KR102213514B1 - Natural composition for inhibiting corona virus activity - Google Patents

Abstract

The present invention relates to a composition derived from a natural product capable of inhibiting the activity of coronavirus, comprising phytoncide extracted from Chamaecyparis obtusa and a substance extracted from a natural product exhibiting antiviral activity. The composition of the present invention is derived from the natural product, thereby having no toxicity, and has high utility since the composition does not cause side effects even when applied to the human body.

Description

Natural composition for inhibiting coronavirus activity {NATURAL COMPOSITION FOR INHIBITING CORONA VIRUS ACTIVITY}

The present invention relates to a composition derived from a natural product capable of inhibiting the activity of coronavirus, and includes a phytoncide extracted from cypress tree and a substance extracted from a natural product exhibiting antiviral activity.

Following the emergence of SARS-CoV with a mortality rate of 9.6% in 2003 and MERS-CoV with a mortality rate of 36% in 2015, COVID-19, a novel coronavirus infection that began in December 2019, has a low mortality rate, but is easily mutated. The number of confirmed patients is exploding due to the high probability of causing infectious diseases and the spreading power.

Accordingly, as the concept of'space defense' has emerged from the concept of'contact defense' such as washing hands and wearing a mask to prevent virus infection, interest in related products is increasing.

'Space defense' refers to a method of lowering the possibility of virus transmission by inactivating or killing the virus in the space itself where the virus may exist in order to minimize the possibility of infection of various bacteria or viruses floating in the air in a specific space. do. In other words, preventive measures to minimize virus droplets and aerosol-type coronavirus infections in daily living spaces such as homes, offices, elevators, restaurants, cars, hotels, and infant care facilities, or in public places such as churches, education centers, schools, and performance halls. to be.

In particular, according to the World Health Organization WHO, H1N1 influenza and coronavirus are transmitted from person to person, and it is reported that there is a high possibility of droplet infection, which is infected by droplets that spread into the air through coughing or sneezing. Therefore, the importance of quarantine against virus particles existing in the atmosphere of a space where many people work or live is increasing day by day.

On the other hand, the cypress ( Chamaecyparis obtusa ) is also known as the cypress, and is an evergreen tree of the coniferous cypress family, and although it is native to Japan, it is widely cultivated as a planted tree in southern Korea through development. It is used as a deodorant and antibacterial agent.

Phytoncide produced from cypress trees is a plant's self-defense material, and is a substance exhaled or secreted by plants to resist pathogens, pests, and fungi, and itself contains sterilization and insecticidal components. The constituents of phytoncide are organic compounds composed of phenolic compounds, alkaloids, glycosides, etc., including terpenes, and are known to have antibacterial, sedative, deodorant, and stress-relieving effects.

Such phytoncide is a natural substance, not a chemical synthetic substance, and is easily absorbed by the human body, and is known to have numerous functions, such as antibacterial, deodorant, sedative, and stress relief. In addition, phytoncide is known to cleanse and moisturize the skin with excellent sterilization, antibacterial, and cleansing effects, and to strengthen immune functions in the body.

As such, various studies have been conducted on the use of phytoncide's antibacterial activity, air purification ability, stress relief, strengthening of intestinal and cardiopulmonary function, indoor sterilization, etc., but it can be sprayed into the atmosphere to show a virus suppression effect. Sufficient research has not yet been conducted on whether or not, and when using a high concentration of phytoncide to exhibit sufficient antibacterial and antiviral effects, there is a problem that the preference is rather low.

Korean Patent Publication No. 10-2017-0027536

It is an object of the present invention to provide a composite composition of cypress water containing extracts derived from natural products and phytoncide having an effect of inhibiting the activity of coronavirus, and a method for preparing the same.

Another object of the present invention is to provide a method for inhibiting or inactivating viruses, including various application forms and uses of the composition.

