KR20210025339A - A COSMETIC COMPOSITION FOR ACNE SKIN CARE COMPRISING Phloretin, Protaetiamycine AND Highly stabilized epidermal growth factor AS EFFECTIVE COMPONENT - Google Patents

Abstract

The present invention relates to an acne skin care cosmetic composition comprising: phloretin, which prevents inflammatory diseases and photoaging of the skin; and an anti-biotic peptide derivative for anti-biotic and anti-inflammatory use for acne bacteria, as active ingredients, and additionally, comprising a highly stable epithelial cell growth factor which promotes skin regeneration and growth and, more specifically, to a cosmetic composition effective for anti-inflammation, an acne-improving effect, anti-bacterial activity, and prevention and treatment of various skin diseases by inhibiting MAPK expression. According to the present invention, it is effective in preventing skin diseases, treating acne and regenerating skin by using the cosmetic composition comprising phloretin, the anti-biotic peptide derivative, and the stabilized epidermal growth factor.

Description

A cosmetic composition for acne skin care containing phloretin, protaetiamycin antibiotic peptide derivative, and high stability epithelial cell growth factor as active ingredients {A COSMETIC COMPOSITION FOR ACNE SKIN CARE COMPRISING Phloretin, Protaetiamycine AND Highly stabilized epidermal growth factor AS EFFECTIVE COMPONENT}

The present invention relates to a cosmetic composition for acne skin care comprising phloretin, an antibiotic peptide derivative, and a high stability epithelial cell growth factor as an active ingredient. In more detail, the present invention provides a cosmetic composition for promoting anti-inflammatory, antibiotics for acne bacteria and skin improvement, cell growth and skin regeneration based on a mechanism of mitogen-activated protein kinases (MAPK) signaling inhibition.

Ultraviolet light is the most important environmental factor that causes skin damage. Continuous exposure to such ultraviolet rays causes skin deformation and causes various inflammatory factors, and promotes the expression of genes that will damage cells such as MAPK by inflammatory factors. Among these skin diseases, acne is an inflammatory disease that occurs in the hair follicles in the skin. In the past, it was considered a disease of adolescents, but it is a skin disease that occurs frequently in adults. If you touch or squeeze the acne area with your hands, various germs may aggravate the inflammation of acne, so it is desirable not to touch it. Cosmetics with a lot of oil in the acne area are concerned with clogging pores, so an essence or cream that does not contain oil. It is better to supply moisture and nutrients without excess.

Effective methods for treating acne bacteria include oral administration and transdermal administration.First, tetracycline antibiotics administered orally are drugs that sterilize acne bacteria and reduce inflammation by preventing the production of free fatty acids, and isotretinoin is a vitamin As a derivative of A, it suppresses the secretion of sebum and causes the sebaceous glands to contract. This action is known to suppress the secretion of hormones by transmitting signals to the brain.

As a transdermal administration method, oil is removed using soap or weakly acidic detergent with excellent cleaning power and low irritation, and ointment containing benzoyl peroxide has antibacterial action and reduces free fatty acids, showing excellent effect on inflammatory acne. . Using a topical antibiotic that has a direct effect on acne bacteria and reduces inflammation by preventing the production of free fatty acids, or using a skin regeneration ointment containing an ingredient called tretinoin, it minimizes the rupture of the hair follicle by removing the dead skin cells that are clogging the pores. It is known to have an effect on various acne by preventing the development of inflammation because it allows it to be discharged well.

However, although the above methods are effective in treating acne, the use of anti-hormonal drugs can cause skin rash, swelling, skin repellency, etc., in the case of salicylic acid preparations, along with side effects due to hormone abuse, and in the case of retinoic acid preparations, contact dermatitis, Side effects such as erythema and dry skin have been reported. In addition, antibiotics such as erythromycin and criindamycin tend to increase the blood concentration of other drugs metabolized by cytochrome p450, and the appearance of resistant strains has reported a case in which the treatment effect decreases, so the appearance of a new acne treatment is urgent. Triclosan, an ingredient that is almost available in acne treatments or toothpaste, reacts with chlorine in tap water to produce chlorinated dioxin, and even a very small amount of triclosan itself is known as a substance that interferes with thyroid function and thyroid hormones, and this substance continues to be used. As a result, resistance is known to affect the production of strains of bacteria. In addition, retinol acid or antibiotics can cause skin irritation, are irritating to the skin, and have side effects such as bacterial resistance, thus limiting their use.

In addition to drug therapy, there are surgical treatments such as skin dermabrasion and cryotherapy, and physical treatments such as ultraviolet irradiation, but it has a disadvantage that it is difficult to correct scars and bear high cost for acne that has started to occur once.

Therefore, it would be urgent to develop a preparation for preventing and treating acne that can prevent inflammation and has no side effects.

The present invention solves the above problems, and is derived from the necessity of the present invention. An object of the present invention is a composition capable of realizing the treatment of acne skin and promotion of skin growth through the regeneration and differentiation of skin cells, anti-inflammatory and anti-acne. Is to provide.

