KR20090084130A - Composition comprising the compound isolated from the flower extract of daphne genkwa for preventing and treating cancer disease - Google Patents

Abstract

A composition containing a compound isolated from a daphne genkwa flower extract as an active ingredient is provided to have cytotoxicity and prevent and treat cancer diseases. A compound isolated from daphne genkwa flower extract is denoted by the chemical formula (I). In the chemical formula (I), R is hydrogen, C2-C10 is alkyl group or C2-C10 alkoxy group. A pharmaceutical composition for preventing and treating cancer diseases comprises 0.1-50 weight% novel compound of the chemical formula (I) or its pharmaceutically allowable salt. A health functional food is used in a form of powder, granule, tablet, capsule, or beverage.

Description

Composition comprising the compound isolated from the flower extract of daphne genkwa for preventing and treating cancer disease}

 The present invention relates to a composition for the prophylaxis and treatment of cancer diseases containing the compound isolated from the original extract as an active ingredient.

[Reference 1] Folksman et al ., Science, 235 , pp. 442-447, 1987

[Reference 2] Cordell et al ., Bioactive Natural Products, CRS Press, pp. 195-220, 1993

[3] Blume-Jensen and Hunter, Nature, 411 , pp. 355-365, 2001

      [Ref. 4] Jung-Seop Shin and Min-Kyo Shin, Antidrug Dictionary, Younglimsa, pp740-741, 1998

[5] Kai et al., Yakugaku Zasshi, 124 , pp. 349-354, 2004

Zhan et al., Bioorgan Med Chem 13 , pp. 645-55, 2005

Lee et al., Chemico-Biol Interact 115 , pp. 215-228, 1998

     Cancer is one of the incurable diseases that humanity has to solve, and huge capital is invested in the development to cure it all over the world.In Korea, it is the number one disease death cause and more than 100,000 people a year It is diagnosed, and about 60,000 or more die. Carcinogens that cause cancer are smoking, ultraviolet rays, chemicals, foods, and other environmental factors. However, the causes of various cancers are difficult to develop the therapeutic agents, and the effects of the therapeutic agents also vary depending on the site of occurrence. Currently, the materials used as therapeutic agents have considerable toxicity, and do not selectively remove only cancer cells. Therefore, there is an urgent need for development of low-toxic and effective anti-cancer agents to prevent the development of cancer as well as its treatment after the occurrence of cancer.

Cancer is characterized by "uncontrolled cell growth," which causes cell clumps called tumors to form, penetrate into surrounding tissues and, in severe cases, metastasize to other organs in the body. Academia is also called neoplasia. Cancer is an intractable chronic disease that, even if treated with surgery, radiation, and chemotherapy, in many cases does not heal fundamentally, suffers the patient, and ultimately leads to death. There are many factors that cause cancer, but they can be divided into internal and external factors. The mechanism by which normal cells undergo transformation into cancer cells has not been precisely identified, but at least 80-90% is known to be influenced by external factors such as environmental factors. Internal factors include genetic factors, immunological factors, and external factors include chemicals, radiation, and viruses. Genes involved in the development of cancer include oncogenes and tumor suppressor genes, which occur when the balance between them is broken down by the internal or external tolerances described above.

Because cancer cells have many similar properties to normal cells, it is not easy to remove cancer cells without damaging normal cells. However, cancer cells have some characteristics that distinguish them from several normal cells. First, cancer cells are not regulated in cell proliferation. Second, they are relatively lacking in differentiation. Third, they penetrate and spread to surrounding tissues. . Normal cells proliferate by receiving signals from growth factors as needed, while cancer cells have low dependence on growth factors and do not have contact inhibition, which inhibits growth by contact with surrounding cells. Secrete angiogenic factor to promote metastasis. In addition, cancer cells are not differentiated, do not cause apoptosis or programmed cell death, and have genetically unstable characteristics. Genetic instability of cancer cells is very important for cancer progression and is known to induce resistance to chemotherapeutic agents (Folksman et al. al ., Science, 235 , pp. 442-447, 1987).