One aspect of the present invention for solving the above problem is 1200 to 1800 parts by weight of cypress water obtained by hot water extraction of leaves and stems of cypress trees; And myrrh ( Commiphora myrrha Holmes ), frankincense tree ( Boswellia Carterii ), gold silver flower ( Lonicera japonica ), baekwoon grass ( Hedyotis diffusa ), haengin (杏仁), doin (桃仁), octagon (八角, Illicium verum Hook.f. ), Bongchul (蓬朮, Curcuma zedoaria ), Mugunghwa ( Hibiscus syriacus ), Tangja tree ( Poncirus trifoliata ) seeds and horse dung mud mushroom ( Phellinus igniarus ) a mixed extract of one or more selected from the group consisting of; It provides an antiviral composition containing; 1 to 100 parts by weight.

Specifically, the cypress water is obtained by adding distilled water to cypress leaves and cypress trunks, hot water extraction at 100°C for 2 to 3 hours, and then concentration under reduced pressure, and the mixed extract is 90°C using 60 to 70% ethanol. It may be obtained by hot water extraction for 3 to 4 hours.

Specifically, the mixed extract is myrrh ( Commiphora myrrha Holmes ) 30 parts by weight, frankincense ( Boswellia Carterii ) 20 parts by weight, gold and silver ( Lonicera japonica ) 15 parts by weight, Baiyun grass ( Hedyotis diffusa ) 15 parts by weight, almond 5 Weight parts, Doin 5 parts by weight, Octagon (八角, Illicium verum Hook. f. ) 10 parts by weight, Bongchul (蓬朮, Curcuma zedoaria ) 10 parts by weight, Hibiscus syriacus 10 parts by weight, Tangja tree ( Poncirus trifoliata ) 5 parts by weight of seeds and 5 parts by weight of horse dung mud ( Phellinus igniarus ) may be mixed and prepared by hot water extraction for 3 to 4 hours at 90°C with 60 to 70% ethanol, but is limited thereto no.

In the present invention, specifically, the extract derived from the natural product may be extracted with water, a C ₁ to C ₄ lower alcohol, or a mixed solvent thereof. More specifically, it may be extracted using distilled water or 60 to 70% ethanol, but is not limited thereto.

Specifically, in the extraction process of the extract, one or more methods selected from the group consisting of cold needle, warm needle, heating, reflux, and ultrasonic extraction may be used alone or in combination, and are not limited thereto, and may be appropriately changed and applied as necessary. I can.

In addition, the extract may be concentrated or the solvent may be removed by performing an extraction or fractionation process, followed by a vacuum filtration process or further concentration and/or lyophilization, and specifically, a process of concentrating under reduced pressure after extraction. The extract may be obtained through the process, but is not limited thereto and may be appropriately changed and applied.

In the present invention, the “phytoncide” is a 100% natural substance that is difficult to artificially synthesize or combine, consisting of phenolic compounds including terpenes, alkaloids, and glycosides. In general, it is rapidly absorbed into the human body through a forest bath. By selectively sterilizing harmful bacteria, it has various effects such as strong antibacterial and antifungal action without resistance, stress relief, skin sedation, sound sleep induction effect, allergic skin disease improvement, and immune function enhancement.

Accordingly, for the busy modern people who cannot frequent forest bathing, the movement of product production to indirectly expect the forest bathing effect by adding phytoncide ingredients to various products that can be encountered in daily life such as fragrances, face wash, cosmetics and medicines has been actively made. Is losing.

As a result of the study on phytoncide, not only the amount of phytoncide emitted from plants, but also the efficacy are all different characteristics.A large amount of phytoncide occurs in coniferous trees such as pine and Japanese pine, especially in the cypress (Chamecyparis obtusa ). It is known that phytoncide, which has the highest content and healing power, is produced.

Currently, products containing phytoncide components mainly use essential oil or cypress water extracted from cypress trees, and the term "essential oil" refers to plant flowers, buds, leaves, stems, roots, and natural resins ( It contains phytoncide as a volatile substance with strong scent obtained from 天然樹脂).

In the present invention, the “antivirus” means inactivation, preferably permanent inactivation of the pathogen virus.