In order to achieve the above object, the present invention relates to floretin (Korean Patent Application No. 10-1755091), antibiotic peptide derivative (Korean Patent Application No. 10-1540638), high stability epithelial cell growth factor (Korean Patent Application No. 10-2013- 0086773) as an active ingredient to provide a cosmetic composition for improving acne skin.

In one embodiment of the present invention, it is preferable to include a complex consisting of the phloletin, an antibiotic peptide derivative, and a high stability epithelial cell growth factor, but is not limited thereto.

In one embodiment of the present invention, the floretin is preferably described in Korean Patent Application No. 10-1755091, but is not limited thereto.

In a preferred embodiment of the present invention, the sequence of the antibiotic peptide derivative is preferably the sequence described in Korean Patent Application No. 10-1540638, but is not limited thereto. In a preferred embodiment of the present invention, the sequence of the high-stability epithelial cell growth factor is preferably Korean Patent Application No. 10-2013-0086773, but is not limited thereto.

In an embodiment of the present invention, the composition comprises 0.010 to 5.114 parts by weight of phloretin, 0.004 to 0.949 parts by weight of an antibiotic peptide derivative, and 0.010 to 5.114 parts by weight of a high stability epithelial cell growth factor based on 70 parts by weight of the lyophilized powder. It is preferable, but is not limited thereto.

In another embodiment of the present invention, the composition is one of Gangwha (Angelica koreana), Gosam (Sophora flavescens), Turmeric (Curcuma longa), Licorice (Licorice), Paeonia japonica, Tea tree extract It is preferable to further include more, and the extract is strong active 0.05 to 15% by weight, gosam 0.05 to 15% by weight, turmeric 0.05 to 15% by weight, licorice 0.002 to 15% by weight, county peony 0.002 to 15% by weight, tea tree 0.002 It is preferably ~ 15% by weight, but is not limited thereto.

According to another aspect of the present invention, the present invention provides a cosmetic composition for improving acne skin, comprising the phloretin, protetiamycin, and high stability epithelial cell growth factor as active ingredients.

The composition of the present invention is very effective in improving anti-inflammatory and acne skin.

Preferably, the composition of the present invention is used for the treatment of acne bacteria and for regeneration and growth of skin cells.

The term "cosmetically effective amount" as used herein means an amount sufficient to achieve the effect of improving the skin of the composition of the present invention described above.

The cosmetic composition of the present invention may be prepared in any formulation commonly prepared in the art, for example, solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing , Oil, powder foundation, emulsion foundation, wax foundation, spray, etc. may be formulated, but is not limited thereto. More specifically, it may be prepared in the form of a flexible lotion, a nutritional lotion, a nutritional cream, a massage cream, an essence, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a pack, a spray, or a powder.

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

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

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

When the formulation of the present invention is a suspension, as a carrier component, a liquid diluent such as water, ethanol or propylene glycol, an ethoxylated isostearyl alcohol, a suspending agent such as polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, and crystallites Castle cellulose, aluminum metahydroxide, bentonite, agar, and the like can be used.

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

Ingredients included in the cosmetic composition of the present invention include ingredients commonly used in cosmetic compositions, in addition to growth factors and carrier ingredients as active ingredients, such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and fragrances. May contain adjuvants.

Since the compositions of the present invention contain the above-described phloretin, protaethiamycin antibiotic peptide derivatives, and high stability epithelial cell growth factors of the present invention as active ingredients, the common content between the two is to avoid excessive complexity of the present specification. The description is omitted.

The features of the cosmetic composition for promoting skin disease prevention, acne skin treatment and regeneration according to the present invention may be understood by examples described in detail below with reference to the accompanying drawings.

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

The cosmetic composition applied in this example contains 0.010 to 5.114 parts by weight of phloletin, 0.004 to 0.949 parts by weight of an antibiotic peptide derivative, and 0.010 to 5.114 parts by weight of a high stability epithelial cell growth factor based on 70 parts by weight of the freeze-dried powder.

Phloretin (2',4',6'-Trihydroxy-3-phydroxyphenylpropiophenone) belongs to the chalcone class of flavonoids and is present in apples, strawberries, and pears as phloretin 2'-0-glucose. Antibiotic effects can be exhibited by inhibiting the formation of cell walls of various bacteria through inhibition of ketoacyl acyl carrier protein syntehase ⅠⅠⅠ (KAS ⅠⅠⅠ). In addition, when stimulated with IFN-γ, the expression of inflammatory mediators can be suppressed through a regulatory mechanism through MAPK, and the effect of inhibiting inflammation when stimulated with IFN-γ or stimulated by UVB can be induced. Photoaging can be prevented and prevented by mediating the signaling pathways of MMP-1 and MAPKs in UVB-stimulated human keratinocytes, particularly inhibiting P.Acnes-induced MMP-1 expression.

Antibiotic peptide derivatives are derivatives with D-type amino acid residues of proteamycin isolated from white-spotted radish larvae. It was confirmed that they inhibit nitric oxide, a substance causing inflammation, and show the best antibacterial activity compared to triclosan and benzoyl peroxide. . In addition, it is an antibiotic peptide that is safe for humans with remarkably low cytotoxicity and hemolytic activity.

