KR20200143824A - Composition for promoting differentiation of skin keratinocyte comprising a SAC derivatives or a SMC derivatives - Google Patents

Abstract

The present invention relates to a composition for promoting differentiation of keratinocytes. More particularly, the present invention relates to a composition which includes an SAC derivative or an SMC derivative as an active ingredient to promote differentiation of keratinocytes, and thus has excellent anti-wrinkle efficacy, has the effect of relieving freckles and spots, regenerating skin, and restoring skin barriers, and is usable in pharmaceuticals, cosmetics, soap, body wash, shampoo and sheet masks.

Description

Composition for promoting differentiation of skin keratinocytes comprising a SAC derivatives or a SMC derivatives}

The present invention relates to a composition for promoting skin keratinocyte differentiation, and more particularly, a composition for promoting skin keratinocyte differentiation comprising a SAC (S-Allyl cysteine) derivative or SMC (S-Methyl cysteine) derivative as an active ingredient It is about.

Skin protects the body from the external environment, stores lipids and moisture, and plays an important physiological function, such as synthesizing vitamin D through interaction with ultraviolet rays. The skin is composed of three layers: the epidermis, the dermis, and the subcutaneous fat layer, and the epidermis, which is located at the outermost part, serves to protect the skin from external invading factors and irritation.

Keratinocytes are the main constituent cells that occupy more than 90% of the epidermis, and keratinocytes derived from the basal layer undergo cell division by receiving nutrients and oxygen from the blood, and this is the outermost cell through the play layer and the granule layer. It moves to the phosphorus stratum corneum and differentiates into structures with characteristics for each layer. This process takes about 14 days, and it takes about 28 days for the keratinization process in which the keratinocytes formed in the basal layer are completely eliminated from the human body. Keratinocytes mainly function as a skin barrier, and in addition, they secrete cytokines that regulate immunological reactions by UV and physiological regulators that induce inflammatory factors, and also play a role in producing and secreting them. The proliferation of keratinocytes promotes the synthesis of skin-constituent proteins such as collagen and fibronectin, induces anti-aging such as skin regeneration and wound healing, and helps to improve skin wrinkles and elasticity.

However, with age, the thickness of the skin components decreases, the skin cells are damaged, and the moisture content of the skin decreases rapidly. In addition, skin elasticity and lubricity decrease due to changes in extracellular matrix components and inhibition of cell proliferation due to aging, and as the tern-over cycle of the skin is prolonged due to the decrease in the function of keratinocytes, spots, Skin pigmentation such as freckles and wrinkles occur. Such aging of the skin is caused by structural and functional changes in the stratum corneum of the skin, and photoaging of the skin is accompanied by irregular arrangements and abnormal changes of keratinocytes. When the differentiation of keratinocytes occurs abnormally, a defect in the skin barrier function occurs, which can lead to skin dryness and chronic skin diseases such as atopy. In addition, when the skin is stimulated by ultraviolet rays and from the outside, melanin-forming cells form melanin, and these melanin are transferred to keratinocytes, and keratinocytes containing melanin move to the stratum corneum and finally fall out of the stratum corneum If the keratinization cycle of keratinocytes is prolonged, a layer of stratum corneum may accumulate on the skin, resulting in pigmentation such as spots and freckles.

Accordingly, in order to suppress and improve skin aging, raw materials with antioxidants, MMPs expression and activity inhibition, collagen production promotion, keratinocyte and fibroblast proliferation, etc. are being developed. However, conventionally developed raw materials include various problems such as toxicity and stability, and the actual effect exhibited is weak compared to known efficacy, and thus, development of a safe material having excellent efficacy and low side effects is required.

As a result of repeated research, the present inventors synthesized novel SAC derivatives and SMC derivatives based on s-allyl cysteine (SAC) or s-methyl-cysteine (SMC). The SAC derivative and SMC derivative are obtained by binding glycine, proline, and niacin to the existing SAC and SMC, which are non-toxic, have good stability, and promote the differentiation of skin keratinocytes The present invention was completed by discovering that it exhibits the effect of improving spots, freckles and wrinkles.

An object of the present invention is to provide a composition for promoting the differentiation of keratinocytes of the skin using an SAC derivative or an SMC derivative as an active ingredient, and to provide a composition that can be used in pharmaceuticals, cosmetics, soaps, body, shampoos and packs. do.

The present invention is a composition for promoting skin keratinocyte differentiation comprising a compound represented by the following Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient, and a composition that can be used in pharmaceuticals, cosmetics, soaps, bodies, shampoos and packs Provides.

<Formula 1>

{In the above formula 1, R ₁ is C ₁ -C ₆ alkyl group or a C ₂ -C ₆ alkylene group, R ₂ is NH ₂ be when R ₃ is hydrogen, if R ₂ is not a NH ₂ R ₂ and R ₃ is bonded to form a C ₂ -C ₆ heterocycle or heteroaryl including N.}

The composition containing the SAC derivative or the SMC derivative according to the present invention is excellent in improving skin wrinkles by promoting the differentiation of skin keratinocytes, and also improving pigmentation such as spots and freckles, skin regeneration, and skin barrier recovery effect. There is.

1 shows the collagen production rate by concentration of the peptide derivative according to the present invention.
Figure 2 shows the tyrosinase inhibition rate by concentration of the peptide derivative according to the invention.
3 shows the stem cell proliferation rate of the peptide derivative according to the present invention.
4 shows the DPPH radical scavenging ability of the peptide derivative according to the present invention.