Specifically, viruses that can be inactivated or permanently inactivated by the composition may be DNA viruses and RNA viruses, and specific examples are influenza A virus (human, bird, swine (new type)), influenza B virus, parainfluenza Virus, (type A~E) hepatitis virus, measles virus, herpes virus, mumps virus, rabies virus, norovirus, HIV virus, SARS, bird flu, swine flu, ebola virus, MERS coronavirus (MERS-CoV) And it may be one or more selected from the group consisting of COVID-19, and most specifically, may be a coronavirus (SARS, MERS, and COVID-19), but is not limited thereto.

In the experimental example of the present invention, the antiviral efficacy of the composition of the present invention was confirmed using the SARS coronavirus (SARS-CoV), MERS coronavirus (MERS-CoV) and COVID-19 (SARS-CoV-2) ( Table 1).

Another aspect of the present invention relates to a cosmetic composition comprising the composition.

The cosmetic composition of the present invention is a solution, external ointment, cream, foam, nutrient lotion, flexible lotion, pack, softening water, emulsion, makeup base, essence, soap, liquid detergent, bathing agent, sunscreen cream, sun oil, suspension, emulsion It can be prepared in a formulation selected from the group consisting of liquid, paste, gel, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation, patch, and spray, specifically soap , It may be a liquid cleaning agent or hand cream, but is not limited thereto.

The cosmetic composition of the present invention may additionally include one or more cosmetically acceptable carriers blended in general skin cosmetics, and as common ingredients, for example, oil, water, surfactants, moisturizers, lower alcohols, thickeners , A chelating agent, a colorant, a preservative, a fragrance, and the like may be appropriately blended, but are not limited thereto.

The cosmetically acceptable carrier included in the cosmetic composition of the present invention varies depending on the formulation of the cosmetic composition.

When the formulation of the present invention is an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. May be used, but is not limited thereto. These may be used alone or in combination of two or more.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, etc. may be used as a carrier component. In particular, in the case of a spray, additional chlorofluorohard It may include a propellant such as locarbon, propane/butane or dimethyl ether, but is not limited thereto. These may be used alone or in combination of two or more.

When the formulation of the present invention is a solution or emulsion, a solvent, a solubilizing agent or an emulsifying agent may be 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, and the like may be used, and in particular, cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan May be used, but is not limited thereto. These may be used alone or in combination of two or more.

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, micro Crystalline cellulose, aluminum metahydroxide, bentonite, agar, or tracant may be used, but is not limited thereto. These may be used alone or in combination of two or more.

In addition, the composition of the present invention may be used as a transdermal administration method such as direct application or spraying on the skin, and the administration route of the composition of the present invention may be administered through any general route as long as it can reach the target tissue.

The amount of use of the composition of the present invention may be appropriately adjusted according to individual differences or formulations such as age and degree of lesions, and may be used from one week to several months by applying one or several times a day to the skin.

It was confirmed that the antiviral composition comprising cypress water containing natural substances and phytoncide provided by the present invention possesses excellent antiviral activity over various types of coronavirus, and is suitable for coronaviruses with frequent mutations through this It is expected to be used as a preventive agent and has excellent formulation safety.

In addition to phytoncide, the composition includes a plurality of extracts having antiviral efficacy derived from natural products, so that it can exhibit excellent virus inhibitory ability even at a low concentration, thereby solving the problem of low preference, such as causing discomfort caused by using a high concentration of phytoncide.

In addition, the composition of the present invention is derived from natural products and is not toxic, and does not cause side effects even when applied to the human body, so its utilization is high.

The antiviral composition of the present invention and the spray using the same have the effect of preventing early damage (contagion) caused by bacteria or viruses from places such as hospitals, lecture rooms, and public transportation, which are the main routes of transmission.

In addition, the antiviral composition of the present invention and the spray using the same, the composition of the present invention remains in the interior walls, exterior walls, ceilings, articles of places such as hospitals, lecture rooms, public transportation, etc. There is no practical effect that is easy to use.

The effects of the present invention are not limited to the above effects, and should be understood to include all effects that can be deduced from the configuration of the invention described in the detailed description or claims of the present invention.

