KR102009604B1 - Antimicrobial composition comprising lonicera japonica extract and magnolia obovata extract - Google Patents

Abstract

The present disclosure provides an antimicrobial composition comprising a fraction of Lonicera japonica extract and a Magnolia obovata extract. In addition, the present disclosure provides a cosmetic composition comprising a Lonicera japonica extract and a Magnolia obovata extract.

Description

ANTIMICROBIAL COMPOSITION COMPRISING LONICERA JAPONICA EXTRACT AND MAGNOLIA OBOVATA EXTRACT

The present disclosure is a honeysuckle extract or fractions thereof; And it relates to an antimicrobial composition comprising a thick extract or a fraction thereof.

Skin is the outermost defense of our body, protecting the body from external stimuli and chemicals such as ultraviolet rays, antigens, and microorganisms. The skin protects the body by adjusting the pH of the skin surface, controlling the amount of moisture, sebaceous glands secretion, or sweating. However, when the barrier function of the stratum corneum is weakened, pathogens and the like easily penetrate the skin, which causes various skin diseases. Therefore, to protect the skin from these pathogens, people use cosmetics to maintain the homeostasis of the skin. Recently, as people's interest in beauty increased, the demand for cosmetics increased rapidly. As a result of the increasing demand and interest in cosmetics, consumer demand for natural and environmentally friendly products has also increased.

The cosmetic product defined in the cosmetics law means that the product is used in the human body to clean and beautify the human body to add charm and brighten the appearance, or to maintain or promote the health of the skin and hair. Since cosmetics contain a lot of water, it has a very suitable environment for microorganisms to inhabit. If cosmetics used to protect the skin are contaminated with microorganisms, preservatives are added to prevent cosmetics from contamination by microorganisms, because they can adversely affect the skin. However, among the preservatives used in cosmetics, in particular, side effects such as the occurrence of breast cancer of parabens preservatives have been reported, the demand for natural cosmetics has increased and accordingly, research on natural preservatives has been actively conducted.

Under these backgrounds, the present inventors made intensive research efforts to develop natural preservatives that can be used in cosmetics, and confirmed the antimicrobial synergistic activity of the honeysuckle extract and thick gourd extract and completed the present invention.

Korean J. Microbiol. Biotechnol. 2014, 42 (4), 361-366

It is an object of the present disclosure to provide natural substances with antimicrobial activity that can be used as preservatives for cosmetics. Specifically, it provides a composition comprising a honeysuckle extract and a thick leaf extract having antibacterial synergistic activity.

As one aspect for achieving the above object, the present disclosure is a honeysuckle extract or fractions thereof; And it provides an antimicrobial composition comprising a thick extract or a fraction thereof.

In the present disclosure, the antimicrobial composition is used in the same sense as an antimicrobial agent, a fungicide, a preservative, a preservative or an antimicrobial agent, and preferably means a substance capable of inhibiting or inhibiting the growth and life function of bacteria.

Honeysuckle ( Lonicera japonica ) is an evergreen broad-leaved shrub that grows in mountains all over Korea. Since ancient times, stems, flowers, and fruits of honeysuckle have been used for high fever, acute hepatitis, and inflammation.

Magnolia obovata is a magnolia and deciduous tree native to Japan and China. The bark of Japanese magnolia is widely used for medicinal purposes such as digestion, diuresis, sodam and vomiting.

In one embodiment, the extracts of the phosphorus vine and the groin are water or alcohol extracts, specifically water, methanol, ethanol, propanol, or butanol extracts, and more specifically ethanol extracts. In one embodiment, the honeysuckle extract is 50% ethanol extract, the thick extract is 70% ethanol extract. In one embodiment, the honeysuckle extract is extracted for 1 to 48 hours, specifically 20 to 30 hours at room temperature. In one embodiment, the thick gourd extract is extracted for 1 to 48 hours, specifically 20 to 30 hours at 20 to 40 ℃, 25 to 35 ℃.

