Composition for treating inflammatory bowel disease comprising a combined extract of Chinese laurel and walnut

Abstract

The present invention relates to a pharmaceutical composition for preventing or treating inflammatory bowel disease, comprising an extract of Ochoa sieboldii and Juglans chinensis.The present invention also relates to a food composition or animal feed composition for preventing or improving inflammatory bowel disease, comprising said extract.

Description

[Technical field]

[0001] The present invention relates to a pharmaceutical composition for preventing or treating inflammatory bowel disease, comprising a complex extract of Chinese walnut and Japanese walnut.The present invention also relates to a food composition or animal feed composition for preventing or improving inflammatory bowel disease, comprising the complex extract. [Background technology]

[0002] Inflammatory bowel disease (IBD) is a disease that causes chronic inflammation of the intestine of unknown cause, and shows a clinical course that progresses with repeated worsening and improvement, and ulcerative colitis and Crohn's disease are representative. The cause and pathophysiology of IBD are still not clearly known, and it is presumed that genetic factors, environmental factors such as intestinal bacteria and food, immunological factors, etc. are involved in the mechanism of onset. Sustained or inappropriate activation of the intestinal immune system plays an important role in the pathophysiology of chronic mucosal inflammation, especially the infiltration of neutrophils, macrophages, lymphocytes and mast cells, which results in mucosal destruction and ulcers. Infiltrated and activated neutrophils are a major source of reactive oxygen species and reactive nitrogen species, which are cytotoxic substances and cause oxidative stress that degrades cross-linked proteins, lipids and nucleic acids, resulting in epithelial dysfunction and damage.

[0003] A fundamental treatment for inflammatory bowel disease has not yet been established, and drugs that alleviate symptoms are used. These include 5-aminosalicylic acid (5-ASA) drugs, such as sulfasalazine, which mainly suppress the production of prostaglandins, and steroid immunosuppressants.

[0004] Sulfasalazine often causes side effects or adverse reactions, such as abdominal fullness, headache, rash, liver disease, leukopenia, agranulocytosis, and male infertility. It is also unclear whether sulfasalazine is effective enough to prevent recurrence in patients who have had diseased bowel removed or who are recovering.

[0005] Steroid immunosuppressants are corticosteroids, and although they have short-term effects, they cannot improve long-term prognosis. In addition, they have side effects such as infectious diseases, secondary adrenal insufficiency, peptic ulcers, diabetes, mental disorders, and steroid-induced nephropathy, so they are limited to being used only in acute cases.

[0006] As there is still no reliable treatment for inflammatory bowel disease, there is a demand for the development of new therapeutic agents that are inexpensive and have excellent therapeutic effects without side effects. For example, a composition for preventing or treating inflammatory bowel disease using a natural product extract (Patent Document 1) has been developed.

[0007] On the other hand, Isatidis Folium is the leaf of Isatis indigotica Fortune, a biennial plant belonging to the Cruciferae family, and is harvested in summer and autumn. The whole plant is called Isatidis and the root is called Banlangen, and is known to be effective against influenza, hepatitis, acute pneumonia, jaundice, etc.

[0008] In addition, the Japanese walnut (Juglans mandshurica Maxim.) is classified as a member of the Juglandaceae family, and exists as a tree or shrub native to America, Eurasia, or South Asia. It resembles the walnut tree native to China, and is mainly grown in the central and northern parts of Korea or the northeastern part of China, where it is used as a medicinal plant and herbal medicine.

[0009] Therefore, the inventors made efforts to develop a new therapeutic agent using natural products that has excellent effects on inflammatory bowel disease, and as a result, they discovered that a mixed extract of Daisy Leaf and Walnut Tree improves inflammatory bowel disease, thereby completing the present invention. [Prior art documents] [Patent documents]

[0010] [Patent Document 1] Korean Patent No. 10-1710730 Summary of the Invention [Problem to be solved by the invention]

[0011] The present invention aims to provide a pharmaceutical composition for preventing or treating inflammatory bowel disease, comprising a complex extract of Radix Ophiopogon and Juglans chinensis.

