KR101406808B1 - Skin care method, skin care composition, and dried microbial cell - Google Patents

Abstract

The present invention provides a cosmetic method, a composition for skin care, and a dried cell which can be suitably used in the production of the composition, which can easily respond to individual differences in the skin environment and selectively reproduce the bacterium.
A step of selectively cultivating a bacterium useful for restoration of the skin of the skin of the skin of the individual, a step of selectively growing the bacterium of the bacterium and / A cosmetic method comprising the step of administering the production material to the skin of the individual, a skin care composition comprising a bacterium which is selectively propagated as described above and / or a production material thereof.

Description

(SKIN CARE METHOD, SKIN CARE COMPOSITION, AND DRIED MICROBIAL CELL)

The present invention relates to a cosmetic method, a composition for skin care, and dried cells suitably used for the production thereof.

It is known that on the surface of human skin, several dozen kinds of skin fungi are on the surface. Some of the inhabits of the skin exist harmful to human body such as Staphylococcus aureus. On the other hand, useful microorganisms useful for maintaining the skin condition are also present, and it is useful for restoring the skin to maintain the microbes in a normal state. For example, Staphylococcus epidermidis, by producing glycerin, contributes greatly to the maintenance of the moisturizing and barrier functions of the skin, and by producing organic acids and maintaining the surface of the skin slightly acidic, (See, for example, Non-Patent Document 1).

It has become useful to keep the surface of the skin clean to prevent the propagation of harmful bacteria in order to prevent infectious diseases such as food poisoning and influenza and to prevent skin diseases such as acne. Therefore, a detergent such as a soap is used. However, if these detergents are used excessively, moisturizing components of the skin such as ceramides may be lost, and the bacterium may also be killed, which may be adversely affected.

Although the presence ratio of each species of the mycelia in the epidermis is affected by various factors, if the growth of the microorganisms is inhibited by any factor, the microorganisms may predominate and cause various diseases. Therefore, in order to relieve the skin flora, researches and developments have been actively conducted on compounds that selectively exhibit a bactericidal action or a bacteriostatic action against only harmful bacteria without affecting the effective bacteria. For example, Japanese Patent Application Laid-Open No. H05-338994 discloses a method for producing a phytolitic extract which is effectively effective only in acnes fungus grown in the skin of the skin, and which does not act on aerobic bacteria, As a component. Patent Document 2 discloses a skin milky lotion improving agent containing a plurality of sugar-linked water-soluble saccharides. Patent Document 3 discloses an ecosystem balance adjusting agent of skin-oncobacteria containing a hinokitiol glycoside as an active ingredient for bacteriolytic and sterilizing Staphylococcus aureus and proliferating Staphylococcus epidermidis. Patent Document 4 discloses a method for producing Staphylococcus epidermidis selected from Ludwigia octovalvis and plant extracts of Pandanus boninensis as an active ingredient having no bactericidal action against Staphylococcus epidermidis and having a sterilization or proliferation inhibiting action against Staphylococcus aureus , And an ecosystem balance adjuster for skin-damaging bacteria containing a wastewater.

Japanese Patent Application Laid-Open No. 2010-202604 Japanese Patent Application Laid-Open No. 2008-50322 Japanese Patent Application Laid-Open No. 2007-246411 Japanese Patent Application Laid-Open No. 2007-153800

Itaru Deco and Hattori Masahira, "100-Shell Broths Living Together with the Human Body," Newton (Newton Press Publishing Co., Ltd.), Nov. 2010, 68-75

However, in the methods of Patent Documents 1 to 4 in which the skin flour microflora is resurfaced by administering a compound of extrinsic nature, the skin flora is constituted by acquiring the drug resistance in the harmful microorganisms, There is a possibility that the effect of restoration can not be exerted when the susceptibility of the drug is changed due to mutation of each species. With this change, it is very difficult to tailor the optimal drug among a plurality of candidate compounds.

SUMMARY OF THE INVENTION The present invention has been made in view of the above problems, and it is an object of the present invention to provide a cosmetic method, a composition for skin care, and a method for manufacturing the same, which can easily adapt to individual differences in skin environment, And to provide a dried cell body which can be used for producing a cell.