Fractions of the phosphorus honeysuckle extract and the thick leaf extract may be fractionated with an organic solvent, specifically ethyl acetate, may be fractionated 2 to 4 times.

The antimicrobial composition of the present disclosure may include the honeysuckle extract or a fraction thereof; and the thick extract or a fraction thereof, respectively, in a weight ratio of 10: 1 to 0.1: 1, specifically, in a weight ratio of 6: 1 to 0.5: 1, more specifically 4: It may be included in a weight ratio of 1 to 1: 1. Since the concentration of each extract or fraction required to inhibit the activity of the strain is different, the weight ratio can also be adjusted according to the strain.

In the present disclosure, it was confirmed that the extracts of the honeysuckle and the grommets or fractions thereof had antimicrobial activity by mixing, among them, Staphylococcus aureus, Bacillus subtilis and yeast, in particular, showed antibacterial synergistic activity. In one embodiment, the bacteria identified as having antimicrobial activity are Staphylococcus aureus , Bacillus subtilis , Gram-negative bacteria Escherichia coli , Pseudomonas aeruginosa , and fungi Yeast, Candida albicans ) And black mold ( Aspergillus niger ). The bacteria may be a cosmetic application regulatory microorganism. In one embodiment, the extracts or fractions thereof of the honeysuckle and thick gourd were confirmed to have antimicrobial activity, respectively, among them, Staphylococcus aureus, Bacillus subtilis, and fungus showed especially synergistic activity. In other words, compared to the case of using only individual extracts or fractions, it was confirmed that a synergistic effect was obtained when the mixtures were used in addition to the increase of the antimicrobial activity due to the simple mixing. In the present disclosure, this was confirmed through a FIC index through a checkboard test (see FIG. 1).

When the antimicrobial composition according to the present disclosure is used as an antimicrobial agent, the formulation of the antimicrobial agent can be appropriately adjusted according to the method of use and use, such as liquids, suspensions, granules, fragrances, powders, syrups, aerosols, extracts, ointments. , May be induced extracts, emulsions, premises, eye drops, injections, spirits, capsules, creams, pills.

On the other hand, the antimicrobial composition of the present disclosure may be provided in the form of a detergent, a pharmaceutical, a food additive, a fragrance, a cosmetic, or a paint as a use for inhibiting the production and growth of a fungus that produces pathogenic bacteria or toxins. When used as a medicament may be used as a disinfectant, therapeutic, preventive, etc., when used as a food additive antimicrobial composition may be appropriately adjusted according to the type of food and the method of use.

The antimicrobial composition of the present disclosure may include 0.001 to 99.99% by weight, preferably 0.1 to 99% by weight of the mixture of each extract or fraction based on the total weight of the composition, and the method of use and purpose of use of the antimicrobial composition. According to the content of the active ingredient can be adjusted appropriately.

Another aspect of the disclosure is a honeysuckle extract or a fraction thereof; And it provides a cosmetic composition comprising a thick extract or a fraction thereof. The honeysuckle extract, fractions thereof, and the hunch extract, fractions thereof are as described above. In addition, the mixed weight ratio thereof is as described above.

The cosmetic composition of the present disclosure may exhibit better antimicrobial activity to protect the skin from bacteria causing various inflammations and diseases on the skin, and to inhibit and sterilize bacteria that may exist in cosmetics.

Cosmetic compositions of the present disclosure are in the group consisting of solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing, oils, powder foundations, emulsion foundations, wax foundations, sprays and packs It may be formulated and provided in the form of choice.

Cosmetic compositions of the present disclosure may further comprise a cosmetically acceptable carrier.

