A composition for preventing, improving, or treating coronavirus infections containing a complex Korean herbal extract as an active ingredient

Abstract

The present invention relates to a composition for preventing, ameliorating or treating coronavirus infection, which contains an Otsujito extract as an active ingredient. The Otsujito extract has excellent effects of protecting cells from coronavirus infection, inhibiting viral proliferation, and reducing the mortality rate due to infection in coronavirus-infected animal models. Therefore, the composition of the present invention, which contains an Otsujito extract as an active ingredient, can be usefully used as a coronavirus infection treatment agent, anti-coronavirus health functional food, quasi-medicinal product, cosmetic, or feed additive.

Description

[Technical Field]

[0001] The present invention relates to a composition for preventing, ameliorating, or treating coronavirus infections, which contains a complex extract of Korean medicinal herbs, particularly an extract of Euljatang, as an active ingredient.

[0002] This result was carried out with the support of the Korea Institute of Oriental Medicine Research and Development (R&D) Project of the Ministry of Science and ICT (Project number: 1711175247). [Background technology]

[0003] Severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) is a virus with a single-stranded positive RNA genome consisting of approximately 30,000 base pairs, and its surface is enveloped in an outer membrane. Since it was first reported in Wuhan, China, at the end of December 2019, the number of infected people has rapidly increased worldwide. The virus is highly efficient at spreading between humans, and the World Health Organization has declared it a pandemic with the potential to spread globally, and is currently exploring how to respond.

[0004] While COVID-19 patients typically experience mild symptoms such as fever, cough, and chest pain, some develop severe illness such as difficulty breathing and pneumonia, which can lead to death.

[0005] To date, approved treatments for COVID-19 include ribavirin, famciclovir, nitazoxanide, nafamostat, and chloroquine. Other drugs used to treat COVID-19 include remdesivir (GS-5734) and favipiravir. However, their efficacy is limited, and due to the nature of nucleoside analogue antivirals, the incidence of mutant viruses is high, necessitating the use of two or more antivirals. Most antivirals used to treat COVID-19 have the ability to inhibit RNA polymerase. However, mouse hepatitis virus, a type of coronavirus resistant to remdesivir, has been discovered, rendering existing antivirals ineffective. Furthermore, the antivirals developed to date have shown serious side effects, necessitating careful application.

[0006] Currently, the most common treatment is symptomatic treatment to alleviate symptoms. Therefore, there is an increasing need to develop anti-coronavirus agents with excellent infection suppression effects and low toxicity to prevent and treat coronavirus outbreaks.

[0007] Meanwhile, Korean Patent Publication No. 2022-0013625 discloses an anti-coronavirus composition characterized by containing honeysuckle extract as an active ingredient, and Korean Patent Publication No. 2022-0018953 discloses an antiviral composition containing bracken extract or a fraction thereof, but there is no disclosure whatsoever of a composition for preventing, ameliorating, or treating coronavirus infections that contains the Etsujito extract of the present invention as an active ingredient. Summary of the Invention [Problem to be solved by the invention]

[0008] The present invention has been derived in response to the above-mentioned needs, and provides an antiviral composition for coronavirus containing an extract of Otsujito as an active ingredient. The present invention has been completed by confirming that the extract protects cells from coronavirus infection, inhibits viral proliferation, and reduces the mortality rate due to infection in coronavirus-infected animal models. [Means for solving the problem]

[0009] In order to solve the above problems, the present invention provides an antiviral composition for coronavirus containing an extract of Otsujito as an active ingredient.

[0010] The present invention also provides a pharmaceutical composition for preventing or treating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0011] The present invention also provides a functional health food composition for preventing or ameliorating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0012] The present invention also provides a quasi-drug composition for preventing or ameliorating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0013] The present invention also provides a cosmetic composition for preventing or ameliorating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0014] The present invention also provides a feed additive for preventing or ameliorating coronavirus infections, which contains an Otsujito extract as an active ingredient. [Effects of the Invention]

[0015] The present invention relates to a composition for preventing, ameliorating or treating coronavirus infections, which contains an extract of Otsujito as an active ingredient, and the extract of Otsujito has the effect of protecting cells from coronavirus infection, suppressing viral proliferation, and reducing the mortality rate due to infection in coronavirus-infected animal models. [Brief explanation of the drawings]