High stability epithelial growth factor is a factor that promotes the growth of epithelial cells, and has been shown to induce growth and division of mammalian cells, particularly epithelial and skin cells, by transmitting signals through EGF receptors in cell membranes. Therefore, it promotes the proliferation of epithelial cells and dermal cells, promotes cell proliferation of fibroblasts that synthesize collagen, which is a component of the dermis, promotes angiogenesis in the damaged area of the skin, and induces the secretion of other regeneration promoting factors. It plays a key role in skin regeneration, such as promoting the synthesis of fibronectin, a substance that forms a network, and minimizing scars on wounds, helping the skin's original function, which decreases with age, and promoting the growth of new skin cells. have.

The high-stability epithelial cell growth factor of the present invention is a high-stability epithelial cell growth factor variant developed by the applicant of the present application [Korea Patent Application No. 10-2013-0086773], which is a problem of the existing epithelial growth factor, which is thermodynamic instability and short An epithelial growth factor variant that overcomes the half-life was used.

The cosmetic composition of the present invention is characterized by 0.010 to 5.114 parts by weight of phloletin, 0.004 to 0.949 parts by weight of antibiotic peptide derivatives, and 0.010 to 5.114 parts by weight of high stability epithelial cell growth factor based on 70 parts by weight of lyophilized powder.

Here, the acne skin care cosmetic composition has a composition as shown in Table 1. It is the same as the composition used in the prior patent [Korea Patent Application No. 10-2015-0099764] and the contents are attached below Table 1.

number Ingredients ingredient function W/W% One Water
Glycerin
Cycloprentasiloxane
Phenoxyethanol
Arginine
Squalane
Polysorbate 60
Carbomer
Cyclohexasiloxane
Dimethicone/Vinyl Dimethicone Crosspolymer
Dimethicone
Ethylhexylglycerin
Allantoin
Sodium Hyaluronate
Lecithin Purified water
glycerin
Cyclopentasiloxane
Phenoxyethanol
Arginine
Squalane
Polysorbate 60
Carbomer
Cyclohexasiloxane
Dimethicone/Vinyl Dimethicone Cross Polymer
Dimethicone
Ethylhexyl glycerin
Allantoin
Sodium hyaluronate
lecithin Basic formulation 2 Phloretin
Antibiotic peptide derivative Floretine
Antibiotic peptide derivative Anti-acne drugs 3 Epidermal growth factor Epithelial cell growth factor Growth factor system 4 Angelica Koreana
Sophora flavescens
Curcuma longa
Licorice
Paeonia japonica
Tea tree curcuma
Gosam
Turmeric
licorice
Count
Tea tree Sedatives

Table 1 is the ingredients and contents of the acne skin care cosmetic composition

In the above table, mannitol, an excipient, is a product of JUNSEI, and all of the ingredients used in hair nutrition and conditioning agents, and others were products of SIGMA-ALDRICH. Growth factor complex and antioxidant were produced by P&P Biopharm and added to the composition. The growth factor produced by PNP Biopharm Co., Ltd. is a recombinant human growth factor produced by E. coli.

The acne care cosmetic composition applied in this embodiment is characterized in that it contains at least one of purified water, a buffer solution or ganghwal, gosam, turmeric, licorice, earl, and tea tree extract.

Kangwha extract contains curcumin and helps to suppress inflammation with anti-inflammatory and antioxidant effects. It protects the skin from free radicals that cause skin aging.

Sophora ginseng extract helps strengthen skin resistance. As it has anti-inflammatory and anti-allergic effects, it has excellent effects in preventing and alleviating inflammatory skin diseases and damage to skin tissues.

Turmeric extract is a component obtained by extracting from the root of turmeric. It helps to soothe the skin tired from external stimuli and stress and make it lively.

Licorice extract smoothes the skin and has antibacterial properties. It has few side effects to the skin and prevents aging with its cell regeneration effect.

Earl extract is suitable for sensitive skin because it has excellent anti-inflammatory properties. In addition, as an ingredient that has anti-aging function, it prevents dermatitis.

Tea tree extract helps to soothe irritation and sensitive skin, and controls the amount of sebum secreted. It has an astringent effect that tightens pores and helps to keep the skin clean and moisturized by catching the shine of the skin.

Here, the base of the cosmetic composition for improving acne skin can be used as long as it is commonly used, such as purified water, butylene glycol, 1,2-hexanediol, glyoxal, modified alcohol, glycerin, sodium hyaluronate Nate, hydroxyethyl cellulose, and allantoin are included. Examples of other ingredients incorporated in the composition include, but are not limited to, stabilizers, preservatives, thickeners, skin moisturizers, and the like.

In addition to the above ingredients, the cosmetic composition for improving acne skin may contain ingredients that can serve to supply nutrients to the skin or supplementary ingredients that are commonly used to promote skin cell growth. For example, vitamins, amino acids, sodium chloride, etc. It may include, but is not limited to these. At this time, the vitamins are characterized by containing at least one selected from the group consisting of vitamin B group, choline, inositol, ascorbic acid, vitamins A, D, E, and K.