[0012] Another object of the present invention is to provide a food composition for preventing or ameliorating inflammatory bowel disease, which comprises a complex extract of Oogi Leaf and Walnut Tree.

[0013] A further object of the present invention is to provide an animal feed composition for preventing or ameliorating inflammatory bowel disease, which comprises a complex extract of Oogi Leaf and Walnut Tree. [Means for solving the problem]

[0014] The present invention provides a pharmaceutical composition for preventing or treating inflammatory bowel disease, comprising a combined extract of Radix Oleifera and Juglans chinensis.

[0015] The present invention also provides a food composition for preventing or ameliorating inflammatory bowel disease, comprising a complex extract of Oolong tea leaves and Walnut.

[0016] Furthermore, the present invention provides an animal feed composition for preventing or ameliorating inflammatory bowel disease, which comprises a complex extract of Oolong tea leaves and Juglans japonica.

[0017] The aforementioned Ooba and Juglans japonica may be contained in a weight ratio of 1:0.1 to 10, preferably in a weight ratio of 1:0.5 to 2, and more preferably in a weight ratio of 1:1.

[0018] The composition may comprise a mixture of extracts from the herbal medicines of Radix Isogai and Juglans nigra, or may comprise an extract extracted from a herbal medicine mixture of Radix Isogai and Juglans nigra.

[0019] The walnut may be a walnut stem, a walnut fruit, a walnut root, or a walnut leaf, and is preferably a walnut fruit.

[0020] The extract may be extracted with a solvent selected from the group consisting of water, C1 to C6 alcohols, and mixed solvents thereof, and is preferably a 0.01% to 90% ethanol extract, more preferably a 60% to 80% ethanol extract, and most preferably a 70% ethanol extract.

[0021] Alternatively, a decrease in the length of the large intestine can be inhibited, and an increase in the weight or length of the spleen can be inhibited.

[0022] The present invention provides a composition for preventing or treating inflammatory bowel disease, comprising a complex extract of Macadamia radiata and Juglans juncea.

[0023] The present invention also provides a use of a composition comprising a complex extract of Mahogany Leaf and Walnut Tree for preventing or treating inflammatory bowel disease. Effect of the Invention

[0024] The composition containing the combined extract of Masago okara and Juglans chinensis of the present invention exhibits excellent therapeutic effects against inflammatory bowel diseases and can be used as a novel therapeutic agent for inflammatory bowel diseases.

[0025] In addition, since the composition of the present invention is a mixed extract of natural products, it has few side effects and is safe, and is very useful for patients with inflammatory bowel diseases of various causes. [Brief description of the drawings]

[0026] [Figure 1] FIG. 1 shows the weight change rate of mice administered the composite extract of the present invention. [Diagram 2] FIG. 1 shows the colon length of mice administered the composite extract of the present invention. [Diagram 3] FIG. 13 is a graph showing the spleen ratio measured by measuring the weight and length of the spleen in each group compared to the negative control group in mice administered the composite extract of the present invention. [Figure 4] FIG. 1 shows the Disease Activity Index (DAI) of mice administered the composite extract of the present invention. [Diagram 5] FIG. 1 is a comparative diagram showing the disease activity index (DAI) of mice administered the combined extract of the present invention when administered with the extract of Daisy Leaf and the extract of Walnut Tree individually. [Figure 6] FIG. 1 shows the results of measuring myeloperoxidase (MPO) activity, which is an indicator of inflammatory response caused by administration of the complex extract of the present invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0027] DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will now be described in detail with reference to the accompanying drawings, in order to enable those skilled in the art to easily carry out the present invention. However, the present invention may be embodied in various forms and is not limited to the embodiments and examples set forth herein.

[0028] Throughout this specification, when a part is said to "comprise" a certain element, this does not mean that it excludes other elements, but that it may further include other elements, unless otherwise specified.

[0029] The present invention provides a composition for preventing, ameliorating or treating inflammatory bowel disease, comprising a complex extract of Radix Ophiopogon and Juglans chinensis.

