KR101122112B1 - A Pharmaceutical composition comprising a polysaccharide having anti-rotavirus activity - Google Patents

Abstract

The present invention relates to a polysaccharide of Panax genus plants having an activity of inhibiting the infection of rotavirus, a method for preparing the same, and a composition containing the same. By inhibiting the activity, the polysaccharide of the present invention can be used as a pharmaceutical, dietary supplement, food additive for the prevention of rotavirus infection, the prevention and treatment of rotavirus diarrhea and enteritis.

Description

A pharmaceutical composition comprising a polysaccharide having anti-rotavirus activity which is effective in preventing and treating rotavirus infection diseases.

The present invention relates to a polysaccharide isolated from a processed Panax genus plant having a prophylactic and therapeutic effect on rotavirus infectious diseases, a preparation method thereof, and a composition for preventing and treating rotavirus infectious diseases containing the same.

Rotavirus belongs to the Reoviridae family and is known as the most important cause of acute colitis in developing and underdeveloped countries.The double capsid of the virus particles is like a cartwheel. It is derived from rota (English wheel) and named rotavirus. In 1943, Light and Hodes reported severe diarrhea in infants and isolated filterable pathogens that cause diarrhea in feces, but their morphological characteristics were revealed by electron microscopy in 1973 and Leovirus-like virus. (reovirus-like virus) was isolated from mammals and birds, and the virus was identified as rotavirus and in 1978 was classified as Genus Rotavirus of the Family Reoviridae family.

The rotavirus genome consists of eleven segments of double-stranded RNA, and the rotavirus particles are spherical with a diameter of 70 nm and the outer and inner capsids of the outer capsid. It has two shells. Inner layer capsid is composed of VP6 protein and contains core part including 11 double RNA fragments and structural protein of VP1, VP2, VP3. Outer capsid is composed of VP7 and VP4. . Of the VP4 and VP7 outer layer capsids, VP4 is a virus hemagglutinin, a host cell adhesion protein that appears as a spike on the surface of the virus and accounts for 2.5% of all viruses. VP7 is a 37kDa glycoprotein that forms a smooth outer capsid shell and accounts for 30% of the total virus. VP6, the main inner capsid protein, determines the subgroup, and VP7 and VP4 in the outer capsid protein determine the serotype. To date, at least two subgroups and four serotypes are known.

Rotaviruses consist of six antigenically different groups (A-F). Rotaviruses in groups A, B, and C are present in both humans and animals, while groups D, E, and F are found only in animals. Groups classified by serum have antigens that are common or crossed with each other. Among them, group A is the most studied and well-characterized since the first reported presence of rotavirus, and occupies most of human isolates.

Rotavirus is known as a major cause of acute diarrhea and enteritis in children and young animals such as cattle, horses, pigs and monkeys. Sporadic diarrheal disease in infants and children, according to World Health Organization statistics show that 5 million children under 5 years of age die from diarrhea every year, 20% of which is 1 million. Diarrhea caused by rotavirus is said to have died. In the United States, 35-50% of infant diarrhea is caused by rotavirus, and the problem is more severe in developing countries, causing about 10 times of rotavirus-borne diarrhea in infants between 6-11 months of age. It is known to have rotavirus antibodies 24 months ago. According to a study of 12 diarrhea-causing agents in pediatric diarrhea in Seoul, Korea, the highest incidence of rotavirus was detected in 47% of diarrhea groups. That's more than double that of 20% (see Table 1).

Pathogens Found in Diarrheal Diseases in Infants and Children Enteropathogenic bacteria Confirmed Enteropathogenic Fungi (%) Korea Bangladesh Venezuela Egypt Rotavirus 46.5 46 31 34 Thermostable enterotoxin producing bacteria (ST) 19.5 17 22 18 Thermolabile enterotoxin producing bacteria (LT) 2.6 4 24 9 LT + ST <1 7 - - Enteropathogenic E. coli 6.5 - 16 9 Shigella 1.7 5 3 One Salmonella <1 <1 2 8 Campylobacter jejuni
( Campylobacter jejuni ) <1 - - One

