KR101086258B1 - Preventive and curative extracts from medicinal plants and their composition for respiratory organ disease - Google Patents

Abstract

The present invention relates to herbal extracts useful for the prevention and treatment of respiratory diseases, and more particularly, herbal extracts extracted from pendulum, ginkgo biloba and wild boar muscles at an appropriate ratio, inhibit airway contraction, airway inflammation inhibitory activity, and 5-lipoxygenase inhibition. Asthma, acute bronchitis, allergic rhinitis, acute lower respiratory tract infections (bronchitis, bronchiolitis, etc.), acute upper respiratory tract infections (pharyngitis, tonsillitis) And laryngitis), and a pharmaceutical composition containing a mixed extract of gingival, ginkgo biloba and tricepsis, which is useful for the prevention and treatment of respiratory diseases such as laryngitis.

Respiratory disease, necrosis, ginkgo biloba, papillary muscle, suppress airway contraction, suppress airway inflammation, 5-lipoxygenase, phosphodiesterase 4, leukotriene D4 antagonism, asthma, bronchitis, allergic rhinitis, sore throat, tonsillitis, laryngitis

Description

Herbal extracts useful for preventing and treating respiratory diseases {Preventive and curative extracts from medicinal plants and their composition for respiratory organ disease}

The present invention relates to herbal extracts useful for the prevention and treatment of respiratory diseases, and more particularly, herbal extracts extracted from pendulum, ginkgo biloba and wild boar muscles at an appropriate ratio, inhibit airway contraction, airway inflammation inhibitory activity, and 5-lipoxygenase inhibition. Asthma, acute bronchitis, allergic rhinitis, acute lower respiratory tract infections (bronchitis, bronchiolitis, etc.), acute upper respiratory tract infections (pharyngitis, tonsillitis) by confirming that the activity, phosphodiesterase 4 inhibitory action, leukotriene D4 antagonism is excellent And laryngitis), and a pharmaceutical composition containing a mixed extract of gingival, ginkgo biloba and tricepsis, which is useful for the prevention and treatment of respiratory diseases such as laryngitis.

Asthma, a representative respiratory disease in respiratory diseases, is a condition in which respiratory distress, cough, and breathing swelling (wheezing) are repeatedly seizures. About 30% of asthmatic patients within 80 years of age Asthma shows asthma symptoms at the age of 4 to 5 years old, and in Korea, the incidence rate is about 10%, and there are cardiac asthma and bronchial asthma. The former is a paroxysmal breathing difficulty seen in heart disease or high blood pressure, and is caused by hypersensitivity of the respiratory tract due to pulmonary congestion or tissue congestion and poor prognosis. The latter is a paroxysmal respiratory distress caused by narrowing of breathing due to bronchial spasm, narrowing, and mucosal swelling. In addition, there are cerebral asthma caused by disorders of the respiratory central region, and uremia asthma occurring in patients with nephropathy.

The incidence of asthma varies according to country, race, and age, but according to a 1991 UK report, about 7% of adults and 13.5% of pediatric age groups reported changes in living conditions, pollution, and stress in Korea. The trend is gradually increasing. Although asthma varies depending on the cause, in modern times when environmental pollution such as pollution is serious, the severity of asthma is maximized, such as the age of onset and the symptoms are prolonged.

The characteristic airway obstruction of asthma occurs by a three-step process. In other words, the contraction of bronchial smooth muscle, thickening of the pulmonary mucosa, and sticky mucus in the bronchus and bronchioles block the airways. Only the contraction of the double bronchial smooth muscle recovers easily.

The exogenous (allergic) asthma attack mechanism is particularly important for antibodies IgE, and some are related to IgG. IgE releases mediators (histamine, SRS-A, ECF-A, NCF, PAF, Kinin, PGs, etc.) that activate mast cells to cause hypersensitivity. Non-allergy Asthma attacks are thought to be mediated by unknown or autonomic nerves. Cholinergic stimulation in endogenous patients not only favors histamine-mediated mediators in mast cells via end organs, but also increases goblet cell secretion, enlarges blood vessels in the lungs, and constricts the trachea, bronchus and giant bronchioles. Bronchospasm and mucus secretion increase.

There is currently no fundamental cure for asthma. Treatment involves several methods and drugs to prevent seizures and prevent complications, but it is not satisfactory. The most important thing is to find and avoid the trigger. Treatment of asthma is mainly for bronchodilators for inhalation, oral or injectable bronchodilators (sympathetic and theophylline drugs), stereoforms (for inhalation, oral, injectables, etc.), leukotriene antagonists (montelukast, fran Lucast, ziluton and the like), and anti-allergic agents (disodium chlorochlorate, chromoline sodium, ketotifen) are used.

