KR102252009B1 - Antiviral or antibacterial composition containing extracts of Torreya nucifera leaf - Google Patents

Abstract

The present invention relates to an antiviral composition comprising an extract of Torreya nucifera leaves rich in a catechin component and a tannic acid component, and to a composition for sterilization harmless to the human body and containing the extract of Torreya nucifera leaves with excellent antibacterial activity.

Description

Antiviral or antibacterial composition containing extracts of Torreya nucifera leaf}

The present invention relates to an antiviral composition comprising a catechin component and a tannic acid component richly containing a birch leaf extract, and a sterilizing composition comprising a birch leaf extract that is harmless to the human body and has excellent antibacterial activity.

Bacterial infectious diseases such as food poisoning, pneumonia, and various viral infectious diseases such as swine flu, avian influenza, and coronavirus, which are recurring every year, cause major social problems, arousing great social interest in effective countermeasures against bacterial and viral diseases. have.

Pathogenic bacteria are 　causing diseases 　, and host pathogenic bacteria infected with host cells infected with tuberculosis bacteria that cause tuberculosis and pneumonia are streptococcus or Pseudomonas that can cause food poisoning, campylobacter, Salmonella, etc. It causes direct or indirect damage. Bacterial infections can generally be treated with antibiotics, and antibiotics are classified as 　disinfectants if they kill bacteria and 　 bacteriostatic agents if they inhibit growth. However, the continued use of antibiotics can result in resistance, and a problem has been raised due to the emergence of multidrug resistant bacteria that have simultaneous resistance to several antibiotics.

Viruses are microscopic infectious pathogenic particles having a size that passes through a bacterial filter paper (0.22 μm). Viruses are classified into bacteriophage, plant virus, and animal virus according to the type of host cell, and are classified into DNA virus, RNA virus, etc. according to the type of nucleic acid. Currently, vaccination is the best way to prevent viral diseases, but interest in the development and distribution of inhibitors for virus prevention that can complement the problems of the potency, safety and stability of the vaccine is increasing day by day.

There is a growing awareness of the importance of hand washing to prevent the spread of bacterial or viral infectious diseases and infectious diseases. Hand sanitizers used for hand washing, also known as hand sanitizers or hand sanitizers, are antimicrobial and antimicrobial substances that replace hand washing with water or wash hands with water after using hand sanitizers. It is a liquid type disinfectant or gel type disinfectant composed of chemical substances, and is mainly used in the form of spraying or pumping. The main component of such a hand sanitizer is composed of ethanol, but there is a limitation in that the sterilization power is lowered depending on the content of ethanol in the hand sanitizer, and additional chemical substances to reinforce the sterilization power may be added to compensate for this. However, as consumers' health-oriented demands are increasing, the avoidance of chemical substances is occurring in society. Especially, in the case of cosmetics that are applied to skin, etc., when considering the phenomenon of consumers checking all ingredients and purchasing and using them, Instead of adding additional chemicals, there is a need for technology development to improve sterilization by using antimicrobial substances obtained from natural materials.

Bismuth tree (scientific name: Torreya nucifera, English name: Nut-bearing torreya, Japanese torreya) is an evergreen tree of the Yew family native to the southern regions of Korea and Japan. , Appetite promotion, digestion promotion, constipation and hemorrhoids, etc., and has been used as an anthelmintic. Although antimicrobial compositions have been reported, it only exhibits antibacterial effects against strains such as acne bacteria, and it is not an invention for sterilizing strains for preventing infectious diseases, and little is known about the antiviral effect of conventional visa extracts.

Unexamined Patent Publication No. 10-2018-0006551

It is an object of the present invention to provide a disinfectant composition comprising a birch leaf extract which is harmless to the human body and has excellent antibacterial activity by using a birch leaf extract rich in a catechin component and a tannic acid component.

In order to achieve the above object, the present invention provides a birch leaf extract, an antiviral composition comprising the same, or a sterilizing composition comprising the same.