Cancer is classified into blood cancer and solid cancer, and it occurs in almost every part of the body such as lung cancer, stomach cancer, breast cancer, oral cancer, liver cancer, uterine cancer, esophageal cancer, and skin cancer. Among the methods used to treat these malignancies, chemotherapy agents, except surgery or radiation therapy, are collectively called anticancer agents, and most of them show anticancer activity by inhibiting the synthesis of nucleic acids. Chemotherapeutic agents are largely classified into antimetabolites, alkylating agents, antimitotic drugs, and hormones, and folates that inhibit metabolic processes necessary for the proliferation of cancer cells. Derivatives (methotrexate), purine derivatives (6-mercaptopurine, 6-thioguanine), pyrimidine derivatives (5-fluorouracil, cytarabine), etc.Introducing alkyl groups to guanine of DNA to modify DNA structure and chain Alkylating agents that exert the effect include nitrogen mustard compounds (chlorambucil, cyclophosphamide), ethyleneimine compounds (thiotepa), alkylsulfonate compounds (busulfan), nitrosourea compounds (carmustine), triazene compounds (dacarbazine) There is. Mitosis inhibitors that inhibit cell division by blocking mitosis as a specific period of mitosis include plant drugs such as actinomycin D, doxorubicin, bleomycin, and anticancer drugs such as mitomycin, vincristine, and wenvillastin. Alkaloids, taxoids including mitosis inhibitors including taxane rings, and the like. In addition, hormones such as corticosteroids and progesterone and platinum-containing compounds such as cisplatin are used as anticancer agents.

     The biggest problem with chemotherapeutic agents is drug resistance, which is a major factor in the initial successful response of anticancer drugs and eventually in the treatment failure. In connection with the problem of overcoming these side effects, efforts have recently been made to find the active ingredient in natural products that are used in the private sector.

Natural products have played an important role as sources of pharmaceutical raw materials and leading substances for the treatment and alleviation of diseases. With the development of science and technology, studies are being actively conducted to separate active ingredients contained in natural products or to synthesize derivatives thereof and to use them as drugs, rather than to use natural products themselves. It is known from the substance itself or its derivatives. Many anti-cancer chemotherapeutic agents currently in clinical use are active substances isolated from natural medicinal plants, and the representative ones are Taxus , a medicinal plant inhabiting the North American Pacific coast. brevifolia) substance Taxol (taxol obtained from bark: a generic name paclitaxel) which is known to be effective, especially women amjung breast and ovarian cancer. As yet another example Lantus decanter Rosedale mouse (Catharanthus vinblastine, alkaloid isolated from roseus ) , Podophyllum peltatum) is a semisynthetic derivative of podophyllotoxin Eto'o (podophyllotoxin) The lignans (lignan) separated from Forsythe (etoposide) and I'll Forsythe (teniposide), kamto teka Oh Kumi appear (Camptotheca Camptothecine isolated from accuminata , Cephalotaxus Haringonia Ba Drupasia ( Cephalotaxus) harringtonia var. The homo haring tonin (homoharringtonin) isolated from drupacea), Fusarium solani (Fusarium 4-ipomeanole isolated from solani ) (Cordell et al. al ., Bioactive Natural Products, CRS Press, pp. 195-220, 1993). Unlike natural materials, natural-derived substances are produced through highly complex metabolic and synthetic processes, and thus have new structures and mechanisms of action and are not yet fully studied for their components or mechanisms of action. It is very worthwhile to separate and study the mechanism of action.

In addition, the recent development of anti-cancer chemotherapeutic agents increases cell selectivity specific to cancer and at the same time reduces the side effects on normal cells, which is the biggest disadvantage of conventional anticancer drugs, and delivers cell signals specifically expressed in cancer cells. There is a trend to develop substances that act on the system. Cancer cells are known to overexpress tyrosine kinase, growth factor receptor, ras farnesyltransferase, telomerase, and cyclin. Thus, it is believed that substances that inhibit these may act as novel anticancer agents selective for cancer cells. Indeed, betulinic acid isolated from natural birch, a natural medicinal plant, is highly selective in melanoma and is currently in clinical trials. Gleevec is a bcr-abl belonging to protein tyrosine kinase. selective inhibition of c-abl and v-abl, which have an excellent effect on chronic myeloid leukemia.Iressa (ZD1839) is an inhibitor of EGF (epidermal grwoth factor) receptor and is currently clinically applied to lung cancer. (Blume-Jensen and Hunter, Nature, 411 , pp. 355-365, 2001).

