 7.8Hz, H -5 ', 5 "), 6.96 (2H, dd, J = 2.1, 8.1Hz, H-6', 6"), 7.05, 7.06 (1H each, d, J = 2.1Hz, H-2 ', 2 ), 7.57, 7.61 (1H each, d, J = 15.9 Hz, H-7 ', 7 "

^{13 C-NMR (125MHz, CD} 3 OD) δ 36.3 (C-2), 38.2 (C-6), 71.1 (C-4), 72.0 (C-5), 72.9 (C-3), 75.2 (C (C-8 '), 116.5 (C-5', 5 "), 123.0, 123.1 (C-6 ' (C-1 ', 1''), 146.7 (C-3', 3 "), 147.0, 147.2 4 "), 168.5, 168.9 (C-9 ', 9"), 178.1 (COOH)

(Example 4-2) Preparation of 4,5-di-CQA

The ethyl acetate fraction obtained in Example 2 was subjected to chromatography using an HP-20 column with 100% ethanol as a mobile phase to obtain fractions 1 to 4. After fraction 3 of the fraction was dissolved in ethanol, high performance liquid chromatography was performed to obtain 4,5-di-CQA. At this time, a mixed solvent of water and acetonitrile was used as a mobile phase, and concentration gradient was performed with polarity gradually from 0 vol% to 20 vol% acetonitrile for 50 minutes. The flow rate was 10 mL / min, Capcell pak UG120 , 5 탆) column was used.

4,5-di-CQA Compound: ¹ H-NMR (500 MHz, CD ₃ OD)? 1.98-2.30 (4H, m, H-2, 6), 4.33 (1H, br s, 1H, dd, J = 2.6, 9.5Hz, H-4), 5.65 (1H, m, H-5), 6.19, 6.27 (1H each, d, J = 15.9Hz, H-8 ', 8 "), 6.72, 6.73 (1H each, d, J = 8.1 Hz, H-5 ', 5''), 6.88, 6.90 (1H each, dd, J = 1.8, 8.1 Hz, H- D, J = 15.9 Hz, H-7 ', 7 "), 7.01 (1H each, d, J = 1.8 Hz, H-

^{13 C-NMR (125MHz, CD} 3 OD) δ 76.1 (C-1), 38.3 (C-2), 69.8 (C-3), 75.8 (C-4), 69.0 (C-5), 39.8 (C C-4 '), 116.1 (C-5', 5 ''), 146.4 (C- ), 122.9 (C-6 ', 6 "), 147.2, 147.3 (C-7', 7"

<Experimental Example 1> Experimental study on Crohn's disease or Ulcerative Colitis of Gold Euphoria Extract

Seven-week-old Sprague Dawley (SD) rats were purchased from Orient bio Korea and stabilized with common solid feeds and used in inflammatory bowel disease drug efficacy studies. During the experimental period, feed and water were freely supplied, and the temperature of the breeding room was kept at 20 ° C and 2 ° C and relative humidity of 50 ° C and 20%. The experimental group was divided into 4 groups according to the randomized block design so that the average body weight was 220 and 20 g per 8 groups.

SD rats fasted for 24 hours were anesthetized with diethyl ether. Then, the polyethylene catheter was placed in a lumen of a colon using a 1 ml syringe, using 2.5% trinitrobenzene diluted with 50% (v / v) ethanol After 0.5 ml of sulfonic acid (trinitrobenzene sulfonic acid, TNBS) was slowly injected, the rat was inverted to prevent 2.5% TNBS from leaking into the anus and allowed to stand until the anesthesia was awakened. Kg of 3,5-di-CQA and 10 mg / kg of 4,5-di-CQA, respectively, for 4 days after the administration of TNBS. 5-ASA (5-aminosalicylic acid), an active metabolite of sulfasalazine known as an agent for the treatment of inflammatory bowel disease, was orally administered at a dose of 100 mg / kg as a positive control.

<Experimental Example 2> Measurement of inflammation area in a rat TNBS model

After 4 days of administration of trinitrobenzene sulfonic acid (TNBS), dissection was performed and an incision was made from the anus to the cecum to observe damage to the colon mucosa. After expanding the large intestine, the area (mm 2) of the colon lesion was measured with a stereomicroscope (× 10) and the colonic lesion cure rate was calculated using the following equation.

As can be seen from FIG. 1, the inhibition of TNBS-induced intestinal damage by about 43% was observed at a dose of 50 mg / kg of the Euglena extract of Example 2, which was superior to that of 5-ASA administered orally. It was also observed that TNBS-induced colonic injury was excellently suppressed at a dose of 10 mg / kg of 3,5-di-CQA and 10 mg / kg of 4,5-di-CQA.

<Experimental Example 3> Measurement of activity of IL-1?

After the autopsy, the esophageal tissue is taken, and 10 times more homogenizing buffer is added, followed by disruption with a homogenizer. After centrifugation at 10,000 rpm for 10 minutes, the homogenized tissue solution (supernatant) was separated and the activity of IL-1 beta in the tissue was measured. The activity of IL-1β was assayed by ELISA kit purchased from Gomavio and detected by ELISA Reader (Bio-Rad).