Hereinafter, the present invention will be described in more detail.

While searching for antiviral and antimicrobial substances obtained from natural materials, the present inventors completed the present invention by discovering that the extract of birch leaves has excellent antiviral and antibacterial effects, and has no toxicity or side effects even when used directly on the skin. Was done.

In the present specification, the'Visa leaf extract' is obtained by extracting from the leaves of the Visaceae tree among various organs of Torreya nucifera, such as fruits, leaves, flowers, stems, and roots.

In one embodiment of the present invention, the extract of birch leaves includes a fat or oil component obtained by extraction with water or an organic solvent. In the above, the organic solvent includes at least one selected from the group consisting of alcohol having 1 to 4 carbon atoms, hexane, acetone, ethyl acetate, diethyl ether, ethyl methyl ketone, and chloroform, but is not limited thereto. The organic solvent may preferably be ethanol.

In one embodiment of the present invention, the concentration of ethanol may be 70% (v/v) to 100% (anhydrous ethanol). When the concentration of the organic solvent is less than 70% (v/v), the extraction amount of the fat-soluble component is reduced relative to the unit weight of the extract.

In one embodiment of the present invention, it is characterized in that the extract is extracted by mixing 10 to 30% by weight of the birch leaf and 70 to 90% by weight of the solvent.

In one embodiment of the present invention, the extracting conditions for the leaves of the birch tree are characterized in that the extraction is performed under pressure, exceeding 0 hours and 4 hours or less at a temperature of 60 to 80°C.

If the extraction temperature is less than 60℃, it is difficult to achieve the target level of extraction, and if the extraction temperature exceeds 80℃ or the extraction time is more than 4 hours, there is a possibility that the volatile components of plant essential oils may be excessively volatilized, resulting in a decrease in productivity. . During extraction, it is extracted under pressure to prevent volatilization of volatile components.

In one embodiment of the present invention, the extract of birch leaves is protocatechuic acid ethyl ester, 4-hydroxy benzoic acid, sirinic acid, coumaric acid, rutin, ferulic acid, naringenin, resveratol, nicotinic acid, gallic acid, Protocatechuic acid, catechin, it provides a sterilizing composition comprising a leaf extract of the birch, characterized in that it comprises at least one selected from tannic acid.

Catechin is a type of polyphenol and is contained in large amounts in green tea and cacao. Catechins are known to have various effects such as cancer inhibition, arteriosclerosis, blood pressure increase inhibition, blood clot prevention, anti-obesity, anti-diabetes, antibacterial, detoxification, anti-inflammatory action, tooth decay prevention, mouth mouth prevention, and normalization of the intestinal flora.

Tannic acid is also called tannin and tannin, and is a kind of polyphenol that gives off a very astringent taste. It is synthesized by plants and is mainly contained in fruits, persimmons, acorns, and tea. It is known to increase the elasticity of blood vessels and to help the enamel formation of teeth to prevent tooth decay and to have antibacterial effects.

The present invention may include such a catechin content of 10000ng / ml ~ 30000ng / ml and tannic acid content of 50000ng / ml ~ 100000ng / ml of the extract of the birch leaves, and the conventional ethanol alone complements the technical limitations of poor sterilization, and has antiviral properties. It provides an effective disinfecting composition further shown.

In one embodiment of the present invention, it provides an antiviral composition comprising the extract of leaves of the birch tree.

In one embodiment of the present invention, the antiviral composition may be an antiviral cosmetic composition or an antiviral pharmaceutical composition.

In one embodiment of the present invention, the target virus of the antiviral composition is a group consisting of a human respiratory syncytical virus, an enteritis virus A, and a human influenza A (H1N1). Can be selected from one or more of.