The kind of the cosmetically acceptable carrier of the present disclosure is not particularly limited as long as it does not impair the activity and properties of the present cosmetic composition, any of the carriers commonly used in the art and cosmetically acceptable may be used. Non-limiting examples of the cosmetically acceptable carrier include saline, sterile water, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol and the like. These may be used alone or in combination of two or more thereof. The cosmetically acceptable carrier of the present application may include a non-naturally occuring carrier.

The cosmetically acceptable carrier of the present disclosure varies depending on the formulation of the cosmetic composition.

When the formulation of the cosmetic composition of the present disclosure is a paste, cream or gel, the carrier component is 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.

When the formulation of the cosmetic composition of the present disclosure is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, etc. may be used as the carrier component, and in particular, in the case of a spray, additionally chloro Propellants such as fluorohydrocarbons, propane / butane or dimethyl ether may be included, but are not limited thereto.

When the formulation of the cosmetic composition of the present disclosure is a solution or emulsion, a solvent, a solubilizer or an emulsifier may be used as the carrier component, for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl Benzoate, propylene glycol, 1,3-butylglycol oil and the like can be used, and in particular, cottonseed oil, peanut oil, corn seed oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or sorbitan Fatty acid ester of may be used, but is not limited thereto.

If the formulation of the cosmetic composition of the present disclosure is a suspension, suspensions such as liquid diluents such as water, ethanol or propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol esters and polyoxyethylene sorbitan esters as carrier components First, microcrystalline cellulose, aluminum metahydroxyde, bentonite, agar or tracant may be used, but is not limited thereto.

When the formulation of the cosmetic composition of the present disclosure is a soap, as a carrier component, alkali metal salts of fatty acids, fatty acid hemiester salts, fatty acid protein hydrolyzates, isethionates, lanolin derivatives, aliphatic alcohols, vegetable oils, glycerol, sugars and the like May be used, but is not limited thereto.

When the formulation of the cosmetic composition of the present disclosure is a pack, a wash off containing a pigment such as kaolin, talc, zinc oxide, or titanium dioxide in a peel-off pack containing polyvinyl alcohol, a general emulsified cosmetic It includes all forms of packs or mask sheet packs, but is not particularly limited thereto.

The cosmetic composition of the present disclosure may further include components included in conventional skin external preparation components such as water, a surfactant, a moisturizer, a lower alcohol having 1 to 6 carbon atoms, a chelating agent, a bactericide, an antioxidant, a preservative, a pigment, and a perfume. Can be.

Another aspect of the disclosure is a honeysuckle extract or a fraction thereof; And it provides a quasi-drug composition comprising a thick extract or a fraction thereof. The honeysuckle extract, fractions thereof, and the hunch extract, fractions thereof are as described above. In addition, the mixed weight ratio thereof is as described above.

The quasi-drug composition of the present disclosure may exhibit better antibacterial activity to protect the skin from bacteria causing various inflammations and diseases on the skin, and to inhibit and sterilize bacteria that may be present in the quasi-drug.

The term "quasi drug" as used in the present disclosure refers to articles that are less active than drugs among those used for the purpose of diagnosing, treating, ameliorating, reducing, treating, or preventing a disease of a human or animal, for example, the Korean Pharmacist Act According to the article, quasi-drugs exclude products used for the purpose of medicines, and include products used for the treatment or prevention of diseases of humans and animals, and products with slight or no direct action on the human body.

The quasi-drug composition of the present disclosure may be prepared in a form selected from the group consisting of body cleanser, foam, soap, mask, ointment, cream, lotion, essence and spray, but is not limited thereto.

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

The antimicrobial composition according to the present disclosure is a natural preservative, and further enhances the antimicrobial activity of each existing extract. In particular, it has antimicrobial activity against the strains included in the cosmetic regulation, and some bacteria have a superior antimicrobial activity by having a synergistic effect than the individual extract. Therefore, it can be used as a natural preservative in cosmetic fields or other fields requiring antimicrobial activity.