Rotavirus is an oral route of infection in feces and causes infectious acute diarrhea in infants under 2 years of age. This infection is often called infantile gastroenteritis or acute gastroenteritis. Acute gastroenteritis caused by rotavirus is accompanied by clinical symptoms such as fever, vomiting, diarrhea and respiratory symptoms, and most cases occur in late temperate to early winter in Korea and other temperate regions. The virus is excreted largely into feces about 8 days after symptoms appear, with 10 ¹¹ virus particles per gram of fecal infection being most likely in infants aged 6 months to 2 years, with dehydration and metabolic acidosis at the time of infection. Occurs. It is known that diarrhea is caused by rotavirus infection, which impairs the absorption of intestinal mucosal cells. Viruses infect the small intestinal chollocytes and proliferate in the cytoplasm of these cells, causing disruption in transport mechanisms. Damaged cells fall into the intestinal tract and release a large amount of virus so that the virus is found in feces. Diarrhea by rotavirus is due to impaired uptake of Na and glucose caused by the replacement of damaged cells with immature cells that are incapable of absorbing.

Most diarrhea patients recover when they receive proper treatment such as water supply, but in developing countries, even these treatments are not common. The incidence of bacterial diarrhea decreases due to clean drinking water supply and sanitary treatment. Does not have a great effect this way. Therefore, it is considered urgent to develop a substance capable of suppressing rotavirus infection. Rotavirus vaccines are currently being developed, but studies have shown that a vaccine that prevents rotorviruses increases the risk of enterotropia causing intestinal blockage, and safety of rotavirus vaccines and therapeutics is required. It's happening.

Panax genus is a member of the Orga family, with about 10 species known worldwide. Panax plants known to date include Panax ginseng, Panax quinquefolia, Panax notoginseng, Panax japonica, Panax trifolia, Himalaya ginseng, Panax pseudoginseng, Panax vietnamensis, etc. (Understanding Korean Ginseng, 9, Korean Ginseng Society, 1995; Advances in Ginseng Research, pp. 127-137, Korean Ginseng Society, 1998). Other Panax plants include Panax elegatior, Panax wangianus, or Panax bipinratifidus.

These Panax plants have many differences in practical use, but their components are very similar. In other words, the Panax genus plants commonly contain saponins of the dharma skeleton, which are rarely found in other plant systems. In addition, the root contains a lot of polysaccharides, especially acidic polysaccharides.

Republic of Korea Patent Registration No. 144130 discloses ginseng protein polysaccharide (Jinsan) that has an immune enhancing effect isolated from ginseng that can be used as cancer prevention or radiation protection agent, and Republic of Korea Patent Registration No. 361187 promotes hematopoietic separation from ginseng A ginseng polysaccharide, a method for preparing the same, and a composition containing the same have excellent activity, bone marrow defense, cancer cell killing immune cell generation and radiation-sensitive action. In addition, Korean Patent Publication No. 2001-88981 relates to a cosmetic composition containing ginseng polysaccharide isolated from ginseng as an active ingredient, and has excellent wrinkle improvement, collagen synthesis promotion effect, damaged skin recovery effect by light contact, or moisturizing effect. Although it has been disclosed, there is no report on the effects of rotavirus treatment on polysaccharides obtained from ginseng as well as other Panax plants.

Republic of Korea Patent No. 355490 describes the rotavirus infection inhibitory activity of the ethyl acetate fraction obtained by sequentially extracting the crust (Citrus aurantium fruit) as an ether and ethyl acetate.

However, there is no mention or suggestion in this document about the rotavirus therapeutic effects associated with ginseng.

Therefore, the present inventors measured the inhibitory activity of rotavirus infection causing infectious acute diarrhea and enteritis in polysaccharides isolated from the Panax genus plants, and confirmed the effects, and in particular, after processing the polysaccharides of the Panax genus plants as they are, The present invention has been completed by finding that extraction is more effective and the yield of polysaccharide is high.

[1] Understanding Korean Ginseng, 9, Korean Ginseng Society, 1995; Advances in Ginseng Research, pp. 127-137, The Korean Ginseng Society, 1998

[Document 2] Korean Patent Registration No. 144130

[Document 3] Korean Patent Registration No. 361187

[Patent 4] Republic of Korea Patent Publication No. 2001-88981

[Reference 5] Republic of Korea Patent No. 355490

The present invention provides a polysaccharide isolated from Panax genus plants having excellent infection inhibitory activity against rotavirus and a method for producing the same.

