KR20040018732A - Composition comprising the extract of Ixeris dentata for therapy against chronic viral hepatitis disease - Google Patents

Abstract

PURPOSE: Provided is a composition comprising an extract of Lxeris dentata for the treatment of chronic viral hepatitis disease. The extract of Lxeris dentata inhibits DNA proliferation of HBV to show antiviral activity, and is thus used for remedies or health foods for the prevention and treatment of chronic viral hepatitis disease. CONSTITUTION: A pharmaceutical composition for the prevention and treatment of chronic viral hepatitis disease is characterized by containing a crude extract of Lxeris dentata having effective inhibition effect on HBV DNA proliferation. The crude extract is obtained by using a polar solvent selected from water, methanol, ethanol, a mixture thereof, and a buffer solution.

Description

Composition comprising the extract of Ixeris dentata for therapy against chronic viral hepatitis disease}

The present invention relates to a composition for preventing and treating viral liver disease, and more particularly, the pharmaceutical composition useful for preventing and treating viral liver diseases such as hepatitis and liver cirrhosis, by using the inhibitory effect of hepatitis B virus transcriptase activity of the rind extract. A composition and a dietary supplement.

More than 300 million people worldwide are infected with hepatitis B virus and are estimated to be chronic carriers, and hepatitis C virus carriers are estimated to be about 100 million, and their numbers continue to increase. Each year, more than 2 million people have hepatitis due to chronic viral infections and develop cirrhosis and liver cancer. Types of viral hepatitis-inducing viruses are known as type A, type B, type C, delta type, GBV, etc., but type B and type C have the highest risk, and their pathological mechanisms are actively studied. It is becoming.

Hepatitis B virus (HBV), a representative virus that causes hepatitis, is a virus that is susceptible to mutation despite being a DNA virus. The HBV gene is about 3.2 Kb of incomplete double strandcyclic DNA and has an extremely compact structure. To date, the HBV DNA has a transcriptase gene (pol), a virus shell gene (capsid), a viral surface antigen gene (HBsAg), and several X genes involved in cancer function. Unlike ordinary DNA viruses that replicate DNA chains intact, HBV is first transcribed with 3.5 Kb of pregenomic RNA chains in the DNA chain by host transcriptase, and then reversed to virus-specific reverse transcriptase (pol). The double chain DNA is replicated by this. This replication process occurs independently of the host chromosome. However, some of the viral DNA has been found to enter the host's chromosome. When HBV DNA gets into the host's chromosome, the gene becomes unstable and the X protein expressed therein promotes transcription of the host cell and inhibits p53 gene function. Is likely to be. If this situation continues, it will develop into HCC.

As described, HBV exhibits a high degree of variability comparable to that of a normal RNA virus, due to its replication pattern similar to that of RNA viruses. HBV gene mutations occur randomly but the frequency is known to be constant. HBV gene mutations result in structural changes in surface antigens in the HBV envelope. Changes in the structure of various surface antigens are factors that can escape the humoral and cellular immune responses of the host, and are important factors in determining disease progression or persistent infection.

Hepatitis B has a latent period of 1-4 months and is more susceptible to infection through blood than oral infections. After hepatitis B, hepatitis B is easily fatigued and tasteless. Acute hepatitis develops into chronic form due to inadequate immune response to the host, or lack of T-cell function, endocrine functions such as steroids, and drugs.

Currently, only a prophylactic vaccine has been developed for the treatment of hepatitis, and no satisfactory treatment has been developed. Recently, a nucleoside analog called lamivudine has been clinically prescribed under the approval of the US FDA, and alpha-interferon is combined with lamivudine. It is becoming. On the other hand, hepatitis C has been shown to be somewhat effective because ribavirin, alpha interferon, etc. is a complex prescription. However, these synthetic drugs and recombinant medicines are still required to develop new drugs because they have physical problems such as side effects and recurrence caused by resistant viruses due to long-term use.