According to the present invention, acne containing phloretin, antibiotics against acne bacteria, and antibiotic peptide derivatives for anti-inflammatory use as an active ingredient, and additionally, high stability epithelial cell growth factor promoting skin regeneration and growth The skin care cosmetic composition is effective in anti-inflammatory, acne improvement effect, antibacterial activity, and prevention and treatment of various skin diseases, and is effective in preventing skin diseases, treating and regenerating acne skin by promoting the regeneration and growth of skin cells.

1 is a picture of measuring toxicity to mammalian cells,
Figure 2 is a picture showing anti-inflammatory activity in RAW264.7 cells stimulated with LPS,
Figure 3 is a picture showing anti-inflammatory activity in HaCaT cells stimulated with acne bacteria,
Figure 4 is a picture showing an analysis of the mechanism of anti-inflammatory action in HaCaT cells stimulated with acne bacteria,
Figure 5 is a picture showing the analysis of the antibacterial mechanism of action,
Figure 6 is a picture showing the anti-inflammatory combination effect for the combination of five mixture ratios (n = 5) of peptide and phloretin,
7 is a picture showing the evaluation of skin sebum by Janus I,
Figure 8 is a picture showing the evaluation of porphyrin by Visiopor,
9 is a picture showing the evaluation of redness by a spectrophotometer,
Figure 10 is an actual photo of the improvement of porphyrin by the present invention.

Hereinafter, in the following description, specific matters such as specific components are described, which are provided to help a more general understanding of the present invention, and the present invention can be practiced without these specific matters. It is self-evident to those who have knowledge. In the following description of the present invention, when it is determined that a detailed description of a related known function or configuration may unnecessarily obscure the subject matter of the present invention, a detailed description thereof will be omitted. The examples given below will aid in understanding the present invention.

Example 1: Identification of the mechanism of action of anti-inflammatory activity of protetiamycin (9Pbw3D-7) and verification of the ability to inhibit inflammation induced by acne bacteria in skin cells and identification of the mechanism of action

Measurement of toxicity to mammalian cells: Measurement of cell proliferation inhibitory activity by MTT assay for RAW264.7 cells, HaCaT.

-To confirm the effect of cytotoxicity, an MTT assay was conducted to confirm the effect of the peptide on RAW264.7, which is a macrophage of a mouse, and on the skin cells of a human.

-In sheep blood, hemolysis was not observed in all samples.

-In the macrophages of mice, the toxicity of 9Pbw3, 9Pbw3D, 9Pbw3D-7 was measured at 50uM in the order of 82%, 53% and 13%, respectively, indicating that 9Pbw3D-7 had low toxicity.

On the other hand, it was confirmed that melittin, a control group, showed hemolytic activity and high cytotoxicity at a very low concentration.

-9Pbw3D-7 is highly applicable due to its low toxicity.

In RAW264.7 cells stimulated with -LPS, the synthesis of nitrite oxide of 9Pbw3D-7 and the inhibitory activities of the pro-inflammatory cytokines TNF-α, IL-6 and IL-1β were measured.

-The inhibitory activity of 9Pbw3D-7 on the production of TNF-α, IL-8, and IL-12 in HaCaT cells induced by acne bacteria was measured by ELISA, and the mechanism of anti-inflammatory action was investigated through western immunoblotting and RT-PCR.

-Quantitatively compare the amount of inflammatory cytokines through ELISA to determine whether or not to suppress the inflammatory expression expressed when LPS, which is known as an agonist of TLR4, in mouse macrophages. It was confirmed that the inhibitory ability for Nitrite, TNF-α, IL-1β and IL-6 production was excellent (FIG. 2).

-Excellent inflammation by observing the inhibition of the production of inflammatory cytokines TNF-α, IL-12 and IL-8, which confirms the inflammatory response by acne bacteria to check whether the inflammatory expression expressed when stimulating skin cells with acne bacteria is suppressed. The inhibitory ability was confirmed (Fig. 3).

-To analyze the mechanism of inhibition of inflammation by acne bacteria, analyzed through western blot and RT-PCR. As a result, it was confirmed at the protein level that 9Pbw3D-7 inhibits the phosphorylation of p38 and ERK in cells by acne bacteria, and produces inflammatory cytokines TNF-α, IL-α, IL-β, IL-8 and IL-12. It was confirmed that the inhibitory ability was excellent (Fig. 4).

Example 2: Protetiamycin (9Pbw3D-7) and phloretin co-administered acne-resistant bacteria, skin clinical-resistant bacteria through the measurement of the antibacterial activity of the combination effect exploration

Protetiamycin and phloretin were administered together against 5 standard acne bacteria, 5 acne-resistant bacteria, and 6 skin disease-resistant bacteria, and the antimicrobial activity was measured to verify the combination effect. Antimicrobial activity was measured by the microdilution method, and the antimicrobial activity for each strain was expressed as MIC (minimal inhibitory concentration) or MBC (minimal bactericidal inhibitory concentration).

-Antimicrobial activities of 9Pbw3D-7 and phloretin were measured against 6 common bacteria, 5 skin disease resistant bacteria, 5 standard acne bacteria and 5 acne resistant bacteria. Antimicrobial activity was measured by the microdilution method, and the antimicrobial activity for each strain was analyzed by MIC (minimal inhibitory concentration).

