KR20110008483A - A composition containing extracts of inula japonica thunberg for preventing and treating infection with highly pathogenic avian influenza virus h5n1 - Google Patents

Abstract

PURPOSE: A composition containing Inula japonica Thunberg extract for preventing and treating avian influenza H5N1 infection is provided to ensure antiviral activity to H5N1. CONSTITUTION: A pharmaceutical composition for preventing and treating highly pathogenic avian influenza H5N1 infection contains Inula japonica Thunberg extract as an active ingredient. The pharmaceutical composition further contains a carrier, excipient, and diluent. A health food for preventing and treating H5N1 avian influenza H5N1 infection contains Inula japonica Thunberg extract as an active ingredient.

Description

A composition containing extracts of Inula japonica Thunberg for preventing and treating infection with Highly Pathogenic Avian Influenza Virus H5N1}

The present invention relates to a capillary extract, and more particularly to a composition for the prevention and treatment of high pathogenic avian influenza H5N1 infection containing a capillary extract having an antiviral activity in high pathogenic avian influenza H5N1.

Influenza viruses are RNA viruses belonging to the Orthomyxoviridae, which are classified into three types, A, B, and C, depending on the possession status of the nuclear protein (NP) and matrix (M) antigens. , Subtypes according to the state of hemagglutinin (HA) and neuraminidase antigen (NA).

The HA binds to the sialic acid site on the surface of the host cell to facilitate the penetration of the virus into the host cell, and the NA separates the binding of the virion surface to the sialic acid to proliferate viral particles. Help release from the host cell. In addition, a small amount of M2 protein in the influenza A virus acts as an ion channel to control the pH inside the virus, and plays an important role in the replication step. Amantadine and rimantadine, used as anti-influenza agents, have antiviral effects by blocking this function.

All avian influenza belongs to type A, and subtypes are classified into a combination of 16 HA and 9 NA. The host in the wild of avian influenza type A is aquatic birds, and the transmission between birds occurs directly or indirectly by water, mist, and food contaminated with feces. Most avian influenza viruses have no symptoms or are mild, but recently, highly pathogenic avian influenza viruses H5 and H7 have been reported in poultry such as chickens and turkeys. In addition, HA of avian influenza selectively binds to SA-α-2 and 3-Gal sites, whereas human influenza virus binds to SA-α-2 and 6-Gal. However, it is known that SA-α-2 and 3-Gal are present in some organs of the human body, and H5N1, a highly pathogenic avian influenza, is also recognized to recognize SA-α-2 and 6-Gal.

Influenza type A is a common infectious disease that infects humans and is highly contagious over a wide area, with many casualties historically. Tens of millions of people were killed by influenza, the so-called “Spanish flu,” which was a pandemic between 1918 and 1919. H5N1, a subtype of the deadly avian influenza A in humans, is currently in widespread worldwide.WHO reported 406 cases and 254 deaths since February 2009. . Although no cases of human infection have been reported in Korea yet, since the first report was reported in Eumseong, North Chungcheong Province in December 2003, it occurred in 18 farms in 10 cities and counties, and sold more than 5 million chickens and ducks. Occurrence, which occurred in 2005 and 2007 in Chungnam, resulted in economic and social costs for farmers due to the disposal of chicken and duck meat. As birds and humans continue to spread around Asia, pigs raised close to the license have been isolated from the highly pathogenic avian influenza H5N1 virus, which is a concern among humans and mammals.

As a preventive measure against influenza virus, vaccine research is difficult to develop due to the facility to secure a high level of safety, high virus yield, and the composition of the vaccine according to the protein variation of the virus. Recently, amantadine, rimantadine, zanamivir, oseltamivir and the like have been used as anti-influenza preparations. Amantadine and rimantadine inhibit the ion channel activity of M2 protein, and oseltamivir and zanamivir have anti-influenza activity by inhibiting neuraminidase.

Recently, however, high pathogenic influenza resistant to these anti-influenza preparations have been reported. Amantadine has side effects on the nervous system and has the disadvantage of rapidly developing resistance to amino acid changes of the M2 protein. Rimantadine has relatively low side effects, but has a narrow range of activity and has disadvantages of developing resistance. Because zanamivir is not suitable for oral administration, it is injected in the form of an inhalant, which can cause bronchial spasms. Mutation of the active region of the virus neuraminidase or erythrocyte aggregates reduces the affinity of HA to host cell receptors, eliminating the need for the neuraminidase, which is essential for release from the host. You will become resistant. In addition, the use of anti-influenza is limited due to the production period and licensing issues.