The respiratory syncytial virus (RSV) is a virus that causes respiratory infection. RSV belongs to the genus 　Pneumovirus of the family 　Paramyxoviridae 　, and the F protein on the viral surface induces membrane fusion with nearby cells. It is the main cause of lower respiratory tract infections and is prevalent in temperate winters and tropical rainy seasons. RSV is a virus with an envelope containing a lipoprotein of 120~200　nm in diameter, and the lipoprotein is composed of F (fusion), G (attachment), and 　SH. The F and G proteins are responsible for binding to the cell membrane. The genome of RSV is about 15000 nucleotides, which is a single negative-stranded RNA.

The rotavirus (Rotavirus A) is a virus that mainly causes diarrhea between infants and children. As a virus belonging to the reoviridae, immunity to the virus occurs with each infection, and subsequent infections have less severe symptoms. Rotavirus does not have an envelope and has an icosahedral capsid consisting of three protein layers, enclosing 11 　dsRNA 　 genome segments. Virus serotypes are determined by the proteins constituting the outermost layer of the capsid, 　VP7 and 　VP4, and both proteins are the main antigens that induce antibody production.

The human influenza A (H1N1) is also referred to as swine flu A (H1N1), and is a new virus caused by mutations of the type A 　 influenza 　 virus, and it became a worldwide epidemic in 2009. Human influenza A is an enveloped virus surrounded by a membrane, and a specific glycoprotein is present in the envelope. Influenza virus A is subtyped according to the types of hemagglutinin (　HA) and neuraminidase (　NA), which are the surface antigens of the virus, and are expressed as H1N1, H3N2, etc. The M1 matrix protein layer is present inside the envelope, and the genetic material is present in the form of binding to the nucleocapsid protein inside the matrix protein. The genetic material is a single-stranded 　RNA, and consists of 7 to 8 segments instead of one.

Regardless of whether the virus is enveloped or not, the extract of the leaves of the birch leaves of the present invention exhibits an excellent antiviral effect of killing more than 90 to 99.9% of cells infected with the RNA genome virus.

In one embodiment of the present invention, the present invention provides a sterilizing composition comprising a birch leaf extract, characterized in that it comprises 10 to 40% by weight of birch leaf extract.

The sterilizing composition may further include 30 to 60% by weight of ethanol, 10 to 30% by weight of purified water, and 10 to 20% by weight of glycerin.

When the amount of the extract of the birch leaves is less than 10% by weight, the bactericidal effect of the composition decreases, and when it exceeds 40% by weight, skin irritation may occur.

When the ethanol content is less than 30% by weight, the effect of dissolving the protein membrane of bacteria may be reduced, and when it is more than 60% by weight, skin irritation may occur.

The purified water is added to adjust the concentration of the sterilizing composition.

The glycerin is a moisturizing factor for solving the dryness caused by ethanol when used on the skin.If it is added less than 10% by weight, the moisturizing effect decreases and you may feel dry after use.If it is used in excess of 20% by weight, it becomes a sticky formulation. It can deteriorate the feeling of use.

In one embodiment of the present invention, the sterilizing composition is characterized in that it is a sterilizing cosmetic composition or a sterilizing pharmaceutical composition.

In one embodiment of the present invention, the target strain of the sterilizing composition is E. coli, Staphylococcus aureus (S. aureus), Pseudomonas aeruginosa (P. aeruginosa), Salmonella (S. typhimurium), Flexneriri. It provides a sterilizing composition comprising a bismuth leaf extract characterized by at least one selected from the group consisting of S. flexneri and K. pneumoniae.

According to the microbial limit test method among general test methods of the Korean Pharmacopoeia or the sterilization test method notified by the Ministry of Food and Drug Safety (No. 2019-71), E. coli ( Escherichia coli ATCC 10536 or Escherichia coli ATCC 11229), yellow Staphylococcus aureus ATCC 6538 is used, and in addition to the standard strain, Cereus bacterium (Bacillus cereus ATCC 21772), Vibrio parahaemolyticus ATCC 27969), Salmonella typhimurium ATCC 13311), Listeria monocytogenes A test strain such as ATCC 19111 (or Listeria monocytogenes ATCC 19115) can be selected and added, and it is disclosed that it is considered suitable when the viable cell number reduction rate is 99.999% or more.