That is, the present invention provides a cosmetic method according to the following [1] to [5], a composition for skin care according to [6] to [9], and dried cells according to [10] to [13] In order to solve the problem.

[1] A method for culturing a skin of a bacterium, comprising the steps of: a step of picking up a skin topical bacterium on the skin surface of an individual; a step of selectively growing an effective bacterium useful for the skin rejuvenation of the skin of the skin topical bacterium; And administering the production material to the skin of the individual.

[2] A process for selectively proliferating the above-

A preliminary culturing step of culturing the above-obtained skin-derived bacteria on a culture medium to form colonies;

A selection step of identifying the colonies and selecting the bacterium;

The method for cosmetic treatment according to the above [1], which comprises a pure culturing step of pure culturing the selected bacterium.

[3] The cosmetic method according to the above [1] or [2], wherein the skin onion is sprouting on the skin surface of the individual in good condition.

[4] The cosmetic method according to any one of [1] to [3], wherein the bacterium is a coagulase negative Staphylococcus aureus.

[5] The method according to [4], wherein the bacterium is Staphylococcus epidermidis.

[6] A composition for skin care comprising a bacterium useful for the restoration of the individual's skin and / or a product thereof, which is selectively proliferated by being separated from the skin flora obtained from the skin surface of an individual.

[7] The composition for skin care according to the above [6], wherein the skin-oncoprotein sprouts on the healthy skin surface of the individual.

[8] The skin care composition according to the above [6] or [7], wherein the bacterium is a coagulase negative staphylococcus.

[9] The skin care composition according to the above [8], wherein the bacterium is Staphylococcus epidermidis.

[10] A dried microbial cell obtained by selectively proliferating an effective microorganism useful for restoration of the skin of the individual among the microbial cells collected from the skin surface of an individual, followed by drying treatment.

[11] The dried cell according to the above [10], wherein the skin onion fungi reproduce on the skin surface of the individual in a healthy state.

[12] The dried cell according to [10] or [11], wherein the bacterium is a coagulase negative staphylococcus.

[13] The dried cell according to [12], wherein the bacterium is Staphylococcus epidermidis.

The fungi of the skin taken from the individual's skin are adapted to the individual differences of the skin environment and are particularly susceptible to proliferation under the skin conditions inherent to the individual. For this reason, when a person who has selectively proliferated only the avian influenza strains picked up on the skin surface of an individual is administered to the skin surface of the individual, the propagation of the harmful bacteria can be effectively inhibited by being dominant on the skin surface, So that the balance of the superficial fungus can be appropriately maintained. Thus, the cosmetic methods and skin care compositions provided by the present invention are useful for maintaining skin conditions healthy, regardless of individual differences in skin conditions. In the cosmetic method and the skin care composition of the present invention, since no medicament is used, it is safe.

Hereinafter, an embodiment of the present invention will be described.

A cosmetic method according to one embodiment of the present invention (hereafter abbreviated as "cosmetic method" in some cases) includes a step of collecting skin-derived bacteria that reproduce on the surface of a person's skin, A step of selectively proliferating an effective bacterium useful for the restoration of an individual's skin and a step of administering to the individual's skin a composition for a skin care comprising the proliferated bacterium of the bacterium and / or the production material thereof.

The skin care composition used in the cosmetic method comprises a bacterium and / or a production material thereof selectively isolated from the skin supernatant collected from the skin surface of an individual, And a step of selectively proliferating only the avian influenza in the collected skin flora. Hereinafter, each step of the production of the composition for skin care will be described in more detail.

(1) Picking up the skin

On the surface of the skin, there are dozens kinds of skin topical bacterium. As the bacterium used in the preparation of the composition for skin care, any one of them may be used alone or in combination with any two or more May be suitably used in combination. Staphylococcus epidermidis can be exemplified as a skin topical bacterium suitable for use in the preparation of a composition for skin care.