Recently, research on safe natural materials has been actively conducted, having similar effects to existing synthetic preparations from plant resources. Choi et al. Reported antiviral activity against influenza A virus of methanol extract of Zanthoxylum piperitum leaves (Choi, HJ, Song, JH, Kwon, DH, Baek, SH, Ahn, YJ (2008) activity of Zanthoxylum Species against influenza virus.Korean J. Medicinal Crop Sci . 16 , 273-278.). Song et al. (-)-Epigallocatechin gallate and (-)-epicatechin gallate} and (-)-epicatechin gallate were the green tea catechins influenza A virus H1N1 in MDCK cells. The effect of inhibiting the replication of H3N1, type B virus has been reported (Song, JM, Lee, KH, Seong, BL (2005) Antiviral effect of catechins in green tea on influenza virus. Antiviral Res. 68 , 66-74.). Vimaranthan et al also reported anti-influenza effects from Echinacea purpurea (1.Vimalanthan, S., Kang, L., Amiguet, VT, Livesey, J., Arnason, JT, Hudson, J.). (2005) Echinacea purpurea aerial parts contain multiple antiviral compounds.Pharm . Biol . 43 , 740-745.). In addition, research on plant resources with anti-influenza activity has been made in China, Korea, Japan, India, and South America. In Korea, various physiological activities such as anti-cancer, anti-oxidation, and anti-thrombosis prevention of traditionally used herbal medicines are verified, and the development of drugs and functional foods using them is being actively performed.

The present inventors searched for a material that has antiviral activity against the highly pathogenic avian influenza virus A / H5N1 in major herbal medicines traditionally used in East Asia such as Korea and China, and the methanol extract of the root of Asarum sieboldii on A / H5N1 was investigated. The patent application has been proved to have excellent antiviral activity (Korean Patent Application No. 10-2008-0111466).

Seonhwa, which shows excellent antiviral activity in the present invention, is a perennial herb of the Campanula Asteraceae, also called Geumbulcho, and is widely distributed in Korea, China, Japan, and grows in wetlands. It is a medicinal plant that has been used traditionally for medicinal origin, and it is originated from the flowers of Inula japonica Thunberg or Inula britannica Linne. The plant contains about 44% of inulin and contains sterols, fats and essential oils. The essential oils contain allantolactone and isoalanantolactone, quercetin, isoquercetin, caffeic acid, and cholorgenic acid), taraxasterol, and many other types of sterols. In oriental medicine, the ablation of the abdomen has the effect of clay, expectoration, bronchospasm, diuretic effect, sputum, gastrointestinal nerve disorders, chest swelling, and indigestion. It is widely used in the condition of daughter and daughter.

Limpning has immunity, anti-inflammatory and antimicrobial activity, and has been used for viral diseases such as hepatitis. On the other hand, various physiological activities have been reported along with the study of volatile components of sunning. Park et al. Reported anticancer activity against colorectal cancer, breast cancer and cervical cancer from Geumbulcho (1. Cha, MR, Kim, JY, Hwang, JH, Park, HR (2006) Cytotoxic activity of the Inula japonica extracts against several human cancer cell line s invitro.Kor. J. Pharmacogn. 37 , 130-135.), Qin et al., four substances with anticancer activity against tumor cell lines (A549, LOVO, CEM, MDA-MB-435). Qin, JJ, Jin, HZ, Fu, JJ, Hu, XJ, Zhu, Y., Yan, SK, Zhang, WD (2009) Japonicones AD, bioactive dimeric sesquiterpenes from Inula japonica Thunb .. Bioorg Med. Chem. Lett. 19 , 710-713. ). Yu et al. Also isolated flavonols with antidepressant effects (Yu, NJ, Zhao, YM, Zhang, YZ, Li, YF (2006) Japonicins A and B from the flowers of Inulajaponica . J. Asian Nat . Prod . Res . 8, 385-390). Jang et al. Also separated anthranilic acid derivatives from gold flames (1. Qin, JJ, Jin, HZ, Fu, JJ, Hu, XJ, Zhu, Y., Shen, YH, Yan, SK, Zhang, WD (2008) Anthranilic acid derivatives from Inula japonica.Chin. Chem. Lett. 19 , 556-558.).