[0016] [Figure 1] The antiviral effect of Otsujito extract against SARS-CoV2 was confirmed by CPE (cytopathic effect) inhibition assay. (A) shows the quantification of the antiviral effect against SARS-CoV2 by treatment with Otsujito extract at different concentrations, and (B) shows the quantification of the antiviral effect against SARS-CoV2 by treatment with remdesivir, the positive control group. NC is the normal group, and VC is the control group treated with SARS-CoV2. [Figure 2] Fluorescence photograph (A) and quantification results (B) confirm the effect of Otsujito extract in suppressing viral infection against SARS-CoV2. NC is the normal group, and VC is the control group treated with SARS-CoV2. ** and *** indicate a statistically significant reduction in viral infection in the Otsujito extract-treated group compared to the SARS-CoV2-treated control group (VC), where ** indicates p<0.01 and *** indicates p<0.001. [Figure 3] The graph shows the results of examining the effects of administration of Otsujito extract on body weight (A), body temperature (B), and survival rate (C) in an animal model infected with SARS-CoV2. Mock is a normal group not infected with SARS-CoV2, Control is a SARS-CoV2-infected group, and Otsujito extract (200 mg / kg) is a group infected with SARS-CoV2 and administered 200 mg / kg of the Otsujito extract of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0017] To achieve the object of the present invention, the present invention provides an antiviral composition for coronavirus containing an extract of Otsujito as an active ingredient.

[0018] The coronavirus is one or more selected from the group consisting of severe acute respiratory syndrome coronavirus 2 (SARS-CoV2), severe acute respiratory syndrome coronavirus (SARS CoV), and Middle East respiratory syndrome (MERS) coronavirus, preferably severe acute respiratory syndrome coronavirus 2 (SARS-CoV2), but is not limited thereto.

[0019] In one embodiment of the present invention, the Etsujitang is a mixture of Angelica acutiloba, Bupleurum Root, Scutellaria Root, Licorice Root, Shengma Root, and Rhubarb, preferably 3.0 to 12.0 parts by weight of Angelica acutiloba, 2.5 to 10.0 parts by weight of Bupleurum Root, 1.5 to 6.0 parts by weight of Scutellaria Root, 1.0 to 4.0 parts by weight of Licorice Root, 0.75 to 3.0 parts by weight of Shengma Root, and 0.5 to 2.0 parts by weight of Rhubarb, more preferably 5.0 to 7.0 parts by weight of Angelica acutiloba. The most preferred mixture is a mixture of 4.0 to 6.0 parts by weight of Bupleurum Root, 2.0 to 4.0 parts by weight of Scutellaria Root, 1.5 to 2.5 parts by weight of Licorice Root, 1.0 to 2.0 parts by weight of Shengma Root, and 0.8 to 1.2 parts by weight of Rhubarb, and the most preferred mixture is a mixture of 6.0 parts by weight of Dong Quai, 5.0 parts by weight of Bupleurum Root, 3.0 parts by weight of Scutellaria Root, 2.0 parts by weight of Licorice Root, 1.5 parts by weight of Shengma Root, and 1.0 part by weight of Rhubarb, but is not limited thereto.

[0020] The Otsujito extract of the present invention has the effect of suppressing the proliferation of severe acute respiratory syndrome coronavirus 2.

[0021] The present invention also provides a pharmaceutical composition for preventing or treating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0022] The pharmaceutical compositions according to the present invention can be formulated into oral dosage forms such as capsules, powders, granules, tablets, suspensions, emulsions, syrups, and aerosols, external preparations, suppositories, and sterile injection solutions by conventional methods.

[0023] The pharmaceutical composition according to the present invention may further comprise a pharmaceutically acceptable carrier, excipient or diluent.

[0024] The carriers, excipients, and diluents contained in the pharmaceutical compositions of the present invention may include a variety of compounds or mixtures thereof, including lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.

[0025] When formulated, the formulation is prepared using commonly used diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants. Solid formulations for oral administration include tablets, pills, powders, granules, capsules, etc., and are prepared by mixing the pharmaceutical composition with at least one or more excipients, such as starch, calcium carbonate, sucrose or lactose, and gelatin. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid formulations for oral administration include suspensions, oral solutions, emulsions, syrups, etc., and may contain various excipients such as wetting agents, sweeteners, flavoring agents, preservatives, and the like in addition to commonly used simple diluents such as water and liquid paraffin. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized formulations, and suppositories. Non-aqueous solvents and suspensions that can be used include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. Suppository bases that can be used include witepsol, macrogol, Tween 61, cacao butter, laurin butter, and glycerogelatin.