Accordingly, the present invention provides a polysaccharide isolated from a Panax genus plant excellent in inhibitory activity against rotavirus and a method for producing the same.

The present invention also provides a composition for the prophylaxis and treatment of rotavirus infectious diseases containing polysaccharides isolated from the genus Panax and the use of the polysaccharides isolated from the genus Panax.

The polysaccharide isolated from ginseng having an inhibitory activity against rotavirus according to the present invention can be used as a pharmaceutical and dietary supplement, or as a food additive for preventing rotavirus infection, preventing and treating rotavirus acute diarrhea and enteritis. Can be.

Figure 1 shows the elution pattern of the polysaccharide separated from the processed Panax genus plants in Sephadex G-75,-■-is 60 ℃-▲-is 100 ℃---of the ginseng heat-treated at 120 ℃ Extract.

In order to achieve the above object, the present invention provides a polysaccharide isolated from raw or processed Panax plants.

The Panax genus plants include Panax ginseng, Panax quinquefolia, Panax notoginseng, Panax notoginseng, Panax japonica, Panax trifolia, Panax pseudoginseng, and Vietnamese ginseng. Roots, stems, or leaves, such as the roots, stems, or leaves of Panax vietnamensis, Panax elegatior, Panax wangianus, or Panax bipinratifidus. The plant of the genus Panax may be prepared by heat treatment in the presence of water for 0.5 to 48 hours at a high temperature of 80 to 180 ℃.

In addition, the polysaccharide has a molecular weight of 3000 to 100000 Daltons.

The present invention provides a method for separating and preparing a polysaccharide from a Panax plant.

The present invention also provides a pharmaceutical composition for the prevention and treatment of rotavirus infection disease containing polysaccharide isolated from processed or unprocessed Panax plants.

Hereinafter, the present invention will be described in detail.

The polysaccharide separable from the genus Panax plant of the present invention can be obtained by the following method, specifically, the present invention,

(1) The raw Panax plant raw material or the root of Panax plant, which has dried the root of Panax plant, is 0.5 to 48 hours at 80-180 ° C, preferably 110-140 ° C in water or with steam. A first step of obtaining a processed Panax plant raw material by heat treatment for 1-4 hours;

(2) about 1 to 20 times the weight of the raw or processed Panax genus plants (kg), preferably about 2 to 10 times the volume (L) of water, C1 to C3 lower alcohols or mixed solvents thereof Extraction methods such as hot water extraction, cold extraction, reflux cooling extraction, or ultrasonic extraction for about 0.5 hours to 2 days, preferably 1 hour to 1 day, at an extraction temperature of from 100 ° C. to 1 to 5 times, preferably 2 Through a manufacturing process consisting of a second step of extracting four times to four times; can be prepared a polysaccharide extract containing a large amount of polysaccharide from the genus Panax plants.

In addition to this, the polysaccharide may be further purified by performing the following steps, and some of the following steps may be omitted or the order may be changed as necessary.

(3) Put the polysaccharide extract of the second step into a cation or anionic ion exchange resin such as Sephadex LH-20, HP-20, Amberlite XAD-2 A third step of adsorbing the non-polar component to the resin and collecting and eluting the fraction of the eluted polysaccharide;

(4) After dissolving the polysaccharide fraction of the third step in a small amount of water and adding C1 to C3 lower alcohol or acetone solvent, the mixture was left at a low temperature of 0 to 10 ° C. for 2 hours to 10 days, and then the resulting precipitate was centrifuged, Or a fourth step of obtaining a polysaccharide precipitate through a method such as filtration;

(5) a fifth step of dissolving the polysaccharide precipitate of the fourth step in water, removing the water-insoluble material by filtration or centrifugation, and then obtaining only an aqueous fraction;

(6) a sixth step of dialysis the water-soluble polysaccharide fraction of the fifth step in flowing water to collect only the fraction remaining in the dialysis membrane;

(7) a seventh step of filtering the polysaccharide fraction of the sixth step through a membrane filter for filtering the molecular weight 2000-5000 Daltons to obtain a purified fraction from which the low molecular weight material is removed;

(8) a purified form separated from the genus Panax plant, characterized in that it comprises an eighth step of collecting the polysaccharide fraction in the range of molecular weight 3000-200000 Daltons by gel filtration of the fraction obtained in step 7. Provided are methods for preparing polysaccharides.

