KR20180034086A - Composition for hair or scalp comprising lysate of Lactobacillus sakei - Google Patents

Abstract

The present specification describes a composition for hair or scalp containing a lysate of Lactobacillus sakei as an active ingredient. The composition can exhibit remarkably excellent hair growth promoting and hair loss preventing effects.

Description

TECHNICAL FIELD The present invention relates to a composition for hair or scalp comprising an eluate of Lactobacillus saccharum Lactobacillus sakei (Composition for hair or scalp comprising lysate of Lactobacillus sakei)

The present specification relates to a composition for hair or scalp comprising an eluate of Lactobacillus sac.

Recently, there is a demand for a material derived from nature according to the tendency to avoid chemical raw materials, insects and animal raw materials. Microbial resources are highly useful materials that utilize inherent characteristics of microorganisms adapted in various environments as sustainable resources in terms of reproducibility. Among them, lactic acid bacteria are used as probiotics in the field of food, which can be used without danger to the human body.

Korean Patent Publication No. 10-2014-0055397

From the viewpoint of one aspect, a problem to be solved by the present invention is to provide a composition excellent in hair growth enhancement and hair loss prevention effect.

In another aspect, a problem to be solved by the present invention is to provide a composition excellent in hair growth and hair loss prevention effect without human toxicity or irritation.

In another aspect, a problem to be solved by the present invention is to provide a composition which is superior in hair growth enhancement and hair loss prevention effect using a naturally derived component.

In another aspect, a problem to be solved by the present invention is to provide a composition which promotes the proliferation of hair follicular dermal papilla cells and has an excellent effect of suppressing the death.

In another aspect, a problem to be solved by the present invention is to provide a composition having an effect of promoting proliferation of outer skin cells and exhibiting an effect of inhibiting its death.

In one aspect, the present invention relates to a composition for hair or scalp comprising, as an active ingredient, a lysate of Lactobacillus sakei .

In another aspect, the present invention relates to a method for producing (a) culturing Lactobacillus sakei ,

(b) centrifuging the cultured Lactobacillus saccharum to remove insoluble components, and

(c) a composition for hair or scalp comprising, as an active ingredient, an eluate of Lactobacillus sacrifice, which comprises eluting the lactobacillus sake obtained in (b) above by heating at 80 to 120 ° C for 10 minutes to 30 minutes And the like.

In one aspect, the present invention can provide a composition having excellent hair growth enhancement and hair loss prevention effect.

In another aspect, the present invention can provide a composition that is excellent in hair growth and hair loss prevention effect without human toxicity or irritation.

From another viewpoint, the present invention can provide an excellent hair growth promoting and hair loss preventing effect using a naturally derived component.

From another viewpoint, the present invention can provide a composition that promotes the proliferation of hair follicular dermal papilla cells and is excellent in the effect of suppressing the death.

In another aspect, the present invention can provide an excellent composition that promotes the proliferation of outer root cells and inhibits its death.

1 is a graph showing the effect of lactobacillus sacae on the promotion of dermal papilla cell proliferation.
FIG. 2 is a graph showing the results of comparing the effect of promoting the proliferation of dermal papilla cells of Lactobacillus sake and its homologous heterologous strain.
FIG. 3 is a graph showing the effect of promoting the proliferation of the root cells of Lactobacillus sac.
Fig. 4 is a graph showing the effect of promoting the proliferation of smooth muscle cells of Lactobacillus sake and an inbred heterologous strain.
FIG. 5 is a graph showing the effect of Lactobacillus casei on the inhibition of appearance-induced apoptotic cell death.
FIG. 6 is a fluorescence microscope photograph showing the effect of Lactobacillus casei on the suppression of apoptotic cell death.

Embodiments of the present application will now be described in more detail with reference to the accompanying drawings. However, the techniques disclosed in the present application are not limited to the embodiments described herein but may be embodied in other forms. It should be understood, however, that the embodiments disclosed herein are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art. In the drawings, the widths and thicknesses of components are slightly enlarged in order to clearly illustrate each component. In addition, although only a part of the components is shown for convenience of explanation, those skilled in the art can easily grasp the rest of the components. It will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the spirit and scope of the invention.