Figure 1 shows the FIC index confirming the antimicrobial activity for each strain at various concentration combinations of the honeysuckle fraction and thick foil fraction confirmed in Experimental Example 2-1. (a) is Staphylococcus aureus, (b) Bacillus subtilis , (c) Escherichia coli , (d) P. aeruginosa , (e) Yeasts ( C. albicans ), and (f) are the results for black mold ( A. niger ).
Figure 2 shows the number of viable cells with time for Bacillus subtilis identified in Experimental Example 2-1. Controls were treated with methyl paraben and treated with honeysuckle fractions alone (Lonicera japonica), thickened fractions alone (Magnolia Obovata), and combinations of honeysuckle fractions and thickened fractions. Respectively.
Figure 3 is a TLC result for the component analysis of each of the honeysuckle fraction and thick thin fractions identified in Experimental Example 3-1.
Figure 4 shows the inhibition ring of each individual component, confirmed in Experimental Example 3-2-1. MP is methylparaben used as a control, EA represents each ethyl acetate fraction.

Hereinafter, exemplary embodiments will be described in detail with reference to the accompanying drawings. However, various changes may be made to the embodiments so that the scope of the patent application is not limited or limited by these embodiments. It is to be understood that all changes, equivalents, and substitutes for the embodiments are included in the scope of rights.

The terminology used herein is for the purpose of description and should not be construed as limiting. Singular expressions include plural expressions unless the context clearly indicates otherwise. In this specification, terms such as "comprise" or "have" are intended to indicate that there is a feature, number, step, action, component, part, or combination thereof described on the specification, and one or more other features. It is to be understood that the present invention does not exclude the possibility of the presence or the addition of numbers, steps, operations, components, components, or a combination thereof.

Unless defined otherwise, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art. Terms such as those defined in the commonly used dictionaries should be construed as having meanings consistent with the meanings in the context of the related art and shall not be construed in ideal or excessively formal meanings unless expressly defined in this application. Do not.

In addition, in the description with reference to the accompanying drawings, the same components regardless of reference numerals will be given the same reference numerals and duplicate description thereof will be omitted. In the following description of the embodiment, when it is determined that the detailed description of the related known technology may unnecessarily obscure the gist of the embodiment, the detailed description thereof will be omitted.

Throughout this specification, unless otherwise indicated, “%” used to indicate the concentration of a particular substance is solid / solid (weight / weight)%, solid / liquid (weight / volume)%, and Liquid / liquid is (volume / volume)%.

Substances used in the experiment

Commercial solvents such as ethanol (EtOH), methanol (MeOH), ethyl acetate (EtOAc) and the like were used. Caffeic acid and luteolin used as comparative materials were Sigma Chemical Co. (Magnolol) and honokiol were purchased from ActivON. The honeysuckle used for the experiment was purchased at Gyeongdong Market in Seoul in April 2017, and Hubak was purchased from Meadow Herb Plus in November 2017.

Preparation Example: Preparation of Extracts and Fractions of Honeysuckle and Twig

400 g of dried honeysuckle was pulverized, immersed in 8 L of 50% ethanol for 24 hours, and filtered. The filtrate was dried under reduced pressure and powder was obtained. The ethyl acetate fraction obtained by distilling 50% ethanol extract three times with ethyl acetate was concentrated under reduced pressure to obtain a powder.

300 g of dried hubak was finely pulverized, stirred and immersed in 3 L of 70% ethanol at 30 ° C. for 24 hours, and then filtered. The filtrate was dried under reduced pressure and powder was obtained. The ethyl acetate fraction obtained by fractionating 70% ethanol extract three times with ethyl acetate in a 1: 1 ratio was concentrated under reduced pressure to obtain a powder.

The yield of 50% ethanol extract obtained using 400 g of dried honeysuckle is 4.85% and the yield of ethyl acetate fraction is 0.41%. The yield of 70% ethanol extract obtained using 300 g of dried thick gourd is 14.52% and the yield of ethyl acetate fraction is 4.47%.