The polysaccharide of the raw or processed Panax genus plants of the present invention obtained through the above production method has a molecular weight in the range of 3000 to 100,000 Daltons, and exhibits inhibitory activity against the infection of Wa virus and SA11 virus, which are one of the rotaviruses. This polysaccharide can be used for the prevention and treatment of rotavirus infection disease.

The present invention also provides a pharmaceutical composition for the prevention and treatment of rotavirus infection disease containing a polysaccharide isolated from the raw or processed Panax genus plants obtained by the above-described manufacturing process.

Specifically, the rotavirus infection disease includes diarrhea caused by rotavirus, infectious acute diarrhea, acute microgitis, and acute gastroenteritis.

The composition for preventing and treating rotavirus infection disease of the present invention comprises 0.5 to 50% by weight of the polysaccharide isolated from the raw or processed Panax genus plants with respect to the total weight of the composition.

Compositions comprising polysaccharides isolated from raw or processed Panax plants of the present invention may further comprise suitable carriers, excipients and diluents according to conventional methods.

Carriers, excipients and diluents that may be included in a composition comprising a polysaccharide isolated from a processed or processed Panax plant 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 oils.

Compositions comprising polysaccharides isolated from raw or processed Panax plants according to the present invention, powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc. It can be formulated in the form of suppositories and sterile injectable solutions.

The amount of the polysaccharide isolated from the processed or processed Panax genus plants of the present invention may vary depending on the age, sex, and weight of the patient, but may be administered once to several times in an amount of 0.1 to 100 mg / kg. In addition, the dosage of the polysaccharide isolated from the raw or processed Panax genus plants can be increased or decreased depending on the route of administration, the severity of the disease, sex, weight, age and the like. Thus, the dosage amounts are not intended to limit the scope of the invention in any manner.

The composition comprising the polysaccharide isolated from the raw or processed Panax genus plants of the present invention can be used in various forms such as pharmaceuticals, foods and beverages for the prevention and treatment of diseases caused by rotavirus in the formulation as described above. Examples of the food to which the polysaccharide separated from the raw or processed Panax plant may be added include, for example, various foods, beverages, gums, teas, vitamin complexes, health functional foods, powdered milk, baby food such as baby food, intestines, and the like. Food or beverages for adults with the disease.

Since the polysaccharide itself isolated from the processed or processed Panax genus plants of the present invention has little toxicity and side effects, it can be used safely even when taken for long periods of time, and can be added to foods or beverages for the purpose of preventing diseases caused by rotavirus. Can be.

The present invention provides a health functional food comprising a polysaccharide isolated from the raw or processed Panax genus plants exhibiting rotavirus infection suppression effect and a food supplement acceptable food supplement.

The health functional food of the present invention is preferably in the form of a pharmaceutical or health beverage such as powders, granules, tablets, capsules, pills, suspensions, emulsions, syrups, aerosols.

The present invention also provides a food additive comprising a polysaccharide isolated from the raw or processed Panax genus plants having rotavirus infection inhibitory activity.

As a food or beverage to which the food additive may be added, food for infants such as formula, formula for growing season, formula for growing season (diet formula), food for adults with diseases caused by enteric diseases, in particular rotavirus infection or Drinks, concrete examples thereof include milk, yoghurt, yoplait, pudding, soy milk, raw food, or baby food.

At this time, in the case of the food additive containing a polysaccharide isolated from the raw or processed Panax genus plants of the present invention, it can be used to add 0.01 to 10% by weight based on the total weight of the food, the raw in functional foods or beverages Alternatively, the amount of polysaccharide isolated from the processed Panax plant may be added at 0.01 to 30% by weight of the total food weight, and the health beverage composition may be added at a ratio of 0.02 to 5 g, preferably 0.3 to 1 g based on 100 ml. have.

The health functional beverage composition of the present invention has no particular limitation on the other ingredients other than the above-mentioned extract as an essential ingredient in the indicated ratio, and may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary 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 ratio of said natural carbohydrate 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 in the range of 0.1 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

The present invention is described in more detail based on the following examples, although the present invention is not limited thereto.