Bronchitis is divided into acute and chronic, and the causes are allergic, infectious, traumatic, etc., and pathologically catarrhal, purulent, obstructive, ulcerative, and invasive. The most common cause is infection by bacteria, viruses, fungi, and the like, and people who are not normally infected are susceptible to infection when the resistance of the whole body is weakened. Allergic bronchitis is a direct reaction from inhaling allergens and partial symptoms from systemic allergic reactions. The trauma may be caused by chemical stimulation such as chlorine or sulfur dioxide, or by physical stimulation such as dust. Contaminated air in large cities is chronically irritating and inflammatory. Pathologically, acute cases include redness, swelling, and dryness in the bronchial mucosa, and mucous or purulent secretions can be seen in the bronchus. It usually recovers without causing complications, but when it is chronically developed, it causes swelling, thickening, and atrophy of the mucous membrane, which can lead to proliferation of fibers, bronchial narrowing, and emphysema. . The most prominent symptoms are cough and sputum. When the cause is caused by infection, fever and chest pain can be seen. In traumatic cases, the mucous membranes of the mouth, nose, and eyes also appear. Coughing is a type of defense in the treatment, so trying to stop it is not a good idea. You must treat the cause at the same time. In winter, increase the room temperature and use a small amount of codeine, atropine, ephedrine, and antihistamines. Steroids or theophylline may be used for inflammation, but the effect is not as expected.

Allergic rhinitis is also referred to as nasal allergy or allergic rhinitis, symptoms of sudden sneezing, watery nasal juice (clear runny nose), large amounts of nasal congestion, and heavy head and tears. The symptoms are very similar, but the antigen is not clear is called vasomotor rhinitis. In other words, when one wakes up in the morning and the body cools down temporarily, the symptoms mentioned above may occur and heal in a few hours. It is often confused with a cold, which is different from the common cold and often involves asthma or hives.

An allergic reaction of allergic rhinitis is a type of antigen-antibody hypersensitivity. Histamine is released from mast cells and basophils, and arachidonic acid is released to release prostaglandins and leukotrienes by cyclooxykenase (COX) and 5-lipooxygenase (5-LO). It produces | generates and mediates the initial reaction which appears between 2 to 90 minutes of antigen exposure, and the late reaction which appears after 4 to 8 hours. The initial response is mainly due to the mediator and the late response is mainly due to the cell infiltration. In addition, allergic and nonallergic rhinitis both act as risk factors for the development of asthma.

Treatment is desensitization (항원 感 作 분명 法) when the antigen is clear, and other drug therapy, surgical therapy, physical therapy, etc., it is difficult to recover completely.

There are many types of airway infections, such as common purulent bacteria, special bacteria (such as Mycobacterium tuberculosis, Diphtheria bacteria, Spiroherta), viruses, and fungi, and are classified into acute and chronic. When the nasal cavity, pharynx, and larynx are individually infected, they are called their respective organ names, but when their entire organs are in an infected state, they are called upper respiratory tract infections. In addition, in case of infection by special bacteria or fungi, it is often called by special names such as upper island tuberculosis, said dodiphtheria, said said candidiasis.

As described above, respiratory diseases such as asthma, allergic rhinitis, and acute bronchitis have some differences in their causes and symptoms, but they have some commonalities.

First, all are aspects of inflammatory diseases. These respiratory diseases are caused by allergies and infections, but inflammation plays an important role in the aggravation and treatment of the disease. That is, influx and activation of leukocytes triggered by allergy, infection, etc. into the respiratory tract and various cytokines and inflammatory mediators released from leukocytes make the disease worse and difficult to treat.

Second, the contraction and relaxation of the airway is not normal, breathing is difficult. In other words, the airway is damaged, excessive response to the general stimulation (asthma) or narrowed airway (bronchitis) is difficult to breathe, so appropriate treatment is necessary.

Third, anti-inflammatory agents, airway contraction inhibitors and dilatants, airway secretion inhibitors play an important role in the main drug therapy, many drugs are commonly used. Antihistamines, anticholinergic agents, beta2 receptor agonists, steroids, leukotrienes D4 receptor antagonists, and phosphodiesterase 4 inhibitors of theophylline are widely used. Nevertheless, airway dilators such as anticholinergic agents and beta2 receptor agonists do not have an effect on the inflammation that aggravates the disease, and thus only relieve symptoms. Steroids, which are known to be effective against inflammation, are problematic for long-term use due to severe side effects and are known to be less effective for chronic bronchitis. Therefore, a combination of the two is often prescribed, due to the side effects of steroids developed in the form of inhalants than the oral drug is difficult to take because there is a problem in taking compliance.

Overcoming the limitations of these current drug therapies, there is a need for the development of new therapeutic agents that can fundamentally cure the cause and effectively improve symptoms. However, respiratory diseases, as described above, are involved with various leukocyte cells and various cytokines and inflammatory mediators released therefrom, so it is difficult to effectively treat chemicals with a single component, and natural extracts with various components and mechanisms may be effective therapeutic agents. Can be.