In one embodiment of the present invention, it is possible to provide a composition for hair or scalp containing, as an active ingredient, an eluate of Lactobacillus sakei .

In one example, the Lactobacillus sake may be derived from gold silver. In one embodiment, the Lactobacillus sake may be isolated and identified from flowers of LONICERA japonica .

The eluate of Lactobacillus casei exhibits remarkably superior hair growth and hair loss prevention effect as compared with the strain of heterologous type.

In one embodiment, the eluate may be a heating eluate obtained by heating and eluting cells of Lactobacillus casei. The heating may be carried out at 80 to 120 ° C for 10 minutes to 30 minutes. The heat-dissolving product is a natural ingredient with little irritation or toxicity when applied to a human body, and can be remarkably excellent in hair growth enhancement and hair loss prevention effect.

In one embodiment, the eluate comprises (a) culturing Lactobacillus sakei ,

(b) centrifuging the cultured Lactobacillus saccharum to remove insoluble components, and

(c) heating the Lactobacillus sake obtained in (b) above by heating at 80 to 120 ° C for 10 minutes to 30 minutes.

The composition according to one embodiment of the present invention is excellent in safety because it has little irritation or toxicity when applied to a human body as a natural ingredient. In addition, it promotes the proliferation of hair follicle dermal papilla cells and outer appearance root cells, suppresses its death, and exhibits remarkably hair growth promoting effect and hair loss prevention effect.

In one embodiment of the present invention, the composition may be an external preparation for skin application.

The composition for external application for skin may be, for example, a cosmetic composition.

The composition according to one embodiment of the present invention may be formulated containing a cosmetically or dermatologically acceptable medium or base. These are all formulations suitable for topical application, for example as a solution, a gel, a solid, a paste anhydrous product, an emulsion obtained by dispersing an oil phase in water, a suspension, a microemulsion, a microcapsule, a microgranule or an ionic form (liposome) In the form of ionic fibrin dispersions, or in the form of creams, skins, lotions, powders, ointments, sprays or conical sticks. It can also be used in the form of a foam or in the form of an aerosol composition further containing a compressed propellant. These compositions may be prepared according to conventional methods in the art.

In addition, the composition according to one embodiment of the present invention may contain a skin absorption promoting substance to increase the skin improving effect.

In one embodiment, the cosmetic composition may contain 0.1% to 20% by weight of the lactobacillus saccharide eluant relative to the total weight of the composition.

In one example, the composition may be used as a scalp or hair composition. Thus, the composition can be used in combination with scalp and hair formulations such as shampoo, rinse, scalp treatment, hair treatment, scalp hair treatment, hair loss hair treatment, damaged hair treatment, leave-in ) Conditioners, wash-off conditioners or hair essences, or scalp or hair, scalp and hair products.

The composition of each of these formulations may contain various components, which may be formulated into conventional scalp or hair compositions, according to the various formulations described above, or to the end purpose, and the type and amount of these components may be readily selected by those skilled in the art .

In one embodiment, the composition may comprise from 0.1% to 20% by weight of the lactobacillus saccharide eluant, based on the total weight of the composition.

In addition, the composition according to an embodiment of the present invention may further comprise at least one selected from the group consisting of fatty substances, organic solvents, solubilizers, thickening agents, gelling agents, softening agents, antioxidants, suspending agents, stabilizing agents, foaming agents, Or any other conventionally used in cosmetics such as nonionic emulsifiers, fillers, sequestering agents, chelating agents, preservatives, vitamins, barrier agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, ≪ RTI ID = 0.0 > cosmetics < / RTI > such as botanical ingredients, or adjuvants conventionally used in the field of dermatology. The adjuvants may be introduced in amounts generally used in the cosmetics or dermatological fields, for example from 0.001% to 5% by weight, for example from 0.001% to 3% by weight, based on the total weight of the composition.