Hereinafter, the present invention will be described in detail by examples. However, the following examples are only illustrative of the present invention, and the present invention is not limited by the following examples.

Example 1. Preparation of extracts derived from natural products having antiviral efficacy

Prior to preparing the antiviral composition of the present invention, an extract derived from a natural product having antiviral efficacy was prepared.

First, in order to prepare the extract derived from the natural product, myrrh ( Commiphora myrrha Holmes ), frankincense tree ( Boswellia Carterii ), gold silver flower ( Lonicera japonica ), baekwoon grass ( Hedyotis diffusa ), almond tree (杏仁), Doin (桃仁), octagon Illicium verum Hook.f. ), Bongchul (蓬朮, Curcuma zedoaria ), Mugunghwa ( Hibiscus syriacus ), Poncirus trifoliata seeds and horse dung mud ( Phellinus igniarus ) were washed and dried to prepare, Hot water extraction was performed at 90° C. for 3 to 4 hours using 70% ethanol.

Specifically, the extract derived from the natural product is myrrh ( Commiphora myrrha Holmes ) 30 parts by weight, frankincense tree ( Boswellia Carterii ) 20 parts by weight, gold and silver flower ( Lonicera japonica ) 15 parts by weight, Baiyun grass ( Hedyotis diffusa ) 15 parts by weight, almond (杏仁) 5 parts by weight, Doin 5 parts by weight, Octagon (八角, Illicium verum Hook.f. ) 10 parts by weight, Bongchul (蓬朮, Curcuma zedoaria ) 10 parts by weight, Hibiscus syriacus 10 parts by weight, Tangja tree ( Poncirus trifoliata ) 5 parts by weight of seeds and 5 parts by weight of horse dung mud ( Phellinus igniarus ) were mixed and prepared by hot water extraction at 90° C. for 3 to 4 hours with 60 to 70% ethanol.

Example 2. Preparation of antiviral composition containing phytoncide

Prior to preparing the antiviral composition containing phytoncide, cypress water for mixing with the extract derived from the natural product prepared in Example 1 was prepared.

To prepare the cypress water, 0.5 kg of dried cypress leaves and 0.5 kg of cypress stems are pulverized and placed in an extraction container, and 15L of distilled water is mixed with 1 kg of cypress leaves and stems and heated at 100°C for 2 to 3 hours And extracted with hot water, filtered through a 400 mesh filter cloth, and concentrated with a vacuum rotary concentrator, and extracted twice more in the same process using the same amount of distilled water to the remaining residue after filtration, filtered and concentrated under reduced pressure to contain phytoncide. Cypress water was prepared.

The extract of Example 1 is 1 to 100 parts by weight, more specifically 10 to 50 parts by weight, and most specifically 30 parts by weight based on 1500 parts by weight of cypress water obtained through the preparation method. Although the virus composition was prepared, it is not limited thereto.

Preparation of Comparative Examples 1 to 4

In the antiviral composition of Example 2, Comparative Examples 1 to 4 were prepared while changing the composition as follows.

Comparative Example 1 was prepared through the same method as in Example 2, except that the frankincense tree was excluded when preparing the composition of Example 1, and Comparative Example 2 further excluded Baiyun grass in Comparative Example 1, and Example 3 was prepared in the same manner as Example 2, except that Example 1 was prepared including 10 parts by weight of the composition of Example 2, and Comparative Example 4 was 100 parts by weight of Example 1 in the composition of Example 2. It was prepared in the same manner as in Example 2, except that it was prepared including.

Experimental Example 1. Cytotoxicity and virus inhibition effect confirmation

The composition of Example 2 and Comparative Examples 1 to 4 was subjected to an experiment to confirm the cytotoxicity and virus inhibitory effect.

Specifically, for the above experiment, the Corona 19 virus (BetaCoV/Korea/KCDC03/2020) isolated in Korea, the MERS virus (MERS-CoV/KOR/KNIH/002_05_2015) and SARS coronavirus (SARS) isolated and cultured from Korean patients -CoV), and cells for culturing it were vero cells (African green monkey kidney cell line), and maintenance system using DMEM (Dulbecco's modified Eagle's medium) medium supplemented with 5% FBS and penicillin/streptomycin. I prepared for it.