The effect of co-administration of protetiamycin (9Pbw3D-7) and phloretin in standard acne bacteria was measured.

Antimicrobial mechanism of action was analyzed using bacterial membranes and animal cell membrane liposomes.

Example 3: Analysis of antibacterial activity and mechanism of action against strains causing skin lesions including acne bacteria

-Antibacterial activity was confirmed through the minimum inhibitory concentration of the bacteria in the standard strains and strains that can cause inflammatory lesions on the skin including acne bacteria. As a result, it was confirmed that the D-form was 4 times more active than 9Pbw3, and 9Pbw3D-7 was observed to be 2 times more active, and excellent antimicrobial activity was confirmed. In addition, it was confirmed that the antibacterial activity was 8-64 times better than benzoyl peroxide, which is the main substance mainly used in acne treatment. (Table 2-5).

MIC(uM) peptide E. coli P.
aeruginosa A.
baumannii S. epidermidis B. Subtilis S. Aureus Melittin 4 4 8 8 8 4 9Pbw3 32 32 8 16 16 16 9Pbw3D 4 4 4 4 4 4 9Pbw3D-7 8 8 8 8 8 8

Table 2 shows antibacterial activity against standard strains

MIC(uM) peptide S. Aureus
0027 S. Aureus
3708 S. Epidermidis
3709 S. Epidermidis
3710 S. Epidermidis 3711 Melittin 8 8 4 4 4 Benzoyl peroxide 64 64 64 64 64 Triclosan 4 4 0.5 0.5 0.5 9Pbw3 4 4 One One One 9Pbw3D 2 2 0.5 One 0.5 Pbw3D-7 2 2 One One One

Table 3 shows antibacterial activity against skin disease-causing strains

MIC(uM) peptide P. acnes
3220 P. acnes
3114 P. acnes
5933 P. acnes
5527 P. acnes
5012 Benzoyl peroxide 64 64 64 64 64 Triclosan 16 16 16 16 16 Melittin 2 2 2 One One 9Pbw3 8 16 16 8 8 9Pbw3D 2 2 2 One 2 Pbw3D-7 4 4 2 4 4

Table 4 shows antibacterial activity against acne bacteria

MIC(uM) peptide P. acnes
0081 P. acnes
9007 P. acnes
9008 P. acnes
9009 P. acnes
9010 Benzoyl peroxide 64 64 64 64 64 Triclosan 32 32 32 32 32 Melittin One 2 2 2 2 9Pbw3 8 16 16 32 16 9Pbw3D 2 2 2 2 2 Pbw3D-7 2 4 4 4 4

Table 5 shows antibacterial activity against resistant acne bacteria

-It shows the same effect, but the combined effect in antibacterial activity was measured to confirm that the antimicrobial effect appeared by treatment with phloretin in combination with the peptide. 9Pbw3D-7: Phloretin= 5.1 ug/ml(1/2MIC): 4.4 ug/ml(1/2MIC). When 1/2 MIC of each substance is used in combination, it shows 100% sterilization activity and 50% combination effect. Excellent combination effect was confirmed. (Table 6)

Table 6 is a table showing the antimicrobial combination effect for the combination of five mixture ratios (n=5) of peptide and phloretin against acne bacteria, (A) in uM units measured in P.acnes KCTC 3314 bacteria, (B) ug The combined effect in units of /ml, (C) the unit of uM measured in P.acnes KCTC 3220 bacteria, and (D) the combined effect in units of ug/ml are shown.

Analysis of antibacterial mechanism of action

-For the analysis of the mechanism of antimicrobial action, LUV (PC:PG=7:3) mimicking the cell membrane of bacteria and LUV (PC:CH=10:1) mimicking the cell membrane of animal cells were used to match the composition of the liposome. Measure the effect on the cell membrane by adding calcein dye. As a result, it was confirmed that 9Pbw3 destroys the membrane without distinction between the cell membrane and the animal cell membrane. In contrast, it was confirmed that the D-form peptides 9Pbw3D and 9Pbw3D-7 act selectively only on bacterial cell membranes, thereby confirming their high applicability.

Example 4: Protetiamycin (9Pbw3D-7) and Floretine Confirmation of enhancement of anti-inflammatory activity when administered in combination

This peptide protetiamycin is significant as a cosmetic material when synergistic or additive effects in anti-inflammatory activity are confirmed when co-administered with phloretin, which has a high synthetic cost and low cost. The combined effect was confirmed by measuring the ability to inhibit the production of various pro-inflammatory cytokines in HaCaT cells that caused inflammation by acne bacteria when the peptide and phloretin were co-administered.

When -9Pbw3D-7 was treated with 0-12.7ug/ml, the amount of nitrite produced at 12.7ug/ml showed 13.5% inhibitory activity from 5052pg/ml to 4366pg/ml.

In order to observe the combination effect with -phloretin, the ability to inhibit Nitrite formation was observed after the combination treatment as shown in FIG. 6. After stimulating RAW 264.7 cells with LPS, add peptide and phloretin at 9Pbw3D-7:Phloretin=12.7ug/ml: 5.5ug/ml to increase nitrite production at 4366pg/ml when treated with only peptide and 2054pg/ml when administered with phloretin. It was confirmed that the combination effect was excellent by confirming that it was inhibited by 53.0%.