* Homogenizing buffer (pH 7.4): 1.15% KCl, 50 mM Tris-HCl, 1 mM EDTA

As can be seen in FIG. 2, the activity of IL-1β increased by TNBS significantly inhibited the activity of IL-1β by about 37% at the dose of 50 mg / kg of the Euglena extract of Example 2. In addition, we observed that the inhibitory activity of IL-1β increased by TNBS at a dose of 10 mg / kg of 3,5-di-CQA and 10 mg / kg of 4,5-di-CQA.

<Experimental Example 4> Measurement of colon length

After 4 days of administration of trinitrobenzene sulfonic acid (TNBS), dissection was performed and the photograph was taken from the anus to the appendix. The result was as shown in Fig. 3, and the length of the colon from the anus to the cecum was measured Is shown in FIG. The colony of 5% TNBS-induced colitis in rats was markedly shortened and hyperemic compared to the normal control group, but the length of the colon was significantly increased at a dose of 50 mg / kg of the Euglena extract of Example 2 by about 20% Able to know. In addition, the effects of increasing the length of the colon in the dose of 10 mg / kg of 3,5-di-CQA and 10 mg / kg of 4,5-di-CQA were observed.

&Lt; Example of pharmaceutical preparation >

(Formulation Example 1) Powdered ethyl acetate fraction

20 mg of the germanium ethyl acetate fraction of Example 2,

Lactose 100 mg,

10 mg of talc

The above components are mixed and filled in airtight bags to prepare powders.

(Formulation Example 2) Preparation of tablets

10 mg of the germanium ethyl acetate fraction of Example 2

Corn starch 100 mg

Lactose 100 mg

2 mg of magnesium stearate

After mixing the above components, tablets are prepared by tableting according to the usual preparation method of tablets.

(Formulation Example 3) Preparation of capsule

10 mg of the germanium ethyl acetate fraction of Example 2

Crystalline cellulose 30 mg

Lactose 14.8 mg

0.2 mg of magnesium stearate

The above components are mixed according to a conventional capsule preparation method and filled in gelatin capsules to prepare capsules.

(Formulation Example 4) Preparation of injections

10 mg of the germanium ethyl acetate fraction of Example 2

180 mg mannitol

2974 mg of sterile distilled water for injection

Na ₂ HPO ₄ , 12 H ₂ O 26 mg

(2 ml) per ampoule in accordance with the usual injection method.

(Formulation Example 5) Production example of liquid agent

20 mg of the germanium ethyl acetate fraction of Example 2

10 g per isomer

5 g mannitol

Purified water quantity

Each component was added and dissolved in purified water according to the usual liquid preparation method, and the lemon flavor was added in an appropriate amount. Then, the above components were mixed and purified water was added thereto to adjust the total volume to 100 ml. The mixture was filled in a brown bottle and sterilized, .

<Example of food production>

(Food Preparation Example 1) Production of beverage

Honey 522 mg

5 mg of chitosanic acid ami

Nicotinamide 10 mg

3 mg of sodium riboflavin hydrochloride

Pyridoxine hydrochloride 3 mg

Inositol 30 mg

Orthoic acid 50 mg

100 mg of gum ethoxide ethyl acetate fraction

Purified water 200 mL

A beverage was prepared using the above-mentioned composition and content by a conventional method.

(Food Preparation Example 2) Vitamin Blend

100 mg of the germanium ethyl acetate fraction

Phenolic acid compound 100 mg

70 [mu] g of vitamin A acetate

Vitamin E 1.0 mg

0.13 mg vitamin B1

0.15 mg of vitamin B2

0.2 [mu] g vitamin B6

10 mg vitamin C

Biotin 10 μg

Nicotinic acid amide 1.7 mg

50 ㎍ of folic acid

Calcium pantothenate 0.5 mg

1.75 mg of ferrous sulfate

0.82 mg of zinc oxide

Magnesium carbonate 25.3 mg

15 mg of potassium phosphate monobasic

55 mg of potassium secondary phosphate

Potassium citrate 90 mg

100 mg of calcium carbonate

24.8 mg of magnesium chloride

The above-mentioned vitamin and mineral mixture is a component suitable for relatively healthy food, and is mixed with a preferred embodiment. However, the compounding ratio is arbitrarily modified, and the above ingredients are mixed according to a conventional method for producing health food, , And can be used in the manufacture of a health food composition according to a conventional method.

Claims (3)

As an extract of C1-C4 alcohol or 50-100% C1-C4 alcohol in Lonicerae Flos, 3,5-di-O-caffeoylquinic acid of the following formula 1 or 4,5-di-O - A pharmaceutical composition for treating or preventing Crohn's disease of large intestine comprising caffeine quinic acid as an active ingredient.

delete As an extract of C1-C4 alcohol or 50-100% C1-C4 alcohol in Lonicerae Flos, 3,5-di-O-caffeoylquinic acid of the following formula 1 or 4,5-di-O - a health functional food composition for improving or preventing colonic disease of Crohn disease, which comprises caffeic oil quinic acid as an active ingredient.

Images