Table 1: Extraction yield of honeysuckle

menstruum Yield (%, w / w) * 50% EtOH Extract 4.85 EtOAc fraction 0.41

* Yield (%, w / w) = (weight of dried extract / weight of dried raw material) x 100

Table 2: Extraction yield of thick cakes

menstruum Yield (%, w / w) * 70% EtOH Extract 14.52 EtOAc fraction 4.47

Experimental Example 1: Antimicrobial Activity of Individual Extracts and Fractions

1-1. Bacterial culture

The antimicrobial activity of 6 strains included in cosmetic microbial regulation was evaluated by using honeysuckle and extract of Tak. Specifically, S. aureus ATCC6538, Bacillus subtilis ATCC19659, E. coli ATCC23736, P. aeruginosa ATCC29336, Yeast ( C. albicans ) ATCC10231 and Black mold ( A. niger) ATCC16404 was distributed by Korea Microbial Conservation Center (KCCM, Seoul, Korea). S. aureus , B. subtilis , E. coli , P. aeruginosa and C. albicans used tryptic-soy medium (Merck) and incubated for 24 hours in a 37 ° C. incubator after inoculation. . For A. niger , Potato-Dextrose medium (Merck) was used and incubated for 48 hours in a 30 ℃ incubator after inoculation.

1-2. Antimicrobial Activity Evaluation Method-Minimum Inhibitory Concentration (MIC) and Minimum Sterilization Concentration (MBC)

In order to measure Minimum Inhibitory Concentration and Minimum Bactericidal Concentration, each strain was adjusted to 3 ~ 5 × 10 ⁶ colony-forming units (CFU) / ml. Used. Methylparaben was used as a control. Samples were used after diluting with DMSO using a 2-fold dilution method and cultured by injecting 20 μl of sample or 20 μl of methyl paraben, 20 μl of bacterial solution, and 160 μl of medium into a 96 well plate. When observed visually after cultivation, the concentration at which each bacteria were not grown was determined as the minimum inhibitory concentration, and the concentration without colonies was determined as the minimum bactericidal concentration by plating on a plate medium with a sterile swab.

1-3. Individual antimicrobial activity evaluation of each extract and fraction

According to the method described in Experimental Example 1-2, the minimum inhibition concentration (MIC) and the minimum sterilization concentration (MBC) were evaluated. The experimental results are shown in Table 3.

Strain MIC (MBC) (μL / ml) Gram positive bacteria Gram negative bacteria leaven Mold S. aureus B. subtilis E. coli P. aeruginosa C. albicans A. niger Honeysuckle
50% EtOH Extract 5000
(10000) -
(-) -
(-) 5000
(-) -
(-) -
(-) Honeysuckle EtOAc Fraction 78
(312) 10000
(10000) 5000
(5000) 312
(1250) 1250
(10000) 10000
(-) Thick 70% EtOH Extract 78
(156) 2500
(2500) 2500
(5000) 1250
(1250) 1250
(5000) 10000
(-) Thick EtOAc fraction 39
(78) 2500
(5000) 2500
(2500) 625
(625) 625
(625) 2500
(-) Methyl paraben 2500 (5000) 2500
(10000) 1250
(5000) 1250
(1250) 1250
(5000) 625
(5000)

Phosphorus 50% ethanol extract showed antimicrobial activity only in S. aureus and P. aeruginosa . However, it did not show high antimicrobial activity compared to methylparaben used as a control. On the other hand, the ethyl acetate fraction of Phosphorus vinegar showed antimicrobial activity in all strains, especially higher than methylparaben in S. aureus and P. aeruginosa . Yeast (C. albicans) also showed antimicrobial activity similar to that of methyl paraben.