[0026] The appropriate dosage of the pharmaceutical composition of the present invention may be variously formulated depending on factors such as the formulation method, administration method, age, weight, sex, pathological condition, diet, administration time, administration route, excretion rate, and reaction sensitivity of the patient.

[0027] The pharmaceutical composition of the present invention can be administered orally or parenterally. 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, etc.

[0028] The present invention also provides a functional health food composition for preventing or ameliorating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0029] The composition may be prepared in any one dosage form selected from the group consisting of powder, granules, pills, tablets, capsules, candies, syrups, and beverages, but is not limited thereto.

[0030] When the functional health food composition of the present invention is used as a food additive, it can be added directly or together with other foods or food ingredients in a conventional manner. The amount of the active ingredient can be appropriately used depending on the purpose of use (prevention or improvement). Generally, when producing a food or beverage, the functional health 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 ingredients. However, when taking it for long-term health purposes, the amount is within the above range, and there is no safety issue, so the active ingredient can be used in an amount greater than the above range.

[0031] There is no particular limitation on the type of the functional health food. Examples of foods to which the functional health food composition can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea drinks, alcoholic beverages, and vitamin complexes, all of which include health foods in the usual sense.

[0032] Furthermore, the functional health food composition of the present invention can be manufactured as a food, particularly a functional food. The functional food of the present invention contains ingredients commonly added during food production, such as proteins, carbohydrates, fats, nutrients, and seasonings. For example, when manufactured as a drink, natural carbohydrates or flavorings may be added as additional ingredients in addition to the active ingredients. The natural carbohydrates are preferably monosaccharides (e.g., glucose, fructose, etc.), disaccharides (e.g., maltose, sucrose, etc.), oligosaccharides, polysaccharides (e.g., dextrin, cyclodextrin, etc.), or sugar alcohols (e.g., xylitol, sorbitol, erythritol, etc.). The flavorings can be natural flavorings (e.g., thaumatin, stevia extract, etc.) or synthetic flavorings (e.g., saccharin, aspartame, etc.).

[0033] In addition to the health functional food composition, the composition may further contain various nutrients, vitamins, electrolytes, flavoring agents, colorants, pectinic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonation agents used in carbonated drinks, etc. The proportion of such additive ingredients is not particularly important, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the health functional food composition of the present invention.

[0034] The present invention also provides a quasi-drug composition for preventing or ameliorating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0035] The quasi-drugs may include skin care products and personal hygiene products, such as, but not limited to, disinfectants, shower foams, gums, wet tissues, detergent soaps, hand washes, or ointments.

[0036] When the quasi-drug composition according to the present invention is used as a quasi-drug additive, the composition can be added as is or together with other quasi-drugs or quasi-drug ingredients, and can be used appropriately in a conventional manner. The amount of the active ingredient to be mixed can be appropriately determined depending on the purpose of use.

[0037] The quasi-pharmaceutical composition of the present invention may be prepared in the form of a general emulsion or solubilized formulation, such as a lotion, cream, ointment, spray, oil gel, gel, oil, or aerosol, but may be used without any particular limitation as long as it has the effect of preventing or ameliorating coronavirus infection of the present invention.

[0038] Furthermore, the quasi-pharmaceutical composition can be used by appropriately blending, as needed, oils, water, surfactants, moisturizers, lower alcohols having 1 to 4 carbon atoms, thickeners, chelating agents, pigments, preservatives, fragrances, and the like that are generally blended in quasi-pharmaceutical compositions in each dosage form.

[0039] The present invention also provides a cosmetic composition for preventing or ameliorating coronavirus infections, which contains an extract of Otsujito as an active ingredient.

[0040] In addition to the extract, the cosmetic composition of the present invention contains ingredients commonly used in cosmetic compositions, such as conventional adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments and fragrances, and carriers.

[0041] The cosmetic composition of the present invention may be prepared in any dosage form commonly used in the art, including, but not limited to, a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleanser, oil, powder foundation, emulsion foundation, wax foundation, spray, etc. More specifically, it may be prepared in the dosage form of a softening lotion (skin), a nourishing lotion (milk lotion), a nourishing cream, a massage cream, an essence, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a shampoo, a pack, a spray, or a powder.

[0042] When the dosage form of the present invention is a paste, cream or gel, the carrier component may be an animal oil, a vegetable oil, wax, paraffin, starch, tragacanth, a cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide.