Hereinafter, preferred embodiments of the present invention will be described in detail. In addition, in the following description, there are many specific matters such as specific elements, etc., 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 the knowledge of In describing 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 present invention provides a composition for promoting skin keratinocyte differentiation comprising a compound represented by the following Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient, wherein the composition includes pharmaceuticals, cosmetics, soaps, body, shampoos and Available in pack.

<Formula 1>

{In the above formula 1, R ₁ is C ₁ -C ₆ alkyl group or a C ₂ -C ₆ alkylene group, R ₂ is NH ₂ be when R ₃ is hydrogen, if R ₂ is not a NH ₂ R ₂ and R ₃ is bonded to form a C ₂ -C ₆ heterocycle or heteroaryl including N.}

In Formula 1, R ₁ is preferably a methyl group or a 1,2-propylene group, R ₂ is NH ₂ and R ₃ is a hydrogen compound.

In Formula 1, it is preferable that R ₂ and R ₃ are bonded to each other to be a C ₂ -C ₆ heterocyclic or heteroaryl containing N.

The compound represented by Formula 1 is preferably any one of the following formulas.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are for illustrating the present invention, and the scope of the present invention is not limited thereto.

[ Synthesis example 1]: SAC- Gly

The SAC-Gly compound according to the present invention may be synthesized by the reaction route of Scheme 1 below, but is not limited thereto.

<Reaction Scheme 1>

Synthesis example of Comp 3 (SAC)

To a 250 ml 3-neck flask were added 110 ml of purified water, 8.39 g (0.2394 mol) of ammonia water (NH ₄ OH), and 5 g (0.0413 mol) of L-cysteine. 7.49 g (0.062 mol) of allyl bromide was slowly added to the flask at 0-5° C., stirred at room temperature for 12 hours, and dried under reduced pressure. When concentrated to about 70%, it was filtered at 5℃ to obtain 6.2 g of crude compound 3 (SAC). 6.2 g of the obtained crude compound 3 (SAC) was added to 7 ml of a solution of H ₂ O:EtOH=2:3, heated, dissolved, and recrystallized to give 5.32 g (yield 80%) of compound 3 (SAC).

Comp 4 synthesis example

In a 250 ml 3-neck flask, 50 ml of methanol and 5 g (0.062 mol) of the compound 3 (SAC) were added, cooled to 0° C., stirred, and then 18.45 g (0.155 mol) of thionyl chloride (SOCl ₂ ) was added to the reaction solution. ) Was slowly added dropwise. When the dropwise addition was completed, the mixture was refluxed and stirred for 12 hours. The reaction solution was cooled to room temperature, concentrated under reduced pressure, and then 30 ml of toluene was added to the concentrated residue, followed by concentration (repeated twice). After completion of concentration, 5.7 g of crude compound 4 (yield 87%) was obtained. Crude compound 4 is used in the next reaction without purification.

Comp 6 synthesis example

Add 3 g (0.04 mol) of compound 5, 40 ml of isopropyl alcohol, and 40 g of 1 N NaOH to a 250 ml 3-neck flask, cool to 0℃, stir, and di-tert-butyl dicarbonate (Boc ₂ O) 11.34 g (0.052 mol) was diluted with 40 ml of isopropyl alcohol and slowly added dropwise to the reaction solution. When the addition was completed, the mixture was heated to room temperature and stirred for 12 hours. When the reaction was completed, 40 ml of n-hexane was added, stirred, and the layers were separated. The aqueous layer was adjusted to pH 3 using 2 NH ₂ SO ₄ solution, and then extracted three times with 40 ml of CHCl ₃ . The obtained organic layer was dehydrated and filtered with anhydrous sodium sulfate (Na ₂ SO ₄ ), and the filtered organic layer was concentrated under reduced pressure to obtain crude compound 6. Crude compound 6 was crystallized with n-hex to give compound 6 5 g (71% yield).

Comp 7 synthesis example

In a 250 ml 3-neck flask, methylene chloride (MC) 20 ml, compound 4 4.35 g (0.021 mol), compound 6 3 g (0.017 mol), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl ( EDCI) 3.94 g (0.021 mol) and 1-hydroxybenzotriazole hyrdate (HOBt) 2.78 g (0.021 mol) were added, cooled to 0°C, stirred for 10 minutes, and then N,N-diisopropyl-ethylamine (DIPEA) 2.66 g (0.021 mol) ) Was slowly added dropwise. After the temperature was raised to room temperature and stirred for 12 hours, 50 ml of H ₂ O and 30 ml of methylene chloride (MC) were added and stirred for 30 minutes, and the resulting solid was filtered and layered. The organic layer was washed with 30 ml of saturated brine, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtrate was concentrated under reduced pressure and purified by column to give 4.3 g (yield 75.5%) of compound 7.

Comp 8 synthesis example

4.3 g (0.013 mol) of the compound 7, 43 ml of tetrahydrofuran (THF), and 40 ml of purified water (H ₂ O) were added to a 100 ml round flask, cooled to 0°C, and stirred for 10 minutes. 0.81 g of lithium hydroxide monohydrate (LiOHH ₂ O) was added to the reaction solution, stirred at room temperature for 3 hours, and concentrated to remove THF. The concentrate was adjusted to pH 3 using 1 NH ₂ SO ₄ aqueous solution, and the aqueous layer was extracted with 50 ml of diethyl ether, and the aqueous layer was re-extracted twice with 50 ml of diethyl ether. The organic layer was collected, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ), and filtered. The filtered organic layer was concentrated under reduced pressure to give 3.2 g of compound 8 (77.7% yield).

SAC- Gly Synthesis example

In a 100 ml round flask, 3.2 g (0.01 mol) of the compound 8 and 20 ml of dioxane were added and stirred for 10 minutes. 2 ml of a 4 M HCl in dioxane solution was added to the reaction solution, followed by stirring at room temperature for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain 2.3 g of a crude compound SAC-Gly.