The original flower (Daphne genkwa SIEB. Et ZUCC) is a bud of the Azalea. The leaves are picked before blooming and dried. The main ingredients are genkwadaphnin, apigenin, cytosterol, benzoic acid, and irritating oily substances. (Information Subsidiary and Shin Min-kyo, Anti-Medical Metabolism, Younglim History, pp740-741, 1998). Wonhwa is a nerve hypersensitivity, heart is not stable, heart beats, surprised, and forgetful symptoms are effective. In addition, there is a clinical report that asthma, sea water, skin swelling, food indigestion is effective, and a significant improvement in schizophrenia, manic depression, neurological disorders. It is also used for diuresis, edema, nephritis. (Kai et al., Yakugaku Zasshi, 124 , pp. 349-354, 2004).

      However, none of the above documents teaches or discloses that a substance that exhibits growth inhibitory effect is more specifically isolated from human lung cancer cells.

Therefore, the present inventors have studied the anticancer drugs of herbal origin, which are widely used in herbal medicine, and showed that the compounds of the present invention isolated from Wonhwa exhibit not only excellent anticancer activity but also high specific selectivity for lung cancer and inhibit cancer cell growth compared to normal cells. The present invention was completed by confirming the effect is excellent.

In order to achieve the above object, the present invention provides a novel compound of formula (I) or a pharmaceutically acceptable salt thereof isolated from the original extract:

                                                     (I)

      Where

R is a substituent group selected from the group consisting of hydrogen atoms, C ₂ to C ₁₀ alkyl group and a C ₂ to C ₁₀ alkoxy.

Preferred compounds of the general formula (I) R is a hydrogen atom, a C ₂ to C and _5-lower alkyl group or a C ₂ to a lower alkoxy group of C ₅ a, in a more preferred compound, R is a hydrogen atom, an ethyl group or a propyl group Compound group.

      The most preferable compound of the general formula (I) is 5 beta-hydroxyresinifenol-6 alpha, 7 alpha-epoxy-12 beta-propanoxyl-9,13,14-ortho-2E, 4E- Dicadienoate or 5beta-hydroxyresinifenol-6alpha, 7alpha-epoxy-12beta-butanoxyl-9,13,14-ortho-2E, 4E-dicadienoate.

       The novel compounds of the present invention represented by general formula (I) may be prepared as pharmaceutically acceptable salts according to methods conventional in the art.

As salts are acid addition salts formed with pharmaceutically acceptable free acids. Acid addition salts are prepared by conventional methods, for example by dissolving a compound in an excess of aqueous acid solution and precipitating the salt using a water miscible organic solvent, such as methanol, ethanol, acetone or acetonitrile. Equal molar amounts of the compound and acid or alcohol (eg, glycol monomethylether) in water can be heated and the mixture can then be evaporated to dryness or the precipitated salts can be suction filtered.

In this case, organic acids and inorganic acids may be used as the free acid, hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid, etc. may be used as the inorganic acid, and methanesulfonic acid, p -toluenesulfonic acid, acetic acid, trifluoroacetic acid, Citric acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, manderic acid, propionic acid, citric acid, lactic acid, glycolic acid, gluconic acid ( gluconic acid), galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, glucuronic acid, aspartic acid, ascorbic acid, carbonic acid, vanic acid, hydroiodic acid, and the like.

Bases can also be used to make pharmaceutically acceptable metal salts. An alkali metal or alkaline earth metal salt is obtained by, for example, dissolving a compound in an excess alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and then evaporating and drying the filtrate. At this time, as the metal salt, it is particularly suitable to prepare sodium, potassium or calcium salts, and the corresponding silver salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (for example, silver nitrate).

Pharmaceutically acceptable salts of the above compounds of formula (I) include salts of acidic or basic groups which may be present in compounds of formula (I), unless otherwise indicated. For example, pharmaceutically acceptable salts include sodium, calcium and potassium salts of the hydroxy group, and other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, hydrogen sulphate, phosphate, hydrogen phosphate, dihydrogen Phosphate, acetate, succinate, citrate, tartrate, lactate, mandelate, methanesulfonate (mesylate) and p -toluenesulfonate (tosylate) salts, which are known in the art. It can be prepared through.

 The present invention provides a pharmaceutical composition for the prevention and treatment of cancer diseases containing a novel compound represented by the general formula (I) or a pharmaceutically acceptable salt thereof as an active ingredient isolated from the original extract.

