KR20070019120A - Anti-Wrinkle Cosmetic Composition Comprising the Extract of Paeonia lactiflora as Active Ingredient - Google Patents

Abstract

The invention peony ( Paeonia It relates to a cosmetic composition containing the lactiflora ) extract as an active ingredient, and more particularly to a cosmetic composition for improving skin wrinkles comprising a peony extract excellent in inhibiting the formation of the SNARE complex. The cosmetic composition of the present invention is excellent in stability in the formulation compared to the conventional skin wrinkle improvers, and exhibits a similar effect of Botox, which has been used in conventional wrinkle improvement procedures, thereby greatly improving the wrinkles of the skin without side effects.

Skin Wrinkles, Peony Extract, Botox, SNARE Complex

Description

Anti-Wrinkle Cosmetic Composition Comprising the Extract of Paeonia lactiflora as Active Ingredient}

FIG. 1 shows immunoprecipitated cells incubated with an untreated group and 0.02% Peony extract with an antibody against syntaxin 1, followed by SDS-PAGE electrophoresis and SNAP-25 or Syntaxin 1 It is a photograph showing the result of Western blotting using the antibody against.

The present invention relates to a cosmetic composition for improving skin wrinkles containing a peony extract as an active ingredient.

Wrinkles in the skin are the most pronounced skin aging manifestations caused by endogenous factors such as age and stress, or external environmental factors such as air pollution or ultraviolet radiation. Wrinkles occur on the face of the forehead, around the eyes, between the legs, and around the mouth, or on different parts of the body, such as the neck, neck, elbow, armpits, hands, and feet. Most are noticeable around 30 years of age, and their number, depth and range increase with age. The main cause of wrinkle formation is that reactive oxygen species generated in skin tissues by skin aging process form crosslinks in collagen fibers and elastic fibers, which occupy most of the skin dermis constituents, and change the formation and decomposition of these fibers. ( Clinics in geriatric medicine, 2001, 17 (4): 617-630).

Antioxidants such as vitamin E, beta-carotene, vitamin C and glutathione used to inhibit skin aging, and substances that enhance dermal collagen synthesis, such as radical scavengers and vitamin A, to date It has been used as a substance to improve skin wrinkles.

However, most of these cosmetic active ingredients have strong skin irritation or are unstable in the formulation itself, resulting in discoloration, discoloration and precipitation, and decrease inherent activity of raw materials.

Wrinkles are formed as a result of muscles moving in a certain direction and are the result of accumulation for a very long time. Botox (botulinum toxin) is a substance used to improve wrinkles through muscle paralysis and is a soluble-N-ethylmaleimide-sensitive factor [NSF] attachment protein [SNAP] receptor involved in neurotransmitters. It inhibits the release of catecholamines by inhibiting the release of catecholamines by inhibiting the formation of the recognition receptor) complex present on the surface of the medium involved in the transport, thereby suppressing muscle contraction and improving wrinkles upon application to the skin.

Botox is a neurotoxin produced by clostridium botulinum , a bacterium that grows well in rotting canned or rotten meat. It has been used for neuromuscular diseases such as stiffness, and has recently been used for the procedure for removing wrinkles. This paralyzes the facial muscles and stretches the wrinkles. However, Botox has to be performed by a specialist due to side effects such as muscle paralysis and has a strong toxicity.

Throughout this specification, a number of articles are referenced and their citations are indicated in parentheses. The disclosure of the cited article is incorporated herein by reference in its entirety, so that the level of the technical field to which the present invention belongs and the contents of the present invention are more clearly explained.

Accordingly, the present inventors produce an extract for a plurality of natural products to develop an anti-wrinkle effect, such as Botox, and to develop an anti-wrinkle substance having excellent stability in the formulation, thereby selecting a substance having a similar effect to Botox. As a result of the operation, Peony extract was selected, and excellent clinical improvement of wrinkles was proved through clinical trials, thus completing the present invention.

The present invention provides a cosmetic composition for improving skin wrinkles containing peony extract as an active ingredient.

The present inventors conducted research to develop a botox-like effect, namely, a substance that inhibits the formation of the SNARE complex, and the peony extract has an excellent SNARE complex formation inhibitory effect and an excellent cell proliferation effect. It was confirmed that the improvement greatly.

Peony as an active ingredient included in the composition of the present invention ( Paeonia lactiflora ) is a perennial dicotyledonous plant belonging to the family Amphitaceae , distributed in Korea, Mongolia, and Siberia.