Example 5: Protetiamycin family Florectin And clinical trials of cosmetic compositions containing growth factors ( KTR Nationally accredited testing institute)

1. Conducted by selecting 22 subjects.

1.1 Number of subjects

Twenty-two subjects who satisfies the criteria for selection of test subjects and did not meet the criteria for exclusion were selected as test subjects. All 22 selected subjects ended the test as final subjects.

1.2 The criteria for selecting test subjects are as described below.

(1) Adult males and females aged 19 to 55 years or younger who have red rashes

(2) Healthy people who do not have chronic physical diseases including skin diseases

(3) A person who voluntarily writes and signs a consent form after receiving a sufficient explanation of the matters to be informed to the test subject from the test manager or the test manager who has been delegated by the test manager.

(4) Those who can follow up during the test period

1.3 The criteria for exclusion from the selection of subjects are as described below.

(1) Pregnant or lactating women and women who are likely to become pregnant

(2) Those who use steroid-containing skin external preparations for more than 1 month to treat skin diseases

(3) People with an allergic peculiar constitution (food, pollen, etc.) or hypersensitivity

(4) People with irritation or allergies to cosmetics, medicines, and daily exposure to sunlight

(5) Those who have used cosmetics and drugs with the same or similar efficacy on the test site within 3 months before the start of the study

(6) Significantly dystrophic, drug or alcoholic

(7) Those who have a spot, acne, tattoos, scars, erythema, capillary dilatation, burn marks, etc. on the test site that affect the test

(8) Those who have not passed 6 months after participating in the same test

(9) If it is deemed inappropriate for the test by the judgment of the person in charge of the test

1.4 The criteria for dropout of test subjects are as described below.

(1) In case of adverse events such as itching or erythema at the test site

(2) In case the test subject has difficulty in evaluating the results due to the application of an external agent other than the test product to the test site, drug use, injection therapy, physical therapy including laser, or drug use during the course of the test

(3) In case the test subject has difficulty in evaluating the results due to medical treatment, application of other products, excessive exposure to ultraviolet rays, excessive drinking and smoking, etc. to the test site during the test process

(4) When the subject is difficult to follow-up due to personal circumstances during the test process

(5) When the subject violates the method of use or schedule without any special reason

2. Methods

2.1 Measurement site

The test subject's face was taken as the measurement site, and the test site was washed with the same cleaning agent at the center at the time of the visit, and then rested for 30 minutes under constant temperature and humidity conditions. For objective measurement, the same tester measured each measurement.

2.2 Measurement schedule

Measurement schedule: Before using the test product (2019-04-29)

: Two weeks after using the test product (2019-05-13)

: After 4 weeks of using the test product (2019-05-27)

2.3 Metrics:

It was carried out in a constant temperature and humidity condition that maintains a constant temperature and humidity.

Temperature: 20.5-22.3 ℃

Humidity: 40.0-57.0% R.H.

Skin sebum

Equipment name: JanusⅠ(Facial Analysis System JANUS, PIE, Korea)

Measurement method: The entire face is photographed in UV mode. The photographed image is not available with the JanusⅠ program.

The average number of sebum on the entire surface was measured. It means that as the result value decreases, it improves.

Porphyrin

Equipment name: Visiopor PP34 (Courage & Khazaka, Germany)

Measurement method: After photographing the test site, the value of porphyrin was measured and analyzed. It means that as the result value decreases, it improves.

Red flag

Equipment name: Spectrophotometer CM700D (Konica Minolta, Japan)

Measurement method: Using a spectrophotometer on the test site, a* value indicating skin redness

It was measured. The measurement was repeated three times on the same area, and divided by the average value of the three.

As the result value decreases, it means that the improvement is improved.

2.4 Compliance assessment

The compliance evaluation was conducted by distributing the product and the compliance log to the test subjects during the test period, so that the subjects directly entered the use or not. Accordingly, compliance was evaluated in the following manner. There were no subjects with less than 80% compliance.

2.5 Survey evaluation

Subjects are requested by the client for skin conditions after 4 weeks of use of the test product. Subjective evaluation of the test product and a 5-point scale (1 = not at all 2 = not so 3 = normal 4 = yes 5 = very yes) / 1 = Not at all good 2 = Not good 3 = Average 4 = Good 5 = Very good).

2.5 safety evaluation

During the use of the test product, the presence or absence of adverse reaction symptoms reported from the test subject was confirmed.

2.7 Adverse reaction evaluation

After using the test product, the tester observed the skin condition and health condition of the test subject and confirmed the presence of skin irritation and body abnormalities caused by the test product through the test subject.

2.8 Adverse Events

In the case report form, the test subject was evaluated for adverse events (erythema, swelling, scales, itchiness, stinging, burning, stiffness, tingling) or other abnormalities with the naked eye. In the case of occurrence, the severity was classified into weak, medium, and severe levels, and the test was checked for discontinuation or dropout, and recorded in the individual case record.