Chronic hepatitis virus is a causative virus that ultimately progresses from hepatitis to cirrhosis, which causes frequent liver failure or liver cancer. Analysis of serological data shows that 10-15% of those infected are infected with hepatitis B and C virus at the same time. These double infected people cause even more serious liver disease. In the United States alone, more than 10,000 people die each year from viral chronic liver disease, and the number of people infected is increasing every year. There are more than 100 million hepatitis B carriers in China, which account for 10-12% of the total population. Especially in Korea, there are more than 3 million chronic carriers as well as acute hepatitis patients. Moreover, in Korea, the proportion of early hepatitis B viral hepatitis virus carriers is known to be significantly higher for serious liver diseases such as chronic hepatitis, cirrhosis and liver cancer. Therefore, it is urgently needed to develop a therapeutic agent with lower side effects and lower cost than lamivudine, which can cure chronic viral hepatitis patients.

Drugs currently being used for the treatment of viral hepatitis and cirrhosis are alpha-interferon, lamivudine, and the like, which are known as immunoactive agents and anti-viral agents. These drugs are commercialized as clinically prescribed antiviral agents, but have many disadvantages. Lamivudine, an organosynthesized nucleoside analogue, is the biggest problem of drug-resistant virus in long-term use, and even if a mutation occurs in the YMDD motif of the site having reverse transcriptase activity of HBV transcriptase protein Adefovir, another nucleoside analogue that exhibits inhibitory activity, causes long-term doses of resistant viruses and renal failure.Also, alpha-interferon, a protein expressed by genetic recombination, is used in chronic C-type infected patients. Only about 30% of patients have a significant response and other side effects. Moreover, the long-term administration of alpha-interferon is a frequent and problematic problem for the emergence of resistant type C viruses against this drug.

In recent years, the combination prescription has become the best to suppress the emergence of drug-resistant mutant virus, so the combination administration of ribavirin and alpha-interferon has become common. Nevertheless, new forms of more complete therapies need to be released, which may be better than organic synthetic antiviral agents found in plant-derived natural products. However, although it is necessary to develop a therapeutic agent to replace the existing antiviral agent as a treatment for liver disease caused by chronic infection of hepatitis virus, it is considered that the time gap is too long until new drug development from these plant natural products is clinically applied.

Therefore, the purpose of prevention or treatment of hepatitis is to prevent the virus proliferation by prescribing a vaccine or to stop the progression of hepatitis by using drugs that inhibit the virus proliferation and to prevent the progression of cirrhosis and liver failure. I'm trying to stop. Therefore, the development of recombinant vaccines and new antiviral agents such as lamivudine and acyclovir, which are synthetic drugs, have been developed, but as a complete treatment for hepatitis, it is still in question, and its use is extremely limited due to side effects and high cost. It is true.

Therefore, researches are actively underway to create new therapeutic agents with low toxicity and side effects from folk remedies or natural products that have been applied for a long time. For example, a composition consisting of a mixed extract of Matari and Hwangbaekpi has been tried as a therapeutic agent for hepatitis B and liver cirrhosis (Korean Patent Application No. 2000-23564), tanshinol B, tanshinone IIB, prochebaquinone A, salvia extract and It has been suggested that the composition consisting of the saliva fraction is effective in the treatment of hepatitis B (Korean Patent Application No. 2000-27306).

Ixeris dentata , used in the present invention, is a perennial herb of Compositae, which is taken or crushed externally using decoction. It is effective in stopping diarrhea, reducing fever, regenerating and detoxifying skin. It is used to treat serpentine, urinary stones, eczema, pneumonia, bruises, and fractures.

Indeed, in the above-mentioned documents and research reports, the biological activity of herbal extracts using viral extract on viral liver disease has not been reported to date.

The present invention relates to a composition for the prevention and treatment of hepatitis, liver cirrhosis, especially caused by hepatitis B virus containing extracts of crust.

The inventors of the present invention have pharmacologically observed the therapeutic effect of viral extracts and cirrhosis of cerebrum extracts and combinations thereof, and whether there is a possibility for the prevention and treatment of hepatitis and cirrhosis with respect to the herbal medicine that has accumulated a large amount of clinical data. The main focus was on.

As a result, the present invention was completed by confirming that the extract extract according to the present invention inhibits the proliferation of hepatitis B virus transcriptase and thus has a prophylactic and therapeutic activity for viral liver disease.

Accordingly, it is an object of the present invention to provide a pharmaceutical composition and a dietary supplement showing a preventive and therapeutic effect of a viral liver disease containing a moth extract as an active ingredient having an inhibitory effect of hepatitis B virus replication enzyme.