Liquor is licensed to be used as a limited raw material for foods in the food industry, so it can be used as a “raw material” for the production of teas, beverages, liquor and spices, which consist of one raw material. When mixed with, it belongs to the "limited raw materials" and can be used in the raw materials up to 50%. The safety and the regulation of the above-mentioned lobulation is very useful industrially for use as a functional food ingredient for the control of highly pathogenic avian influenza H5N1.

Accordingly, it is an object of the present invention to provide a mussel extract having antiviral activity against highly pathogenic avian influenza H5N1.

Another object of the present invention is to provide a composition for the prevention and treatment of highly pathogenic avian influenza H5N1 infection containing the above-mentioned extract.

The purpose of the present invention as described above is to prepare a variety of herbal extracts, and to verify the antiviral activity against the highly pathogenic avian influenza H5N1 of the herbal extracts, among them, the extract is showing antiviral activity against the highly pathogenic avian influenza H5N1. This was achieved by confirming the fact.

The present invention provides a pharmaceutical composition for the prevention and treatment of highly pathogenic avian influenza H5N1 infection containing the extract ( Inula japonica Thunberg) as an active ingredient.

In another aspect, the present invention provides a health food composition for the prevention and improvement of high pathogenic avian influenza H5N1 infection containing the extract ( Inula japonica Thunberg) as an active ingredient.

In another aspect, the present invention provides a feed or feed additive composition for the prevention and improvement of high pathogenic avian influenza H5N1 infection containing the extract ( Inula japonica Thunberg) as an active ingredient.

In the present invention, the term "active ingredient" refers to a substance or a group of substances (a medicinal agent whose pharmacologically active ingredient is not known, etc.) which is expected to express the efficacy or effect of the medicine directly or indirectly by inherent pharmacological action. It means containing a main component).

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

Dosage forms of the pharmaceutical compositions of the present invention may be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection.

In the present invention, "health functional food" means a food manufactured and processed using raw materials or ingredients having functional properties useful for the human body according to Act No. 6767 of the Health Functional Food Act, and "functional" means It means ingestion for the purpose of obtaining useful effects on health use such as nutrient control or physiological action on structure and function.

In the present invention, after preparing the prepared methanol extract, it was added to MDCK cells inoculated with H5N1 avian influenza virus, and the virus titer was measured by erythrocyte agglutination (HA) analysis to verify antiviral activity. .

As a result, the virus titer decreased by 4 log to 2 ³ HAunit, and it was confirmed that the perennial extract exhibited antiviral activity against H5N1 avian influenza virus.

The extract according to the present invention has an excellent antiviral activity against high pathogenic avian influenza H5N1, which is a very useful invention in the pharmaceutical industry that can provide a pharmaceutical composition for the prevention and treatment of high pathogenic avian influenza H5N1 infection.

Hereinafter, preferred embodiments of the present invention will be described in more detail with reference to the following Examples and Experimental Examples. However, the scope of the present invention is not limited to these examples.

Example: Herbal Medicine Extract Preparation

27 kinds of herbal medicines used in the present invention were purchased from Gyeongdong market in Seoul (see Table 1). 30 g of each dried herbal material was immersed in 300 mL of methanol, extracted at room temperature for 24 hours, and then filtered using Whatman filter paper no. The filtrate was concentrated at 40 ° C or less using a vacuum condenser and then lyophilized and stored at 4 ° C.