As used herein, the term 'peony extract' refers to an extract obtained from various organs (eg, leaves, flowers, stems and roots, etc.) of the peony, and preferably means an extract obtained from the root of a mature peony.

The peony extract in the present invention may be prepared using a variety of extraction solvents such as (a) water, (b) anhydrous or hydrous lower alcohols having 1 to 4 carbon atoms (methanol, ethanol, propanol and butanol, etc.), and (c) the lower alcohol. A mixed solvent with water, (d) acetone, (e) ethyl acetate, (f) chloroform or (g) 1,3-butylene glycol can be obtained from the peony, preferably an aqueous alcohol solution, More preferably, 70% ethyl alcohol can be obtained as an extraction solvent. On the other hand, it will be apparent to those skilled in the art that the extract of the present invention can be obtained not only the above-described extraction solvent but also a peony extract having substantially the same effect using other extraction solvents.

In addition, the extract of the present invention includes not only the extract by the above-described extraction solvent, but also the extract that has undergone a conventional purification process. Obtained by various additional purification methods, such as, for example, separation using ultrafiltration membranes having a constant molecular weight cut-off value, separation by various chromatography (manufactured for separation according to size, charge, hydrophobicity or affinity). The fraction is also included in the peony extract of the present invention.

The peony extract of the present invention may be prepared in powder form by an additional process such as distillation under reduced pressure and freeze drying or spray drying.

According to the most preferred embodiment of the present invention, the peony extract in the present invention is dried and powdered mature root, and extracted at 15-35 ℃ by adding 70% ethyl alcohol corresponding to 10 times of this, then filtered 5- It can be obtained by standing at 10 ° C., aging, filtration and drying at 65 ° C. under reduced pressure.

According to a preferred embodiment of the present invention, the content of the peony extract is 0.0025-5.0 wt%, more preferably 0.02-1.0 wt% based on the total weight of the composition. If the content is less than the expected effect is difficult to appear, if the content is exceeded there is a problem in color and appearance in the cosmetic formulation.

The ingredients included in the cosmetic composition of the present invention include ingredients commonly used in cosmetic compositions in addition to peony extracts as active ingredients, and include, for example, conventional auxiliaries such as antioxidants, stabilizers, solubilizers, vitamins, pigments and flavorings, and Carrier.

Cosmetic compositions of the present invention may be prepared in any formulation conventionally prepared in the art and include, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing , Oils, powder foundations, emulsion foundations, wax foundations and sprays, and the like, but are not limited thereto. More specifically, it may be prepared in the form of a flexible lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.

When the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, trakant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide, etc. may be used as carrier components. Can be.

When the formulation of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used, in particular in the case of a spray, additionally chlorofluorohydrocarbon, propane Propellant such as butane or dimethyl ether.

When the formulation of the present invention is a solution or emulsion, a solvent, solubilizing agent or emulsifying agent is used as the carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 Fatty acid esters of, 3-butylglycol oil, glycerol aliphatic ester, polyethyleneglycol or sorbitan.

When the formulation of the present invention is a suspension, liquid carrier diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystals Soluble cellulose, aluminum metahydroxy, bentonite, agar or tracant and the like can be used.

When the formulation of the present invention is a surfactant-containing cleansing, the carrier component is aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide. Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters and the like can be used.

The cosmetic composition of the present invention exhibits the use of skin wrinkle improvement. The cosmetic composition of the present invention is less toxic than botox and includes a peony extract having an effect of improving skin wrinkles as an active ingredient. Peony extract included in the cosmetic composition of the present invention by the SNARE complex formation inhibitory effect and cell growth promoting effect, to improve the wrinkles of the skin and excellent stability in the formulation. This fact is specified in detail in the following examples.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, and it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples according to the gist of the present invention. .

Preparation Example 1 Preparation of Peony Extract

The peony extract contained in the cosmetic composition of the present invention was prepared as follows: Peony powder, first washed with purified water and dried, and then broken into small pieces into 10-fold 70% ethyl alcohol, and extracted at 15-35 ° C. for 5 days. It was. It was filtered through a 300 mesh filter cloth, then aged for 7-10 days at 5-10 ° C., and filtered with Whatman filter paper. This extract was then dried on a rotary vacuum evaporator at 65 ° C. to obtain a peony extract powder. The peony extract powder was dissolved in a solution in which water and 1,3-butylene glycol were mixed in the same amount to obtain a peony extract of the present invention. The weight of the peony extract of the following examples is expressed as the concentration of the final extract powder.