      The original picture as defined herein is characterized by the bud, stem, leaf or root of the adzuki bean, preferably the bud of the adzuki bean.

      The cancer diseases include gastric cancer, colon cancer, breast cancer, lung cancer, non-small cell lung cancer, bone cancer, pancreatic cancer, skin cancer, head or neck cancer, skin or intraocular melanoma, uterine cancer, ovarian cancer, colon cancer, small intestine cancer, rectal cancer and anal muscle cancer , Fallopian tube carcinoma, endometrial carcinoma, cervical carcinoma, vaginal carcinoma, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, lymph gland cancer, bladder cancer, gallbladder cancer, endocrine cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue Sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocyte lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system (CNS) tumor, primary CNS lymphoma , Spinal cord tumors, brainstem glioma or pituitary adenoma, preferably lung cancer, colon cancer, breast cancer, prostate cancer, gastric cancer or liver cancer, more preferably lung cancer.

        Hereinafter, the method for obtaining the compound of the present invention will be described in detail.

       The compound of the present invention is a mixed solvent of chloroform: acetone: methanol (1: 1: 1) of about 1 to 50 times the dry weight, preferably 8 times the dry weight by dipping the dried Won, and about 1 to 20 days at room temperature. A first step of extracting crude extract, hot water extraction, ultrasonic extraction, reflux extraction, and the like, preferably using a cold needle extraction method for about 7 days, and concentrating under reduced pressure; A second step of dissolving the crude extract in a ultrasonic solvent in a mixed solvent of methanol: chloroform (2: 8), filtering the supernatant and the precipitate, and concentrating and fractionating the supernatant among them; The methanol and chloroform fractions obtained in the second step were subjected to silica gel column chromatography with methanol: chloroform (1: 1 (v / v) eluting solvent) to be divided into three fractions; among the fractions of the third step. A fourth step of dividing the second fraction into five fractions by performing a reversed-phase ODS flash column with a solvent of 50 to 100% methanol; the highly active fraction of the fourth fraction is acetone: methylene chloride (8-30%). ) Can be divided into six fractions by performing a silica open column as an eluting solvent, and the compounds of the present invention can be obtained from the highly active fractions.

      The composition for preventing and treating cancer diseases of the present invention comprises 0.1 to 50% by weight of the compound based on the total weight of the composition.

      However, the composition as described above is not necessarily limited thereto, and may vary according to the condition of the patient and the type and extent of the disease.

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

  The compositions comprising the compounds according to the invention are each formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories, and sterile injectable solutions according to conventional methods. Carriers, excipients and diluents which may be used in combination with the extract, and which may be included in the composition comprising the extract include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin , Calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and the solid preparations may include at least one excipient such as starch, calcium carbonate, sucrose in the extract. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium styrate talc are also used. Oral liquid preparations include suspending agents, liquid solutions, emulsions, and syrups, and may include various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. . Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

Preferred dosages of the compounds of the present invention depend on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, but may be appropriately selected by those skilled in the art. However, for the desired effect, the compound of the present invention is preferably administered at 0.01 mg / kg to 10 g / kg, preferably 1 mg / kg to 1 g / kg per day. Administration may be administered once a day or may be divided several times. Therefore, the above dosage does not limit the scope of the present invention in any aspect.

       The composition of the present invention can be administered to mammals such as rats, mice, livestock, humans, etc. by various routes. All modes of administration can be expected, for example by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

      The present invention also provides a health functional food for the prevention and improvement of cancer diseases containing the compound represented by the general formula (I) isolated from the original extract as an active ingredient.

      The composition containing the compound of the present invention can be used in various ways, such as drugs, food and beverages for the prevention and improvement of cancer diseases. Examples of the food to which the compound of the present invention may be added include various foods, beverages, gums, teas, vitamin complexes, health supplements, and the like, and may be used in the form of powders, granules, tablets, capsules, or beverages. have.

       Since the compound of the present invention has little toxicity and side effects, it is a drug that can be used safely even when taken for a long time for the purpose of prevention.

       The compounds of the present invention may be added to food or beverages for the purpose of preventing and ameliorating cancer diseases. In this case, the amount of the compound in the food or beverage is generally added to the health food composition of the present invention to 0.01 to 15% by weight of the total food weight, the health beverage composition is 0.02 to 10 g, preferably based on 100 ml Can be added in a ratio of 0.3 to 1 g.