[ Synthesis example 2] : SMC - Gly

The SMC-Gly compound according to the present invention may be synthesized by the reaction route of Scheme 2 below, but is not limited thereto.

<Reaction Scheme 2>

Comp 10 ( SMC ) Synthesis example

Compound 2 10 g (0.0825 mol), ethanol 90 ml, and 2N NaOH aqueous solution 72 g (0.144 mol) were added to a 3-neck 250 ml flask, followed by stirring at room temperature for about 10 minutes. 11.7 g (0.0825 mol) of iodomethane was slowly added to the reaction solution, followed by stirring at room temperature for 1 hour. After the reaction is complete, conc. The pH was adjusted to 6-7 with HCl, stirred for 30 minutes, and filtered. After washing with 40 ml of H ₂ O:EtOH (1:1), the wet body was dried under reduced pressure to obtain 4.28 g (yield 38%) of crude compound 10 (SMC).

Comp 11 synthesis example

50 ml of methanol and 5 g (0.037 mol) of Compound 10 (SMC) were added to a 250 ml 3-neck flask, followed by cooling and stirring at 0°C. Trimethylsilane chloride (TMSCl) 8.04 g (0.074 mol) was slowly added dropwise to the reaction solution, followed by stirring at room temperature for 12 hours, and then concentrated under reduced pressure. 30 ml of toluene was added to the concentrated residue, followed by concentration (repeated twice). After completion of concentration, 6.11 g (yield 89.0%) of crude compound 11 was obtained. The crude compound 11 is used in the next reaction without purification.

Comp 6 synthesis example

It was synthesized in the same manner as in the synthesis method of Comp 6 in the synthesis reaction route of SAC-Gly.

Comp 12 synthesis example

In a 250 ml 3-neck flask, methylene chloride (MC) 30 ml, compound 11 3.82 g (0.021 mol), compound 6 3 g (0.017 mol), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl ( EDCI) 3.94 g (0.021 mol) and 1-hydroxybenzotriazole hyrdate (HOBt) 2.78 g (0.021 mol) were added, cooled to 0°C, stirred for 10 minutes, and then N,N-diisopropyl-ethylamine (DIPEA) 2.66 g (0.021 mol) ) Was slowly added dropwise. After the temperature was raised to room temperature and stirred for 12 hours, 50 ml of H ₂ O and 30 ml of methylene chloride (MC) were added and stirred for 30 minutes, and the resulting solid was filtered and layered. The organic layer was washed with 30 ml of saturated brine, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtrate was concentrated under reduced pressure and purified by column to obtain 4.1 g (78.1% yield) of compound 12.

Comp 13 synthesis example

4.1 g (0.013 mol) of the compound 12, 41 ml of tetrahydrofuran (THF), and 41 ml of purified water (H ₂ O) were added to a 100 ml round flask, cooled to 0°C, and stirred for 10 minutes. Lithium hydroxide monohydrate (LiOHH ₂ O) 0.84 g was added to the reaction solution, stirred at room temperature for 3 hours, and then concentrated to remove THF. The concentrate was adjusted to pH 3 using 1 NH ₂ SO ₄ aqueous solution, and the aqueous layer was extracted with 40 ml of diethyl ether, and the aqueous layer was re-extracted twice with 40 ml of diethyl ether. The organic layer was collected, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ), filtered, and then the filtered organic layer was concentrated under reduced pressure to obtain 3.1 g of compound 13 (yield 79.2%).

SMC - Gly Synthesis example

Compound 13 3.2 g (0.011 mol) and dioxane 20 ml were added to a 100 ml round flask, followed by stirring for 10 minutes. 2 ml of a 4 M HCl in dioxane solution was added to the reaction solution, stirred at room temperature for 12 hours, and then concentrated under reduced pressure to obtain 2.33 g of a crude compound SMC-Gly. Ethylacetate and n-Hexane were used for crystallization to obtain 2.1 g (yield 84%) of the compound SMC-Gly.

[ Synthesis example 3]: SAC-Pro

The SAC-Pro compound according to the present invention may be synthesized by the reaction route of Scheme 3 below, but is not limited thereto.

<Reaction Scheme 3>

Synthesis example of Comp 3 (SAC)

88 ml of purified water, 6.71 g (0.191 mol) of ammonia water (NH ₄ OH), and 4 g (0.033 mol) of L-cysteine were added to a 250 ml 3-neck flask. Allyl bromide 5.99 g (0.05 mol) was slowly added to the mixture at 0 to 5°C, stirred at room temperature for 12 hours, and dried under reduced pressure. When concentrated to about 70%, it was filtered at 5°C to obtain 4.79 g of crude compound 3 (SAC). 4.79 g of the obtained crude compound 3 (SAC) was added to 48 ml of a H ₂ O:EtOH=2:3 solution, heated, dissolved, and recrystallized to give 4.3 g (yield 81%) of compound 3 (SAC).

Comp 4 synthesis example

To a 250 ml 3-neck flask, 43 ml of methanol and 4 g (0.025 mol) of the compound 3 (SAC) were added, followed by cooling and stirring at 0°C. Thionyl chloride (SOCl ₂ ) 18.45 g (0.124 mol) was slowly added dropwise to the reaction solution, refluxed and stirred for 12 hours, and the reaction solution was cooled to room temperature and concentrated under reduced pressure. After adding 30 ml of toluene to the concentrated residue, it was concentrated (repeated twice). After concentration was completed, 4.5 g of crude compound 4 (yield 85.7%) was obtained. Crude compound 4 is used in the next reaction without purification.