Thick leaf extract showed effective antimicrobial activity in 6 strains of both 70% ethanol extract and ethyl acetate fraction. In particular, S. aureus , Bacillus subtilis , P. aeruginosa , and C. albicans showed similar or higher antimicrobial activity. In other strains except B. subtilis and E. coli , the antimicrobial activity of 70% ethanol extract and ethyl acetate fraction was found to be about 2 to 4 times different.

Experimental Example 2: Antimicrobial Synergy Activity of Honeysuckle and Twig Extract

The antimicrobial synergistic effect between the two extracts was confirmed by using ethyl acetate fractions of the phosphorus vine and the groin, which showed antibacterial activity in all 6 strains. The checkerboard test was used to identify the antimicrobial synergies. Specifically, the FIC index was used to determine synergistic, additive, and irrelevant effects. In addition, Kill-time analysis was performed using the concentration confirming the synergistic effect of Bacillus subtilis among the six strains.

2-1. Checkboard test

10 μl of the ethyl acetate fraction of each of the phosphorus vine and the thick foil was mixed and used as a sample. Except for this point, the same procedure as in the evaluation of the minimum inhibitory concentration of Experimental Example 1-2 was performed. At this time, all wells in the Checkerboard test included two samples in different concentration combinations. After diluting sample A by 2 times to make a dilution, sample B diluted 2 times was added to make a concentration combination and incubated. After incubation, the compounding ratio of the concentration at which bacterial growth was not visually confirmed was determined. This was confirmed using Fractional inhibition concentration (FIC) and FIC index.

Synergistic effects when FIC index ≦ 0.5; An additive effect when 0.5 <FIC index ≤ 1; Independent if 1 <FIC index ≦ 4; If the FIC index> 4, it was determined that there is an antagonistic effect.

Experimental results confirming the antimicrobial activity against all six strains are shown in FIG. As a result, the antimicrobial synergistic effect was confirmed in Staphylococcus aureus (a), Bacillus subtilis (b) and yeast (e).

Specifically, for Staphylococcus aureus, the FIC value was 0.5 for 20 ppm and 10 ppm thick, indicating that it had an antimicrobial synergistic effect. In addition, the four concentration combinations showed additional effects. In the case of Bacillus subtilis, the FIC value of 0.5 was obtained at 2500 ppm of the honeysuckle, and 625 ppm of the thick vines. Two mixed concentration combinations showing total antibacterial synergy effect and eight combinations showing additional effect were found. The yeast showed a FIC value of 0.5 at 313 ppm and 156 ppm of honeysuckle, and three and seven concentration combinations representing total antimicrobial synergy and additional effect combinations, respectively. Antimicrobial synergistic effects of honeysuckle and peppermint extracts against E. coli and black mold were not observed, but additional effects were found at some concentration combinations. Pseudomonas aeruginosa did not show synergistic and additive effects. Therefore, it was confirmed that the mixture of the phosphorus vine and the thick ethyl acetate fraction showed antimicrobial synergistic effect in Staphylococcus aureus, Bacillus subtilis, and Yeast.

2-2. Kill-time analysis

As a method of measuring the bacterial count of the culture medium added with the sample over time, the experiment was carried out by selecting a specific concentration based on the results of the checkerboard test. At this time, the strain was selected among the Staphylococcus aureus, Bacillus subtilis, Yeast, which showed the antimicrobial synergistic activity, the Bacillus subtilis showing the most synergy and added effect.

0.5 ml of the sample was added to 4.5 ml of the bacterial solution suspended at 10 ⁵ CFU / ml, and then cultured for 18 hours at 3 hour intervals to confirm the change in the number of bacteria. The number of bacteria cultured for a certain time was confirmed after culturing by diluting with physiological saline until visual confirmation.

The control group was treated only with DMSO, and the sample of phosphorus vinegar acetate acetate was 2500 ppm, the thick ethyl acetate fraction was 625 ppm, and the combination of phosphorus vine and peppermint extract was 2500 ppm and 625, respectively. Each was processed to ppm. The number of viable cells over time was confirmed. The experimental results are shown in FIG.