The cytotoxicity of host cells to the compositions of Example 2 and Comparative Examples 1 to 4 was measured by a cell attachment test. Specifically, 4.0×10 ⁴ cells/100 μL of vero cells were suspended in culture medium and inoculated into each well of a 96-well plate, followed by incubation for 24 hours in 5% CO ₂ , 37°C conditions, and 70-80% monolayer ( monolayer) was formed, and then washed once with PBS and inoculated with 100 μL of the culture solution and 100 μL of the composition. Thereafter, the cytopathic effect (CPE) was observed under a microscope for 2 days to confirm whether the cells adhered and proliferated and did not cause toxicity.

The virus for inoculation was cultured in a 75cm ² tissue culture flask (Corning, USA), and when the attached cells fell off due to the cell lesion effect, freezing and thawing were repeated twice, followed by centrifugation at 1,500×g for 5 minutes. Cell debris was removed, and the supernatant containing the virus was subdivided into a cryogenic vial (Nalgene, USA) and stored at -80°C.

Infectious titer TCID ₅₀ (50% tissue culture infective dose) of the inoculation virus was calculated. As a result, COVID-19 virus was 2.19 × 10 ² /mL, MERS-CoV virus was 2.35 × 10 ⁷ /mL, and SARS-CoV virus Was confirmed to be 3.24×10 ⁴ /mL.

Antiviral efficacy test for the extract was inoculated with 100 μL of each 100 μL of inoculation culture solution containing the compositions of Example 2 and Comparative Examples 1 to 4, and 100 μL of 10 times the amount of virus of TCID ₅₀ , and cultured for 2 days and then tested for cytotoxicity. On the contrary, the degree of inhibition of the cellular lesion effect of the composition was observed under a microscope. A 96-well plate was set as a control with 2 wells each of normal cells per sample, cells inoculated with only virus, and cells to which only the compositions of Example 2 and Comparative Examples 1 to 4 were added.

As a result of the cytotoxicity test, it was confirmed that all of Example 2 and Comparative Examples 1 to 3 did not exhibit cytotoxicity under the above conditions, except for Comparative Example 4 prepared by including 100 parts by weight of the composition of Example 1.

In addition, the test results of confirming the virus suppression ability were classified into failure to suppress the virus (-), partial suppression (+), and complete suppression (++), and the results are shown in Table 1 below.

Restraint ability Example 2 Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 COVID-19 ++ + - + ++ MERS-CoV ++ + - - ++ SARS-CoV ++ + - + ++

As shown in Table 1, it was confirmed that the composition of Example 2 of the present invention completely inhibited the virus, and Comparative Example 1 except for the frankincense tree slightly decreased the virus inhibitory ability, and the comparative example except for the frankincense tree and the Baiyun grass. 2 confirmed that the effect was significantly decreased compared to other experimental groups. Comparative Example 3, by adding a small amount of the composition of Example 1, caused a decrease in the effect, so that the antiviral effect was not exhibited on MERS-CoV, and Comparative Example 4 showed an excellent virus inhibitory effect, but the only cell toxicity test result As it has been confirmed to be toxic, it seems to be somewhat unreasonable to apply it to humans.

Experimental Example 2. User's sensory evaluation check when spraying

As confirmed in Experimental Example 1, the composition of Example 2 is expected to be able to prevent virus infection by applying an automatic sprayer or a humidifier to suppress viruses existing in the atmosphere of a certain space using excellent virus suppression ability. Sensory evaluation was performed when the compositions of Example 2 and Comparative Example were sprayed into the air.

At this time, Comparative Example 4, which may exhibit cytotoxicity, was excluded, and only Comparative Examples 1 to 3 were targeted.