In addition, respiratory diseases such as asthma, bronchitis, allergic rhinitis, acute lower respiratory tract infections (bronchitis, bronchiolitis, etc.), acute upper respiratory tract infections (sore throat, tonsillitis, laryngitis) are very diverse and cause only conventional treatment. The reality is that there is a problem in the treatment, such as recurrence after treatment. Therefore, prevention and treatment of respiratory diseases have emerged as one of the important tasks of the medical community, and the development of fundamental prevention and treatment is urgently needed.

On the other hand, the herbal medicine used in the present invention (Gentianae macrophyllae radix) is a perennial herb, the height is 40 ~ 60 cm, the root is firm and not elastic. It is distributed in Heilongjiang, Liaoning, North China, Henan, and Sichuan in China. It is collected in spring and autumn. Remove the foliage, beard root, black, and dry in the sun or in pairs to dry in the sun, and when it is reddish yellow or yellowish gray, spread it out and dry it again in the sun. In oriental medicine, it is effective in driving away wind, removing moisture, harmonizing blood, releasing muscles and tendons, removing heat, and diuresis. Pain caused by customs, spasms of musculature, jaundice, blood transfusion, and urine It has been used for the purpose of treating. The major components known so far are gentianine, gentianidine, gentianol, gentiopicroside, and the like. 5171-5177; In 1998, the Korean Medicine and Herbal Medicines Standards Collection, Korea Medical Index, p. 352; 1988].

Ginkgo biloba L. is a deciduous arboreous tree, 40 m high and 4 m in diameter. Leaves are fan-shaped, 4 to 8 cm long, 5 to 10 cm wide, and the ends are shallow to two in the middle. It is split, its base is wedge-shaped, and the leaf veins are parallel and bifurcated. Ginkgo biloba is called encyclopaedia. It is a leaf of Ginkgo biloba and is harvested in September ~ October and used to dry it in the sun. The taste is sweet, bitter and bitter, and the nature is flat. In oriental medicine, there are yeasts that replenish the core and converge the lungs to remove moisture and stop diarrhea. The main components of the ginkgo leaves known to date are flavonoids such as isorhamnetin, kaempferol and quercetin and their glycosides, ginkgetin, isoginkgetin and gincoride (ginkgolide) A, etc. [Hyangkae Dictionary, Book Publishing Younglim, pp. 122-124, Chinese Medicinal Dictionary, Book Publishing Jungdam, pp. 2028-2030; In 1998, the Korean Medicine and Herbal Medicines Standards Collection, Korea Medical Index, p. 303; 1988].

Sophorae Radix is shrub and 1 to 2 m high. Roots usually 2-5, cylindrical, yellowish brown. Stems are cylindrical, grooved on the surface, tightly covered with short, soft hairs, and the upper part of the stem is usually bent in an "之" shape. Dried roots, which are used as medicinal herbs, are long, cylindrical, slightly bent, 10-25-35 cm in length, and 0.3-1 cm in diameter. The surface is brown to blackish brown with longitudinal wrinkles, and has a long horizontal and slightly bumped crust. The main production area is the 廣西 of China. Herbal medicine has long been used to treat tumors, edema, pain, jaundice, diarrhea, and hemorrhoids. Major ingredients known to date include alkaloids such as matrine, oxymatrine, anagyrine, methylcytisine, and sophoranone, and sophoradine. Flavonoids such as sophoradin, sophoranochromene, and sophoradochromene are known [Chinese Dictionary, Book Publishing, Jeongdam, pp. 2627-2632 1998].

The effects of these extracts on the respiratory diseases have not been studied.

Therefore, the present inventors have studied to develop a drug for treating respiratory diseases, the results of the mixture of jinkyo, ginkgo biloba and wild boiled root extracts inhibit airway contraction activity, airway inflammation inhibitory activity, 5-lipoxygenase inhibitory activity, phosphodiesterase 4 The present invention was completed by confirming that the inhibitory action and leukotriene D4 antagonistic activity are excellent.

Therefore, an object of the present invention is to provide a drug for the prevention and treatment of respiratory diseases, which contains a mixture of jinyokgyo, ginkgo biloba and wild boar muscle as an active ingredient.

The present invention is characterized by a drug for the prevention and treatment of respiratory diseases containing a mixture of jinyokgyo, ginkgo biloba and wild bovine root as an active ingredient.

Hereinafter, the present invention will be described in detail.

According to the present invention, herbal extracts extracted from orthodontic gingiva, ginkgo biloba and wild erectus extracts at an appropriate ratio have anti-airway contraction inhibitory activity, airway inflammation inhibitory activity, 5-lipoxygenase inhibitory activity, phosphodiesterase 4 inhibitory action, and leukotriene D4 antagonist. By confirming its excellent function, ginseng leaves and acid are useful for the prevention and treatment of respiratory diseases such as asthma, acute bronchitis, allergic rhinitis, acute lower respiratory tract infection (bronchitis, bronchiolitis, etc.), acute upper respiratory tract infection (sore throat, tonsillitis, laryngitis) The present invention relates to a medicament containing a mixture of triceps.