The skin fungus is present on the surface of the skin of the whole body but is particularly present on the surface of the face or armpit. Therefore, it is preferable to collect them from these sites. Particularly, in the case of using a composition for skin care as a cosmetic, it is particularly preferable to collect the skin topical bacterium from the skin surface of the face. Since it is considered that the balance of the skin suppositories is appropriately maintained on the healthy skin surface, it is preferable that the skin suppositories are collected on a healthy skin surface.

Collection of the skin microbes can be carried out using any known method. For example, there is a method of collecting by wiping the surface of the skin with a sterilized cotton swab soaked with physiological saline solution or buffer solution, a method of pressing the sterilized agar medium on the surface of the skin and transferring the skin microbes to the surface of the medium. Since the ratio of the number of species and the number of species contained in the supernatant is an index for diagnosing the health condition of the skin, it is preferable that the supernatant is collected at a certain area on the skin. In the case of collecting the skin-damaging bacteria by a method of wiping the surface of the skin with a sterilizing cotton swab, in order to keep the area wiped with the sterilizing cotton swab constant, a synthetic resin, glass, or the like, Or a prismatic sampling tool may be gently adhered to the skin, and the inside of the opening may be scoured to wipe the skin-derived bacteria. By this method, it is always possible to collect the skin-damaging bacteria from the skin surface within a predetermined area range.

On the other hand, when the sterilized agar medium is squeezed onto the surface of the skin, the skin-infective bacteria are collected. By using the medium having the predetermined area (radius) Can be collected. Examples of the medium that can be used for the collection of the skin fungi include selective agar medium for staphylococci such as standard agar medium, tryptic case medium agar medium for egg yolk, mannit salt medium for egg yolk addition, In order to contact the surface of the skin, it is preferable to use a medium having a composition which does not contain a substance likely to act as an allergen.

(2) selective proliferation of bacilli

As described above, the skin uptake bacteria collected from the surface of the skin also contain harmful bacteria. Of these, only the desired bacterium is selectively propagated. As a selective proliferation method, there are a method of separating and culturing only the desired effective bacteria, and a method of culturing under the condition that only the desired bacteria are grown. Any method may be used.

As an example of the former method, the step of selectively growing an effective microorganism may include the following steps (i) to (iii).

(i) a preliminary culturing step of culturing the collected skin flora on a medium and forming colonies

(ii) Identification of colonies and selection of an effective bacteria

(iii) a pure culture process for purely culturing the selected bacilli

(i) preliminary culture process

The supernatant of the skin collected using a sterilizing cotton swab or the like is diluted with a dilution liquid such as phosphate buffered saline (PBS) or the like to a predetermined magnification (for example, 10 to 100,000 times), and is applied to the surface of the culture medium for culture , And cultivation is carried out. The culture conditions are appropriately set in accordance with the kind of the bacterium for the purpose of separation and the conditions of the medium to be used.

In addition, in the case of collecting the inoculum of the skin by pressing the culture medium on the surface of the skin, the preliminary culture process may be carried out directly on the surface of the culture medium that has been collected, but it is preferable to use a sterilized cotton swab Diluted, applied onto the surface of the culture medium for culture, and pre-cultured using the same procedure as described above. Examples of the medium for culture include a medium for general bacteria such as a tryptic case small agar medium (TS medium), a medium for selective separation for the above staphylococci, and the like.

(ii) Selection process

In the preliminary culturing step, the colonies formed on the culture medium are identified and the selection of the bacterium is carried out. The selection process is performed, for example, in the following procedure. First, the culture medium is visually observed, the number of colonies is counted, and preliminary classification is performed by appearance if possible. Then, the identification of the species is carried out for each of the individual colonies or representative ones of them. For example, in the case of Staphylococcus epidermidis, identification is performed by combining microscopic observation, Gram stain and coagulase activity. Especially, since Staphylococcus epidermidis is a negative coagulase (CNS) unlike pathogenic bacteria such as Staphylococcus aureus, the measurement of the activity of coagulase is important for discrimination from Staphylococcus aureus.