Comp 15 synthesis example

In a 250 ml 3-neck flask, 3 g (0.026 mol) of compound 14, 60 ml of methylene chloride (MC), and 3.05 g (0.03 mol) of triethylamine (TEA) were added, followed by cooling and stirring at 0°C. 8.4 g (0.038 mol) of di-tert-butyl dicarbonate (Boc ₂ O) was slowly added dropwise to the reaction solution, and then heated to room temperature and stirred for 5 hours. After the reaction was completed, 15 ml of saturated citric acid aqueous solution was added and stirred, followed by layer separation. The organic layer was washed with saturated NaCl aqueous solution, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtered organic layer was concentrated under reduced pressure to obtain crude compound 15. The crude compound 15 was crystallized with ethyl acetate and n-hex to give 4.7 g of compound 15 (yield 84%).

Comp 16 synthesis example

In a 250 ml 3-neck flask, 35 ml of methylene chloride (MC), 4.13 g (0.02 mol) of compound 4, 3.5 g (0.016 mol) of compound 15, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl ( EDCI) 3.74 g (0.02 mol) and 1-hydroxybenzotriazole hyrdate (HOBt) 2.64 g (0.02 mol) were added, cooled to 0℃, stirred for 10 minutes, and then N,N-diisopropyl-ethylamine (DIPEA) 2.52 g (0.02 mol) ) Was slowly added dropwise. After heating to room temperature and stirring for 12 hours, 50 ml of H ₂ O and 30 ml of methylene chloride (MC) were added and stirred for 30 minutes, and the resulting solid was filtered and layered. The organic layer was washed with 30 ml of saturated brine, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtrate was concentrated under reduced pressure and purified by column to give 3.93 g (82% yield) of compound 16.

Comp 17 synthesis example

Compound 16 was added to THF and purified water in a 100 ml round flask, cooled to 0°C, and stirred for 10 minutes. LiOHH ₂ O was added to the reaction solution, stirred at room temperature for 3 hours, and then concentrated to remove THF. The concentrate was adjusted to pH 3 using 1 NH ₂ SO ₄ aqueous solution, and the aqueous layer was extracted with diethyl ether, and the aqueous layer was re-extracted twice with 40 ml of diethyl ether. The organic layer was collected, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ), filtered, and then the filtered organic layer was concentrated under reduced pressure to obtain a compound SAC-Pro (yield 80%).

[ Synthesis example 4] : SMC -Pro

The SMC-Pro compound according to the present invention may be synthesized by the reaction route of Scheme 4 below, but is not limited thereto.

<Reaction Scheme 4>

Comp 10 ( SMC ) Synthesis example

15 g (0.124 mol) of compound 2, 135 ml of ethanol, and 108 g (0.216 mol) of 2N NaOH aqueous solution were added to a 250 ml 3-neck flask, followed by stirring at room temperature for about 10 minutes. 17.55 g (0.124 mol) of iodomethane was slowly added to the reaction solution, and after stirring at room temperature for 1 hour, conc. The pH was adjusted to 6-7 with HCl. After stirring for 30 minutes, it was filtered and washed with 60 ml of H ₂ O:EtOH=1:1 solution. The wet body was dried under reduced pressure to obtain 6.3 g (yield 37%) of crude compound 10 (SMC).

Comp 11 synthesis example

It was synthesized in the same manner as in the synthesis method of Comp 11 in the synthesis reaction route of SMC-Gly.

Comp 15 synthesis example

It was synthesized in the same manner as in the synthesis method of Comp 15 in the synthesis reaction route of SAC-pro.

Comp 18 synthesis example

In a 250 ml 3-neck flask, methylene chloride (MC) 39 ml, compound 11 3.99 g (0.021 mol), compound 15 3.85 g (0.018 mol), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl ( EDCI) 4.11 g (0.021 mol) and 1-hydroxybenzotriazole hyrdate (HOBt) 2.9 g (0.021 mol) were added, cooled to 0°C, stirred for 10 minutes, and then N,N-diisopropyl-ethylamine (DIPEA) 2.77 g (0.021 mol) ) Was slowly added dropwise. After heating to room temperature and stirring for 12 hours, 50 ml of H ₂ O and 35 ml of methylene chloride (MC) were added and stirred for 30 minutes, and the resulting solid was filtered and layered. The organic layer was washed with 30 ml of saturated brine, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtrate was concentrated under reduced pressure to obtain 4.9 g of crude compound 18 (crude yield: 93%).

Comp 19 synthesis example

It was carried out in the same manner as the method of preparing the compound SAC-Pro, and after producing compound 19, the final compound SMC-Pro was obtained in 82% yield.

[ Synthesis example 5]: SAC- Nico

The SAC-Nico compound according to the present invention may be synthesized by the reaction route of Scheme 5 below, but is not limited thereto.

<Reaction Scheme 5>

Synthesis example of Comp 3 (SAC)

To a 250 ml 3-neck flask, add 110 ml of purified water, 8.39 g (0.2394 mol) of ammonia water (NH ₄ OH), and 5 g (0.0413 mol) of L-cysteine, and then 7.49 g (0.062 mol) of allyl bromide at 0~5℃. Slowly added at, and stirred at room temperature for 12 hours. After the reaction was completed, dried under reduced pressure and concentrated to about 70%, filtered at 5°C to obtain 6.4 g of crude compound 3 (SAC). 6.4 g of the obtained crude compound 3 (SAC) was added to 64 ml of a solution of H ₂ O:EtOH=2:3, heated, dissolved, and recrystallized to give 5.4 g of compound 3 (yield 81.2%).