In one embodiment of the invention, the composition may be a pharmaceutical composition.

The pharmaceutical compositions according to the present disclosure may be of various oral or parenteral formulations. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid form preparations for oral administration include tablets, pills, powders, granules, soft or hard capsules, etc. These solid preparations may contain one or more excipients such as starch, calcium carbonate, sucrose, Or lactose, gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate, talc, and the like are also used. Liquid preparations for oral administration include suspensions, solutions, emulsions, syrups and the like. Various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included in addition to water and liquid paraffin, which are simple diluents commonly used. have. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used as the non-aqueous solvent and suspension agent. Examples of the suppository base include witepsol, macrogol, tween 61, cacao paper, laurin, glycerogelatin and the like.

The pharmaceutical dosage forms of the compositions herein may be used in the form of their pharmaceutically acceptable salts and may also be used alone or in combination with other pharmaceutically active compounds, as well as in a suitable set. The salt is not particularly limited as long as it is pharmaceutically acceptable so long as it is pharmaceutically acceptable and includes, for example, hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrofluoric acid, hydrobromic acid, formic acid acetic acid, tartaric acid, lactic acid, citric acid, fumaric acid, , Benzenesulfonic acid, toluenesulfonic acid, and naphthalenesulfonic acid.

The composition of the present invention may be administered parenterally or orally, and may be administered in one to several divided doses in an amount of 0.1 to 500 mg, preferably 1 to 100 mg per kg of body weight per day have. The dosage for a particular patient may vary depending on the patient's body weight, age, sex, health condition, diet, time of administration, administration method, excretion rate, severity of disease, and the like.

The pharmaceutical composition according to the present invention may be formulated into various forms such as powders, granules, tablets, soft or hard capsules, oral formulations such as suspensions, emulsions, syrups and aerosols, external preparations such as ointments and creams, suppositories, Sterile injectable solutions, and the like, and may be formulated and used in any form suitable for pharmaceutical preparations.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are merely illustrative of the present invention and that the scope of the present invention is not construed as being limited by these embodiments.

[Preparation Example 1] Preparation of Lactobacillus saccharide eluate

1. Isolation of Lactobacillus casei strain

Lactobacillus sake obtained from Eunhwa was washed twice with primary distilled water to remove foreign matter. The washed ginseng was dipped in distilled water 10 times by weight and reacted at a speed of 250 rpm for 30 minutes with shaking. Decimal dilution of the reaction solution in deionized water of 1 10 -10 ⁵ times, and deep nose Bashile Lactobacillus MRS agar and incubated divides the (Difco Lactobacilli MRS Agar®) medium. At this time, the culture was carried out in a chamber of anaerobic condition at 32 ° C. for 2 days, and a colony showing white colonies was taken. Lactobacillus casei (accession number: KCTC12675BP) was isolated from Ganoderma lucidum in the same manner as above.

2. Culture and elution of Lactobacillus casei

The isolated Lactobacillus sake was stored in a cryotube at 10% by weight in skim milk, and inoculated into a medium (MRS Broth) and cultured at 37 ° C for 24 hours. When the bacterial concentration in the liquid medium reached 5 × 10 ⁸ to 1 × 10 ⁹ CFU / ml, nutrient cells were harvested at 4 ° C by centrifugation at 4000 rpm and washed with PBS (phosphate buffered saline). The washed cells were resuspended in PBS to a concentration of 5 × 10 ⁸ to 1 × 10 ⁹ CFU / mL and heated at 100 ° C. for 10 minutes to obtain an eluate.

[Preparation Example 2] Preparation of eluate of homologous heterologous strain

Lactobacillus brevis (Lactobacillus brevis) and Lactobacillus plantarum (APsulloc 331263, Accession No. KCCM11180P) were heated and eluted by the same method as in Production Example 1 as a strain of Lactobacillus sake, .

[Experimental Example 1] Effect of proliferation of dermal papilla cells

The proliferation effect in human dermal papilla cells (hDPCs) was confirmed by using the eluates prepared in Preparation Examples 1 and 2.