Example  1. Raw Panax  Preparation of Plant Polysaccharides

5 g of methanol was added to 0.5 g each of fine ginseng, hwagi ginseng, and chile, respectively, and extracted by leaving it at room temperature for 1 day. This operation was repeated twice. The supernatant was removed from the extract, and 10 times of water (5 liters) was added to the remaining residue. The resulting filtrate was extracted by filtration for 4 hours at 80 ° C. (molecular weight 5000). It was dialyzed with distilled water for 7 days, and the dialysis solution was precipitated with 50% ethanol, the precipitate was dissolved in water, and the supernatant obtained by centrifugation was used as a crude Panax plant polysaccharide.

Example  2. Machining Panax  Preparation of plant polysaccharides 1

To 0.5 kg of fine ginseng, 5 liters of methanol was added and left at room temperature for 1 day for extraction. This operation was repeated twice. The supernatant was removed from the extract and the remaining residue was heat-treated at 60, 100 and 120 ° C. for 2 hours, and 10 times of water (5 liters) was added thereto. The filtrate obtained by extracting and filtering for a period of time was put in a dialysis membrane (molecular weight 5000), and dialyzed against flowing distilled water for 7 days.The precipitated solution was precipitated with 50% ethanol, and the precipitate was dissolved in water. It was used as a ginseng polysaccharide.

The elution pattern of the ginseng polysaccharide solution obtained by treatment at temperatures of 60 ° C., 100 ° C. and 120 ° C., respectively, was identified using a Sephadex column (Sephadex G-75, 1.6 × 50 cm, Pharmacia). The column was eluted with distilled water and the flow rate was 10 mL / h. The eluted fractions were fractionated by 0.5 ml. The BSA marker and ferritin were measured at 280 nm and the polysaccharides eluted from the column were measured by carbazole reaction and then absorbance at 540 nm.

Figure 1 shows the elution pattern of ginseng polysaccharide in Sephadex G-75 column-■-is the extract of ginseng heat-treated at 60 ℃-▲-is 100 ℃-●-is 120 ℃. As shown in FIG. 1, the heat treatment at 120 ° C. showed a narrower molecular weight range and better yield than the other two cases.

Example  3. Processing Panax  Production of polysaccharides of plants 2

0.5 kg each of fine ginseng, Hwagi ginseng, and chile were put into an autoclave and heated at 130 ° C. for 2 hours using steam, and then extracted with 100 ° C. for 3 hours. The extract was filtered, injected into an HP-20 or Amberlite XAD-2 column, and eluted with water. The fraction eluted with water was concentrated under reduced pressure to obtain about 130 g of each of the polysaccharide fractions.

Example  4. Processing Panax  Production of plant polysaccharides 3

0.5 kg of fine ginseng was put into an autoclave and heated at 130 ° C. for 2 hours using steam, and then extracted with 5 liters of water at 100 ° C. for 3 hours. Water was removed under reduced pressure and 80% ethanol was added to the remaining residue to dissolve the residue as much as possible, and left overnight at 4 ° C, followed by centrifugation to obtain 120 g of the precipitated polysaccharide fraction.

Example  5. Processing Panax  Production of plant polysaccharides 4

0.5 kg of fine ginseng is added to an autoclave and heated at 130 ° C. for 2 hours using steam. Then, 3 liters of 80% methanol is added, refluxed for 3 hours, cooled, left for 8 hours, and the supernatant is removed. 3 liters of water was added to the mixture, and the extract was extracted three times for 3 hours. The extract was concentrated under reduced pressure to obtain 160 g of a polysaccharide extract.

Example  6. Processing Panax  Purification of Plant Polysaccharides

The polysaccharide extract prepared in Example 4 was purified by performing the following procedure.

The polysaccharide extract was placed in Sephadex LH-20 ion exchange resin and flowed into the water to adsorb the nonpolar component to the resin, and the eluted polysaccharide fraction was collected and concentrated.

The concentrated polysaccharide fraction was dissolved in a small amount of water, left at 4 ° C. for 4 days with 100% ethanol, and the resulting precipitate was collected by filtration. After the filtered polysaccharide precipitate was dissolved in water, the insoluble material was removed by filtration or centrifugation, and only the soluble portion was collected.