TABLE 1

Herbal Medicine Used in the Examples

Botany Name Used part Poncirus trifoliata Rafinesqul Fruit Belamcanda chinensis Leman Root Chelidonium majus Linne var. asiaticum (Hara) Ohwi Aerial Smilax china linne Root Lindera strichnifolia Villars Root Inula japonica Thunberg Flower Schizomepeta tenuifolia Briquet Aerial Patrinia villosa Jussieu All parts Gentiana macrophylla Pallas Root Citrus unshiu markovich Fruit Scutellaria baicalensis Georgi Root Phellodendron amurense Ruprecht Bark Torilis japonica Decandolle Fruit Angelica tenuissima Nakai Root Saposhnikovia divaricata Schiskin Root Cibotium barometz J. Smith Root Lonicera japonica Thunberg Flower Xanthium strumarium Linne Fruit Magnolia denudata Desrousseaux Flower Clematis mandshurica Maximoxicz Root Vitex rotundifolia Linne Fruit Corydalis ternata Nakai Stem Zizyphus jajaba var. inermis Rehder Fruit Bupleurum falcatum Linne Root Taraxacum platycarpum H. Dahlstedt Aerial Chaenomeles sinensis (Thouin) Koehne Fruit Chrysanthemum indicum Linne Flower

Experimental Example: Verification of Virus Propagation and Antiviral Activity

Avian influenza virus (A / Vietnam / 1194/04 (H5N1) -NIBRG-14) used in this experiment was introduced under the permission of Korea Disease Control Center, World Health Organization (WHO) and US Disease Control Center (CDC). Was used. Virus propagation was performed at BL-3 facility using an allantoic fluid of fertilized eggs hatched for 11 days.

To verify the antiviral activity of Chinese medicinal herbs, MDCK (Madin-Darby canine kidney) cells were cultured in 96-well plates using MEM medium containing antibiotics and 10% FBS. After 24 hours, monolayer-cultured MDCK cells were inoculated with 100TCID ₅₀ H5N1 avian influenza virus and incubated again for 1 hour for virus infection. After washing the virus infected MDCK cells, medium (1: 100) containing methanol extract of each herbal medicine was added. After 48 hours, the virus titer of the supernatant was measured by hemagglutination (HA) analysis to verify antiviral activity.

The virus titer was 2 ⁷ HAunit when no sample was added, and it was found to have distinct antiviral activity when the virus titer was reduced by 4 log or more than when no sample was added. The results are shown in Table 2 below.

TABLE 2

Inhibitory Activity of Medicinal Herbal Extracts of Herbal Medicine against Influenza Virus A / H5NI

Botany Name Inhibitory activity Poncirus trifoliata Rafinesqul - Belamcanda chinensis Leman - Chelidonium majus Linne var. asiaticum (Hara) Ohwi - Smilax china linne - Lindera strichnifolia Villars - Inula japonica Thunberg + (4 log reduction) Schizomepeta tenuifolia Briquet - Patrinia villosa Jussieu - Gentiana macrophylla Pallas - Citrus unshiu markovich - Scutellaria baicalensis Georgi - Phellodendron amurense Ruprecht - Torilis japonica Decandolle - Angelica tenuissima Nakai - Saposhnikovia divaricata Schiskin - Cibotium barometz J. Smith - Lonicera japonica Thunberg - Xanthium strumarium Linne - Magnolia denudata Desrousseaux - Clematis mandshurica Maximoxicz - Vitex rotundifolia Linne - Corydalis ternata Nakai - Zizyphus jajaba var. inermis Rehder - Bupleurum falcatum Linne - Taraxacum platycarpum H. Dahlstedt - Chaenomeles sinensis (Thouin) Koehne - Chrysanthemum indicum Linne -

+: Detected,-: not detected

As shown in Table 2 above, the antiviral activity against the highly pathogenic avian influenza virus H5N1 of the methanol extract of Inula japonica Thunberg called mucin in the 27 herbal medicines used in the experiment was verified. The virus without sample was 2 ⁷ HAunit, while the methanol extract of Inula japonica Thunberg added reduced 4 log to 2 ³ HAunit.

Although anti-cancer activity and major physiologically active substances have been reported, the present invention is the first to confirm the antiviral activity of the invasion ( Inula japonica Thunberg) against the highly pathogenic avian influenza H5N1 through the present invention is excellent research results.

Claims (3)

Pharmaceutical composition for the prevention and treatment of high pathogenic avian influenza H5N1 infection containing the extract of Inula japonica Thunberg as an active ingredient. Health functional food composition for the prevention and improvement of high pathogenic avian influenza H5N1 infection containing the extract of Inula japonica Thunberg as an active ingredient. Feeding or feed additive composition for the prevention and improvement of high pathogenic avian influenza H5N1 infection containing the extract of Inula japonica Thunberg as an active ingredient.