1. Isolation and Culture of hDPCs

The dermal papilla tissues were separated from the scalp tissues using a microscope and cultured for 14 days in a collagen type I coated culture dish. The cells were cultured for 72 hours in a culture medium containing 5% CO _{2 and} 37 ° C in DMEM (Dulbecco's modified eagles medium; HyClone GE Healthcare) supplemented with antibiotics (1%), fungizone After incubation, the medium was replaced with DMEM medium supplemented with 10% FBS. When the cells grew over 90% of the dish, the cells were collected using 0.25% trypsin / 10 mM EDTA (Welgene) The cells were cultured in DMEM medium supplemented with 5% CO _{2 at} 37 ° C for 72 hours (until cells grew to 90% or more of the dish), and were then given to the experiment.

2. Culture for confirmation of hDPCs cell proliferation effect

HDPCs were inoculated into 96 well plates at 2000 cells / well and cultured at 37 ° C in 5% CO ₂ for 24 hours. The eluates prepared in Preparation Examples 1 and 2 were each treated at 10% by volume and cultured at 37 ° C for 5 hours in 5% CO ₂ for 72 hours. As a control group, untreated control group was used, and 5% FBS as a positive control group was treated with the eluate in the same amount.

3. Cell proliferation assay: 120% or more viability

MTT solution (Sigma, 70 μg / well) was added to each well of the 96-well plate and incubated at 37 ° C. for 3 hours in 5% CO ₂ . After that, formazan was dissolved in DMSO and absorbance was measured at 570 nm using a microplate reader. The relative cell proliferation rate of the control group is shown in FIG. 1 and FIG. 2, where the control value is 100.

1 and 2, it can be confirmed that the strain of Preparation Example 1 exhibits excellent hair follicle dermal papilla cell proliferation effect, while the strain of Preparation Example 2 does not show significant effect even when it is a homologous strain of Bacillus sake.

[Test Example 2] Effect of proliferation of root cells

The proliferation effect of outer root sheath keratinocyte (ORS keratinocyte) was confirmed by using the eluates prepared in Preparation Examples 1 and 2.

1. Isolation and culture of outward-looking cells (ORS keratinocytes)

From the scalp tissues of the occipital lobe, the outer skin cells were separated using a microscope and cultured for 14 days in a collagen type I coated culture dish. The cells were cultured for 3 days at 37 ° C in 5% CO _{2 in} a culture medium containing antibiotics (1%), fungizone (0.1%) and DMEM supplemented with 20% FBS (Dulbecco's modified eagles medium (EPILIFE MEPI500CA, Invitrogen). Confluent cells were treated with trypsin / EDTA (T / E) for subculture

2. Cultivation for confirmation of cell proliferation effect of cultured root cells

Cells were seeded at 20000 cells / well in 96 well plates and cultured for 24 hours at 5% CO _{2 and} 37 ° C. The eluate prepared in Preparation Examples 1 and 2 was treated at 10% by volume and cultured at 37 ° C for 5 hours in 5% CO ₂ for 72 hours. As a control group, non-treated group was used. As a positive control group, growth medium (Epilife MEPI500CA, invitrogen) supplemented with growth factor (EpiLife S-012-5, invitrogen) Respectively.

3. Cell proliferation assay: 120% or more viability

MTT solution (Sigma, 70 μg / well) was added to each well of the 96-well plate and incubated at 37 ° C. for 3 hours in 5% CO ₂ . After that, formazan was dissolved in DMSO and absorbance was measured at 570 nm using a microplate reader. Relative cell proliferation rates are shown in FIGS. 3 and 4 with the value of the control (control) being 100 in the measured results.

3 and 4 show that the strain of Preparation Example 1 exhibits a superior appearance root growth cell proliferation effect, while the strains of Preparation Example 2 do not show a significant effect even when they are heterologous strains of Bacillus sake and the same species.

[Test Example 4] Inhibitory effect on the appearance of root cells

Using the eluates prepared in Preparation Example 1, the inhibitory effect of DKK-1-induced outer root sheath keratinocyte (ORS keratinocyte), known as hair loss inducing factor, was confirmed.