As a result, when the honeysuckle extract was added, the bacteria gradually increased over time. In addition, the bacterium gradually increased with time, but showed a slightly lower bacterial count compared to honeysuckle. Both the honeysuckle and the hunch extract showed a lower bacterial count than the control group, but the antimicrobial effect of the extract itself was confirmed. On the other hand, the addition of a mixture of honeysuckle and thick gourd extract showed about 10 times lower bacterial counts than the addition of the extract, respectively, and it can be seen that the antibacterial effect is increased when the two extracts are used in combination. Therefore, the antimicrobial synergistic effect of honeysuckle and peppermint extract against Bacillus subtilis was confirmed by Kill-time analysis.

Experimental Example 3: Evaluation of Components of Honeysuckle and Thick Extract

3-1. Component analysis of each extract

The ethyl acetate fraction in the honeysuckle and thick leaf extract was dissolved in 100% ethanol, filtered using a syringe filter (Milliopore 0.45 μm), and used for the polar TLC (normal phase) analysis using the filtered extract solution. The developing solvent used for the TLC analysis is shown in Table 4. The components were identified by spectroscopic data already reported, ultraviolet rays, NP-PEG (natural products-polyethylene glycol, 2-aminoethyl diphenylborinate), and the color of the band using vanillin-sulfuric acid coloration. TLC chromatogram results are shown in FIG. 3.

Elution system Honeysuckle Hexane: ethyl acetate: acetic acid = 21: 14: 5 (v / v) Thick Benzene: methanol = 27: 1 (v / v)

TLC chromatogram analysis of the phosphorus ethyl acetate fraction showed four bands when NP was developed. The darkest bands were caffeic acid and luteolin, respectively. Caffeic acid, a phenolic compound, is mainly found in herbs such as coffee beans, fruits, thyme, etc., and has antioxidant and anticancer effects. Luteolin is a flavone structure, which contains not only honeysuckle, but also peanut shells, moths, chrysanthemums, parsley and celery. It has antioxidant, anti-inflammatory and cell protective effects.

TLC chromatogram analysis of the thick ethyl acetate fraction showed two thick bands when sulfuric acid was developed. This was shown as magnolol and honokiol, respectively. Magnolol and Honokiol are classified as lignans and exist as isomers of each other. It is found mainly in the bark of magnolia and has antioxidant and antibacterial effects.

3-2. Evaluation of antimicrobial activity of individual components in each extract

The antimicrobial activity of each extract identified through the component analysis was evaluated to confirm the antimicrobial activity relationship between the extract and the single component. Antimicrobial activity was assessed by disc diffusion assay and minimal inhibitory concentration.

3-2-1. Disc diffusion assay

The antimicrobial activity of the extract according to each extraction condition was measured by disc diffusion assay in the strain. The cultured strain was used after adjusting to 3 ~ 5 × 10 ⁶ CFU / ml. Strains cultured in plate medium were prepared by smearing 100 μl using a sterile swab, and the sample was slowly absorbed into a paper disc (diameter 8 mm, Roshi kaisha. Ltd., Tokyo, Japan) and dried. The solvent was volatilized. At this time, luteolin was 0.05 mg per disc, and the remaining material was 5 mg per disc. After incubating the paper disks in which each sample was absorbed onto the plated plate medium, the antimicrobial activity was compared by measuring the inhibitory ring (clear zone, mm) generated around the disks.

As a result of the disc diffusion experiments of the honeysuckle and Thubark extracts for the six strains, Pseudomonas aeruginosa, the strain showing the most obvious inhibition ring, is shown in FIG. 4. In Pseudomonas aeruginosa, 50 mm ethanol extract and ethyl acetate fraction showed 8 mm and 9 mm inhibition rings, respectively, which showed less activity than methylparaben (18 mm), but caffeine, a component of honeysuckle, was 10 mm inhibitory ring. Luteolin did not show an inhibitory ring. The thick extract showed less activity than the control methylparaben (16 mm), but the inhibitory ring of 11 mm and 10 mm in the extract of 70% ethanol and ethyl acetate fraction, respectively. In addition, magnolol and honokiol, which are the main components of the extract, showed no inhibitory ring.