Comp 4 synthesis example

To a 250 ml 3-neck flask, 50 ml of methanol and 5 g (0.062 mol) of the compound 3 (SAC) were added, followed by cooling and stirring at 0°C. Thionyl chloride (SOCl ₂ ) 18.45 g (0.155 mol) was slowly added dropwise to the reaction solution, refluxed and stirred for 12 hours, and then the reaction solution was cooled to room temperature and concentrated under reduced pressure. 30 ml of toluene was added to the concentrated residue, followed by concentration (repeated twice). After the concentration was completed, 5.83 g (yield 89.0%) of crude compound 4 was obtained. Crude compound 4 is used in the next reaction without purification.

Comp 21 synthesis example

In a 250 ml 3-neck flask, methylene chloride (MC) 22 ml, compound 4 4.54 g (0.021 mol), compound 20 2.20 g (0.018 mol), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl (EDCI ) 4.11 g (0.021 mol), 1-hydroxybenzotriazole hyrdate (HOBt) 2.9 g (0.021 mol) was added, cooled to 0°C, stirred for 10 minutes, and then N,N-diisopropyl-ethylamine (DIPEA) 2.77 g (0.021 mol) Was slowly added dropwise. After the temperature was raised to room temperature and stirred for 12 hours, 50 ml of H ₂ O and 30 ml of methylene chloride (MC) were added and stirred for 30 minutes. The resulting solid was filtered and layered. The organic layer was washed with 30 ml of saturated brine, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtrate was concentrated under reduced pressure and purified by column to give 4.2 g (yield 87.7%) of compound 21.

Comp 22 synthesis example

4.17 g (0.013 mol) of the compound 21, 42 ml of tetrahydrofuran (THF), and 42 ml of purified water (H ₂ O) were added to a 100 ml round flask, cooled to 0°C, and stirred for 10 minutes. Lithium hydroxide monohydrate (LiOHH ₂ O) 0.79 g was added to the reaction solution, stirred at room temperature for 3 hours, and concentrated to remove THF. After washing the aqueous layer with 50 ml of diethyl ether, the aqueous layer was adjusted to pH 3 using 1 NH ₂ SO ₄ aqueous solution. The resulting solid was filtered, washed sufficiently with purified water, and the wet body was concentrated under reduced pressure to give 2.3 g (yield 68.8%) of compound 22.

SAC- Nico Synthesis example

Compound 22 was added to a mixed solvent of 200 ml of methylene chloride and 150 ml of t-butanol to dissolve, and then t-sodium butoxychloride was slowly added at room temperature. After stirring for about 30 minutes, the suspension was removed by filtering through a filter paper. The compound SAC-Nico was obtained in 88% yield.

[ Synthesis example 6]: SMC - Nico

The SMC-Nico compound according to the present invention may be synthesized by the reaction route of Scheme 6 below, but is not limited thereto.

<Reaction Scheme 6>

Comp 10 ( SMC ) Synthesis example

15 g (0.124 mol) of compound 2, 135 ml of ethanol, and 108 g (0.216 mol) of 2N NaOH aqueous solution were added to a 250 ml 3-neck flask, followed by stirring at room temperature for about 10 minutes. 17.55 g (0.124 mol) of iodomethane was slowly added to the reaction solution, and after stirring at room temperature for 1 hour, conc. The pH was adjusted to 6-7 with HCl. After stirring for 30 minutes, filtered, washed with 60 ml of H ₂ O:EtOH=1:1 solution, and dried under reduced pressure to give crude compound 10 (SMC) 6.5 g (yield 38%).

Comp 11 synthesis example

50 ml of methanol and 5 g (0.037 mol) of compound 10 (SMC) were added to a 250 ml 3-neck flask, followed by cooling and stirring at 0°C. Trimethylsilane chloride (TMSCl) 8.04 g (0.074 mol) was slowly added dropwise to the reaction solution, and the mixture was stirred at room temperature for 12 hours and then concentrated under reduced pressure. After adding 30 ml of toluene to the concentrated residue, it was concentrated (repeated twice). After completion of concentration, 5.97 g (yield 87.0%) of crude compound 11 was obtained. Crude compound 11 is used in the next reaction without purification.

Comp 23 synthesis example

In a 250 ml 3-neck flask, methylene chloride (MC) 22 ml, compound 11 3.98 g (0.021 mol), compound 20 2.20 g (0.018 mol), 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl (EDCI ) 4.11 g (0.021 mol), 1-hydroxybenzotriazole hyrdate (HOBt) 2.9 g (0.021 mol) was added, cooled to 0°C, stirred for 10 minutes, and then N,N-diisopropyl-ethylamine (DIPEA) 2.77 g (0.021 mol) Was slowly added dropwise. After heating to room temperature and stirring for 12 hours, 50 ml of H ₂ O and 30 ml of methylene chloride (MC) were added and stirred for 30 minutes, and the resulting solid was filtered and layered. The organic layer was washed with 30 ml of saturated brine, dehydrated with anhydrous magnesium sulfate (MgSO ₄ ) and filtered. The filtrate was concentrated under reduced pressure and purified by column to obtain 3.6 g of compound 23 (79.2% yield).

Comp 24 synthesis example

4.17 g (0.013 mol) of the compound 23, 42 ml of tetrahydrofuran (THF) and 42 ml of purified water (H ₂ O) were added to a 100 ml round flask, cooled to 0°C, and stirred for 10 minutes. Lithium hydroxide monohydrate (LiOHH ₂ O) 0.79 g was added to the reaction solution, stirred at room temperature for 3 hours, and concentrated to remove THF. After washing the aqueous layer with 50 ml of diethyl ether, the aqueous layer was adjusted to pH 3 using 1 NH ₂ SO ₄ aqueous solution. The resulting solid was filtered, washed sufficiently with purified water, and the wet body was concentrated under reduced pressure to obtain compound 24.

