KR102516639B1 - Composition for preventing, ameliorating or treating coronavirus infectious disease comprising herbal medicine extract as effective component - Google Patents

Abstract

The present invention relates to a composition for preventing, alleviating or treating a coronavirus infection containing a Spirodela polyrhiza extract as an active component. The Spirodela polyrhiza extract has effects of protecting cells from coronavirus infection, and has an excellent effect of inhibiting the intracellular viral replication and viral intracellular infection, so that the composition containing the Spirodela polyrhiza extract of the present invention can be usefully used as a treatment for coronavirus infection, health functional food, quasi-drug, cosmetic or feed additive for anti-coronavirus.

Description

Composition for preventing, ameliorating or treating coronavirus infectious disease comprising herbal medicine extract as effective component}

The present invention relates to a composition for the prevention, improvement or treatment of coronavirus infection comprising an herbal medicine extract, particularly a bupyeong extract as an active ingredient.

The new type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) has a single-stranded positive RNA genome consisting of about 30,000 base pairs, and the virus surface is surrounded by an envelope. After it was first reported in Wuhan, China at the end of December 2019, infections worldwide exploded. The virus spreads very efficiently between people, so the World Health Organization has declared it as a pandemic that can spread worldwide and is seeking countermeasures.

Normal Covid-19 patients show relatively mild symptoms such as elevated body temperature, coughing and chest pain, but some patients develop serious illnesses such as shortness of breath and pneumonia, leading to death.

To date, approved treatments for coronavirus infections include ribavirin, famciclovir, nitazoxanide, nafamostat, and chloroquine. In addition, remdesi Vir (remdesivir, GS-5734) and favipiravir are used for the treatment of coronavirus infections, but their effectiveness is low and the incidence of mutant viruses is high due to the nature of nucleic acid analog antiviral agents, so two or more antiviral agents are applied together. . Most of the antiviral drugs used for coronavirus infection have RNA polymerase inhibitory ability, but Mouse hepatitis virus, a type of coronavirus resistant to remdesivir, was discovered, and the existing antiviral The current situation is that there is no effect of the virus agent. In addition, since antiviral agents developed to date have severe side effects, great care is required in their application.

In addition, the most common treatment method at present is symptomatic therapy to relieve symptoms. Therefore, there is an increasing need for the development of new anti-coronavirus agents with excellent infection suppression effect and low toxicity for preventing and treating the outbreak of coronavirus.

On the other hand, Korean Patent Publication No. 2016-0115372 discloses a composition for enhancing innate immunity and antiviral activity containing Bupyeong extract as an active ingredient, and Korean Patent Publication No. 2022-0044084 discloses a coronavirus infection containing an extract of Hovenia japonica. Although a composition for prevention or treatment is disclosed, there is no disclosure of a composition for preventing, improving or treating coronavirus infection comprising the herbal extract of the present invention, particularly the bupyeong extract, as an active ingredient.

The present invention was derived from the above needs, by providing an antiviral composition against coronavirus containing bupyeong extract as an active ingredient, and confirming that the bupyeong extract inhibits coronavirus infection and proliferation, The invention was completed.

In order to solve the above problems, the present invention provides an antiviral composition for coronavirus comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for preventing or treating coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

In addition, the present invention provides a health functional food composition for preventing or improving coronavirus infection comprising a bupyeong ( Spirodela polyrhiza ) extract as an active ingredient.

In addition, the present invention provides a quasi-drug composition for preventing or improving coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

In addition, the present invention provides a cosmetic composition for preventing or improving coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

In addition, the present invention provides a feed additive for preventing or improving coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

The present invention relates to a composition for preventing, ameliorating or treating a coronavirus infection comprising bupyeong extract as an active ingredient, wherein the bupyeong extract has the effect of protecting cells from coronavirus infection, replication of the virus in cells and cells of the virus It has the effect of suppressing my infection.