[0030] The large green leaves include dried leaves of Isatis indigotica Fortune.

[0031] The walnut may be a walnut stem, a walnut fruit, a walnut root or a walnut leaf, and preferably a walnut fruit.

[0032] The term "composite extract" or "composite herbal extract" as used herein refers to the active ingredient of the pharmaceutical composition of the present invention, which may include an extract extracted from each of the herbal medicines of Daphne Root or Walnut, or an extract extracted from a mixture of these herbal medicines.

[0033] The extract used in the present invention can be obtained by using a conventional extraction solvent known in the art. A polar solvent or a non-polar solvent can be used as the extraction solvent. Examples of polar solvents include water, C1-C6 alcohols (e.g., methanol, ethanol, propanol, butanol, normal propanol, isopropanol, normal butanol, etc.), acetic acid, and mixtures of the above polar solvents. Examples of non-polar solvents include acetone, acetonitrile, ethyl acetate, methyl acetate, butyl acetate, fluoroalkanes, hexane, ether, chloroform, dichloromethane, and mixtures of the above non-polar solvents.

[0034] The extract of the present invention may be extracted with a solvent selected from the group consisting of water, C1 to C6 alcohols, and mixed solvents thereof, and is preferably an extract with 0.01% to 90% ethanol, more preferably an extract with 60% to 80% ethanol, and most preferably an extract with 70% ethanol.

[0035] The extract used in the present invention may be extracted by hot water extraction, cold maceration extraction, reflux cooling extraction, ultrasonic extraction or any other conventional extraction method known in the art.

[0036] The term "crude extract" refers to what is commonly known in the art, but in a broader sense also includes fractions obtained by further fractionation of the extract. In other words, the extract includes not only those obtained using the above-mentioned solvents, but also those obtained from them through a purification process. For example, fractions obtained from the above extracts by passing through an ultrafiltration membrane with a certain molecular weight cutoff value, and fractions obtained by various further purification methods such as separation by various chromatographs (designed for separation by size, charge, hydrophobicity or affinity) are also included in the extract of the present invention.

[0037] The term "prevention" as used herein means any action of suppressing or delaying a gastrointestinal disorder or the like by administering a composition according to the present invention, and the term "treatment" means any action of administering said composition to improve or beneficially alter the symptoms of an individual suspected of or suffering from a gastrointestinal disorder. The term "amelioration" as used herein means any action of at least reducing a parameter related to the condition being treated by administering a composition comprising an extract of the present invention, e.g., the severity of the symptoms.

[0038] The pharmaceutical composition of the present invention can be administered parenterally or orally depending on the intended method, and the dosage range varies depending on the patient's body weight, age, sex, health condition, diet, administration time, administration method, excretion rate, severity of disease, etc. In addition, the therapeutically effective amount of the composition varies depending on the administration method, target site, and patient condition, and when used in the human body, the dosage should be determined appropriately taking into consideration both safety and efficiency.

[0039] The food composition of the present invention may be a functional food, a dairy product, a fermented product or a food additive.

[0040] The functional food refers to food manufactured and processed using raw materials or ingredients that have functional properties useful to the human body, and "functional" means that it is consumed for the purpose of regulating nutrients for the structure and function of the human body or obtaining useful effects for health purposes such as physiological actions.

[0041] The animal feed compositions of the present invention can be ingested by any non-human animal, such as non-human primates, sheep, dogs, cattle, horses, and the like.

[0042] In the present invention, "prevention" means any action of suppressing or delaying a disease, etc. by administering a composition according to the present invention, "improvement" means any action of at least reducing a parameter related to the condition being treated by administering a composition of the present invention, such as the severity of a symptom, and "treatment" means any action of improving or favorably changing the symptoms of an individual who has developed a disease or an individual suspected of having such a disease by administering a composition of the present invention. EXAMPLES

[0043] The present invention will be described in more detail below with reference to examples. However, these examples are merely for illustrating the present invention, and the present invention is not limited thereto.