SMC - Nico Synthesis example

Compound 24 was added to a mixed solvent of 200 ml of methylene chloride and 150 ml of t-butanol to dissolve, and then t-sodium butoxychloride was slowly added at room temperature. After stirring for about 30 minutes, the suspension was removed by filtering through a filter paper. The compound SMC-Nico was obtained in a yield of 85%.

[ Example 1] Keratinocytes Measurement of differentiation promoting efficacy

Was introduced into human keratinocytes in the culture flask culture grown to about 80%, SAC peptides and specific concentration of each of the peptide derivative (10 ^-7, 10 ^-8, 10 ^-9, 10 ^-10, 10 ^{- 11} , 10 ^-12 , 10 ^-13 M) and incubated for 5 days. Then, after washing the cultured cells with PBS (Phosphate burrered saline), 10 mM Tris-HCl buffer (Tris-HCl, pH 7.4) containing 2% SDS (sodium dodecyl sulfate) and 20 mM DTT (Dithiothreitol) was added. After sonication and centrifugation, the precipitate was suspended again in 1 ml of PBS, and the absorbance was measured at 420 nm. Based on the negative control, distilled water (0%), the amount of increase by each sample was compared, and it is shown in Table 1 below. As shown in Table 1 below, it was confirmed that the peptide derivative of the present invention has a better effect of promoting keratinocyte differentiation compared to SAC, and it was found to promote keratinocyte differentiation in an overall concentration-dependent manner.

Standard solution concentration (M) Effect of promoting keratinocyte differentiation (%) SAC SAC-Gly SMC-Gly SAC-Pro SMC-Pro SAC-Nico SMC-Nico 10 ^-7 142 157 140 161 149 162 145 10 ^-8 140 157 143 172 153 170 145 10 ^-9 145 154 151 166 157 164 151 10 ^-10 139 149 142 153 143 156 140 10 ^-11 112 124 134 142 132 141 120 10 ^-12 97 101 108 109 107 107 103 10 ^-13 80 87 84 81 88 79 80

[ Example 2] Measurement of collagen biosynthesis promotion efficacy

Human dermal fibroblasts were inoculated in a 6-well microplate containing DMEM (Dulbecco's Modified Eagle's Medium) medium at a concentration of 2×10 ⁵ cells/well and incubated at 37°C, 5% CO ₂ for 24 hours. Cultured. After 24 hours, the DMEM culture medium containing 0.05% FBS (fetal bovine serum) After removal of the medium in each well the concentration of SAC peptides and each peptide-specific derivative (10 ^-7, 10 ^-8, 10 ^{-9, 10- 10} , 10 ^-11 , 10 ^-12 , 10 ^-13 M), and then incubated again for 24 hours. After 24 hours, the medium was collected to prepare a sample, and a C-terminal peptide of procollagen type I was prepared using a collagen measurement kit (Procollagen Type I C-peptide (PIP) EIA kit (MK101), Takara, Kyoto, Japan). The amount of (procollagen type I C-peptide; PIP) was measured. The standard solution included in the collagen measurement kit was diluted by concentration, absorbance was measured at 450 nm, and a standard concentration curve was prepared. 100 µl of an antibody solution with POD (peroxidase) enzyme attached to each well was added, and 20 µl of each of the standard solution diluted by concentration and the supernatant of the sample were added and mixed. After allowing the mixture to stand at 37° C. for 3 hours, all contents were removed, and each well was washed 4 times with 400 μl of PBS (phosphate buffered saline). After adding 100 µl of the substrate solution to the well, it was allowed to stand at room temperature for 15 minutes. To stop the reaction, 100 µl of 1 N sulfuric acid solution was added and mixed, and the absorbance was measured at 450 nm. This experiment was performed twice at the same time to obtain the average value, which is shown in Table 2 and FIG. 1 below. As a result of the experiment, each peptide derivative promoted collagen biosynthesis, and each peptide derivative showed the highest collagen production rate between 10 ^-9 ~ 10 ^-7 M concentration. Through this, it was confirmed that the anti-wrinkle effect of the peptide derivative according to the present invention is excellent.

Standard solution concentration (M) Relative collagen synthesis (of control, %) SAC SAC-Gly SMC-Gly SAC-Pro SMC-Pro SAC-Nico SMC-Nico 10 ^-7 143 150 138 176 161 188 158 10 ^-8 142 154 154 191 180 186 153 10 ^-9 145 160 163 180 178 165 160 10 ^-10 130 152 163 158 170 152 161 10 ^-11 130 134 164 150 127 150 130 10 ^-12 120 113 122 122 120 120 122 10 ^-13 100 117 111 103 112 100 102

[ Example 3] Tyrosinase activity Inhibitory ability Measure

Tyrosinase is an enzyme that promotes the production of melanin by oxidizing tyrosine in melanocytes present in the base layer of the epidermis of the skin.