Briefly explaining the manufacturing process of the mixture of Jingyo, Ginkgo biloba and wild bovine root according to the present invention,

1) After reflux extraction with water or alcohol solution of 10-15 times the weight of mixed herbal medicine with ginkgo leaf, ginkgo biloba and wild boiled root, it is filtered, and the residue is added with 7-12 times water or alcohol aqueous solution of the weight of mixed herbal medicine and warmed again. Filtering after extraction and mixing with the previous filtrate and filtering;

2) separating the filtrate obtained in 1) with the same amount of lower alcohol or nonpolar solvent, and then concentrating under reduced pressure at 60 to 70 ° C; And

3) azeotropically concentrating with water of 25 to 50 times the total amount of the concentrate of 2), homogeneously suspending with the same amount of water, followed by freeze drying.

To explain this in detail, water or alcohol solution is added to the mixture of Chinese herbal medicine, ginkgo biloba and wild boiled root, and refluxed for 2 to 5 hours. 10-15 times of is suitable. In addition, it is preferable that the mixed herbal medicine is mixed in a weight ratio of 1: 0.5 to 2.0: 0.5 to 2.0 of jinchuan, ginkgo biloba, and wild rump. If it is out of the above range is the problem that the extraction efficiency of the components effective in the prevention and treatment of respiratory diseases is poor and is not preferable. Then, the filtrate was collected by filtration, and the residue was added with an aqueous solution of water or alcohol (preferably ethanol, methanol, butanol) 7 to 12 times the weight of the raw material, and re-extracted for 2 to 5 hours after heating and filtered. To increase the extraction efficiency. If the amount of water is too small, the stirring becomes difficult and the solubility of the extract is low, so that the extraction efficiency is reduced. If the amount is too large, the use of lower alcohols and nonpolar solvents used in the next purification step is not economical, causing problems in handling. Can be.

In the present invention, the method of re-extracting after the first extraction is adopted. Even in the case of mass production of the herbal extracts, even though the filtration is effective, the loss occurs because the water content of the herbal medicine itself is high, so the extraction efficiency is reduced only by the first extraction. This is to prevent this. In addition, as a result of verifying the extraction efficiency of each step, it was found that about 80 to 90% of the total extraction amount is extracted by the second extraction, and the third or more multistage extraction is not economical.

As described above, the extract obtained by extraction with an aqueous solution of water or alcohol over the first and second stages is filtered and concentrated, and then purified from impurities such as unnecessary proteins, polysaccharides and fatty acids contained in the filtrate. The impurities are purified by performing layer separation 2-4 times with alcohol or nonpolar solvent to obtain a solvent fraction. The lower alcohol is preferably alcohol having 1 to 6 carbon atoms, more preferably butyl alcohol, propyl alcohol or isopropyl alcohol, and the nonpolar solvent is ethyl acetate, dichloromethane, chloroform, carbon tetrachloride and methyl ethyl ketone. When the amount of the lower alcohol or the nonpolar solvent is less than that of the filtrate, the particles are formed by unnecessary components such as fatty acids, resulting in poor layer separation and inefficient extraction of the active ingredient. .

The lower alcohol or nonpolar solvent fraction obtained after layer separation is concentrated under reduced pressure at 60 to 70 ° C. to remove the solvent remaining in the sample. The concentrate thus obtained is azeotropically concentrated two to three times with water of 25 to 50 times the total amount of the concentrate, followed by the addition of an equal amount of water and suspended homogeneously. The reason for azeotropic concentration with water at the time of concentrated drying is to effectively control the content of the remaining lower alcohol in order to use the obtained herbal extract as a pharmaceutical raw material.

The extract thus obtained is lyophilized to obtain a powdery final extract, which is expected to be very useful as a prophylactic and therapeutic agent for respiratory diseases because of its excellent airway contraction inhibitory activity and airway inflammation inhibitory effect.

Tablets, capsules, injections and the like are prepared by formulating in a conventional manufacturing method containing a mixture extract of jinchuan, ginkgo biloba and wild boiled roots according to the present invention, among which lactose, microcrystalline cellulose, magnesium stearate used as a base in the manufacture of tablets In combination with the present invention and the ratio of the present invention in a ratio of 2: 1, tablets having an activity for the prevention and treatment of respiratory diseases can be prepared.

In the manufacture of such pharmaceuticals, herbal extracts may be used as such, but may also be mixed with pharmaceutically acceptable carriers, excipients, diluents and the like to prepare powders, granules, capsules or injections. Is possible. In addition, the extract of the jinchuan according to the present invention has been used for food and medicinal use since ancient times, and there is no particular restriction on its dosage, body absorption, weight, age of patients, sex, dietary condition, administration time, administration method, excretion Rate, the severity of the disease, and the like. In general, it is preferable to administer about 0.1-10 mg / kg body weight of the mixture of jinchuan, ginkgo biloba and wild bovine root. Therefore, the extract containing the active ingredient of the present invention is to be prepared in consideration of the effective amount range, and the unit dosage form formulated in this way according to the judgment of experts and the needs of the individual to monitor or observe the administration of the drug as needed Specialized medications can be used or administered at regular intervals.