FIG. 1 is a photograph showing the effect of inhibiting HBV transcriptase activity through the priming reaction after treatment with the extract of HBV transcriptase and five kinds of herbal extracts during the expression of activated HBV transcriptase in insect cell lines.

FIG. 2 is a photograph showing the inhibitory effect of HBV transcriptase activity of the compound medicinal herb through the priming reaction after treatment with the concentration of the compound containing the fungus upon expression of activated HBV transcriptase in insect cell lines.

Figure 3a shows the inhibition of viral replication activity of the compound containing the fungus, in the cells 4 days, 8 days after treatment with 0.15% of the drug extract to the cell line infected with Woodchuck hepatitis B virus (WHBV) Table comparing the amounts of pregenonomic RNA and WHBV surface antigen mRNA,

Figure 3b is treated with 0.15% complex drug to the cell line infected with Woodchuck hepatitis B virus in order to observe the inhibition of viral replication activity of the compound extract containing the moth, the viral DNA in the nucleus, intracellular virion DNA and DNA replication intermediates Southern blot analysis of the photographs.

In accordance with the above object, the present invention provides a crude extract of Ixeris dentata effective for inhibiting hepatitis B virus proliferation.

The crude extract means water, methanol, ethanol, lower alcohols, mixed solvents thereof and extracts soluble in buffers.

Hereinafter, the present invention will be described in detail.

In order to prepare a crude extract of Ixeris dentata effective for inhibiting the hepatitis B virus proliferation of the present invention,

The crude crude extract is washed and peeled after cutting off the bark, and then naturally dried at a temperature of 10 to 40 ° C., preferably 20 to 30 ° C. for about 1 to 30 days, preferably about 5 to 20 days. Powder, and then dissolved in an extraction solvent such as water, lower alcohol, buffer, and the like at a concentration of 1 to 40%, preferably 1 to 20%, at about 1 hour to 50 to 100 ° C, preferably 70 to 80 ° C. One day, preferably about 1 to 5 hours of hot water, then centrifuged to obtain the supernatant extract. In the present invention, as a preferred embodiment, crude extract is obtained by bathing the moth powder using phosphate buffer.

The present invention provides a pharmaceutical composition effective for the prevention and treatment of viral liver disease containing the moth extract obtained in the extraction process.

The composition for the prevention and treatment of viral liver disease of the present invention comprises 0.5 to 50% by weight of the extract based on the total weight of the composition.

It was confirmed that the activity of HBV transcriptase was significantly reduced when the mixed herbal medicinal solution containing the fungus obtained in the above manner was treated with an HBV replication gene-expressing insect cell line by concentration.

The composition comprising the moth extract of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

The composition comprising the extract of the present invention may further comprise a suitable carrier, excipient and diluent according to conventional methods.

Carriers, excipients and diluents that may be included in the composition comprising the extract of the present invention 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.

Compositions comprising extracts according to the invention are formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories and sterile injectable solutions, respectively, according to conventional methods. Can be used.

The pharmaceutical preparations thus prepared may be administered by a route selected from oral, parenteral (including transdermal, intradermal, intramuscular and intravenous), rectal and inhalation.

The amount of the extract of the present invention may vary depending on the age, sex, and weight of the patient, but in general, the amount of 0.1 to 1000 mg / kg, preferably 10 to 100 mg / kg, per adult It may be administered once to several times daily. In addition, the dosage of the extract can be increased or decreased depending on the route of administration, the degree of disease, sex, weight, age and the like. Therefore, the above dosage does not limit the scope of the present invention in any aspect.

In addition, the present invention provides a dietary supplement containing the moth extract.

The composition containing the moth extract of the present invention can be used in a variety of drugs, food and beverages for the prevention and treatment of viral liver disease. Examples of the food to which the compound herbal medicine containing the present extract may be added include various foods, beverages, gums, teas, vitamin complexes, and health supplements.

The moth extract itself of the present invention is a drug that can be used with confidence even for long-term use for the purpose of prevention because there is little toxicity and side effects.

The extract of the present invention may be added to food or beverage for the purpose of preventing viral liver disease. At this time, the amount of the extract in the food or beverage may generally be added at 0.01 to 30% by weight of the total food weight, the health beverage composition is 0.02 to 30 g, preferably 0.3 to 3 g based on 100 ml Can be added.