Experimental Example 1: Inhibitory Effect of SNARE Complex Formation

The effect of peony extract on neuronal signal transduction of SNARE complex formation was measured by immunoprecipitation. Neurotransmitters are mediated by the regulation of exocytosis of synaptic vesicles. The SNARE complex is involved in neuronal exocytosis and consists of the vesicle protein VAMP (synaptobrevin) and the plasma membrane binding protein syntaxin 1 and SNAP-25.

In this experiment, to determine whether Peony extract inhibits the binding of SNAP-25 and Syntaxin 1, SNARE complex constituent proteins were isolated from PC-12 cells and immunoprecipitated using antibody against Syntaxin 1 and immunoprecipitated protein. After purification, Western blotting was performed using antibodies against SNAP-25 and Syntaxin 1.

PC-12 cells (Korea Cell Line Bank) were seeded in T75 flasks containing DMEM medium (Sigma-aldrich) containing 10% fetal bovine serum (3 x 10 ⁶ cells / well), 5% concentration (CO ₂ ) Was incubated at 37 ° C. for 24 hours in a CO ₂ incubator. The culture was replaced with DMEM medium containing 10% fetal calf serum, and the peony extract of Preparation Example 1 was added at a concentration of 0.02%, followed by incubation for 48 hours, and each cell was collected.

The collected cells were washed twice with PBS buffer, and then homogenization buffer containing a complete protease inhibitor mixture (Roche): 150 mM NaCl, 2.5 mM KCl, 10 mM Na ₂ HPO ₄ And homogenized 50 times with 23-gauge needle in 2 mM KH ₂ PO ₄ , pH 7.4). The homogenate was centrifuged at 2,500 rpm for 10 minutes to remove nuclei and unbroken cells. Thereafter, centrifuged at 120,000 xg for 1 hour to obtain plasma membrane pellets. The pellet was dissolved in a solution containing the complete protease inhibitor mixture (1% Triton X-100, 20 mM HEPES-KOH, pH 7.2, 2 mM EDTA, 100 mM KCl and 1 mM DTT) and reacted at 4 ° C. for 10 minutes. , The supernatant was taken by centrifugation for 20 minutes at 13,000 rpm. After measuring the protein concentration of the supernatant, 200 μg of protein was taken and reacted with monoclonal syntaxin 1 antibody (Gottingen) at 4 ° C. for 30 minutes. Protein A-Sepharose (Amersham) was added and reacted with gentle shaking at 4 ° C. for 12 hours. Sepharose beads were precipitated by centrifugation and washed four times with lysis buffer. The precipitate was treated with SDS-sample buffer and SDS-PAGE and immunoblotting were performed as standard methods. Electroblotting was performed using an immobilon-P membrane (Millipore). As a primary antibody, syntaxin 1 antibody or SNAP-25 antibody (Gottingen) was used. Horseradish peroxidase-conjugated IgG (horseradish peroxidase-conjugated IgG) was used. The experimental results are shown in FIG.

As shown in the experimental results of FIG. 1, the untreated group had almost similar band intensities of Syntaxin 1 and SNAP-25, whereas the experimental group incubated with Peony extract had a band of SNAP-25 compared to that of Syntaxin 1. The intensity was much weaker. It is confirmed that the specific component of the peony extract is excellent in binding to Syntaxin 1 with SNAP-25. Therefore, the peony extract was found to inhibit the composition of the SNARE complex, that is, the mechanism by which SNAP-25 binds to Syntaxin 1.

Experimental Example 2: Proliferation Effect of Peony Extract

The cell growth promoting effect of the peony extract was measured. The peony extract prepared in Preparation Example 1 was dissolved in an appropriate concentration in a buffer solution and then used as a sample solution. Mouse-derived NIH / 3T3 fibroblasts (Korea Cell Line Bank) were inoculated in DMEM medium containing 4.5 g / l glucose, 10% serum, and 1% antibiotics and cultured at 37 ° C. in a 75 cm 2 T-flask. After incubation for 24 hours under 5% CO ₂ conditions, cells were isolated by treatment with 0.05% Trypsin containing 0.02% EDTA, and then inoculated in 96 well-flask and incubated for 6 hours. The cell number is then 2 × 10 ³ cells / well. The sample solution of appropriate concentration was diluted in DMEM medium, treated with fibroblasts cultured to a final concentration of 25, 50, 100 and 200 µg / ml and incubated at 37 ° C for 4 days. After incubation, the medium was obtained, and the absorbance was measured at 570 nm to compare the degree of cell proliferation.