3-2-2. MIC assessment

In order to evaluate the exact antimicrobial activity of the single components of the honeysuckle and Thubark extract, the minimum inhibitory concentration was evaluated. The experimental results are shown in Table 5.

Strain MIC (μL / ml) Gram positive bacteria Gram voice bacteria leaven Mold S. aureus B. subtilis E. coli P. aeruginosa C. albicans A. niger Methyl paraben 1250 1250 1250 1250 1250 625 Caffeic acid 312 5000 5000 2500 5000 > 10000 Luteolin 25 200 200 50 200 200 Mixture of magnolol and honokiol (1: 1) 31.2 10000 10000 1250 5000 39

Caffeic acid showed high antimicrobial activity in Staphylococcus aureus and Pseudomonas aeruginosa, and also in Bacillus subtilis, Escherichia coli, and yeast. However, black mold did not show antimicrobial activity even at high concentrations. On the other hand, luteolin showed antibacterial activity against all strains at low concentrations, especially against Staphylococcus aureus and Pseudomonas aeruginosa. This suggests that luteolin did not show inhibitory ring in the previous disc diffusion assay because it was treated at low concentration. Phosphorus 50% ethanol extract showed antimicrobial activity in Staphylococcus aureus and Pseudomonas aeruginosa (Table 3), which can be predicted by relatively polar caffeic acid activity. Antibacterial activity of the phosphorus ethyl acetate fraction was shown in 6 strains, which can be expected as a result of the influence of luteolin, a nonpolar component.

Magnolol and Honokiol showed antimicrobial activity in all strains, especially in Staphylococcus aureus, Pseudomonas aeruginosa and Black mold. Through this, it can be estimated that the antimicrobial effect of the extract of habakku on six strains is a result of the influence of magnolol and honokiol. In addition, the antimicrobial synergistic effect of the extract can be predicted by the interaction between magnolol, honokiol and other components through the superior effect in the extract than the antimicrobial effect of a single component.

Although the embodiments have been described with reference to the accompanying drawings, those skilled in the art may apply various technical modifications and variations based on the above. For example, the described techniques may be performed in a different order than the described method, and / or components of the described systems, structures, devices, circuits, etc. may be combined or combined in a different form than the described method, or other components. Or even if replaced or substituted by equivalents, an appropriate result can be achieved.

Therefore, other implementations, other embodiments, and equivalents to the claims are within the scope of the following claims.

Claims (5)

An antimicrobial composition comprising a honeysuckle extract and a thick gourd extract, wherein the honeysuckle extract comprises rufeoline and caffeic acid as active ingredients, and the thick gourd extract comprises honokiol and magnolol as active ingredients,
The composition is an antimicrobial composition having an antimicrobial activity against any one selected from the group consisting of Staphylococcus aureus , Bacillus subtilis , and fungi yeast ( Candida albicans ).
A cosmetic composition comprising a honeysuckle extract and a thick gourd extract, wherein the honeysuckle extract contains rufeoline and caffeic acid as active ingredients, and the thick gourd extract contains honokiol and magnolol as active ingredients,
The composition is a cosmetic composition having an antimicrobial activity against any one selected from the group consisting of Staphylococcus aureus , Bacillus subtilis , and fungi yeast ( Candida albicans ).
The antimicrobial composition of claim 1 wherein the extract is a water, methanol, ethanol, propanol or butanol extract.
delete According to claim 1, The honeysuckle extract; And the thick extract extracts each comprising a weight ratio of 10: 1 to 0.1: 1.

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