1 is a result of confirming the antiviral effect of bupyeong extract against SARS-CoV2 by CPE (Cytopathic effect) inhibition analysis. (A) is a photograph of a plate stained with crystal violet for the antiviral effect against SARS-CoV2 according to the Bupyeong extract treatment at each concentration, and (B) is the result of quantification thereof. NC is a normal group, and VC is a control group treated with SARS-CoV2.
Figure 2 is a fluorescence photograph (A) confirming the viral infection inhibitory effect of Bupyeong extract on SARS-CoV2 and a result (B) of quantification thereof. NC is a normal group, and VC is a control group treated with SARS-CoV2. **** means that the viral infection of the SARS-CoV2 and Bupyeong extract treatment groups was statistically significantly reduced compared to the control group (VC) treated with SARS-CoV2, p<0.0001.
3 is a result of confirming the viral replication inhibitory effect of Bupyeong extract on SARS-CoV2. NC is a normal group, and VC is a control group treated with SARS-CoV2. *** means that the number of viral copies of the SARS-CoV2 and Bupyeong extract treatment groups decreased statistically significantly compared to the control group (VC) treated with SARS-CoV2, p<0.001.

In order to achieve the object of the present invention, the present invention bupyeong ( Spirodela polyrhiza ) Provides an antiviral composition for coronavirus containing an extract as an active ingredient.

The coronavirus may be one or more selected from the group consisting of type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2), severe acute respiratory syndrome coronavirus (SARS CoV) and Middle East respiratory syndrome (MERS) coronavirus, It is preferably type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2), but is not limited thereto.

The extraction solvent of the bupyeong extract is preferably water, C ₁ ~ C ₄ lower alcohol or a mixture thereof, more preferably ethanol, but is not limited thereto.

The bupyeong is preferably using an outpost of duckweed, but is not limited thereto.

The bupyeong extract of the present invention has the effect of inhibiting the proliferation of type 2 severe acute respiratory syndrome coronavirus.

In addition, the present invention provides a pharmaceutical composition for preventing or treating coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

The pharmaceutical composition according to the present invention is formulated according to conventional methods into oral formulations such as capsules, powders, granules, tablets, suspensions, emulsions, syrups, aerosols, external preparations, suppositories and sterile injection solutions. can

The pharmaceutical composition according to the present invention may further include a pharmaceutically acceptable carrier, excipient or diluent.

Carriers, excipients and diluents that may be included in the pharmaceutical composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate , cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and various compounds or mixtures including mineral oil and the like.

When formulated, it is prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc. These solid preparations include at least one excipient in the pharmaceutical composition, for example, starch, calcium carbonate, sucrose or lactose, gelatin, etc. is prepared by mixing In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid formulations for oral use include suspensions, solutions for oral use, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, aromatics, and preservatives may be included. . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried formulations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents. As a base for the suppository, Witepsol, Macrogol, Tween 61, cacao butter, laurin paper, glycerogelatin, and the like may be used.

A suitable dosage of the pharmaceutical composition of the present invention will be prescribed in various ways depending on factors such as formulation method, administration method, patient's age, weight, sex, morbid condition, food, administration time, administration route, excretion rate and reaction sensitivity. can

The pharmaceutical composition of the present invention can be administered orally or parenterally, and in the case of parenteral administration, it can be administered by topical application to the skin, intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, and the like.

In addition, the present invention provides a health functional food composition for preventing or improving coronavirus infection comprising a bupyeong ( Spirodela polyrhiza ) extract as an active ingredient.

The composition may be prepared in any one formulation selected from powder, granule, pill, tablet, capsule, candy, syrup and beverage, but is not limited thereto.

When using the health functional food composition of the present invention as a food additive, the health functional food composition may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to conventional methods. The amount of the active ingredient can be appropriately used depending on the purpose of its use (prevention or improvement). In general, when preparing food or beverage, the health functional food composition of the present invention is added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less, based on the total amount of raw materials. However, in the case of long-term intake for health purposes, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.

There is no particular limitation on the type of health functional food. Examples of foods to which the health functional food composition can be added include meat, sausages, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea There are drinks, alcoholic beverages, and vitamin complexes, and includes all health foods in a conventional sense.

In addition, the health functional food composition of the present invention can be made into food, particularly functional food. The functional food of the present invention includes components commonly added during food preparation, and includes, for example, proteins, carbohydrates, fats, nutrients, and seasonings. For example, when prepared as a drink, natural carbohydrates or flavoring agents may be included as additional ingredients in addition to active ingredients. The natural carbohydrate is a monosaccharide (eg, glucose, fructose, etc.), a disaccharide (eg, maltose, sucrose, etc.), an oligosaccharide, a polysaccharide (eg, dextrin, cyclodextrin, etc.) or a sugar alcohol (eg, For example, xylitol, sorbitol, erythritol, etc.) is preferable. As the flavoring agent, natural flavoring agents (eg, thaumatin, stevia extract, etc.) and synthetic flavoring agents (eg, saccharin, aspartame, etc.) may be used.