In the antibacterial test using the sterilizing composition of the present invention, in addition to the test strains notified by the Ministry of Food and Drug Safety, it is possible to find out the sterilization ability of 99.999% or more against P. aeruginosa, S. flexneri, and K. pneumoniae. there was. In that the sterilization effect for each test strain may vary depending on the composition of the disinfectant and the antibacterial active ingredient, the sterilization ability for the following strains corresponds to the unique effect of the sterilization composition containing the extract of the leaves of the present invention. .

Pseudomonas aeruginosa is an aerobic 　gram-negative bacteria, belongs to bacillus, and does not form spores. Because this fungus is inherently resistant to antibiotics, it is widely used in medical research. It is also opportunistic, so it does not cause great harm to healthy people, but it can cause very serious infections in people with cystic fibrosis or people with reduced immunity. Pseudomonas aeruginosa can grow even under the lowest nutrient conditions, so its growth range is very wide. Especially in humid environments in hospitals, instruments and even disinfectant solutions are everywhere.

Four species are included in the bacterial Shigella (hereinafter referred to as Shigella), and the Flexneri Shigella is also called Flexner Shigella, Group B Shigella, Shigella, and Group B Shigella. It is one of the most common causative agents of acute diarrhea in humans. Shigella is a Gram-negative, painless anaerobic bacteria that does not have flagella and is non-motile. Vaccines have not been developed yet, and cross-immunity does not occur between serotypes within the species as well as each species, making it difficult to develop a vaccine. Shigella is transmitted through the fecal-oral route, and because Shigella is resistant to an acidic environment and can be passed on to the intestine by surviving in the stomach, it is highly contagious enough to cause disease even by ingesting 10 to 100 bacteria.

Klebsiella pneumonia is a rod-shaped gram-negative bacterium, and is conditionally anaerobic and oxidase-positive. This bacterium belongs to enteric bacteria and is a symbiotic bacteria present in the skin and oral cavity. It is considered as a bacteria that causes opportunistic infections such as sepsis, pneumonia, urinary tract infection, and soft tissue infection in patients with reduced immunity. In particular, this bacterium accumulates extensive beta-lactam-degrading enzyme [extended-spectrum β-lactamase (ESBL)] and also transfers related genes to other bacteria. In the case of K. pneumoniae , which produces these ESBLs, it becomes resistant to various antibiotics, making it very difficult to treat infectious diseases.

Pseudomonas aeruginosa, which is not killed by a low level of disinfection, Shigella aeruginosa, which is resistant to an acidic environment, and pneumonia bacilli, which are resistant to various antibiotics, can be killed only by a complex disinfection method of medium or higher levels in the past. Corresponds to the strain that is difficult to do. In the present invention, the fact that 99.999% of the strain can be killed only by using a hand sanitizer corresponds to a novel discovery of the present invention that did not exist before.

In one embodiment of the present invention, it provides a birch leaf extract, characterized in that the sterilizing composition can be used directly on human skin.

Since the sterilizing composition does not have cytotoxicity and does not irritate the skin because it has been confirmed that it is safe for human skin, the sterilizing composition can be used directly on the human skin.

The cosmetic composition according to the present invention may be provided in any formulation suitable for topical application. For example, it may be provided in the form of a solution, emulsion, suspension, solid, gel, powder, paste, foam or aerosol composition, and the composition of such a formulation may be prepared according to conventional methods in the art.

The cosmetic composition according to the present invention may further include a moisturizing agent, an emollient agent, a surfactant, a UV absorber, a preservative, a fungicide, an antioxidant, a pH adjuster, an organic and inorganic pigment, a fragrance, a cooling agent or a limiting agent.

As a natural substance containing high catechin and tannic acid according to the present invention, the extract of the leaves of the birch tree according to the present invention has an antiviral activity of up to 99% depending on the virus species, and thus can be used as an antiviral composition.