[0044] [Example] Preparation of a complex extract containing Ooba and Juglans japonica A sample of washed and dried Isatidis Folium and Juglans mandshurica Maxim. mixed in a 1:1 weight ratio was added to a 10-fold 70% ethanol solution and extracted at room temperature for 72 hours. After extraction, the mixture was filtered under reduced pressure using 5μm filter paper and then concentrated under reduced pressure at 50-65℃ to obtain a powdered herbal extract. EXAMPLES

[0045] Evaluation of body weight changes in mice following administration of the composite extract Six-week-old C57BL / 6N male mice were purchased and bred for one week, after which they were randomly divided into five groups of eight mice each, and experiments were carried out as shown in Table 1 below.

[0046] 2.5% dextran sulfate sodium (DSS) diluted with sterile water was administered in drinking water for a total of 8 days, and 5-aminosalicylic acid (5-ASA), a therapeutic agent for inflammatory bowel disease, or the composite extract of the present invention was orally administered for a total of 10 days starting from 2 days before the administration of DSS.

[0047] [Table 1]

[0048] Weight loss is a typical symptom of inflammatory bowel disease (Non-Patent Document 1), and the mice were weighed once a day at the same time (10:00 a.m.) using an electronic scale. The weight change rate for each group was calculated by adding up the weights of eight mice per group and dividing by eight. The average weight immediately before the start of administration of 2.5% DSS in drinking water was set as 100%, and the average weight of each group was calculated daily to measure the weight change rate. Student's T test was performed on each group of negative control groups and experimental groups that received 2.5% DSS in drinking water to verify the significance.

[0049] As shown in Figure 1, the weight change rate of mice administered the composite extract for 7 days was measured, and the normal group had a weight change rate of 105.64 ± 0.54%, while the negative control group had a weight change rate of 89.31 ± 2.75%, which was lower than the normal group. In addition, the positive control group had a weight change rate of 91.38 ± 2.01%, while the group administered 30 mg / kg of the composite extract of the large green leaf and the Japanese walnut fruit had a weight change rate of 93.18 ± 1.67%, and the group administered 300 mg / kg of the composite extract of the large green leaf and the Japanese walnut fruit had a weight change rate of 97.15 ± 0.89%, which was lower than the positive control group. In particular, the group administered 300 mg / kg of the composite extract of the large green leaf and the Japanese walnut fruit showed the most excellent effect, which confirmed that the effect was far superior to that of the positive control group.

[0050] Therefore, it can be seen that the composite extract of the sclerotium macrocarpa leaves and the fruit of the Japanese walnut according to the present invention has the effect of suppressing weight loss in mice. EXAMPLES

[0051] Evaluation of colon length in mice following administration of the composite extract It is known that there is a good correlation between colon shortening and histological changes, and colon length is usually used as a morphological parameter of the degree of inflammation. Therefore, in order to evaluate the colon length of mice administered the composite extract of the present invention, the mice in Example 1 were sacrificed on the last day of each administration, and the colon length was measured. The average length is shown in Figure 2.

[0052] As shown in Figure 2, the colon length of mice administered the composite extract was measured, and the normal group had a length of 7.48±0.30cm, while the negative control group had a length of 5.44±0.12cm, indicating that the colon length was reduced compared to the normal group. The positive control group had a length of 5.28±0.08cm, the group administered 30mg / kg of the composite extract of oak leaves and walnut fruit had a length of 5.15±0.12cm, and the group administered 300mg / kg of the composite extract of oak leaves and walnut fruit had a length of 6.09±0.06cm. In particular, the reduction in colon length was suppressed in the group administered 300mg / kg of the composite extract of oak leaves and walnut fruit, which confirmed that the effect was far superior to that of the positive control group.

[0053] Therefore, it can be seen that the combined extract of the oyster leaves and the walnut fruit of the present invention has the effect of suppressing the decrease in the length of the large intestine in mice. EXAMPLES

[0054] Evaluation of the level of splenic inflammation in mice following administration of the composite extract A typical symptom associated with inflammatory bowel disease is splenitis, and when inflammation occurs in the spleen, the spleen size increases due to its characteristics (Non-Patent Document 2), and it is actually known that the spleen increases in size in an IBD model induced after DSS treatment (Non-Patent Document 3). Therefore, in order to evaluate the splenic inflammation level of mice administered the composite extract of the present invention, the mice of Example 1 were sacrificed on the last day of each administration, and the spleen weight and length of each group were measured in comparison with the negative control group. The results are shown as spleen ratios.