The health beverage composition of the present invention has no particular limitations on the liquid components other than the above-mentioned extracts as essential ingredients in the indicated proportions, and may contain various flavors or natural carbohydrates as additional ingredients, as in general beverages. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And conventional sugars such as polysaccharides such as dextrin, cyclodextrin, 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 said 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 salts thereof. , 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.

Hereinafter, the present invention will be described in more detail based on the following Examples and Experimental Examples, but the content of the present invention is not limited by the following Examples and Experimental Examples.

Example 1. Preparation of Crude Crude Extract

The moths (cultivated at the Suwon University laboratory) were washed with roots, air-dried at 20 ° C for 7 days, and then powdered with a blender.

Example 2. Preparation of a compound herbal extract containing bitter

84 g of the composite herbal powder containing 6 g of the bark powder of Example 1 was prepared in a phosphate buffered saline at a concentration of 10%, followed by agitation at 80 ° C. for 2 hours, and then at 10,000 rpm for 5 minutes. By centrifugation, the supernatant was taken and used as a sample for cell experiments.

Example 3. Preparation of a compound herbal extract containing crust

100 g of the complex herbal powder containing 7.1 g of the bark powder of Example 1 was added with 1 L of an aqueous 80% ethanol solution and refluxed four times at 50 ° C. for 1 hour, and then combined and filtered with a filter paper (Watman, USA). This was concentrated under reduced pressure with a reduced pressure concentrator (Eyela, N-1000, Japan) to obtain a 25g extract was used as a sample.

Reference example. Hepatitis B Virus Enzyme Expression

In order to express the hepatitis B virus polase (pol) protein in insect cell lines, an HBV cloning gene (Invitrogen) was cloned into baculovirus (FLAG-pol-stem-loop: FPL-pol). The Sf9 cell line was infected and transformed. Transformed FPL-pol insect cell lines were cultured in T-75 flasks, with 5% FCS (fetal calf serum, Gibco), 0.1% Pluronic F-68 in TNM-FH medium. And Grace medium (Grace's medium, Gibco) containing 50 μg / ml gentamicin was used and incubated at 26 ° C. for 48 hours.

Experimental Example 1. Determination of HBV DNA replication inhibitory ability

In order to examine the hepatitis B virus (HBV) proliferation inhibitory effect of the hepatitis B extract of the present invention, the degree of DNA replication enzyme activity of the hepatitis B virus was measured.

When expressing the hepatitis B virus transcriptase of the reference example, 10% complex herbal extract (supernatant) containing the fungus of Example 2 from the beginning of culture was added to the Sf9 cell culture medium at a final concentration of 500 µg / ml, respectively. After hours the transcriptase activity was measured.

1st group: Control group which added only saline solution.

Group 2: Add 10% apple extract (final concentration 500 µg / ml)

Group 3: Add 10% bitter extract (final concentration 500㎍ / ㎖)

4th group: 10% of the extract of Algae extract (final concentration 500µg / ml)

Group 5: 10% kelp extract added group (final concentration 500 μg / ml)

Group 6: 10% 칡 extract addition group (final concentration 500 µg / ml)

The seventh group: 10% royal broth extract added group (final concentration 500 ㎍ / ㎖)

Group 8: alpha-interferon addition group (10 ⁶ IU, Roche)

HBV transcriptase protein activity expressed in Sf9 cell line infected with FPL-pol recombinant baculovirus was measured by the priming reaction method (Lanford et al., J. Virology , 69 , 4331, 1995). The expressed transcriptase protein was purified by immunoaffinity chromatography by binding to M2 Bead (Sigma) to which an anti-FLAG monoclonal antibody was bound, and then pre-prepared with HBV transcriptase. HBV transcriptase-primer complexes were allowed to form on genomic RNA. The activity of HBV transcriptase was measured to the extent that ³² P-TTP (3,000 Ci / mmol, NEN Co., Inc.) was inserted during the primer reaction, and the amount of ³² P-TTP-labeled transcriptase protein was measured on the SDS-PAGE gel. It was confirmed. At this time, if the herbal composition inhibited the synthesis of the primer by inhibiting the synthesis of the primer, it was determined that the herbal composition had the effect of inhibiting the HBV DNA synthesis ability.