The collected polysaccharide fractions were dialyzed in running water to collect the remaining fractions in the dialysis membrane. The remaining fractions were collected by filtration through a membrane filter filtering a molecular weight of 2000-5000 Daltons, and then the remaining fractions were collected. A fraction of 200000 daltons was obtained.

Reference Example  One. MA Culture of -104 Cells

MA-104 cells (Toyama Prefectural Hygiene Testing Institute) contained 10% FBS (fetal bovine serum, Biofluids Inc., USA), 1% antibiotics-antimycotics (Gibco BRL, USA) and 3.5 g / liter sodium bi Culture was performed using DMEM medium (Dulbecco's modified eagle's medium, Sigma, USA) to which carbonate (sodium bicarbonate, Wako Pure Chem. Co., Japan) was added. When the cells proliferated about 80% of the flask width, the cells were detached and passaged after treatment with 0.25% trypsin, and the culture of the cells was performed using a CO ₂ incubator saturated with 5% CO ₂ gas. Used.

Reference Example  2. Rotavirus stocks ( stock solution Manufacturing

Dispense 2 × 10 ⁶ MA-104 cells into a tissue culture flask (25 cm 2), incubate in a CO ₂ incubator at 37 ° C. for 1 hour, attach, remove the medium and wash without FBS The cell surface was washed twice with DMEM medium. In addition, 20 μl of infectious DMEM medium containing trypsin 100 μg / ml was added to 400 μl of Wa virus solution (ATCC, USA) and SA11 virus solution (National Institute of Health, Korea), which is a rotavirus, and activated at 37 ° C. for 30 minutes. activation.

The virus solution was spread well on the cell surface, infected at 37 ° C. for 1 hour, then removed and 5 ml of infectious DMEM medium containing 5 μg / ml trypsin without FBS was added. Incubated for 5 days in ₂ incubators.

After incubation, the cytopathic effect (CPE) was confirmed, and the process of freezing and dissolving was repeated three times to completely break the cell membrane and centrifuged at 4 ° C and 5000rpm for 20 minutes to obtain the supernatant of the virus, and then frozen and stored at -70 ° C. Was used.

Experimental Example  1. Measurement of Rotavirus Inhibitory Effect of Polysaccharides

MA-104 cells were treated with 0.25% trypsin and removed from the flask, washed twice with DMEM medium for washing, and then the cell number was diluted to 3 × 10 ⁵ cells / ml.

Wa virus stock or SA11 virus stock was used at 1000- or 10000-fold dilutions in DMEM medium for infection, respectively.

The concentration of the samples in Example 1 was adjusted to 0.4 mg / ml, so that the final concentration in the treatment was 0.1 mg / ml.

50 μl of 0.4 mg / ml sample extract, 50 μl of the above-mentioned Wa virus dilution or SA11 virus dilution and 100 μl of MA-104 cells of 3 × 10 ⁵ cells / ml were plated and incubated in a 37 ° C. CO ₂ incubator. .

After 3-5 days of virus infection in the cells, the cytopathic effect (CPE) was observed to determine the inhibitory activity against rotavirus infection, and Table 2 shows the IC ₅₀ for the virus of each sample.

Plaque assay is to determine whether the virus is infected by observing that the monolayer of MA104 cells is infected with the virus, resulting in the CPE (cytopathic effect). In the results of the plaque assay, there was no significant difference between the LT polysaccharides treated at 60 ° C, the MT polysaccharides treated at 100 ° C and the HT polysaccharides treated at 120 ° C.

Inhibitory Effect of Ginseng Polysaccharide on Rotavirus Infection sample IC ₅₀ (μg / ml) Wa virus SA11 virus LT polysaccharide 10.4 12.5 MT polysaccharide 8.3 9.3 HT polysaccharide 12.6 8.4

 From the above results, it was confirmed that the ginseng polysaccharide according to the present invention has inhibitory activity against rotavirus. Therefore, the ginseng polysaccharide according to the present invention can be used for the purpose of preventing and treating rotavirus. In particular, considering that rotavirus diarrhea occurs during weaning, ginseng polysaccharides can be added to baby food to prevent rotavirus diarrhea, which accounts for 50% of weaning diarrhea.

The ginseng polysaccharide of the present invention may be administered in the following formulations, and the following formulation examples are merely illustrative of the present invention, thereby not limiting the contents of the present invention.