In addition to the health functional food composition, various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonic acid It may further contain a carbonation agent used in beverages and the like. The ratio of the components to be added is not very important, but is generally selected in the range of 0.01 to 0.1 parts by weight based on 100 parts by weight of the health functional food composition of the present invention.

In addition, the present invention provides a quasi-drug composition for preventing or improving coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

The quasi-drugs may include external skin preparations and personal hygiene products. For example, it may be a disinfectant cleanser, shower foam, gargreen, wet tissue, detergent soap, hand wash, or ointment, but is not limited thereto.

When the quasi-drug composition according to the present invention is used as a quasi-drug additive, the composition may be added as it is or used together with other quasi-drugs or quasi-drug ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use.

The quasi-drug composition of the present invention may be prepared in the form of, for example, a general emulsified formulation and a solubilized formulation. For example, it may have formulations such as emulsions, creams, ointments, sprays, oil gels, gels, oils, and aerosols such as lotions, but it may be used without limitation as long as it exhibits the effect of preventing or improving coronavirus infection of the present invention. there is.

In addition, the quasi-drug composition may appropriately contain oil, water, a surfactant, a moisturizer, a lower alcohol having 1 to 4 carbon atoms, a thickener, a chelating agent, a colorant, a preservative, or a flavoring agent, which are generally formulated in quasi-drug compositions, in each dosage form. Can be used in combination.

In addition, the present invention provides a cosmetic composition for preventing or improving coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

The cosmetic composition of the present invention includes components commonly used in cosmetic compositions in addition to the extract, for example, antioxidants, stabilizers, solubilizers, vitamins, pigments and flavoring agents, and carriers.

The cosmetic composition of the present invention can be prepared in any formulation conventionally prepared in the art, for example, a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing , It may be formulated as an oil, powder foundation, emulsion foundation, wax foundation and spray, but is not limited thereto. More specifically, it may be formulated into a flexible lotion (skin), nourishing lotion (milk lotion), nourishing cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, shampoo, pack, spray or powder. can

When the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. are used as carrier components. It can be.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and in particular, in the case of a spray, additional chlorofluorohydrocarbon, propane / May contain a propellant such as butane or dimethyl ether.

When the formulation of the present invention is a solution or emulsion, a solvent, solubilizing agent or emulsifying agent is used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 ,3-butyl glycol oil, fatty acid esters of glycerol, polyethylene glycol or sorbitan.

When the formulation of the present invention is a suspension, as a carrier component, a liquid diluent such as water, ethanol or propylene glycol, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystals Star cellulose, aluminum metahydroxide, bentonite, agar or tragacantha and the like may be used.

When the formulation of the present invention is surfactant-containing cleansing, as carrier components, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyl taurate, sarcosinate, fatty acid amide Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters and the like can be used.

In addition, the present invention provides a feed additive for preventing or improving coronavirus infection comprising an extract of Bupyeong ( Spirodela polyrhiza ) as an active ingredient.

The feed additive of the present invention corresponds to supplementary feed under the Feed Management Act. In the present invention, the term 'feed' may refer to any natural or artificial diet, one meal, etc., or a component of the one meal meal, suitable for or suitable for consumption by animals. The type of feed is not particularly limited, and feeds commonly used in the art may be used. Non-limiting examples of the feed include vegetable feeds such as grains, root fruits, food processing by-products, algae, fibers, pharmaceutical by-products, oils and fats, starches, meal or grain by-products; Animal feeds such as proteins, inorganic materials, oils and fats, mineral oils, oils and fats, single cell proteins, zooplankton, or food may be mentioned. These may be used alone or in combination of two or more.

Hereinafter, the present invention will be described in more detail using preparation examples and examples. These Preparation Examples and Examples are only for explaining the present invention in more detail, and it is obvious to those skilled in the art that the scope of the present invention is not limited thereto.

Preparation Example 1. Preparation of bupyeong extract

Bupyeong was purchased as a standard product from OmniHub Co., Ltd., and Bupyeong extract was specifically, 10 parts by weight of 30% (v/v) ethanol was added to 1 part by weight of Bupyeong dry raw material, extracted under reflux for 3 hours, and then filtered through a sieve. After filtering through (extraction cloth), the passage through a 53 μm filter was concentrated under reduced pressure and then lyophilized to prepare.