The results of Figure 1 shows that the addition of the moth extract, inhibits the virus replication enzyme compared to the control group, showing the same degree of inhibition as the alpha-interferon treatment, effective in inhibiting the proliferation of hepatitis B virus Could confirm.

Experimental Example 2. Determination of HBV DNA replication inhibitory ability of the extract of the complex herbal containing the moth

In order to examine the effect of inhibiting hepatitis B virus (HBV) proliferation which causes viral hepatitis of the herbal extract containing 3.8% of the invention, the degree of DNA replication enzyme activity of hepatitis B virus was measured.

When expressing the hepatitis B virus transcriptase of Example 3, the 10% complex herbal extract (supernatant) of Example 2 was added to 10 ml cell culture medium in the amounts of 50 µl, 100 µl and 500 µl, respectively, from the beginning of culture. After 48 hours, the transcriptase activity was measured according to the method of Experimental Example 1 above.

1st group: Control group which added only saline solution.

2nd group: 50 microliters of 10% multiple herbal extracts (final concentration 50 micrograms / ml).

3rd group: 100 microliters of 10% multiple herbal extracts (final concentration 100 micrograms / ml).

4th group: 500 microliters of 10% multiple herbal extracts (final concentration 500 microgram / ml).

Referring to the results of FIG. 2, lane 1 is a control group treated only with saline, and the activity of basic replication enzymes can be observed. On the other hand, lanes 2, 3, and 4 were treated with 10% complex herbal extracts by concentration. As the concentration was increased, it was confirmed that the transcriptase activity of hepatitis B virus was suppressed and the transcriptase activity was significantly decreased when the concentration of lane 4 was treated at 500 μg / ml.

Therefore, it was confirmed that the composite herbal composition containing the fungus of the present invention can be useful for screening the activity of HBV virus transcriptase protein expressed in insect cell lines according to the difference in concentration.

Experimental Example 3. Determination of anti-viral activity of a complex herbal containing crust against hepatitis B virus expressed in HepG2.2.15 cell line.

HepG2.2.15 cell line (Georgetown University, Dr. Korba, USA) is a cell line in which HBV is propagated by transforming the liver cancer cell line HepG2 with HBV DNA. Observed.

Efficacy inhibitory concentration (EC ₅₀ ) for the production of intracellular HBV DNA replication intermediates of the compound containing the fungus and the inhibitory concentration (EC ₉₀ ) against HBV virion DNA secreted into the cell line culture were determined. The inhibition of HBV production by the combined herbal powders and the bath solution was measured in comparison with lamivudine (3TC, Glaxo Wellcome) used as a positive control.

Cell lines were incubated in each of the T-96 well plates, and the compound containing the fungus was added thereto at a predetermined ratio, and the lamivudine solution was diluted and added as a control. HBV virus particles and HBV DNA produced by each cell line were measured during 7 days of incubation, and cell death was measured in dilutions showing cell characteristics.

As shown in the results of Table 1, the concentrations showing the same level of activity of EC ₅₀ and EC ₉₀ were observed at about one-third of the concentrations of the compound herbal powder compared to the bath solution. EC ₅₀ and EC ₉₀ of the combined herbal powder was found to be 2.8 times higher than the activity of the bath liquid of the combined herbal medicine. In addition, there was almost no cytotoxicity by this combination drug, and the selectivity index (SI) was calculated using the highest concentrations of the tested compound drugs and drugs (SI = CC ₅₀ / EC ₉₀ ).

EC ₅₀ EC ₉₀ CC ₅₀ S.I. Complex Herb Powder 90 µg / ml 254 µg / ml > 3700 µg / ml 14.6 Complex Herbal Thermal Solution 259 µg / ml 697 µg / ml > 1000 µg / ml 1.4 Lamivudine (3TC) 0.06 μM 0.166 μM 2165 μM 13042

Experimental Example 4. Determination of the combined herbal effect containing crust in WHBV virus proliferating cell line

Containing bark in primary hepatocytes (Dr. Jacobs, Cornell University, Cornell University, USA) selected from Woodchuck laboratory animals with hepatitis caused by infection with the Woodchuck HBV virus (WHBV), which is genetically similar to human HBV virus. Anti-viral activity by one combination drug powder was measured by comparing RNA amounts. Alpha-interferon is a drug currently used to inhibit the hepatitis virus proliferation, and used as a positive control group to see the efficacy of the combined herbal medicine.