Hereinafter, the present invention will be described in more detail based on the following examples, but the present invention is not limited thereto.

Preparation Example 1 Preparation of Mixed Extract of Orthodontic Ginkgo Biloba Leaves

The mixture was washed with water to remove impurities, 100 g of dried finely pulverized gyoza, 100 g of ginkgo biloba, and 100 g of triceps were evenly mixed, followed by hot water extraction for 5 hours with 2 L of water. The filtrate was collected and collected, and the residue was added with 2 L of water and extracted by reheating for 3 hours. The filtrates were combined and concentrated to 1 L. An equal amount of saturated n-butyl alcohol was added thereto, followed by three layer separations, and only n-butyl alcohol was collected and concentrated under reduced pressure until the herbal extract was dried at 65 ° C. Most of n-butyl alcohol and water are evaporated, 0.1 L of water is added, the azeotropic concentration is concentrated, and then repeated two times. Bicep mixed extract was obtained.

Preparation Example 2 Preparation of Mixed Extract of Orthodontic Ginkgo Biloba Leaves

Remove impurities by washing with water, mix 100 g of dried finely pulverized oak, 100 g of ginkgo biloba, and 100 g of wild boil, and add 2 L of 50% (v / v) ethanol in water and stir well. Time reflux extraction. The filtrate was collected and collected. The residue was added to 1.5 L of 30% (v / v) ethanol aqueous solution and re-extracted for 3 hours, and then the filtrates were combined and concentrated to 1 L. An equal amount of saturated n-butyl alcohol was added thereto, followed by three layer separations, and only n-butyl alcohol was collected and concentrated under reduced pressure until the herbal extract was dried at 65 ° C. Most of n-butyl alcohol and water were evaporated, 0.3 L of water was added, and then azeotropically concentrated. Then, the resultant was repeated twice. Bicep mixed extract was obtained.

Comparative Example 1: Preparation of Orthodontic Extract

The mixture was washed with water to remove impurities, and 300 g of well-dried finely pulverized granules were evenly mixed. Then, 2 L of water was added thereto, followed by hot boiling extraction with stirring. The filtrate was collected and collected, and the residue was added with 2 L of water and extracted by reheating for 3 hours. The filtrates were combined and concentrated to 1 L. An equal amount of saturated n-butyl alcohol was added thereto, followed by three layer separations, and only n-butyl alcohol was collected and concentrated under reduced pressure until the herbal extract was dried at 65 ° C. After most of n-butyl alcohol and water were evaporated, 0.1 L of water was added, the azeotropic concentration was repeated, and then repeated twice. Finally, the same amount of distilled water was added and suspended, followed by freeze-drying to obtain powdery powdered extract.

Comparative Example 2: Preparation of Ginkgo Leaf Extract

The mixture was washed with water to remove impurities, 300 g of well-dried ginkgo biloba were mixed evenly, and 2 liters of water was added thereto, followed by hot water extraction with stirring. The filtrate was collected and collected, and the residue was added with 2 L of water and extracted by reheating for 3 hours. The filtrates were combined and concentrated to 1 L. An equal amount of saturated n-butyl alcohol was added thereto, followed by three layer separations, and only n-butyl alcohol was collected and concentrated under reduced pressure until the herbal extract was dried at 65 ° C. After most of n-butyl alcohol and water were evaporated, 0.1 L of water was added, the azeotropic concentration was repeated, and then repeated twice. Finally, the same amount of distilled water was added, suspended, and freeze-dried to obtain a powdered ginkgo leaf extract.

Comparative Example 3: Preparation of wild boar extract

The mixture was washed with water to remove impurities, 300 g of well-dried wild boar, and then evenly mixed. Then, 2 L of water was added thereto, followed by hot boiling extraction with stirring. The filtrate was collected and collected, and the residue was added with 2 L of water and extracted by reheating for 3 hours. The filtrates were combined and concentrated to 1 L. An equal amount of saturated n-butyl alcohol was added thereto, followed by three layer separations, and only n-butyl alcohol was collected and concentrated under reduced pressure until the herbal extract was dried at 65 ° C. After most of n-butyl alcohol and water were evaporated, 0.1 L of water was added to the azeotropic condensate, and the resultant was repeated twice. Finally, the same amount of distilled water was added and suspended. .

Example 1: Airway Shrinkage Inhibition Test in vitro )

In order to evaluate the airway contraction inhibitory activity of the extracts prepared in Preparation Examples 1 to 2 and Comparative Examples 1 to 3 were carried out by the following method using an extraction bronchus, the results are shown in Table 1 below.