[0043] When the dosage form of the present invention is a powder or spray, the carrier component may be lactose, talc, silica, aluminum hydroxide, calcium silicate, or polyamide powder, and, particularly when the dosage form is a spray, may further contain a propellant such as a chlorofluorohydrocarbon, propane / butane, or dimethyl ether.

[0044] When the dosage form of the present invention is a solution or emulsion, a solvent, solubilizer or emulsifier 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, glycerol aliphatic esters, polyethylene glycol or fatty acid esters of sorbitan.

[0045] When the dosage form of the present invention is a suspension, the carrier component may be a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth, or the like.

[0046] When the dosage form of the present invention is a surfactant-containing cleanser, the carrier component may be a fatty alcohol sulfate, a fatty alcohol ether sulfate, a sulfosuccinic acid monoester, isethionic acid, an imidazolinium derivative, methyl tauric acid, sarcosinic acid, a fatty acid amide ether sulfate, an alkylamidobetaine, a fatty alcohol, a fatty acid glyceride, a fatty acid diethanolamide, a vegetable oil, a lanolin derivative, or an ethoxylated glycerol fatty acid ester.

[0047] The present invention also provides a feed additive for preventing or ameliorating coronavirus infections, which contains an Otsujito extract as an active ingredient.

[0048] The feed additive of the present invention corresponds to a supplemental feed under the Feed Management Act. In the present invention, the term "feed" refers to any natural or artificial diet, meal, or component of such meal, intended for, or suitable for, animal consumption, ingestion, and digestion. The type of feed is not particularly limited, and any feed commonly used in the art can be used. Non-limiting examples of the feed include plant-based feeds such as grains, roots and fruits, food processing by-products, algae, fibers, pharmaceutical by-products, oils and fats, starches, meal, or grain by-products; and animal-based feeds such as proteins, minerals, oils and fats, minerals, single-cell proteins, zooplankton, or food and drink. They may be used alone or in combination of two or more types.

[0049] The present invention will be described in more detail below using Production Examples and Examples. It will be obvious to those skilled in the art that these Production Examples and Examples are merely for the purpose of explaining the present invention in more detail, and that the scope of the present invention is not limited thereto.

[0050] Manufacturing Example 1. Manufacturing of Otsujito extract The Otsuji-to used in the present invention was a complex herbal extract consisting of 6.0 g of Angelica acutiloba, 5.0 g of Bupleurum Root, 3.0 g of Scutellaria Baicalensis, 2.0 g of Licorice Root, 1.5 g of Shengma Root, and 1.0 g of Rhubarb, provided by Kracie Pharma Ltd., JAPAN.

[0051] Example 1. Confirmation of antiviral activity of Otsujito extract through CPE (cytopathic effect) inhibition assay Vero E6 cells were cultured in a 96-well plate and inoculated with SARS-CoV2 at an moi of 0.01. After inoculation, the cells were treated with various concentrations of the Etsujitang solution (prepared in Preparation Example 1) diluted with DMEM containing 2% (v / v) FBS and 1% (v / v) penicillin-streptomycin to match the treatment concentration, followed by incubation for 3 days. After incubation, the cell culture medium was removed, and the cells were treated with a 1:1 volumetric mixture of methanol and acetone and fixed for 10 minutes at room temperature. The solution was then removed, and 0.4% (w / v) crystal violet was added to each well. The cells were then stained at room temperature for 10 minutes, after which the solution was removed and the cells were dried. After drying, 100 μl of a 1:1 volumetric mixture of methanol and acetic acid, diluted 5-fold with triple-distilled water, was added to each well. Crystal violet was extracted for 2 hours, and its absorbance was measured at 550 nm.

[0052] As a result, as shown in Figure 1, it was confirmed that when the Otsujito extract of the present invention was treated, the proliferation of SARS-CoV2 was inhibited in a concentration-dependent manner, and therefore the Otsujito extract of the present invention has excellent antiviral effects that protect cells from the SARS-CoV2 virus and inhibit viral proliferation.

[0053] Example 2: Confirmation of the viral infection suppression effect of Otsujito extract The inhibitory effect of Otsujito extract on SARS-CoV2 virus infection was confirmed by immunofluorescence analysis. 4 Cells were dispensed into a 96-well plate at 1 / well and treated with Otsujito extract. Then, SARS-CoV2 diluted 1:1000 in DMEM containing 2% (v / v) FBS and 1% (v / v) antibiotics was added and cultured for 24 hours. After culture, the cells were fixed with 4% (v / v) formaldehyde solution and reacted with a primary antibody (N-protein) diluted 2:2000, followed by a secondary antibody diluted 2:200. After incubation, the cells were observed under a fluorescence microscope and their fluorescence intensity was analyzed.