1. Cell culture

The cells were cultured in a 96-well plate and an 8-chamber slide in 20000 cells / well at 5% CO _{2 and} 37 ° C for 24 hours. After starvation for 24 hours, the cells were treated with 50ng / ml of DKK and 50ng / ml of DKK + 10% by volume of the eluate prepared in Preparation Example 1, respectively. The control group was a control group and the positive control group (Epilife MEPI500CA, invitrogen) supplemented with growth factor (EpiLife S-012-5, invitrogen)

2. MTT assay

MTT solution (Sigma, 70 ㎍ / well) was added to each well of the 96-well plate treated with the above test substance and incubated at 37 ° C for 3 hours at 5% CO ₂ . After that, formazan was dissolved in DMSO and absorbance was measured at 570 nm using a microplate reader. As a result, the relative cell proliferation rate (I) of the group treated with DKK-1 alone was 100, and the relative cell proliferation rate (I) was 100 when the negative control value was 100.

3. TUNEL assay (8 chamber slide)

The 8-chamber slides were incubated for 24 h, fixed with 4% paraformaldehyde, and washed with 0.05% triton-X100 in PBS. TUNEL reaction mixture (Roche kit), washed and stained with DAPI (4 ', 6-diamidino-2-phenylindole) and observed with fluorescence microscope. A fluorescence microscope photograph of the cells is shown in Fig.

5 and 6, it can be confirmed that the lactobacillus sake of Production Example 1 exhibits a superior suppression effect on smooth muscle cell death.

Formulation examples of compositions according to one aspect of the present disclosure are described below, but are also applicable to various other formulations, which are intended to be illustrative only and not for purposes of limitation.

[Formulation Example 1] Tablets

After mixing 100 mg of the eluate of Production Example 1 of the present invention, 400 mg of lactose, 400 mg of corn starch and 2 mg of magnesium stearate, tablets were prepared by tableting according to a conventional preparation method of tablets.

 [Formulation Example 2]

100 mg of the eluate of Preparation Example 1 of the present invention, 400 mg of lactose, 400 mg of corn starch and 2 mg of magnesium stearate were mixed and filled in gelatin capsules according to the conventional preparation method of capsules to prepare capsules.

[Formulation Example 3]

50 mg of the eluate of Preparation Example 1 of the present invention, 250 mg of anhydrous crystalline glucose and 550 mg of starch were mixed and granulated into granules using a fluidized bed granulator, followed by filling the pellets.

[Formulation Example 4] Drinking agent

50 mg of the eluate of Preparation Example 1 of the present invention, 10 g of glucose, 0.6 g of citric acid and 25 g of liquid oligosaccharide are mixed, 300 ml of purified water is added, and each bottle is filled with 200 ml each. And the mixture was sterilized at 130 DEG C for 4 to 5 seconds to prepare a drink.

[Formulation Example 5]

Injections were prepared according to the compositions shown in the following table in a conventional manner.

Compounding ingredient content The eluate of Preparation Example 1 10-50 mg Sterile sterilized water for injection Suitable amount pH adjusting agent Suitable amount

[Formulation Example 6] Softening lotion (skin lotion)

The softening longevity was prepared by a conventional method according to the composition shown in the following table.

Compounding ingredient Content (% by weight) The eluate of Preparation Example 1 0.2 glycerin 3.0 Butylene glycol 2.0 Propylene glycol 2.0 Carboxyvinyl polymer 0.1 Phage-12 nonylphenyl ether 0.2 Polysorbate 80 0.4 ethanol 10.0 Triethanolamine 0.1 Preservative, coloring, fragrance Suitable amount Purified water Balance

[Formulation Example 7] Nutritional lotion (milk lotion)

Nutrition lotion was prepared according to the conventional method according to the composition shown in the following table.