(iii) a pure culture process

As described above, the bacterium selected (separated) from the submerged bacteria is cultivated in a pure manner. Since the bacterium obtained in this manner is considered to be adapted to the skin environment of an individual who has taken the skin biocide, it is contained in the composition for skin care and is returned to the skin of an individual, thereby predominantly growing on the surface of the skin, And the effect of inhibiting the growth of harmful bacteria can be expected. The conditions of the culture or the culture used for the pure culture are appropriately set in accordance with the kind of the bacterium for the purpose of separation and the conditions of the medium to be used as in the case of the preliminary culture process.

(3) Preparation of composition for skin care

When the composition for skin care is prepared, the bacterium selectively propagated as described above can be directly used. However, since it is difficult to keep the bacterium in a long-term storage state, dried bacteria of the bacterium are prepared, And dispersing the composition in a base containing water to restore the bacterium and to prepare a skin care composition containing the surviving bacteriostatic bacteria.

The base material of the composition for skin care is not particularly limited as long as the bacterium can survive and / or propagate therein, and the base material of the skin care composition may be any of special cosmetics such as lotion, milky lotion, serum, gel, cream, lotion, A hair care product, a hair tonic, a hair lotion, a hair growth agent, a hair treatment, a hair conditioner, a hair pack, a scalp care article such as a hair dye, or a hair cosmetic. The blending amount of the bacterium per unit volume is appropriately selected according to the type of the substrate and the bacterium of the bacterium.

As a method of producing the dried cells, any known method can be used without particular limitation, and specific examples thereof include a freeze drying method and a hot air drying method. For example, in the case of producing dried cells by the freeze-drying method, useful microorganisms are dispersed in a dispersion medium (usually water is used) in the presence of a suitable protecting agent such as skim milk or soy milk, frozen with liquid nitrogen or the like, The water is removed under reduced pressure with a freeze dryer. The dried cells thus obtained can be stored in sterilized ampoules or the like.

In the case where the dead cells of the bacterium of the bacteriostatic organism or the material for producing the bacterium of the bacterium have a specific proliferation promoting activity against the bacteriostatic organism, the skin care composition may contain them instead of the live bacteria of the bacterium. Instead of feeding live bacteria of Bacillus thuringiensis, it is possible to increase the ratio of bacillus to skin lacunae by supplying a substance that specifically promotes the growth of Bacillus bacillus on the surface of the individual to the skin of the individual, This is because it is possible to maintain soundness. When a bacterium of Bacillus subtilis or a production material of Bacillus subtilis is used in place of the live bacteria of Bacillus subtilis, the composition for skin care may be a skin cleanser such as a shampoo, a body shampoo and a head sponge containing a surfactant.

The composition for skin care may contain any additive commonly used in cosmetics and the like insofar as it does not inhibit the production and / or proliferation of the live bacteria of the bacterium. Specific examples of such additives include flavoring agents, preservatives, antioxidants, bactericides (those having no antimicrobial activity against bacteriostats or limited to weak ones), emulsifiers, and stabilizers. The composition for skin care may further comprise a substance that specifically promotes the proliferation of avian bacteria.

By administering a skin care composition containing the bacterium of the above-mentioned bacterium and / or the production material thereof to the skin of an individual, the bacterium which is adapted to the environment of the surface of the individual skin rapidly proliferates from the surface of the skin, Can be improved. In such a cosmetic method, as a method of administering the composition for skin care, any method depending on the properties of the composition for skin care, such as application of a liquid preparation such as lotion, lotion, lotion, spraying of a spray agent, have. The administration may be performed using a finger or the like, but may be performed using a device such as a sprayer or an applicator.

(Example)

Hereinafter, examples for the purpose of confirming the effects of the present invention will be described.

[1] Picking up the skin

The following operation was performed while wearing sterile gloves.

After washing, a cylindrical (25 mmφ) cylinder made of polycarbonate sterilized in an autoclave was lightly pressed against the surface of the subject's skin, and the surface of the skin exposed inside the cylinder was immersed in phosphate buffered saline (PBS) for 20 seconds It was thoroughly wiped with a damp sterile swab. The cylinder was removed and the remaining water on the skin surface was wiped off with a sterile cotton swab. Moisture adhered to the inner side of the skin contact surface of the cylinder was similarly wiped with a sterilizing cotton swab. Place these sterile swabs in the PBS-containing tube (the part of the donor's hand touching them), put the cap on it, and move it into the clean bench.