As tyrosinase, a product of Sigma, extracted from mushrooms, was used. First, the substrate tyrosine was dissolved in a sodium phosphate buffer solution (0.1 M, pH 6.8) at a concentration of 0.3 mg/ml, and then 334 µl of the solution was taken and put into a test tube. Then, a sodium phosphate buffer solution (0.1 M, pH 6.8) 576 μl were added. SAC peptide and each of the peptide derivatives of the present invention by concentration (10 ^-7 , 10 ^-8 , 10 ^-9 , 10 ^-10 , 10 ^-11 , 10 ^-12 , 10 ^-13 M) by 70 µl each in the test tube It was added to the solution and reacted for 10 minutes in a 37° C. constant temperature bath, and 70 μl of purified water was used as a control. 20 µl of 1700 unit/ml tyrosinase solution was added to the reaction solutions of the experimental group and the control group, so that the total volume was 1 ml, and the reaction was carried out in a 37°C incubator for 30 minutes. The test tube containing the reaction solution was quenched in ice water to stop the activity of tyrosinase, and the absorbance was measured at a wavelength of 470 nm. The blank of the experimental group is a reaction product excluding the tyrosinase solution, that is, a reaction product of tyrosine, each peptide derivative, and a sodium phosphate buffer solution, and the blank of the control group was added purified water instead of the peptide derivative from the blank of the experimental group. The inhibition rate of tyrosinase for each concentration of the peptide derivative and the control group was calculated using the following equation, and the results are shown in Table 3 and FIG. 2 below.

Tyrosinase inhibition rate (%) = 100-{[(A-B)-(C-D)]/(A-B)} × 100 (%)

A: absorbance of the control group, B: blank absorbance of the control group, C: absorbance of the experimental group, D: the blank absorbance of the experimental group

As shown in Table 3 below, it was found to inhibit tyrosinase activity in an overall concentration-dependent manner.

IC ₅₀ (M) Relative tyrosinase activity inhibition (of control, %) 10 ^-7 M 10 ^-8 M 10 ^-9 M 10 ^-10 M 10 ^-11 M 10 ^-12 M 10 ^-13 M SAC <10 ^-11 94 88 78 61 55 46 40 SAC-Gly <10 ^-11 95 79 78 66 43 50 35 SMC-Gly <10 ^-10 95 87 70 52 55 45 40 SAC-Pro <10 ^-10 96 87 71 67 57 51 44 SMC-Pro <10 ^-11 89 88 76 62 57 54 53 SAC-Nico <10 ^-11 96 91 75 61 56 44 45 SMC-Nico <10 ^-11 96 50 63 62 57 51 41

[ Example 4] Stem cell proliferation promotion efficacy evaluation

Stem cell proliferation promoting efficacy was evaluated by BrdU proliferation assay (Calbiochem), which is a method of comparing the amount of cells dividing with a BrdU (bromodeoxyuridine) label. Cord-stem-derived skin stem cells were inoculated at a concentration of 3×10 ³ cells/cm ² , and SAC peptide and each peptide derivative according to the present invention were added at a concentration of 10 ^-9 M and cultured for 3 to 5 days, and stem cell proliferation The degree was measured. As a control, cultured in DMEM-serum free was used. As a result, as shown in Table 4 and FIG. 3, each of the peptide derivatives showed an effect of promoting skin stem cell proliferation of about 1.6 to 2.1 times, and it was confirmed that the skin regeneration effect was excellent.

SAC SAC-Gly SMC-Gly SAC-Pro SMC-Pro SAC-Nico SMC-Nico Control Relative Stem cell viability (of control, %) 213 200 176 164 181 210 196 100

[ Example 5] Antioxidant efficacy evaluation

The antioxidant activity of the peptide derivative was measured using the DPPH radical scavenging ability assay. 20 µl of SAC peptide (5 μM) and each peptide derivative by concentration (1, 5, 10, 20 mg/ml) were taken, and 800 µl of 0.2 mM 1,1-diphenyl-2-picryhydrazyl (DPPH) solution was added. After mixing well, it was reacted in the dark for 30 minutes to measure absorbance at 518 nm, and DPPH radical scavenging ability according to the concentration was confirmed. As shown in FIG. 6 below, it was confirmed that DPPH free radical scavenging ability increased as the concentration of all of the peptide derivatives increased, and showed similar efficacy compared to the SAC peptide 5 μM.

[ Example 6] Cream production

A cream containing the peptide derivative of the present invention was prepared according to the composition shown in Table 5 below. First, the aqueous phase and the oil phase were heated, uniformly mixed and dissolved, respectively, and then the oil phase was added to the aqueous phase at 75° C. and mixed and solubilized. After cooling to 50° C., the solubilized additive phase 1 was added to the phase in which the peptide derivative was dissolved, and then the additive phase 2 was mixed.

Prize ingredient content (%) Awards Purified water to 100 Moisturizing ingredients 10 to 25 Thickener Appropriate amount Paid PEG-100 Stearate 0.1 to 2 Glyceryl stearate 0.1 to 2 Polysorbate 60 0.1 to 2 Stearic Acid 0.1 to 2 Cetearyl alcohol 0.1 to 2 Capric Caprylic Triglyceride 10 to 30 Tocopheryl Acetate 0.1 to 0.5 Additive phase 1 Dipropylene glycol 0.23 glycerin 0.02 Western Rape Sterol 0.05 PEG-5 Lepe Seed Sterol 0.05 cholesterol 0.05 Setes-3 0.03 Setes-5 0.03 1,2-hexanediol 0.05 Hydrogenated Lecithin 0.0.5 Sodium Stearoyl Glutamate 0.02 Disodium EDTA 0.01 Octyldodecanol 0.25 Purified water 1.56 Peptide derivative 0.10 Additive phase 2 incense Appropriate amount Preservative Appropriate amount Other additives Appropriate amount

[ Example 7] Measurement of skin keratin turnover promotion effect

The effect of promoting exfoliation was measured in 20 healthy adult men and women. First, before applying the sample, measure the inner color of the upper arm of the test subject with a chroma meter (CR300, Minolta, Japan), and then add 0.4 ml of dihydroxyacetone (DHA) at a concentration of 10% by weight into the heel top chamber and place the upper arm of the arm. Attached to the inside for 6 hours. After 24 hours, the color of the area colored brown by DHA was measured to compare the color difference from before the sample application. Thereafter, the cream of Example 6 was applied twice a day, and the degree of bleaching every day was measured with a chromameter to measure the time taken to return to the original skin color, and the exfoliation rate was calculated by the following equation. At this time, the turnover period means the time it takes for the existing stratum corneum to be replaced with the new stratum corneum, and in this embodiment, it means the time it takes for the skin colored by DHA to completely return to its original state. As a comparative example, a cream prepared by adding SAC instead of the peptide derivative of the present invention was used.