       In addition to containing the extract as an essential ingredient in the indicated proportions, the health beverage composition of the present invention has no particular limitation on the liquid component, and may contain various flavors or natural carbohydrates as additional ingredients, such as ordinary drinks. Examples of the above-mentioned natural carbohydrates are conventional monosaccharides such as disaccharides such as glucose and fructose, such as maltose, sucrose and the like, and polysaccharides such as dextrin, cyclodextrin and the like. Sugars and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (tauumatin, stevia extract (for example, rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of natural carbohydrates is generally about 1-20 g, preferably about 5-12 g per 100 ml of the composition of the present invention.

      In addition to the above, the composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and its Salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like. The compositions of the present invention may also contain pulp for the production of natural fruit juices and fruit juice beverages and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical but is generally selected from the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

The composition containing the compound isolated from the original extract of the present invention as an active ingredient exhibits cytotoxicity against various types of human cancer cells, and has a selective growth inhibitory activity against human lung cancer cells and has a great effect on the growth of normal cells. By confirming that it has excellent anticancer activity, it can be usefully used as a pharmaceutical composition and health functional food for the prevention and treatment of cancer diseases.

        Hereinafter, the present invention will be described in detail by Examples and Experimental Examples.

       However, the following Examples and Experimental Examples are only illustrative of the present invention, and the content of the present invention is not limited to the following Examples and Experimental Examples.

Example  1. Isolation of Compound

1-1. Preparation of Original Extract

       10 kg of dried Wonhwa (Herbal Medicine) was extracted three times in a water bath three times using an ultrasonic extractor device with chloroform: acetone: methanol (1: 1: 1). The extract was filtered and concentrated under reduced pressure to obtain 386 g of the original extract. The original extract was dispersed in methanol: chloroform (2: 8), filtered and concentrated under reduced pressure to obtain 216 g of the original methanol and chloroform extract (hereinafter referred to as 'WH'), which were obtained in Examples 1-2 below. Was used.

1-2. Isolation of Compounds from Native Extracts

WH obtained in Example 1-1 was subjected to silica gel column chromatography using chloroform, methylene chloride (1: 1 (v / v)) and methanol: chloroform (1: 1 (v / v)) solvent. To give three fractions. Among them, the second fraction having the highest cell growth inhibitory effect on human lung cancer cells (A549) was taken, and four fractions were obtained by ODS flash chromatography with 50-100% methanol. The third fraction having the highest activity was taken and six fractions were obtained by silica column chromatography using acetone: methylene chloride solvent gradient (5: 100 → 30: 100 (v: v)). The third fraction of this was 93% methanol using high performance liquid chromatography (ODS HPLC) to obtain novel substance (1), 5beta-hydroxyresinifenol-6 alpha, 7 alpha-epoxy-12 beta-butanoxyl. -9,13,14-ortho-2E, 4E-dicadienoate (40 mg) was isolated (hereinafter referred to as "yuanhualine" or "YH1"), and the fifth fraction was New material (2) 5beta-hydroxyresinifenol-6alpha, 7alpha-epoxy-12beta-propanoxyl-9,13, using high performance liquid chromatography (ODS HPLC) with 93% methanol 14-Orso-2E, 4E-dicadienoate (40 mg) was isolated (hereinafter referred to as "yuanhuahine" or "YH2").

1-3. Determine the structure of the compound

Analytical HPLC was used to evaluate the purity of the components isolated in Examples 1-2 above. Various spectroscopic experiments were carried out in the state of being identified as a single component, that is, H ¹ -NMR, C ¹³ -NMR, DEPT, H ¹ -H ¹ COZY, H ¹ -C ¹³ COZY, HMBC, and HRESIMS. The structure was determined. Yuanhuahin, a novel compound, was obtained in a white amorphous state and showed molecular ions of m / z 601 [M + H] + and 623 [M + Na] + in HRESIMS, yielding a molecular formula of C ₃₃ H ₄₄ O ₁₀ could be predicted. As shown in Table 1, huahuahin was similar to the result of huahuadin, except for one additional methylene by ¹ H NMR spectral analysis. Signals for five methyl groups were observed in the ¹ H NMR spectrum and six apliphatic methylenes, one exo-methylene, seven aliphatic methines and five olefinic methines were observed. According to the ¹³ C NMR spectrum with DEPT and HMQC spectra, 33 carbon atoms consist of 5 methyl carbon atoms, 7 methylene carbon atoms, 12 methine carbon atoms and 9 quaternary carbon atoms. The new compound huahuarin was obtained in a white amorphous state and showed molecular ions of m / z 615 [M + H] + and 637 [M + Na] + in HRESIMS, yielding a molecular formula of C ₃₄ H. ₄₆ O ₁₀ was identified (see Table 1).