Formulation Example 1 Preparation of Injection

Ginseng polysaccharide of Example 1 ............ 100 mg

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

Methylparaben ............... 0.8 mg

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

Sterile distilled water for injection ..............

The above ingredients are mixed and made into 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

Ginseng polysaccharide of Example 1 ............ 200 mg

Lactose 100 mg

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

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

The above components are mixed and tableted according to a conventional method for producing tablets to produce tablets.

Formulation example  3. Preparation of Capsule

Ginseng polysaccharide of Example 2 ............................ 100 mg

Lactose 50 mg

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

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

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

Capsules are prepared by mixing the above ingredients and filling into gelatin capsules according to a conventional method for preparing capsules.

Formulation Example 4 Preparation of Liquid

Ginseng polysaccharide of Example 2 ... 1000 mg

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

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

Lemon Flavor ......................

Purified water was added to make a total of 1000 ml.

According to the conventional method for preparing a liquid, the above components are mixed, and then filled into a brown bottle and sterilized to prepare a liquid.

Formulation Example 5 Preparation of Healthy Food

Ginseng Polysaccharide of Example 1 ............................ 1000 mg

Vitamin Blend .....

Vitamin A Acetate ......... 70 μg

Vitamin E ..................... 1.0 mg

Vitamin B1 ..................... 0.13 mg

Vitamin B2 ..................... 0.15 mg

Vitamin B6 ......................................... 0.5 mg

Vitamin B12 .................. 0.2 μg

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

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

Nicotinamide ... 1.7 mg

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

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

Inorganic mixtures ...............

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

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

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

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

Dibasic calcium phosphate ........................ 55 mg

Potassium Citrate ..... 90 mg

Calcium Carbonate ... 100 mg

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

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

Formulation example  6. Manufacture of health drinks

Ginseng Polysaccharide of Example 1 ...................................... 1000 mg

Citric Acid .................................... 1000 mg

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

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

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

Purified water is added to the whole ........ 900 ㎖

The above components were mixed according to a conventional health drink manufacturing method, and the mixture was heated at 85 DEG C for about 1 hour with stirring, and the solution thus prepared was filtered to obtain a sterilized 2-liter container, which was sealed and sterilized, &Lt; / RTI &gt;

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

Claims (2)

delete A first step of heat-treating the roots of Panax ginseng or Panax notoginseng in water or with steam at 110 to 140 ° C. for 0.5 to 48 hours; 0.5 hours to 2 days of hot water extraction, cold soaking at an extraction temperature of 10 to 100 ° C. with water having a volume of 2 to 10 times the weight of the first heat-treated plant Panax genus, C1 to C3 lower alcohols, or a mixed solvent thereof A second step of extracting the extraction, reflux cooling extraction, or ultrasonic extraction by repeating the extraction method one to five times; The polysaccharide extract of the second step is placed in Sephadex LH-20, HP-20 or Amberlite XAD-2 ion exchange resin, and water is flowed to adsorb the nonpolar component to the resin. A third step of collecting and concentrating the eluted polysaccharide fraction; After dissolving the polysaccharide fraction of the third step in a small amount of water and adding a lower alcohol or acetone solvent of C1 to C3 and leaving it at a low temperature of 0 to 10 ℃ for 2 hours to 10 days, the resulting precipitate is centrifuged or filtered. A fourth step of obtaining a polysaccharide precipitate through; A fifth step of dissolving the polysaccharide precipitate of step 4 in water, removing the water-insoluble material by filtration or centrifugation, and then obtaining only a water-soluble polysaccharide fraction; A sixth step of dialyzing the fifth step of water-soluble polysaccharide fraction in flowing water to collect only the fraction remaining in the dialysis membrane; A seventh step of filtering the sixth polysaccharide fraction through a membrane filter for filtering a molecular weight of 2000-5000 daltons to obtain a purified fraction from which low molecular weight material is removed; The polysaccharide extract containing a large amount of the processed Panax genus plant polysaccharide obtained through the eighth step of collecting the polysaccharide fraction of the molecular weight 3000-200000 Daltons by gel filtration of the fraction obtained in step 7 as an active ingredient Pharmaceutical composition for the prevention and treatment of diarrhea caused by rotavirus.

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