The composition for sterilization comprising the extract of birch leaves according to the present invention complements the shortcomings of the existing ethanol disinfectant, and provides a stable and high sterilization of 99.999% or more against various infectious bacteria, as well as bacteria according to the Korean Pharmacopoeia General Test Method and KFDA notice. Show ability.

Even if the composition for sterilization according to the present invention is applied directly to the skin, there is no concern about the risk of side effects or toxicity.

Hereinafter, the present invention will be described in detail based on examples and the like, but the present invention is not limited by the following examples.

Example 1. Extract of birch leaves

1 kg of birch leaves collected from southern regions such as Jeju Island, Jangseong or Goheung were crushed, mixed with 100% ethanol, heated, and extracted for 3 hours at 70°C.

Example 2. Preparation of a composition for sterilization containing the extract of birch leaves

Based on the total volume of the sterilizing composition 1L, 0.6L of ethanol, 0.25L of purified water, 0.1L of glycerin, and 0.05L of birch leaf extract are mixed.

<Experimental Example 1> Ingredients of the extract of the leaf extract of the present invention

The components of the extract of the leaf extract prepared in Example 1 were analyzed.

The gumrang range of Table 1 below is 1 to 1000 ng/ml (2 to 2000 ng/ml for catechin), and the results obtained by converting the sample dilution factor are as follows.

number Ingredient name Calculated concentration (ng/mL) One Protocatechuic acid ethyl ester 8.491 2 4-hydroxy benzoic acid 289.115 3 Caffic acid N/A 4 Syringic acid 465.46 5 Vanillic acid N/A 6 Coumaric acid 48.235 7 Rutin 573.768 8 Ferulic acid 333.126 9 Oxyresveratrol N/A 10 Naringein 126.909 11 Resveratol 17.293 12 Benzoic acid N/A 13 Nicotinic acid 11.144 14 Gallic acid 11.211 15 Protocatechuic acid 64.417 16 Verbadoside N/A 17 Bergenin N/A 18 Catechin 16300

* N/A: Not detected

The range of gumrang in Table 2 below is 0.25-50 ug/ml, and it is the result obtained by converting the sample dilution factor (100 times). number Ingredient name Calculated concentration (ug/mL) One Tannic acid 86.4

As can be seen in Tables 1 and 2, the extract of the leaves of the birch tree of the present invention contains 16300 ng/ml of catechin, and contains more than 86400 ng/mL of tannic acid. Due to the high catechin and tannin content of the extract of the extract of the leaves of the extract of the present invention, the composition including the extract of the extract of the leaf of the extract of the present invention exhibits excellent bactericidal and antiviral activity.

<Experimental Example 2> Antimicrobial effect test for each strain on a sterilizing composition containing a birch leaf extract

In order to investigate the antimicrobial spectrum of the composition for sterilization comprising the extract of the leaves of the present invention in Example 2,

The antibacterial test method was ASTM E2315-16, and the test conditions were at the reaction temperature (22±2)°C at the concentration of the stock solution, and the reaction time was 5 minutes.

As an experimental strain

Escherichia coli (E. coli) ATCC 10536,

Staphylococcus aureus (S.aureus) ATCC 6538,

Pseudomonas aeroginosa (P. aeroginosa) ATCC 15522,

Salmonella typhimurium (S. typhimurium) ATCC 13311,

Shigella flexneri (S. flexneri) ATCC 12022, and

Klebsiella pneumoniae (K. pneumoniae) ATCC 4352 was used.

The results of the sterilization experiment are shown in Table 3 below.