Were cultured for 2 weeks the stem cells treated with interferon (Roche Co.) week-second embodiment of the composite herbal powder containing alpha sseumbagwi of liver state, and control the processing unit 10 ⁶ IU to the culture to a concentration of 0.15%. After 4 days of incubation, virus-related DNA and RNA products were extracted from the cell line, and after 8 days, the same amount of DNA and RNA extracts were obtained and Southern blot hive with hepatitis B virus surface antigen (HBsAg) DNA probe. Southern blot hybridization was performed to measure DNA and RNA levels through phosphoimage analysis.

Samples were used based on the amount of 18S ribosomal RNA of the hepatocyte cell line at the time of hybridization, and the analysis result was expressed as a percentage of 18S RNA.

In the results of Figure 3a, WHBV RNA was reduced by about 20% in the cell line treated with multi-dose powder containing the fungus four days, indicating a reduction in the infectious virus particles in the subsequent replication cycle. In the cell line treated with the compound powder for 8 days, the WHBV pregenonomic RNA was reduced by 25% compared to the untreated group and the amount of RNA corresponding to the WHBV surface antigen was also reduced (see FIG. 3A).

Compared with the amount of intranuclear WHBV DNA, out-secreted virion DNA, and intermediate of intermediate DNA produced in the culture of the cell line without the multi-drug powder, the multi-drug-containing multi-drug powder and alpha-interferon were treated. The amount of DNA was significantly decreased in the cell line, especially in the cell line treated with the compound powder, the viral DNA in the nucleus almost disappeared, and the amount of the intermediate DNA and virion DNA was also greatly reduced (see FIG. 3b). Through this result, it was confirmed that the compound herbal powder containing bitter bark also has the effect of inhibiting the replication of Woodchuck hepatitis B virus.

Clinical Example 1. Verification of therapeutic effect by the administration of multi-drug containing herbal supplements to patients (volunteers) with liver disease of hepatitis and cirrhosis.

Changes in biochemical and molecular biological values of humans related to viral hepatitis and cirrhosis (tested for the presence of HBV DNA in the body) were examined.

The compound herbal containing the moth of Example 2 was evaluated by oral administration for 1.0 to 1.5g per adult per day for 25 days or more. These indicators include changes in GPT and GOT levels in the blood, and indicator changes such as HBs and anti-HBs antibodies and HBe and anti-HBe antigens to measure signs of liver disease and the presence of HBV DNA in serum. Was measured by the PCR method. GOT and GPT refer to the levels of aspartic acid and alanine degrading enzymes, respectively, and when liver problems occur and hepatocytes are destroyed, this component enters the blood and is detected and sensitively reflects hepatocellular necrosis. -40 UI.

Table 2 shows the results of clinical treatment by the combination drug administration containing the moth. Remaining HBV DNA may be a problem for recurrence in hepatitis patients. The results of Table 2 show that after the combination drug administration, HBV DNA disappeared and was not detected in 5 of 6 patients. It was confirmed that the herbal medicine can be effectively used for the treatment and prevention of hepatitis B by inhibiting HBV DNA transcriptase activity and inhibiting HBV DNA proliferation. In particular, in hepatitis B patients who received 60 days of combined herbal medicine, it was confirmed that GOT and GPT levels were significantly reduced, which could be effectively used for the treatment of hepatitis B.

Symptom Treatment period (days) Diagnosis Before after GOT GPT HBV DNA HBsAg Anti-HBsAg HBeAg Anti-HBeAg Cirrhosis and ascites 35 I'm × × × + - + - after 28 15 - - + - + Hepatitis and Cirrhosis 90 I'm 52 44 × + - × - after 62 85 + + - - + Hepatitis B 25 I'm × × + + - + - after 23 31 - + - - + Fatty liver 25 I'm 81 97 + + - - - after 37 67 - + - - + Hepatitis B 155 I'm 34 51 + + - - + after 35 53 - + - - - Hepatitis B 60 I'm 213 290 + + - + - after 32 34 - + + + +

Hereinafter, an example of the preparation of the pharmaceutical composition will be described, but it is not intended to limit the present invention but merely to explain in detail.