Experimental Method

Hartely male guinea pigs (400-450 g, SLC, Japan) were sensitized by intravenous injection of anti-ovalbumin anti-serum at a volume of 1.5 ml / kg. 48 hours after sensitization, guinea pigs were bled and bronchus was removed. After removing the other tissues attached to the bronchus from the liquid Krebs-Heseleit, the cartilage was incised in a ring shape to include 2-3 cartilage. The cartilage part of the ring was cut while preserving the bronchial muscles, the threads were connected to both sides, and suspended in an organ bath. After stabilization for a certain time, 10 μg / ml of carbachol was added to induce maximum contraction. The bronchus was washed and stabilized with Krebs-Heseleit fluid. One minute after the extract to test was added 2 μmol indomethacin (indomethacin). After 5 minutes, 10 μg / ml of ovalbumin was added to cause contraction. Shrinkage was calculated by comparing the contraction with carbacol and egg albumin. Bronchoconstriction was measured using a physiological activity meter (MP150, BioPAC system) connected to a force transducer (FT4, BioPAC system).

division density Bronchoconstriction inhibition rate (%) Preparation Example 1 0.25 mg / ml 76 0.5 mg / ml 82 Production Example 2 0.25 mg / ml 82 0.5 mg / ml 88 Comparative Example 1 0.25 mg / ml 70 0.5 mg / ml 73 Comparative Example 2 0.25 mg / ml 73 0.5 mg / ml 78 Comparative Example 3 0.25 mg / ml 48 0.5 mg / ml 74 Positive control group Montelukast 10 μM 24 Rolipram 10 μM 72

As shown in Table 1, it can be seen that the extract prepared as in the present invention has a large degree of suppression of airway contraction of the extracted bronchus.

Example 2: Airway Shrinkage Inhibition Test in vivo )

In order to evaluate the airway contraction inhibitory activity of the extracts prepared in Preparation Examples 1 and 2 and Comparative Examples 1 and 3 were carried out by using the airway decay reaction induced when the antigen was exposed to the sensitized guinea pig, The results are shown in Table 2 below.

Experimental Method

Anti-ovalbumin antiserum was intravenously injected into a vein of Hartley male guinea pigs (400-450 g, SLC, Japan) at a volume of 1.5 ml / kg. After 48 hours the extract to be tested was orally administered. After 30 minutes, pyrilamine maleate (10 mg / kg), indomethacin (10 mg / kg), and propranolol (Propranolol, 0.1 mg / kg) were pretreated subcutaneously. Then, the various airway indicators of the guinea pigs were measured in a double chamber plethysmograph box equipped with a plethysmometer (HSE, German) and the basic airway resistance value was measured. After 30 minutes of pretreatment, 1% ovalbumin was aerosolized using high pressure compressed air and sprayed for 2 minutes. After 4 hours the extract to be tested was orally administered again. After 24 hours, the waste broth was washed with 1.5 ml of phosphate buffer solution to collect the washing solution. The white blood cell count and eosinophil count in the wash solution were counted.

division density Bronchoconstriction inhibition rate (%) Preparation Example 1 200 mg / ml 65 400 mg / ml 82 Production Example 2 200 mg / ml 67 400 mg / ml 85 Comparative Example 1 200 mg / ml 42 400 mg / ml 52 Comparative Example 2 200 mg / ml 60 400 mg / ml 78 Comparative Example 3 200 mg / ml 51 400 mg / ml 70 positivity
Control Montelukast 100 mg / kg 47 Franlukast 100 mg / kg 27

As shown in Table 2, it can be seen that the extract prepared as in the present invention has a large degree of suppressing airway contraction of the sensitized guinea pig.

Example 3: Airway Inflammation Inhibition Test

In order to evaluate the airway inflammation inhibitory activity of the extracts prepared in Preparation Examples 1 and 2 and Comparative Examples 1 to 3, Was carried out by the method, and the results are shown in Table 3 below.

Experimental Method

BALB / c female mice (6 weeks old, SLC, Japan) were sensitized by intraperitoneal administration of 0.2 ml of a mixture of 10 μg of egg albumin and alum on days 0, 7, and 14. After 8 and 10 days of final sensitization, 0.7% egg albumin was aerosolized using high-pressure compressed air and sprayed for 50 minutes to induce airway inflammation. After 24 hours of induction of inflammation, the waste bronchus was washed with 1.5 ml of phosphate buffer solution to collect the wash solution. The white blood cell count and eosinophil count in the wash solution were counted. The extract to be tested was orally administered 7 times at the final sensitization 7-10 days.

division density Airway inflammation inhibition rate (%) Preparation Example 1 400 mg / ml 31 800 mg / ml 58 Production Example 2 400 mg / ml 36 800 mg / ml 65 Comparative Example 1 400 mg / ml 26 800 mg / ml 35 Comparative Example 2 400 mg / ml 21 800 mg / ml 30 Comparative Example 3 400 mg / ml 25 800 mg / ml 55 Positive control group Rolipram 10 mg / ml 35

As shown in Table 3, the extract prepared as in the present invention can be seen that the degree of inhibition of airway inflammation is large.

Example 4: 5-lipoxygenase (5-LO) inhibitory action

In order to evaluate the inhibitory activity of the extracts prepared according to Preparation Examples 1 to 2 and Comparative Examples 1 to 3, the primary pathogenesis of asthma, the rucotriene producing enzyme was carried out by the following method using the cells, as a result Is as shown in Table 4 below.