[0055] As shown in Figure 3, the weight and length of the spleen of mice administered the composite extract were measured, and the negative control group had an increased spleen weight and length compared to the normal group. In addition, the positive control group, the group administered 30 mg / kg of the composite extract of Chinese leopard leaves and Chinese walnut fruit, and the group administered 300 mg / kg of the composite extract of Chinese leopard leaves and Chinese walnut fruit had a decreased spleen weight and length compared to the negative control group. In particular, the group administered 300 mg / kg of the composite extract of Chinese leopard leaves and Chinese walnut fruit had the greatest decrease, suggesting that the compound extract has a more excellent effect compared to the positive control group.

[0056] Therefore, it can be seen that the combined extract of the oak leaf and the walnut fruit of the present invention has the effect of improving the symptoms of splenic enlargement caused by splenic inflammation associated with inflammatory bowel disease in mice. EXAMPLES

[0057] Evaluation of Disease Activity Index (DAI) in Mice by Administration of Complex Extracts-Part I To measure the severity of inflammatory bowel disease in the inflammatory bowel disease animal model treated with the administration method of Example 1, changes in body weight, stool consistency, and the presence or absence of bloody stool macroscopically observed in the stool or anus were checked every day during the administration period according to the Disease Activity Index (DAI) rank in Table 2, and the disease activity index was measured. The results are shown in Figure 4.

[0058] [Table 2]

[0059] As shown in Figure 4, the disease activity index of mice administered the composite extract was significantly increased in the negative control group. In addition, the disease activity index was reduced in the group administered 30 mg / kg of the composite extract of Chinese leopard leaves and Chinese walnut fruit and the group administered 300 mg / kg of the composite extract of Chinese leopard leaves and Chinese walnut fruit compared to the negative control group. In particular, the group administered 300 mg / kg of the composite extract of Chinese leopard leaves and Chinese walnut fruit showed a much more excellent reduction effect than the positive control group. EXAMPLES

[0060] Evaluation of Disease Activity Index (DAI) in Mice by Administration of Complex Extracts-Part II In order to evaluate whether the combined extract has a synergistic effect compared to when the extracts of Daisy Leaf and Walnut are used individually, 6-week-old C57BL / 6N male mice were purchased and pre-bred for one week, after which they were randomly divided into 6 groups of 7 mice each, and the experiment was carried out as shown in Table 3 below.

[0061] [Table 3]

[0062] 2.5% dextran sulfate sodium (DSS) diluted with sterilized water was administered in drinking water for a total of 5 days, and 5-aminosalicylic acid (5-ASA), a single extract of Daisy leaf, a single extract of Walnut fruit, or the composite extract of the present invention, which are therapeutic agents for inflammatory bowel disease, were orally administered for a total of 7 days starting from 2 days before the administration of DSS.

[0063] In addition, disease activity index was measured using the same method as used in Table 2. The results 5 days after administration are shown in Figure 5.

[0064] As shown in Figure 5, the disease activity index increased significantly in the negative control group, and compared to the negative control group, the disease activity index decreased in the positive control group (5-ASA), the group administered 50 mg / kg of a single extract of Daphne digitata, and the group administered 100 mg / kg of the composite extract of the present invention, while the group administered 50 mg / kg of a single extract of Japanese walnut fruit showed almost no effect of reducing the disease activity index.