Exfoliation rate (%) = (Turnover cycle of blank-Turnover cycle when applying sample)/Turnover cycle of blank × 100

As shown in Table 6 below, the keratin exfoliation rate of the SAC, which is a comparative example, is 14.06%, whereas the peptide derivative of the present invention has an average keratin exfoliation rate of about 27%, so the turnover period is short and the exfoliation effect is excellent. As can be seen, it was confirmed that it has an excellent effect on improving pigmentation such as freckles and spots as it induces rapid peeling of melanin.

Turnover cycle (days) Exfoliation rate (%) Control (no treatment group) 19.2 - SAC 16.5±2.4 14.06 SAC-Gly 14.2±2.5 26.04 SMC-Gly 14.0±2.8 27.08 sAC-Pro 13.8±3.1 28.13 SMC-Pro 13.6±3.4 29.17 SAC-Nico 13.6±3.6 29.17 SMC-Nico 14.1±3.4 26.56

[ Example 8] Efficacy evaluation of skin barrier function

20 healthy adult men and women were selected, and acetone was applied twice a day for 4 days in a size of 2×2 cm ² on the lower arm of both arms, and then the skin barrier was damaged on the 5th and 6th day with acetone. After damaging the skin area, the cream of Example 6 was applied at 300 μL per 2×2 cm ^2, and the degree of recovery of Transepidermal Water Loss (TEWL) was measured. The degree of recovery of transdermal water loss was calculated by the following method. As a comparative example, a cream prepared by adding SAC instead of the peptide derivative of the present invention was used.

Percutaneous moisture loss recovery (%) = (TEWL immediately after injury-TEWL after cream application)/(TEWL immediately after injury-Initial TEWL)×100

As shown in Table 7 below, in the case of the cream containing the peptide derivative of the present invention compared to SAC, it was confirmed that the recovery rate of the skin barrier was high.

Transdermal water loss recovery (%) Day 5 Day 6 SAC 25.2±0.34 52.9±0.21 SAC-Gly 30.0±0.35 71.1±0.47 SMC-Gly 31.9±0.21 72.8±0.71 SAC-Pro 35.4±0.65 75.3±0.41 SMC-Pro 32.0±0.42 74.2±0.48 SAC-Nico 33.1±0.28 73.1±0.24 SMC-Nico 32.8±0.34 71.2±0.33

The above description is only illustrative of the present invention, and those of ordinary skill in the art to which the present invention pertains will be able to make various modifications without departing from the essential characteristics of the present invention. Accordingly, the embodiments disclosed in the present specification are not intended to limit the present invention, but to explain the present invention, and the spirit and scope of the present invention are not limited by these embodiments. The scope of protection of the present invention should be interpreted by the claims, and all technologies within the scope equivalent thereto should be interpreted as being included in the scope of the present invention.

Claims (12)

A composition for promoting skin keratinocyte differentiation, comprising a compound represented by the following Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient
<Formula 1>

{In the above formula 1, R ₁ is C ₁ -C ₆ alkyl group or a C ₂ -C ₆ alkylene group, R ₂ is NH ₂ be when R ₃ is hydrogen, if R ₂ is not a NH ₂ R ₂ and R ₃ is bonded to form a C ₂ -C ₆ heterocycle or heteroaryl including N.}
The composition of claim 1, wherein the composition has an effect of improving spots, freckles and wrinkles.
The composition according to claim 1, wherein R ₁ in Formula 1 is a methyl group or a 1,2-propylene group.
The composition of claim 1, wherein in Formula 1, R ₂ is NH ₂ and R ₃ is hydrogen.
The composition according to claim 1, wherein in Formula 1, R ₂ and R ₃ are bonded to each other to form a C ₂ -C ₆ heterocyclic or heteroaryl containing N
The composition of claim 1, wherein the compound represented by Chemical Formula 1 is any one of the following Chemical Formulas

A cosmetic composition for promoting skin keratinocyte differentiation, comprising a compound represented by the following Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient
<Formula 1>

{In the above formula 1, R ₁ is C ₁ -C ₆ alkyl group or a C ₂ -C ₆ alkylene group, R ₂ is NH ₂ be when R ₃ is hydrogen, if R ₂ is not a NH ₂ R ₂ and R ₃ is bonded to form a C ₂ -C ₆ heterocycle or heteroaryl including N.}
The cosmetic composition according to claim 7, wherein the cosmetic composition has an effect of improving spots, freckles and wrinkles.
The cosmetic composition according to claim 7, wherein R ₁ in Formula 1 is a methyl group or a 1,2-propylene group.
The cosmetic composition according to claim 7, wherein in Formula 1, R ₂ is NH ₂ and R ₃ is hydrogen.
The cosmetic composition according to claim 7, wherein R ₂ and R ₃ are bonded to each other in Formula 1 to form a C ₂ -C ₆ heterocycle or heteroaryl containing N
The cosmetic composition of claim 7, wherein the compound represented by Chemical Formula 1 is any one of the following Chemical Formulas

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