Treatment Concentration of Peony Extract Absorbance (570 nm) 0 0.952 25 μg / ml 0.996 50 μg / ml 1.143 100 μg / ml 1.305 200 μg / ml 1.579

As can be seen in the table above, the peony extract promoted the proliferation of cells in a treatment concentration-dependent manner.

Preparation Example 2: Preparation of Peony Extract-Containing Cosmetic Composition

The cosmetic composition of the present invention containing the obtained peony extract was prepared, and a cosmetic composition containing no peony extract was prepared according to the composition ratios of the following tables.

ingredient Content (% by weight) Example 1 Comparative Example 1 Peony extract 0.1 - Butylene glycol 2.0 2.0 Propylene glycol 6.0 6.0 Carboxy Vinyl Polymer 0.5 0.5 Beeswax 2.0 2.0 vaseline 7.0 7.0 Polysorbate 60 2.0 2.0 Sorbitan sesquioleate 2.5 2.5 Squalane 3.0 3.0 Liquid paraffin 10.0 10.0 Triethanolamine 0.5 0.5 Tocopheryl Acetate 0.1 0.1 Spices, preservatives a very small amount a very small amount Purified water Remaining amount Remaining amount Sum 100 100

Experimental Example 3: Clinical effect of skin wrinkle improvement of peony extract-containing cosmetic composition

Clinical effects of skin wrinkle improvement on cosmetics containing the peony extract of the present invention prepared in Preparation Example 2 were measured.

20 women (40 years old and older) (average age 44.5 years) were divided into two groups (A and B), and the cosmetic composition of Example 1 was applied to group A, and the cosmetic composition of Comparative Example 1 was compared to group B. Formulated according to the composition of Table 2 was applied for 12 weeks (2 times / day, 0.2 g / time) around the wrinkled eyes. After the cosmetic application (T ₀ ) and 12 weeks after the cosmetic application (T ₁₂ ) through the objective evaluation of the expert and the subject's subjective evaluation by classifying into 7 grades in the following Table 3, the degree of improvement of skin wrinkles (Δ W) Was measured. The results are shown in Tables 4 and 5 below.

Skin Wrinkle Evaluation Criteria score Exacerbate extremely badly -3 worse -2 Slightly worse -One No change 0 Slightly improved One Improving 2 Very improved 3

Objective evaluation of the skilled person division Average wrinkle improvement (ΔW) Example 1 Comparative Example 1 T ₀ T ₁₂ T ₀ T ₁₂ Subject 1 0 2 0 0 Subject 2 0 2 0 0 Subject 3 0 3 0 One Subject 4 0 One 0 0 Subject 5 0 2 0 0 Subject 6 0 2 0 0 Subject 7 0 2 0 One Subject 8 0 3 0 0 Subject 9 0 One 0 0 Subject 10 0 3 0 One ΔW (T ₁₂ -T ₀ ) 2.1 0.3

Subjective Evaluation of the Subject division Average wrinkle improvement (ΔW) Example Comparative Example 1 T ₀ T ₁₂ T ₀ T ₁₂ Subject 1 0 One 0 0 Subject 2 0 3 0 0 Subject 3 0 3 0 One Subject 4 0 2 0 0 Subject 5 0 One 0 0 Subject 6 0 2 0 0 Subject 7 0 2 0 0 Subject 8 0 2 0 0 Subject 9 0 One 0 0 Subject 10 0 3 0 One ΔW (T ₁₂ -T ₀ ) 2.0 0.2

As a result of the test, the group coated with the cosmetic composition of Example 1 containing the peony extract after 12 weeks had a significant wrinkle improvement effect as the average wrinkle improvement was 2.1 and 2.0 in the objective evaluation of the expert and the subjective evaluation of the subject, respectively. Confirmed.

Experimental Example  4: containing peony extract Cosmetics  Stability of the Formulations of the Composition

For the cosmetic composition of Example 1 and Comparative Example 1 prepared in Preparation Example 2 4 weeks, 1 month and 6 months in a thermostat and room temperature of 4 ℃, 45 ℃, 50 ℃, Cycle (4 ℃-45 ℃) The long-term preservation stability was evaluated in terms of the following criteria, and the results are shown in Table 6 below.