Experimental Method

Human peripheral blood mononulear leukocytes (PBML) were stabilized at 37 ° C on Hank's balanced salt solution (HBSS), and then reacted for 15 minutes with the extract to be tested. Arachidonic acid (arachidonic acid) was added thereto as a substrate to produce rucotriene B4 (LTB4) for 15 minutes. The amount of LTB4 produced was measured using an Enzyme immunoassay (EIS) kit.

division density 5-lipoxygenase inhibition rate (%) Preparation Example 1 0.03 mg / ml 98 0.1 mg / ml 100 Production Example 2 0.03 mg / ml 100 0.1 mg / ml 100 Comparative Example 1 0.03 mg / ml 75 0.1 mg / ml 82 Comparative Example 2 0.03 mg / ml 85 0.1 mg / ml 97 Comparative Example 3 0.03 mg / ml 97 0.1 mg / ml 100 * Positive control NDGA (nordihydroguaiatinic acid) IC ₅₀ = 87.4 nM

As shown in Table 4, the extract prepared as in the present invention can be seen that the degree of 5-lipoxygase inhibitory action is large.

Example 5: Phosphodiesterase 4 (PDE4) Inhibitory Activity

In order to evaluate the inhibitory activity against phosphodiesterase 4, which plays an important role in the asthma onset of the extracts prepared in Preparation Examples 1 to 2 and Comparative Examples 1 to 3, cells were used in the following manner, The results are shown in Table 5 below.

Experimental Method

Human U937 cells were stabilized at 25 ° C. on a mixed solution of 50 mM Tris-HCl and 5 mM MgCl ₂ (pH 7.5). The amount of adenosine produced by adding a mixture of 1.01 μM ([ ³ H] cAMP + cAMP) as a substrate and reacting for 20 minutes was quantified by measuring the amount of [ ³ H] adenosine.

division density % Phosphodiesterase 4 inhibition Preparation Example 1 0.03 mg / ml 82 0.1 mg / ml 100 Production Example 2 0.03 mg / ml 88 0.1 mg / ml 100 Comparative Example 1 0.03 mg / ml 65 0.1 mg / ml 74 Comparative Example 2 0.03 mg / ml 70 0.1 mg / ml 92 Comparative Example 3 0.03 mg / ml 78 0.1 mg / ml 100 Positive control IBMX (3-isobutyl-1-methylxanthine) IC ₅₀ = 20.7 μM

As shown in Table 5, the extract prepared according to the present invention can be seen that the degree of phosphodiesterase 4 inhibitory action is large.

Example 6: Lukotriene D4 Receptor (Leukotriene D4, LTD4) Antagonism

In order to evaluate the inhibitory activity against the LTD4 receptor, which is the main pathogenesis of asthma, of the extracts prepared in Preparation Examples 1 and 2 and Comparative Examples 1 and 3, cells were used in the following methods, and the results are shown in Table 6 below. As shown in.

Experimental Method

LTD4 receptors isolated from lung tissue of Duncan Hartely series guinea pigs were 25 ° C. in 50 mM Tris-HCl buffer (5 mM CaCl ₂ , 5 mM MgCl ₂ , 100 μg / ml bacitracin, 1 mM benzamidine, 0.1 mM phenylmethylsulfonyl fluoride). Stabilized. 0.2 nM [ ³ H] LTD4 was added and reacted with the extract to be tested. Binding rate was analyzed by radioligand binding analysis, and nonspecific binding was determined using 0.1 μM LTD4. Specific binding rate 85%, Kd 0.2 nM, Bmax 0.24 pmol / mg protein.

division density LTD4 receptor inhibition rate (%) Preparation Example 1 0.03 mg / ml 80 0.1 mg / ml 92 Production Example 2 0.03 mg / ml 85 0.1 mg / ml 95 Comparative Example 1 0.03 mg / ml 41 0.1 mg / ml 58 Comparative Example 2 0.03 mg / ml 5 0.1 mg / ml 8 Comparative Example 3 0.03 mg / ml 78 0.1 mg / ml 89 Positive control LTD4 IC ₅₀ = 1.12 nM

As shown in Table 6, the extract prepared according to the present invention can be seen that the degree of LTD4 receptor antagonism is large.

Example 7 Antihistamine Action

In order to evaluate the inhibitory activity against histamine, which is the main pathogenesis of asthma, of the extracts prepared in Preparation Examples 1 and 2 and Comparative Examples 1 and 3, cells were used in the following methods, and the results are shown in Table 7 below. As shown.