Example 1. Evaluation of antiviral activity: CPE inhibition assay system construction

After culturing Vero E6 cells in a 96-well plate, SARS-CoV2 was inoculated with m.o.i 0.01. After inoculation, the bupyeong extract of Preparation Example 1 was diluted with DMEM containing 2% (v/v) FBS and 1% (v/v) penicillin-streptomycin and cultured for 3 days. After incubation, the cell culture medium was removed and treated with a mixed solution of methanol:acetone in a volume ratio of 1:1, followed by fixation at room temperature for 10 minutes. Thereafter, the solution was removed, and 0.4% (w/v) crystal violet was added to each well, followed by staining at room temperature for 10 minutes, and then the solution was removed and dried. After drying, methanol:acetic acid was mixed at a volume ratio of 1:1, and 100 μl of a 5-fold diluted solution with tertiary distilled water was added to each well, crystal violet was extracted for 2 hours, and absorbance was measured at 550 nm.

As a result, as shown in FIG. 1, the bupyeong extract of the present invention effectively inhibits the proliferation of SARS-CoV2 and has an excellent antiviral effect of protecting cells from the SARS-CoV2 virus.

Example 2. Verification of viral infection inhibitory effect of bupyeong extract

The inhibitory effect of SARS-CoV2 virus infection was confirmed by immunofluorescence analysis using Bupyeong extract. Specifically, after the Vero E6 cells were dispensed into a plate at 1×10 ⁴ cells/well, and the sample of Preparation Example 1 was treated by concentration, 2% (v/v) FBS and 1% (v/v) antibiotics were included. 1000-fold diluted SARS-CoV2 was treated with DMEM and cultured for 24 hours. After culturing, the cells were fixed with a 4% (v/v) formaldehyde solution, reacted with a 2000-fold diluted primary antibody (Nucleocapsid protein), and then reacted with a 200-fold diluted secondary antibody. After the reaction, the cells were observed under a fluorescence microscope and the fluorescence intensity was analyzed.

As a result, as shown in FIG. 2, intracellular infection of the SARS-CoV2 virus was effectively inhibited by the treatment with the bupyeong extract.

Example 3. Verification of viral replication inhibitory effect of bupyeong extract

The viral replication inhibitory effect of bupyeong extract was confirmed by qPCR. Specifically, after culturing Vero E6 cells in a 96-well plate, SARS-CoV2 was inoculated with m.o.i 0.01, treated with the sample of Preparation Example 1, and cultured for 24 hours. After culturing, viral RNA released outside the cell was extracted using AccuPrep Viral RNA Extraction kit from Bioneer. Subsequently, cDNA was synthesized using Toyobo's ReverTra Ace qPCR RT kit, PCR was performed with specific primers for the RBD of spike protein, and viral RNA copy number was measured using spike protein gene plasmid standard samples. Calculated.

As a result, as shown in FIG. 3, the bupyeong extract was excellent in inhibiting the replication of SARS-CoV2 virus.

Claims (10)

Bupyeong ( Spirodela polyrhiza ) An antiviral composition for type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) containing an ethanol extract as an active ingredient. delete delete Bupyeong ( Spirodela polyrhiza ) A pharmaceutical composition for preventing or treating type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) infection comprising an ethanol extract as an active ingredient. [Claim 5] The pharmaceutical composition for the prevention or treatment of type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) infection according to claim 4, further comprising a pharmaceutically acceptable carrier, excipient or diluent in addition to the bupyeong ethanol extract. composition. Bupyeong ( Spirodela polyrhiza ) Health functional food composition for preventing or improving type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) infection containing an ethanol extract as an active ingredient. The method of claim 6, wherein the composition is prepared in any one formulation selected from powder, granule, pill, tablet, capsule, candy, syrup and beverage, characterized in that type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2 ) Health functional food composition for preventing or improving infectious diseases. Bupyeong ( Spirodela polyrhiza ) A quasi-drug composition for preventing or improving type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) infection containing an ethanol extract as an active ingredient. Bupyeong ( Spirodela polyrhiza ) A cosmetic composition for preventing or improving type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) infection containing an ethanol extract as an active ingredient. Bupyeong ( Spirodela polyrhiza ) A feed additive for preventing or improving type 2 severe acute respiratory syndrome coronavirus (SARS-CoV2) infection containing an ethanol extract as an active ingredient.

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