The sensory test for the antiviral composition was conducted by selecting 24 adult men and women in their 20s to 50s. Specifically, 6 people in each group were evenly distributed by age and gender, and 1ml was sprayed into a sealed 1 pyeong space, The subjects' preference for scent, discomfort (strong odor), dizziness, uncomfortableness, and overall preference were evaluated.

A 10-point scoring method was used for the rating, and the higher the preference, the higher the score was given. When the evaluation was completed once for each person, they were completely ventilated while taking a sufficient rest for 30 minutes, and the average value was calculated by repeating evaluation three times. The results are shown in Table 2 below.

division Example 2 Comparative Example 1 Comparative Example 2 Comparative Example 3 Fragrance preference 9.1 8.3 7.7 8.2 Discomfort (strong smell) 9.0 7.4 7.2 8.5 Dizziness 8.9 8.3 8.1 8.5 Uncomfortable 8.7 8.0 7.8 8.2 Overall preference 8.9 7.9 7.5 8.1

As shown in Table 2 above, it was confirmed that the frankincense tree extract and the gold silver flower extract had a large effect on the overall flavor preference, and the result was significantly inferior to that of Example 2 in both the resulting discomfort and overall preference. Confirmed. In addition, the result of Comparative Example 3 was confirmed that when the composition of Example 1 was included in less than the numerical range suggested by the present invention, the preference fell so that a person could perceive the overall sensory feeling.

Experimental Example 3. Inhalation toxicity test

Inhalation toxicity was confirmed when the composition of Example 2 of the present invention was sprayed into the air.

Specifically, the composition of Example 2 was exposed for 4 hours daily at a concentration of 0, 125, 250 and 500 mg/m ³ for 5 males and females of mice (BALB/c) for 14 days of mortality, general symptoms, and body weight. Changes and autopsy findings were observed.

Experimental standards were conducted in accordance with the'Standards for Toxicity Tests for Drugs, etc.' and'Standards for Drug Safety Test Management' announced by the KFDA.

As a result of the experiment, even at the highest concentration of 500 mg/m ³ , no abnormal signs were observed in mortality, general symptoms, weight change, autopsy findings, etc., and it was confirmed that it is safe because it is not toxic even if inhaled at a high concentration (500 mg/m ³ ).

The above description of the present invention is for illustrative purposes only, and those of ordinary skill in the art to which the present invention pertains will be able to understand that it can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not limiting. For example, each component described as a single type may be implemented in a distributed manner, and similarly, components described as being distributed may also be implemented in a combined form.

The scope of the present invention is indicated by the claims to be described later, and all changes or modified forms derived from the meaning and scope of the claims and the concept of equivalents thereof should be construed as being included in the scope of the present invention.

Claims (6)

1200 to 1800 parts by weight of cypress water obtained by adding distilled water to the leaves and stems of cypress trees, hot water extraction at 100°C for 2 to 3 hours, and then concentration under reduced pressure; And
Myrrh ( Commiphora myrrha Holmes ) 30 parts by weight, Frankincense ( Boswellia Carterii ) 20 parts by weight, Gold and silver ( Lonicera japonica ) 15 parts by weight, Baekwoon grass ( Hedyotis diffusa ) 15 parts by weight, almond (杏仁) 5 parts by weight, Doin (桃仁) 5 parts by weight, octagonal (八角, Illicium verum Hook.f. ) 10 parts by weight, Bongchul (蓬朮, Curcuma zedoaria ) 10 parts by weight, Hibiscus syriacus 10 parts by weight, Poncirus trifoliata seeds 5 parts by weight And horse dung mud ( Phellinus igniarus ) 5 parts by weight of the mixture, and 1 to 100 parts by weight of a mixed extract prepared by hot water extraction for 3 to 4 hours at 90°C with 60 to 70% ethanol, and containing, coronavirus activity. Inhibitory composition. delete delete The method of claim 1,
The composition for inhibiting coronavirus activity is one or more selected from the group consisting of COVID-19, MERS-CoV virus and SARS-CoV virus, antiviral composition. A cosmetic composition comprising the composition for antiviral of claim 4. An antiviral aqueous dispersion composition for spray comprising the composition for antiviral of claim 4.