[0065] In conclusion, the combined extract of Walnut and Chinese walnut leaves shows a significant effect of reducing disease activity indexes compared to the single extract of Chinese walnut leaves, which is a remarkable effect that cannot be predicted from the single extracts. Therefore, it can be seen that the combined extract of the present invention has a superior synergistic effect of improving disease activity indexes in an inflammatory bowel disease-induced mouse model compared to the single extract of Chinese walnut leaves and the single extract of Walnut. EXAMPLES

[0066] Evaluation of myeloperoxidase (MPO) activity by administration of the complex extract Myeloperoxidase (MPO) is an enzyme found mainly in neutrophils, and MPO activity in tissues is an indicator of neutrophil infiltration, and therefore an indicator of inflammatory response, and correlates with the level of intestinal damage caused by inflammatory bowel disease. Therefore, to measure the MPO activity by administration of the composite extract of the present invention, an experiment was carried out as shown in Table 3 of Example 5.

[0067] 2.5% dextran sulfate sodium (DSS) diluted with sterilized water was administered in drinking water for a total of 5 days, and 5-aminosalicylic acid (5-ASA), a single extract of Daisy leaf, a single extract of Walnut fruit, or the composite extract of the present invention, which are therapeutic agents for inflammatory bowel disease, were orally administered for a total of 7 days starting from 2 days before the administration of DSS. A mouse MPO ELISA kit (HK210-02, Hycult biotechnology) was used to measure MPO activity. Colon tissues were washed with cold PBS, and 200 μL of lysis buffer (200 mM NaCl, 5 mM EDTA, 10 mM Tris, 10% glycerol, 1 mM PMSF, 1 mg / mL leupeptin, and 28 mg / mL aprotinin (pH 7.4)) was added per 10 mg of tissue weight, followed by homogenization for 30 seconds using a tissue homogenizer (Mixer mill MM400, Retsch). The homogenized samples were centrifuged at 1500Xg for 15 minutes to obtain the supernatant, which was then used to measure MPO activity.

[0068] As shown in Figure 6, MPO activity was significantly higher in the negative control group (DSS-administered group) compared to the normal group, and MPO activity was reduced by 9.6% in the positive control group (5-ASA 100 mg / kg-administered group) compared to the negative control group.

[0069] In addition, MPO activity decreased by 16.9% and 6.3% in the groups administered a single extract of Chinese walnut leaves and a single extract of Chinese walnut fruit, respectively, and the most significant decrease in MPO activity was 41.2% in the group administered 100 mg / kg of the composite extract of Chinese walnut leaves and Chinese walnut fruit, which is the composite extract of the present invention.

[0070] Therefore, it can be seen that the combined extract of Chinese walnut leaves and Chinese walnut fruit of the present invention has an outstanding synergistic effect in suppressing MPO activity in an inflammatory bowel disease-induced mouse model, compared to groups administered an extract of Chinese walnut leaves alone or an extract of Chinese walnut fruit alone.

Claims

1. A pharmaceutical composition for preventing or treating inflammatory bowel disease, comprising an extract of Daicho and Juglans digitata.

2. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the weight ratio of the oenophilous leaf and the walnut is 1:0.1-10.

3. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the weight ratio of the oenophilous leaf and the walnut is 1:0.5-2.

4. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, comprising: 1:1 weight ratio of Chinese walnut and oak leaf.

5. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, characterized in that the composition comprises a mixture of extracts from the herbal medicines of Chinese laurel leaf and Chinese walnut, or an extract extracted from a mixture of the herbal medicines of Chinese laurel leaf and Chinese walnut.

6. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the Juglans japonica is a fruit of Juglans japonica.

7. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the extract is extracted with a solvent selected from the group consisting of water, C1 to C6 alcohols, and mixtures thereof.

8. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the extract is a 0.01% to 90% ethanol extract.

9. 2. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the extract is a 60% to 80% ethanol extract.

10. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the extract is a 70% ethanol extract.

11. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the composition inhibits a decrease in body weight or colon length.

12. The pharmaceutical composition for preventing or treating inflammatory bowel disease according to claim 1, wherein the composition inhibits an increase in spleen weight or length.

13. A food composition for preventing or improving inflammatory bowel disease, comprising extracts of Daigoba and Juglans japonica.

14. An animal feed composition for preventing or improving inflammatory bowel disease, comprising an extract of Daigoba and Juglans japonica.

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