As shown in Table 2, huahuarin was similar to the results of huahuadin, except for two additional methylenes by ¹ H NMR spectral analysis. It can be seen that the molecular weight of yuanhuarin is 2 × 14 mmu higher than yuanhuadine (see Table 2). (Zhan et al., Bioorgan Med Chem 13 , pp. 645-55, 2005).

NMR data of Yuan Hua Hin Assignment ¹³ C (ppm) ¹ H (ppm) One  160.7  7.57 (1H, broad singlet) 2  137.1 3  209.7 4   72.7 5   72.4  4.26 (1 H, s) 6   60.7 7   64.3  3.56 (1H, s) 8   35.6  3.51 (1H, d, 2.7) 9   78.3 10   47.7  3.92 (H, br s) 11   44.3  2.36 (1H, g, 7.2) 12   78.3  4.99 (1H, s) 13   83.9 14   80.7  4.76 (1H, d, 2.7) 15  143.3 16  113.6  5.01 and 4.95 (2H, broad singlet) 17   18.9  1.84 (3H, s) 18   18.5  1.31 (3H, d, 7.2) 19   10.1  1.80 (3H, dd, 2.7, 1.2) 20   65.2  3.94 and 3.81 (2H, d, 11.7) One'  117.2 2'  122.5  5.65 (1H, d, 16.5) 3 '  135.3  6.67 (1H, dd, 16.5, 10.5) 4'  128.8  6.05 (1H, 15.0, 10.5) 5 '  139.6  5.87 (1 H, m) 6 '   32.9  2.10 (2H, q, 7.2) 7 '   28.9  1.39 (2H, m) 8'   31.5  1.26 (2H, m) 9 '   22.7  1.31 (2H, overlapped) 10 '   14.2  0.89 (3H, t, 6.9) One''  173.4 2''   28.0  2.25 (2H, q, 7.5) 3 ''    9.2  1.09 (3H, t, 7.5)

NMR data of yuanhuarin Assignment ¹³ C (ppm) ¹ H (ppm) One  159.4  7.36 (1H, broad singlet) 2  136.8 3  208.9 4   72.7 5   70.2  4.00 (1H, d, 2.1) 6   61.5 7   63.8  3.33 (1H, d, 6.6) 8   35.2  3.32 (1H, 3.9) 9   78.3 10   47.3  3.70 (1H, br s) 11   43.8  2.24 (1 H, m) 12   78.1  4.08 (1 H, s) 13   83.6 14   80.3  4.61 (1 H, d, 2.7) 15  143.0 16  113.2  4.85 and 4.79 (2H, br s) 17   18.4  1.67 (3H, s) 18   18.0  1.12 (3H, d, 7.2) 19 9.6  1.61 (3H, br s) 20 64.7  3.56 and 3.76 (2H, broad doublet, 12.3) One'  116.9 2' 122.2  5.46 (1H, d, 15.3) 3 ' 135.0  6.49 (1H, doublet, 15.3, 10.5) 4' 128.4  5.88 (1H, m) 5 ' 139.3  5.70 (1H, m) 6 ' 32.5  1.94 (2H, q, 6.9) 7 ' 28.5  1.21 (2H, m) 8' 31.2  1.14 (2H, m) 9 ' 22.3  1.10 (2H, m) 10 ' 13.7  0.72 (3H, t, 6.6) One'' 172.9 2'' 36.2  2.06 (2H, t, 7.5) 3 '' 18.0  1.42 (2H, m) 13.3  0.75 (3H, t, 7.5)

Experimental Example 1. Measurement of human cancer cell growth inhibition efficacy in vitro

The effect on the cell growth of the compounds obtained in Example 1-2 was determined by sulforhodamine B (SRB method) (Lee et al., Chemico-Biol Interact 115 , pp. 215-228, 1998 ).