3. Statistics

The statistical processing of this test was performed using the SPSS (Ver. 25) program. The significance of the measured value compared to before using the test product was confirmed when the significance probability p <0.05 in the 95% confidence interval. For statistics before and after use, a parametric method of Repeated measures ANOVA and a non-parametric method of Friedman test were performed after normality test.

4. Results

4.1 Subject information

Information on subjects who participated in this human use test is as follows. (Table 7)

4.2 Compliance assessment

All the subjects who completed the test had a compliance rate of 80% or more with the test product, and all data were analyzed as a result.

4.3 Skin sebum evaluation (JanusⅠ)

The results of measuring the average number of sebum before, 2 and 4 weeks after use of the product using JanusⅠ are as follows (Table 9). As a result of testing with Repeated measures ANOVA, a parametric method according to the normality test, the measured values of skin sebum decreased statistically significantly after 2 and 4 weeks of use compared to before use.

4.4 Porphyrin evaluation (Visiopor)

Porphyrin was measured before, 2 and 4 weeks after use of the product using Visiopor, and the results are as follows (Table 10). According to the normality test, as a result of the nonparametric Friedman test and post-mortem analysis with the Wilcoxon signed rank test, the porphyrin measurement value decreased statistically significantly after 2 and 4 weeks of use compared to before use.

4.5 Redness evaluation (Spectrophotometer)

The results of measuring redness before, 2 and 4 weeks after use of the product using a spectrophotometer are as follows (Table 11). According to the normality test, as a result of testing with the nonparametric Friedman test and post-analysis with the Wilcoxon signed rank test, the measured value of redness decreased statistically significantly after 2 and 4 weeks of use compared to before use.

4.6 Survey evaluation

4.7 Safety evaluation

During the use of the test product, the adverse reaction symptoms reported in the subject were evaluated. There were no symptoms of adverse reactions that occurred during the study period.

4.7.1 Adverse reaction evaluation by a dermatologist

No adverse reactions to allergic contact dermatitis or irritant contact dermatitis were observed after using the test product in the subjects.

4.7.2 Report of adverse skin reactions by test subjects

As a result of confirming the subjects, no specific symptoms related to adverse skin reactions were reported after using the test product.

5. Conclusion

The final 22 subjects were subjected to a human application test for the effect of improving skin sebum, porphyrin, and redness of the acne skin care cosmetic composition of the present invention.

This test was conducted on 22 healthy adults aged 19 to 51 years old, and the results before and after 2 and 4 weeks of use of the test product are as follows.

1) The average age of 22 subjects who ended this test was 32.6 years old. The selected subjects had no specific skin symptoms and had no history of disease or drug use that could affect the study.

2) As a result of checking the skin sebum, the average number of sebum showed improvement rates of 34.37% and 83.83%, respectively, after 2 and 4 weeks of use compared to before, and statistically significantly decreased (P<0.001).

3) As a result of confirming porphyrin, the porphyrin result showed an improvement rate of 31.02% and 61.22%, respectively, after 2 and 4 weeks of use compared to before use, and statistically significantly decreased (P<0.001).

4) As a result of confirming the redness, the redness result showed improvement rates of 14.18% and 24.37%, respectively, after 2 and 4 weeks of use compared to before use, and statistically significantly decreased (P <0.05, 0.001).

6) Subjective questionnaire evaluation by test subjects: After using the test product, all items of the efficacy evaluation were evaluated as'normal' or higher.

7) Evaluation of adverse reactions by a dermatologist: During the use of the test product, there were no reports of specific adverse skin reactions in the test subjects, and no abnormal findings were observed even on the physical examination by a dermatologist.

Therefore, the acne skin care cosmetic composition of the present invention is a product that helps improve skin sebum, porphyrin, and redness, and clinical trial results have been shown.