Compounding ingredient Content (% by weight) The eluate of Preparation Example 1 1.0 glycerin 3.0 Butylene glycol 3.0 Propylene glycol 3.0 Carboxyvinyl polymer 0.1 Wax 4.0 Polysorbate 60 1.5 Caprylic / capric triglyceride 5.0 Squalane 5.0 Sorbitacecequioleate 1.5 Liquid paraffin 0.5 Cetearyl alcohol 1.0 Triethanolamine 0.2 Preservative, coloring, fragrance Suitable amount Purified water Balance

[Formulation Example 8] Massage cream

Massage creams were prepared in a conventional manner according to the composition shown in the following table.

Compounding ingredient Content (% by weight) The eluate of Preparation Example 1 2.0 glycerin 8.0 Butylene glycol 4.0 Liquid paraffin 45.0 Beta Glucan 7.0 Carbomer 0.1 Caprylic / capric triglyceride 3.0 Wax 4.0 Cetearyl glucoside 1.5 Sesquioleic acid sorbitan 0.9 Vaseline 3.0 paraffin 1.5 Preservative, coloring, fragrance Suitable amount Purified water Balance

[Formulation Example 9] Production of hair lotion

A hair lotion is prepared according to a conventional method with the composition shown in the following table.

ingredient Content (% by weight) The eluate of Preparation Example 1 2.0 Purified water Balance glycerin 3.0 Butylene glycol 3.0 Liquid paraffin 7.0 Beta Glucan 7.0 Carbomer 0.1 Edelweiss extract 2.0 Caprylic capric triglyceride 3.0 Squalene 5.0 Cetearyl glucoside 1.5 Sorbitan stearate 0.4 Polysorbate 1.2 antiseptic Suitable amount incense Suitable amount Pigment Suitable amount Triethanolamine 0.1

[Formulation Example 10] Preparation of shampoo composition

A shampoo composition was prepared according to a conventional method with the composition shown in the following table.

Component (% by weight) Content (% by weight) Polyquaternium-10 0.5 Sodium laureth sulfate 20 The eluate of Preparation Example 1 0.3 incense 1.0 Methyl paraben 0.1 Cetyl alcohol 0.5 Sodium chloride 0.8 Purified water To 100

[Formulation Example 11] Manufacture of hair treatment

Hair treatments were prepared according to a conventional method with the compositions shown in the following table.

Category (% by weight) Content (% by weight) The eluate of Preparation Example 1 0.5 Stearamidopropyl dimethylamine 2 glycerin 0.5 Spices 0.5 antiseptic 0.03 Cetostearyl alcohol 5 Lactic acid One Distilled water to 100

Korea Biotechnology Research Institute KCTC12675BP 20140911 Korea Microorganism Conservation Center (overseas) KCCM11180P 20110328

Claims (10)

A composition for hair or scalp comprising, as an active ingredient, an eluate of Lactobacillus sakei . The method according to claim 1,
Wherein said Lactobacillus sake is isolated and identified from gold silver. The method according to claim 1,
Wherein the eluted product of Lactobacillus saccharide is obtained by heating and lyophilizing the cells of Lactobacillus casei. The method of claim 3,
Wherein the heating is carried out at a temperature of from 80 DEG C to 120 DEG C for 10 minutes to 30 minutes. The method according to claim 1,
Wherein the composition is for hair growth enhancement or hair loss prevention. The method according to claim 1,
Wherein said composition is for proliferating and inhibiting the proliferation of hair follicular dermal papilla cells. The method according to claim 1,
Wherein the composition is for promoting proliferation and suppressing the death of the endometrial superficial cells. The method according to claim 1,
Wherein the composition is a cosmetic composition. The method according to claim 1,
Wherein the composition is a pharmaceutical composition. (a) Lactobacillus sakei was cultured,
(b) centrifuging the cultured Lactobacillus saccharum to remove insoluble components, and
(c) a composition for hair or scalp comprising, as an active ingredient, an eluate of Lactobacillus sacrifice, which comprises eluting the lactobacillus sake obtained in (b) above by heating at 80 to 120 ° C for 10 minutes to 30 minutes ≪ / RTI >

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