Results of sterilization activity test by strain Test Items unit Sample classification Result Test Methods Sterilization experiment (E. coli) Log reduction - >5.73 ASTM E2315-16 Sterilization experiment (S. aureus) Log reduction - >5.38 ASTM E2315-16 Sterilization experiment (P. aeruginosa) Log reduction - >5.68 ASTM E2315-16 Sterilization experiment (S. typhimurium) Log reduction - >5.77 ASTM E2315-16 Sterilization experiment (S. flexneri) Log reduction - >5.78 ASTM E2315-16 Sterilization experiment (K. pneumoniae) Log reduction - >5.76 ASTM E2315-16

Log reduction(LR) = mean log(microbial population)-mean log(surviving test population)

log reduction 1 or more: 90% or more

log reduction 2 or more: 99% or more

log reduction 3 or more: 99.9% or more

log reduction 4 or more: 99.99% or more

log reduction 5 or more: 99.999% or more

As shown in Table 3, the hand sanitizer containing the extract of the leaf extract of the present invention is S. aureus, which is a Gram-positive bacteria, and E. coli, which is a gram-negative bacteria, P. aeruginosa, and Salmonella It exhibits a high antibacterial activity of 99.999% or more against all of S. typhimurium, S. flexneri, and K. pneumonia, which exceeds that of satisfying the standards for disinfectants notified by the Ministry of Food and Drug Safety, and is low. It can be seen that it shows excellent antibacterial activity even in strains that are not sterilized by level of disinfection.

<Experimental Example 3> Antiviral effect test on a composition containing a birch leaf extract

ASTM E1052-20 method was used to investigate the antiviral spectrum of the extract of the extract of the leaves of the present invention in Example 2, and the test conditions were 0.1% (v/v) of the extract of the extract of the leaf of the present invention in 100% ethyl alcohol. The diluted solution diluted with was used at the reaction temperature (22±2)℃, and the reaction time was 5 minutes.

Viruses include Human respiratory syncytial virus, Strain: Long, ATCC VR-26

Rotavirus A, Strain: Wa (TC adapted), ATCC VR-2018 and

Human influenza A (H1N1), Strain: KUMC-76, KBPV-VR-76 were used.

Host cells include Hep-2, KCLB 10023,

CV-1, KCLB 10070 and

MDCK (NBL-2) and ATCC CCL-34 were used.

The results of the sterilization experiment are shown in Table 4 below.

Antiviral activity test result Test Items unit Sample classification Result Test Methods Antiviral test
(Human respiratory syncytial virus) Log reduction - ≥3.63 ASTM E1052-20 Antiviral test
(Rotavirus A) Log reduction - 1.62 ASTM E1052-20 Antiviral test
[Human influenza A(H1N1)] Log reduction - 2.50 ASTM E1052-20

Log reduction (LR) = LU (untreated viral control group)-LT (treated viral test group)

log reduction 1 or more: 90% or more

log reduction 2 or more: 99% or more

log reduction 3 or more: 99.9% or more

log reduction 4 or more: 99.99% or more

log reduction 5 or more: 99.999% or more

As shown in Table 4, the leaf extract of the present invention has a reduction rate of 99.9% or more of virus proliferation level in the human respiratory syncytial virus, and 90% or more of virus proliferation level reduction in enteritis virus A (Rotavirus A). In human influenza A (H1N1), the virus proliferation level decreased by more than 99%. This indicates that the extract of the leaf extract of the present invention has high antiviral activity in reducing infectious viruses.

Although preferred embodiments of the present invention have been described above, the embodiments described above are illustrative in all respects and should be understood as non-limiting, and the scope of the present invention is indicated by the claims to be described later rather than the detailed description. , All changes or modified forms derived from the meaning and scope of the claims and their equivalent concepts should be interpreted as being included in the scope of the present invention.

Claims (7)

delete A pharmaceutical composition for antiviral comprising extracts of leaves of birch leaves,
The group consisting of protocatechuic acid ethyl ester, 4-hydroxybenzoic acid, ringic acid, comaric acid, rutin, ferulic acid, naringenin, resveratol, nicotinic acid, gallic acid, protocatechuic acid, catechin, and tannic acid. Antiviral pharmaceutical composition comprising at least one selected from. delete The method of claim 2, wherein the target virus of the antiviral pharmaceutical composition is one or more selected from the group consisting of human respiratory syncytical virus, enteritis virus A, and human influenza A. Antiviral pharmaceutical composition, characterized in that the. delete delete delete