<110> PnP Biopharm Co., Ltd. <120> A COSMETIC COMPOSITION FOR ACNE SKIN CARE COMPRISING Phloretin, Protaetiamycine AND Highly stabilized epidermal growth factor AS EFFECTIVE COMPONENT <130> P19-0125HS <160> 3 <170> KopatentIn 2.0 <210> 1 <211> 53 <212> PRT <213> Homo sapiens <400> 1 Asn Ser Asp Ser Glu Cys Pro Leu Ser His Asp Gly Tyr Cys Leu His 1 5 10 15 Asp Gly Val Cys Met Tyr Ile Glu Ala Leu Asp Lys Tyr Ala Cys Asn 20 25 30 Cys Val Val Gly Tyr Ile Gly Glu Arg Cys Gln Tyr Arg Asp Leu Lys 35 40 45 Trp Trp Glu Leu Arg 50 <210> 2 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> peptide <400> 2 Arg Leu Trp Leu Ala Ile Trp Arg Arg 1 5 <210> 3 <211> 208 <212> PRT <213> Artificial Sequence <220> <223> Mutant <400> 3 Met Ala Pro Leu Gly Glu Val Gly Asn Tyr Phe Gly Val Gln Asp Ala 1 5 10 15 Val Pro Phe Gly Asn Val Pro Val Leu Pro Val Asp Ser Pro Val Leu 20 25 30 Leu Ser Asp His Leu Gly Gln Ser Glu Ala Gly Gly Leu Pro Arg Gly 35 40 45 Pro Ala Val Thr Asp Leu Asp His Leu Lys Gly Ile Leu Arg Arg Arg 50 55 60 Gln Leu Tyr Cys Arg Thr Gly Phe His Leu Glu Ile Phe Pro Asn Gly 65 70 75 80 Thr Ile Gln Gly Thr Arg Lys Asp His Ser Arg Phe Gly Ile Leu Glu 85 90 95 Phe Ile Ser Ile Ala Val Gly Leu Val Ser Ile Arg Gly Val Asp Ser 100 105 110 Gly Leu Tyr Leu Gly Met Asn Glu Lys Gly Glu Leu Tyr Gly Ser Glu 115 120 125 Lys Leu Thr Gln Glu Cys Val Phe Arg Glu Gln Phe Glu Glu Asn Trp 130 135 140 Tyr Asn Thr Tyr Ser Ser Asn Leu Tyr Lys His Val Asp Thr Gly Arg 145 150 155 160 Arg Tyr Tyr Val Ala Leu Asn Lys Asp Gly Thr Pro Arg Glu Gly Thr 165 170 175 Arg Thr Lys Arg His Gln Lys Phe Thr His Phe Leu Pro Arg Pro Val 180 185 190 Asp Pro Asp Lys Val Pro Glu Leu Tyr Lys Asp Ile Leu Ser Gln Ser 195 200 205

Claims (12)

A cosmetic composition for acne skin care comprising phloretin, a protaethiamycin antibiotic peptide derivative, and a high stability epithelial cell growth factor as active ingredients. The method of claim 1, wherein the composition comprises Insulin-like Growth Factor (IGF-1), Vascular Endothelial Growth Factor (VEGF), and Keratinocyte Growth Factor-2 (Keratinocyte Growth). Factor, KGF-2), stem cell growth factor (SCF), and an acne skin care cosmetic composition further comprising at least one of a growth factor complex composed of a noggin peptide. The cosmetic composition for acne skin care according to claim 1, wherein the composition further comprises superoxide dismutase (SOD). The method of any one of claims 1 to 3, wherein the composition is a growth factor complex of 0.010 to 5.114 parts by weight, and a superoxide dismutase (SOD) of 0.004 to 0.949 parts by weight based on 70 parts by weight of the lyophilized powder. Acne skin care cosmetic composition comprising a portion. The method of claim 1 or 2, wherein the growth factor complex is 100 to 625 parts by weight of high stability fibroblast growth factor-9 variant, based on 100 parts by weight of insulin-like growth factor-1, stabilized basic fibroblast growth factor variant 100 to 625 parts by weight, 100 to 625 parts by weight of vascular endothelial cell growth factor, 100 to 625 parts by weight of keratinocyte growth factor-2, 100 to 625 parts by weight of stem cell growth factor, and 100 to 3,200 parts by weight of Nogin peptide. Acne skin care cosmetic composition, characterized in that. The method of claim 1, wherein the epidermal growth factor (EGF) variant is an amino acid residue in which isoleucine, which is the 38th amino acid of SEQ ID NO: 1, is substituted with cysteine, and aspartic acid, which is the 46th amino acid, is substituted with cysteine. A cosmetic composition for acne skin care comprising a mutant epidermal growth factor (EGF) substituted variant. The method of claim 5, wherein the high-stability fibroblast growth factor-9 variant comprises a variant in which at least one amino acid in the amino acid sequence of SEQ ID NO: 3 is substituted with cysteine, and at least one amino acid is substituted with tyrosine. Cosmetic composition for acne skin care. The method of claim 5, wherein the high-stability fibroblast growth factor-9 variant comprises a variant in which 117th glycine of SEQ ID NO: 3 is substituted with cysteine, and phenylalanine at 184th of SEQ ID NO: 1 is substituted with tyrosine. A cosmetic composition for acne skin care. The method of claim 5, wherein the stabilized basic fibroblast growth factor variant is that the 69th and 87th cysteine of SEQ ID NO: 3 is substituted with serine, the 75th alanine is substituted with cysteine, and the 50th histidine is substituted with tyrosine. Acne skin care cosmetic composition, characterized in that. The method of claim 1, wherein the amino acid sequence of the protaethiamycin peptide is Arg-Leu-Trp-Leu-Ala-Ile-Trp-Arg-Arg-NH2 of SEQ ID NO: 2, and the 7th amino acid Trp is D Acne skin care cosmetic composition, characterized in that the -form amino acid. 4. tree) acne skin care cosmetic composition, characterized in that it further comprises at least one of the extracts. According to any one of claims 1 and 3, wherein the composition is turmeric extract 0.05 to 15% by weight, gosam extract 0.05 to 15% by weight, turmeric extract 0.05 to 15% by weight, licorice extract 0.05 to 15% by weight, earl Acne skin care cosmetic composition, characterized in that it further comprises at least one extract of 0.002 to 5% by weight of the extract, and 0.002 to 5% by weight of tea tree extract.

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