[0054] As a result, as shown in Figure 2, treatment with Otsujito extract effectively inhibited intracellular infection of the SARS-CoV2 virus.

[0055] Example 3. Efficacy evaluation of Otsujito extract in an animal model of SARS-CoV2 infection To evaluate the inhibitory effect on SARS-CoV2 infection in a hACE2-transformed mouse model, Otsujito extract was orally administered once daily from two days before SARS-CoV2 infection until the day of death. On the day of infection (day 0), the Otsujito extract of the present invention was orally administered, and one hour later, SARS-CoV2 was inoculated intranasally at 2,000 pfu / mouse (20 μl / mouse) for infection. Body weight and temperature were measured daily until the end of the experiment.

[0056] As a result, as shown in Figure 3, the body weight of the group administered the Otsujito extract of the present invention was shown to be at a higher level than the SARS-CoV2-infected group from day 6 onwards, and while the body temperature of the SARS-CoV2-infected group dropped sharply on day 6, the group administered the Otsujito extract of the present invention maintained a nearly normal level. The survival rate of the SARS-CoV2-infected group dropped sharply on day 10, but the survival rate of the group administered the Otsujito extract of the present invention remained at approximately 60%. Therefore, it was determined that the Otsujito extract of the present invention can reduce the mortality rate of the severe SARS-CoV2-infected group.

Claims

1. An antiviral composition against severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) containing as an active ingredient an extract of Otsujito consisting of 5.0 to 7.0 parts by weight of Angelica Root, 4.0 to 6.0 parts by weight of Bupleurum Root, 2.0 to 4.0 parts by weight of Scutellaria Root, 1.5 to 2.5 parts by weight of Glycyrrhiza Root, 1.0 to 2.0 parts by weight of Shengma Root, and 0.8 to 1.2 parts by weight of Rhubarb.

2. A pharmaceutical composition for the prevention or treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection, comprising as an active ingredient an extract of Otsujito consisting of 5.0 to 7.0 parts by weight of Angelica Root, 4.0 to 6.0 parts by weight of Bupleurum Root, 2.0 to 4.0 parts by weight of Scutellaria Root, 1.5 to 2.5 parts by weight of Glycyrrhiza Root, 1.0 to 2.0 parts by weight of Shengma Root, and 0.8 to 1.2 parts by weight of Rhubarb.

3. The pharmaceutical composition for the prevention or treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection according to claim 2, characterized in that it further contains a pharmaceutically acceptable carrier, excipient or diluent in addition to the Otsujito extract.

4. A health functional food composition for preventing or ameliorating severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection, comprising as an active ingredient an extract of Otsujito consisting of 5.0 to 7.0 parts by weight of Angelica Root, 4.0 to 6.0 parts by weight of Bupleurum Root, 2.0 to 4.0 parts by weight of Scutellaria Root, 1.5 to 2.5 parts by weight of Glycyrrhiza Root, 1.0 to 2.0 parts by weight of Shengma Root, and 0.8 to 1.2 parts by weight of Rhubarb.

5. The health functional food composition for preventing or ameliorating severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection according to claim 4, characterized in that the composition is manufactured in any one dosage form selected from the group consisting of powder, granules, pills, tablets, capsules, candies, syrup, and beverages.

6. A quasi-pharmaceutical composition for preventing or ameliorating severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection, comprising as an active ingredient an extract of Otsujito consisting of 5.0 to 7.0 parts by weight of Angelica Root, 4.0 to 6.0 parts by weight of Bupleurum Root, 2.0 to 4.0 parts by weight of Scutellaria Root, 1.5 to 2.5 parts by weight of Glycyrrhiza Root, 1.0 to 2.0 parts by weight of Shengma Root, and 0.8 to 1.2 parts by weight of Daio.

7. A feed additive for preventing or ameliorating severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infection, comprising as an active ingredient an extract of Otsujito consisting of 5.0 to 7.0 parts by weight of Angelica Root, 4.0 to 6.0 parts by weight of Bupleurum Root, 2.0 to 4.0 parts by weight of Scutellaria Root, 1.5 to 2.5 parts by weight of Licorice Root, 1.0 to 2.0 parts by weight of Shengma Root, and 0.8 to 1.2 parts by weight of Rhubarb.

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