Human Cancer Cell Line (Korea Cell Line Bank) (A549, Human Lung Cancer Cell; HCT116, Human Colon Cancer Cell; T47D, Human Breast Cancer Cell; PC-3, Human Prostate Cancer Cell; SNU-638, Human Gastric Cancer Cell; SK-HEP-1, Human Liver cancer cells) were passaged in RPMI / MEME / DMEM medium containing 10% FBS, 1% PSF and the like. In each well of a 96-well plate, 10 μl of sample dissolved in 10% DMSO and 190 μl of the cell suspension (5 × 10 ⁴ cells / ml) were added and incubated for 3 days. 190 μl of the cell suspension was added to at least 16 wells and incubated for 30 minutes to be used as a zero-day control before the experiment. The cultured cells were fixed with 10% trichloroacetic acid (TCA) and stained with SRB solution, and the dye solution was dissolved with 10 mM Tris-base, and the absorbance was measured at 515 nm. In the case of incubation in 10% DMSO as a control cell viability according to each test material treatment was measured using the following equation (1).

When the control group was not treated with the sample at 100%, the value of the sample treatment group was expressed as a percentage of the control group, and each test substance treatment was calculated as the average value ± SEM of the double or triple test. IC ₅₀ value is the concentration of test substance for 50% survival. The effects of yuan hua hin and yuan hua rin on human cancer cells and human lung cells are shown in Table 3 and Tables 4 to 1 (see Fig. 1).

As shown in Table 3, the compounds showed the effect of selectively inhibiting the growth of human lung cancer cells in a concentration-dependent manner. The IC ₅₀ for human lung cancer cells was 0.7-53 nM and the IC ₅₀ was around 10 μM for other cancer cell tumors, indicating that the selectivity for lung cancer cells was more than 10,000 times higher (see FIG. 1).

     On the other hand, as shown in Table 4, the compounds showed a low cell growth inhibitory effect on human normal lung cells compared to human lung cancer cells was observed that the relatively low toxicity. (See Figure 1).

Experimental Example 2 Observation of Cell Morphology in Vitro

Human lung cancer cells A549 cells were suspended in 10% FBS-MEME at 2 x 10 ⁵ cells / ml. 10 ml of cell suspension was attached to a 10 cm cell culture dish. Yuan Hua Hin of Example 1-2 (5-100 nM concentration) was added and then incubated for a certain time at 37 ℃, 5% CO ₂ . First, the cells were incubated for 48 hours to observe the morphological changes of the cells, and then the culture medium was removed and washed twice with PBS (phosphate buffered saline). The morphology of the cells was observed at 100 times magnification under the microscope. The results are shown in FIG. 2 (see FIG. 2).

     As a result, as shown in Figure 2, it was confirmed that the number of cells treated with hua hua hin compared to the control cells.

 Hereinafter, the preparation examples of the composition containing the compound of the present invention will be described, but the present invention is not intended to limit the present invention, but is intended to be described in detail.

Formulation example  One. Powder  Produce

Yuanhuahin 20 mg

Lactose 100 mg

Talc 10 mg

The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

Formulation example  2. Preparation of Tablets

Yuanhuarin 10 mg

Corn starch 100 mg

Lactose 100 mg

2 mg magnesium stearate

After mixing the above components, tablets are prepared by tableting according to a conventional method for preparing tablets.

Formulation example  3. Manufacture of capsule

Yuanhuahin 10 mg

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium Stearate 0.2 mg

According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

Formulation example  4. Preparation of Injectables

Yuanhuarin 10 mg

Mannitol 180 mg

Sterile distilled water for injection 2974 mg

Na ₂ HPO _{4 ,} 12H ₂ O 26 mg

According to the conventional method for preparing an injection, the amount of the above ingredient is prepared per ampoule (2 ml).

Formulation example  5. Liquid  Produce

Yuanhuahin 20 mg

10 g of isomerized sugar

5 g of mannitol

Purified water

According to the conventional method of preparing a liquid solution, each component is added and dissolved in purified water, lemon flavor is added to the mixture, and then the above ingredients are mixed, purified water is added to adjust the total amount to 100 ml, and then filled in a brown bottle. The solution is prepared by sterilization.