[2] preliminary culture

100 占 퐇 of the mycelial liquid obtained as described above was added to a microtube containing 900 占 퐇 of PBS and subjected to swirling to prepare a 10-fold diluted solution. The same procedure was repeated using a diluent instead of the bacterial solution to prepare a 100 to 100,000-fold diluted solution. Each of the obtained dilutions was added to a tryptic case small agar medium (TS medium) and a selective medium for Staphylococcus aureus (hereinafter referred to as " selective medium "), uniformly spread and cultured at 37 DEG C for 48 hours.

[3] Isolation of Staphylococcus epidermidis

The colonies formed on the selective medium were visually observed, and each colony was numbered. One drop of deionized water was placed on the slide glass, and the cells collected from the colonies were dispersed using a platinum wire, and then baked with a gas burner on the back side, and the cells were flame-fixed on the surface of the slide glass. To this, a crystal violet solution, an iodine solution, and a Bipel solution were sequentially reacted to perform Gram stain. Thereafter, the cells contained in each colony were observed under a microscope for dyeability and morphology. Among them, Gram-positive staphylococci were cultured on selective medium (37 ° C, 48 hours). In addition, when the species was identified using an API Staph plate, it was confirmed that it was Staphylococcus epidermidis.

[4] Pure culture of Staphylococcus epidermidis and production of dried cells

The coagulase activity was negative (CNS) according to the above measurement, and pure culture was carried out on a selective medium. Cells were collected from the obtained colonies using platinum ears, dispersed in skim milk as a protecting agent, injected into an ampoule, and rapidly frozen at a very low temperature of -80 캜. This was placed in a freeze dryer to obtain a dried cellulosic powder.

[5] restoration of dried cells

The ampoule containing the dried cells was opened and a small amount of a TS liquid medium was added to suspend the dried cells. The obtained suspension was added to a liquid TS medium, a flat-plate TS medium, and a mannitol medium, and cultured at 37 占 폚 for 48 hours. As a result, it was confirmed that bacteria were detected in all media and that colonies were formed on a plate medium. These bacteria were identified, and it was confirmed that it was Staphylococcus epidermidis.

Claims (13)

Claims [1] A method for treating skin, comprising the steps of:
A step of selectively growing an avian bacterium useful for maintaining the moisturizing function of the skin of the individual,
And a step of administering to the skin of the individual at least one of the propagated bacterium of the bacterium or the production material thereof. The method according to claim 1,
The step of selectively growing the above-
A preliminary culturing step of culturing the above-obtained skin-derived bacteria on a culture medium to form colonies;
A selection step of identifying the colonies and selecting the bacterium;
And a pure culture step of purely culturing the selected bacterium. The beauty treatment method according to claim 1 or 2, wherein the skin onion fungi reproduce on the normal skin surface of the individual. The cosmetic method according to claim 1 or 2, wherein the bacterium is a coagulase negative staphylococcus. The method of claim 4, wherein the bacterium is Staphylococcus epidermidis. Which comprises at least one of a bacterium useful for maintaining the moisturizing function of the skin of the individual or a product thereof produced by selectively growing the bacterium from the skin of the skin of the individual and applied to the individual Wherein the composition is applied to a skin. [Claim 7] The composition for skin care according to claim 6, wherein the skin onion fungi reproduce on the normal skin surface of the individual. The skin care composition according to claim 6 or 7, wherein the bacterium is a coagulase negative staphylococcus. 9. The composition for skin care according to claim 8, wherein the bacterium is Staphylococcus epidermidis. The present invention is applied to an individual who has picked up the above-mentioned skin-derived microorganism obtained by selectively growing an effective microorganism useful for maintaining the moisturizing function of the skin of the individual, . 11. The dried microbial cell according to claim 10, wherein the skin-derived microbes reproduce on the normal skin surface of the individual. 12. The dried cell of claim 10 or 11, wherein the bacterium is a coagulase negative staphylococcus. 13. The dried cell of claim 12, wherein the bacterium is Staphylococcus epidermidis.