* Evaluation standard

Good: ○, Unstable: ×, Separation: △

Storage temperature and period Example 1 Comparative Example 1 4 ℃ 1 week 0 0 1 month 0 0 6 months 0 0 45 ℃ 1 week 0 0 1 month 0 0 6 months 0 0 50 ℃ 1 week 0 0 1 month 0 0 6 months 0 0 Cycle (4-45 ℃) 1 week 0 0 1 month 0 0 6 months 0 0 Room temperature 1 week 0 0 1 month 0 0 6 months 0 0

As shown in the above table, when the cosmetic composition of Comparative Example 1 containing no peony extract was found to have good stability, it was found that there was no problem in stability by the other ingredients of the formulation, and the peony extract contained The cosmetic composition of Example 1 was found to be very good long-term storage stability.

Hereinafter, based on the results of the above test examples, a cosmetic composition containing peony extract is presented. However, the composition of the present invention is not limited to the following formulation examples.

Formulation Example 1: Softener (Skin Lotion)

ingredient Content (% by weight) Peony extract 0.1 1,3-butylene glycol 6.0 glycerin 4.0 Oleyl alcohol 0.1 Polysorbate 20 0.5 ethanol 15.0 Benzophenone-9 0.05 Incense, preservative a very small amount Purified water up to 100

Formulation Example 2: Nutrients (Milk Lotion)

ingredient Content (% by weight) Peony extract 0.1 Propylene glycol 6.0 glycerin 4.0 Triethanolamine 1.2 Tocopheryl Acetate 3.0 Floating paraffin 5.0 Squalane 3.0 Macadamia Nut Oil 2.0 Polysorbate 60 1.5 Sorbitan sesquioleate 1.0 Carboxyvinyl Polymer 1.0 Incense, preservative a very small amount Purified water up to 100

Formulation Example 3: Nutrition Cream

ingredient Content (% by weight) Peony extract 0.1 vaseline 7.0 Liquid paraffin 10.0 Beeswax 2.0 Polysorbate 60 2.0 Sorbitan sesquioleate 2.5 Squalane 3.0 Propylene glycol 6.0 glycerin 4.0 Triethanolamine 0.5 Carboxy Vinyl Polymer 0.5 Tocopheryl Acetate 0.1 Incense, preservative a very small amount Purified water up to 100

Formulation Example 4: Massage Cream

ingredient Content (% by weight) Peony extract 0.1 Propylene glycol 6.0 glycerin 4.0 Triethanolamine 0.5 Beeswax 2.0 Tocoberyl Acetate 0.1 Polysorbate 60 3.0 Sorbitan sesquioleate 2.5 Cetearyl Alcohol 2.0 Liquid paraffin 30.0 Carboxy Vinyl Polymer 0.5 Incense, preservative a very small amount Purified water up to 100

Formulation Example 5: Pack

ingredient Content (% by weight) Peony extract 0.1 Propylene glycol 2.0 glycerin 4.0 Carboxy Vinyl Polymer 0.3 ethanol 7.0 Fiji-40 Hydrogenated Castor Oil 0.8 Triethanolamine 0.3 Incense, preservative a very small amount Purified water up to 100

As described in detail above, the present invention provides a cosmetic composition for improving skin wrinkles containing peony extract as an active ingredient. The cosmetic composition of the present invention is safer than conventional skin care using botox, and exhibits a similar effect to Botox, thereby reducing the activity of skin muscles and improving skin wrinkles. In addition, the stability in the formulation of skin wrinkle improvers, which has been raised as a conventional problem, has been solved by using plant extracts.

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that such a specific technology is only a preferred embodiment, and the scope of the present invention is not limited thereto. Therefore, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

Claims (5)

Cosmetic composition for improving skin wrinkles containing peony extract as an active ingredient. According to claim 1, wherein the peony extract is water, anhydrous or hydrous lower alcohol having 1 to 4 carbon atoms, a mixed solvent of the lower alcohol and water, acetone, ethyl acetate, butyl acetate, chloroform and 1,3-butylene glycol Cosmetic composition for improving skin wrinkles, characterized in that extracted by using a solvent selected from the group consisting of as an extraction solvent. The cosmetic composition for improving skin wrinkles according to claim 2, wherein the peony extract is extracted using an alcohol as an extraction solvent. The cosmetic composition for improving skin wrinkles according to claim 1, wherein the peony extract is contained in an amount of 0.0025-5.0 wt% based on the total weight of the composition. The composition of claim 1 wherein the cosmetic composition consists of a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation and spray Cosmetic composition for improving skin wrinkles, characterized in that the formulation is selected from the group.

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