Formulation example  6. Manufacture of healthy food

Yuanhuarin 1000 mg

Vitamin mixture proper amount

70 μg of Vitamin A Acetate

Vitamin E 1.0 mg

Vitamin B1 0.13 mg

Vitamin B2 0.15 mg

Vitamin B6 0.5 mg

0.2 μg of vitamin B12

Vitamin C 10 mg

10 μg biotin

Nicotinic Acid 1.7 mg

50 μg folic acid

Calcium Pantothenate 0.5mg

Mineral mixture

Ferrous Sulfate 1.75 mg

Zinc Oxide 0.82 mg

Magnesium carbonate 25.3 mg

Potassium monophosphate 15 mg

Dibasic calcium phosphate 55 mg

Potassium Citrate 90 mg

Calcium Carbonate 100 mg

Magnesium chloride 24.8 mg

Although the composition ratio of the above-mentioned vitamin and mineral mixtures is mixed with a component suitable for a health food in a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional health food manufacturing method. The granules may be prepared and used for preparing a health food composition according to a conventional method.

Formulation example  7. Manufacture of health drinks

Yuan Hua Hin 1000 mg

Citric acid 1000 mg

100 g oligosaccharides

Plum concentrate 2 g

1 g of taurine

Add 900 ml of purified water

After mixing the above components in accordance with a conventional healthy beverage production method, and stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained in a sterilized 2 L container, sealed sterilization and then refrigerated and stored in the present invention For the preparation of healthy beverage compositions.

Although the composition ratio is a composition that is relatively suitable for the preferred beverage in a preferred embodiment, the compounding ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and usage.

      1 is a diagram showing the growth inhibitory effect of hua hua hin on human lung cancer cells and normal lung cells of the present invention,

Figure 2 is a view showing a micrograph of the human lung cancer cell growth inhibitory effect of yuan hua hin of the present invention.

Claims (9)

Novel compounds of formula (I) or pharmaceutically acceptable salts thereof, isolated from the original extract:

                                                     (I) Where R is a substituent group selected from the group consisting of hydrogen atoms, C ₂ to C ₁₀ alkyl group and a C ₂ to C ₁₀ alkoxy. The method of claim 1 wherein the general formula (I) compound R is hydrogen atom, C ₂ to C ₂ lower alkyl group or a lower alkoxy group to a compound or a pharmaceutically acceptable salt thereof of the C ₅ C _5. The compound of formula (I) according to claim 2, wherein the compound of formula (I) is 5beta-hydroxyresinifenol-6alpha, 7alpha-epoxy-12beta-propanoxyl-9,13,14-ortho-2E , 4E-dicadienoate or 5 beta-hydroxyresinifenol-6 alpha, 7 alpha-epoxy-12beta-butanoxyl-9,13,14-ortho-2E, 4E-dicadienoate or Pharmaceutically acceptable salts thereof. A pharmaceutical composition for the prevention and treatment of cancer diseases, comprising as an active ingredient a novel compound represented by the general formula (I) of claim 1 or an pharmaceutically acceptable salt thereof isolated from the original extract. The pharmaceutical composition according to claim 4, wherein the original flower comprises buds, stems, leaves, or roots of red bean flowers.       The method of claim 4, wherein the cancer disease is gastric cancer, colon cancer, breast cancer, lung cancer, non-small cell lung cancer, bone cancer, pancreatic cancer, skin cancer, head or neck cancer, skin or eye melanoma, uterine cancer, ovarian cancer, colon cancer, small intestine cancer , Rectal cancer, anal muscle cancer, fallopian tube carcinoma, endometrial carcinoma, cervical carcinoma, vaginal carcinoma, vulvar carcinoma, Hodgkin's disease, esophageal cancer, small intestine cancer, lymph gland cancer, bladder cancer, gallbladder cancer, endocrine cancer, thyroid cancer, Parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system (CNS) A pharmaceutical composition that is a tumor, primary CNS lymphoma, spinal cord tumor, brain stem glioma, or pituitary adenoma. 5. A pharmaceutical composition according to claim 4, comprising 0.1 to 50% by weight of said compound relative to the total weight of the composition. A health functional food for the prevention and improvement of cancer diseases containing the novel compound represented by the general formula (I) of claim 1 separated from the original extract as an active ingredient. The dietary supplement of claim 8 in the form of a powder, granule, tablet, capsule or beverage.

Images