Formulation Example 1 Preparation of Injection

Example 1 Dry Extract ............... 100 mg

Sodium metabisulfite ......................................... 3.0 mg

Methylparaben ...... 0.8 mg

Propylparaben ...................................... 0.1 mg

Sterile Distilled Water for Injection ...

The above ingredients were mixed and adjusted to 2 ml by a conventional method, and then filled into 2 ml ampoules and sterilized to prepare an injection.

Formulation Example 2 Preparation of Tablet

Example 1 Dry Extract ............... 200 mg

Lactose 100 mg

Starch ......................................... 100 mg

Magnesium Stearate ...............

The tablets were prepared by mixing the above components and tableting according to a conventional method for producing tablets.

Formulation Example 3 Preparation of Capsule

Example 1 Dry Extract ............... 100 mg

Lactose ...... 50 mg

Starch ............................ 50 mg

Talc ........................................ 2 mg

Magnesium Stearate ...............

The capsules were prepared by mixing the above components and filling the gelatin capsules according to a conventional method for preparing capsules.

Formulation Example 4 Preparation of Liquid

Example 1 Dry Extract ............... 1000 mg

Sugar ........................... 20 g

Isomerized sugar ......................................... 20 g

Lemon scent .........................

Purified water was added to adjust the total volume to 100 ml.

The above-mentioned components were mixed according to a conventional method for preparing a liquid solution, filled into a 100 ml brown bottle, and sterilized to prepare a liquid solution.

In addition, health foods and health drinks were prepared in the following manner.

Formulation Example 5 Preparation of Health Food

Example 1 Dry Powder ......................................... 1000 mg

Vitamin Blend ......................... 20 g

Vitamin A Acetate ............... 70 μg

Vitamin E ......................... 1 mg

Vitamin B ₁ ................................. 0.13 mg

Vitamin B ₂ ................................. 0.15 mg

Vitamin B ₆ ....................... 0.5 mg

Vitamin B ₁₂ ................................. 0.2 μg

Vitamin C ......................................... 10 mg

Biotin .......................... 10 μg

Nicotinic Acid Amide ...................................... 1.7 mg

Folic acid ......................................... 50 μg

Calcium Pantothenate ......................................... 0.5 mg

Mineral mixture ...............

Ferrous Sulfate ................................. 1.75 mg

Zinc Oxide ........................ 0.82mg

Magnesium Carbonate ......................................... 25.3 mg

Potassium monophosphate ......................................... 15 mg

Dicalcium Phosphate Dioxide ..................... 55 mg

Potassium citrate ..... 90 mg

Calcium Carbonate ... 100 mg

Magnesium Chloride .............. 24.8 mg

Although the composition ratio of the vitamin and mineral mixture is a composition suitable for a relatively healthy food in a preferred embodiment, the composition ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional health food manufacturing method, Granules may be prepared and used to prepare health food compositions according to conventional methods.

Formulation Example 6 Preparation of Health Beverage

Example 1 Dry Powder ......................................... 1000 mg

Citric Acid ..................... 100 mg

Oligosaccharide ......................................... 100 g

Plum concentrate ..................... 2 g

Taurine ..................................... 1 g

Purified water is added.

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 suitable for a 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 use purpose.

As described above, the moth extract according to the present invention is excellent in inhibiting the replication enzyme activity of hepatitis B virus and consequently inhibiting the growth of the virus, thereby preventing and treating viral liver diseases such as hepatitis and cirrhosis. It can be usefully used.

Claims (8)

Crude extract of Ixeris dentata , effective for inhibiting hepatitis B virus proliferation. A pharmaceutical composition for the prevention and treatment of viral liver disease containing crude extracts of moth. The composition of claim 1 or 2, wherein the crude extract is extracted with at least one polar solvent selected from water, lower alcohols such as methanol, ethanol, mixed solvents and buffers thereof. The composition of claim 3 wherein the polar solvent is a buffer. The composition of claim 1 or 2, wherein the dosage is administered in an amount of 10 to 100 mg per kg body weight of the adult. 3. A composition according to claim 1 or 2 comprising 0.5 to 50% of said extract relative to the total weight of the composition. A dietary supplement comprising crude extracts from the bark and food-acceptable food supplements that have prophylactic and therapeutic effects on viral liver disease. 8. The dietary supplement of claim 7, in the form of a powder, granule, tablet, capsule or beverage.