Experimental Method

Human recombinant CHO-K1 cells were stabilized at 25 ° C. on 50 mM Tris buffer (including 2 mM MgCl ₂ , 100 mM NaCl, 250 mM sucrose). The extract to be tested and 1.2 nM [ ³ H] pyrylamine were added and reacted. Binding rate was analyzed by radioligand binding analysis, and nonspecific binding was determined using 1 μM pyrylamine. Specific binding rate 94%, Kd 1.1 nM, Bmax 6.7 pmol / mg protein.

division density Histamine activity inhibition rate (%) Preparation Example 1 0.1 mg / ml 52 Production Example 2 0.1 mg / ml 60 Comparative Example 1 0.1 mg / ml 13 Comparative Example 2 0.1 mg / ml 10 Comparative Example 3 0.1 mg / ml 45 * Positive control pyrilamine IC ₅₀ = 30.4 nM

As shown in Table 7, it can be seen that the extract prepared as in the present invention has a large degree of antihistamine action.

Example 8: Acute Toxicity of Oral Administration in Rats

Acute toxicity test was performed using 6-week-old SPF SD rats as follows.

Two animals per group were dissolved orthogonal, ginkgo biloba leaves and wild bovine root extracts prepared in Preparation Examples 1 and 2 and administered orally at a dose of 1 g / kg / ml. After administration of the test substance, mortality, clinical symptoms, and changes in body weight were observed. Hematological and hematological examinations were performed. Necropsy was performed to observe abdominal and thoracic organ abnormalities. As a result, no significant clinical symptoms or dead animals were noted in all animals treated with the test substance, and no toxic changes were observed in weight changes, blood tests, blood biochemical tests, and autopsy findings. Therefore, the extract according to the present invention did not show a toxicity change up to 1,000 mg / kg in all rats and the minimum lethal dose (LD ₅₀ ) orally administered was determined to be a safe substance more than 1,000 mg / kg.

Formulation Example 1 Preparation of Tablet

Tablets for oral administration were prepared using the wet granules method and the dry granules method with the following compositions using the extracts of orthodontic gingiva, ginkgo biloba and wild boar muscles of the present invention.

[Furtherance]                     

200 mg of mixed extract of ginkgo, ginkgo biloba and wild boil, hard mg of silicic anhydride, 2 mg of magnesium stearate, 50 mg of microcrystalline cellulose, 25 mg of starch glycolate, 113 mg of corn starch, ethanol anhydride.

Formulation Example 2: Preparation of Ointment

Using ointment, ginkgo biloba leaves and wild boar root extract of the present invention, an ointment was prepared in the following composition.

[Furtherance]

5 g of jinkyo, ginkgo biloba and wild boar root extract, 20 g of cetyl palmitate, 40 g of cetanol, 40 g of stearyl alcohol, 80 g of myristanisopropyl, 20 g of sorbitan monostearate, 60 g of polysorbate, paraoxy 1 g of benzoate, 1 g of methyl paraoxybenzoate, phosphoric acid and purified water

Formulation Example 3 Preparation of Injection

Injectables were prepared by the following compositions using the extract of jinjak, ginkgo biloba and wild bovine root.

[Furtherance]

100 mg of mixed extract of gyokyo, ginkgo biloba and wild boar, mannitol 180 mg, sodium dihydrogen phosphate 25 mg, water for injection 2974 mg

Formulation Example 4 Preparation of Transdermal

Transdermal preparations were prepared by the following methods using the mixed extract of gingival, ginkgo biloba and wild boar muscle of the present invention.

[Composition 1]

0.4 g of jinchuan, ginkgo biloba and wild boar mixed extract, 1.3 g sodium polyacrylate, 3.6 g glycerin, 0.04 g aluminum hydroxide, 0.2 g methyl paraben, 14 g water.

[Composition 2]

0.8 g of jinchuan, ginkgo biloba and wild boar mixed extract, 1.6 g propylene glycol, 0.8 g liquid paraffin, 0.4 g isopropyl myristate, 16.4 g gel bar 1430.

As described above, the extracts of orthodontic, ginkgo biloba and wild bovine root according to the present invention show good effects in various respiratory disease models, whereas the conventional synthetic medicines are effective only in some limited respiratory disease models. In other words, respiratory diseases such as asthma, acute bronchitis and allergy due to excellent airway contraction inhibitory activity, airway inflammation inhibitory activity, 5-lipoxygenase inhibitory activity, phosphodiesterase 4 inhibitory activity, leukotriene D4 antagonistic action It is very useful for the prevention and treatment of rhinitis, acute lower respiratory tract infections (bronchitis, bronchiolitis, etc.), acute upper respiratory tract infections (pharyngitis, tonsillitis, laryngitis).

Claims (3)

A drug for the prevention and treatment of respiratory diseases selected from asthma, acute bronchitis, allergic rhinitis, acute lower respiratory tract infection and acute upper respiratory tract infection, characterized in that it contains an extract of a mixture of orthodontic, ginkgo biloba and wild bovine root as an active ingredient. According to claim 1, wherein the extract is a drug, characterized in that the extract is extracted by mixing the weight ratio of 1: 1 ~ 0.5 ~ 2.0: 0.5 ~ 2.0 of the jinchuan, ginkgo leaves and wild bovine root. delete