KR102608639B1 - Manufacturing method of encapsulated keratin and Cosmetic composition for improving skin barrier with encapsulated keratin manufactured therefrom - Google Patents
Manufacturing method of encapsulated keratin and Cosmetic composition for improving skin barrier with encapsulated keratin manufactured therefrom Download PDFInfo
- Publication number
- KR102608639B1 KR102608639B1 KR1020210011431A KR20210011431A KR102608639B1 KR 102608639 B1 KR102608639 B1 KR 102608639B1 KR 1020210011431 A KR1020210011431 A KR 1020210011431A KR 20210011431 A KR20210011431 A KR 20210011431A KR 102608639 B1 KR102608639 B1 KR 102608639B1
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- KR
- South Korea
- Prior art keywords
- keratin
- weight
- encapsulated
- present
- skin
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Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/02—Cosmetics or similar toiletry preparations characterised by special physical form
- A61K8/11—Encapsulated compositions
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/49—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds
- A61K8/494—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with more than one nitrogen as the only hetero atom
- A61K8/4953—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with more than one nitrogen as the only hetero atom containing pyrimidine ring derivatives, e.g. minoxidil
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Abstract
본 발명은 캡슐화케라틴의 제조방법 및 이의 방법으로 제조된 캡술화케라틴을 포함하는 피부장벽 개선용 화장료 조성물에 관한 것으로, 불용성의 케라틴을 사이클로덱스트린 및 그 유도체들로부터 안정화시키고, 캡슐화를 통해 세포독성이 낮고, 항염증효과 및 세포재생 개선 효과가 우수하며, 각질형성세포 분화 유도에 따른 피부 장벽 강화 효과를 발휘하는 화장료 조성물을 제공한다.The present invention relates to a method for producing encapsulated keratin and a cosmetic composition for improving the skin barrier containing encapsulated keratin produced by this method, which stabilizes insoluble keratin from cyclodextrin and its derivatives and prevents cytotoxicity through encapsulation. Provides a cosmetic composition that has a low, excellent anti-inflammatory effect and cell regeneration improvement effect, and has a skin barrier strengthening effect by inducing keratinocyte differentiation.
Description
본 발명은 캡슐화케라틴의 제조방법 및 이의 방법으로 제조된 캡술화케라틴을 포함하는 피부장벽 개선용 화장료 조성물에 관한 것으로, 구체적으로는 불용성의 케라틴을 사이클로덱스트린 및 그 유도체들로부터 안정화시키고, 캡슐화를 통해 세포독성이 낮고, 항염증효과 및 세포재생 개선 효과가 있는 화장료로 이용하는 기술에 관한 것이다.The present invention relates to a method for producing encapsulated keratin and a cosmetic composition for improving the skin barrier containing encapsulated keratin produced by the method. Specifically, the present invention relates to stabilizing insoluble keratin from cyclodextrin and its derivatives through encapsulation. This relates to technology for use as a cosmetic with low cytotoxicity, anti-inflammatory effects, and improved cell regeneration effects.
피부 장벽은 피부의 가장 바깥쪽에 있는 각질층으로, 외부 자극으로부터 피부를 보호하며, 보습효과를 유지하는 기능을 한다. 각질층은 호산성 각화세포가 겹겹이 쌓여있는 형태이며, 케라틴(keratin) 58%, 천연보습인자(NMF, Natural Moisturizing Factor) 38%, 세포간지질(intercellular lipid) 11%로 구성되어 있다. 케라틴 단백질로 구성된 각질세포는 자외선 차단, 물리적 장벽의 역할을 담당하고, 각질세포간의 접착제 역할을 하는 세포간지질은 외부로부터의 물질 흡수 방지, 항균 장벽 기능을 담당한다.The skin barrier is the outermost stratum corneum of the skin, and its function is to protect the skin from external stimuli and maintain a moisturizing effect. The stratum corneum is a layer of eosinophilic keratinocytes and is composed of 58% keratin, 38% natural moisturizing factor (NMF), and 11% intercellular lipid. Keratinocytes, composed of keratin protein, block ultraviolet rays and act as a physical barrier, and intercellular lipids, which act as adhesives between keratinocytes, prevent the absorption of substances from the outside and function as an antibacterial barrier.
케라틴은 각질형성세포로부터 생성되는데, 각질형성세포는 기저층(표피의 가장 아래층)에서 형성되어 세포분열을 하면서 표피의 위층으로 이동하며 피부표면에서 탈락한다. 세포 내의 케라틴은 세포골격(cytoskeleton)을 이루는 주요단백질로, 외부에서 가해지는 손상으로부터 세포를 보호해 주는 역할을 하며, 세포생존에 관여하는 다른 단백질들과 상호작용을 통하여 신호전달체계의 조절기능을 수행한다. 케라틴은 표피에 가장 풍부하게 존재하는 세포간 섬유상 연결 단백질로서 케라틴은 분자량이 크며 시스틴 이중황화결합(cystine disulfide bonds)에 의해서 연결되어 있다. 이에 따라 오래된 각질 세포가 정상적으로 탈락하며 새로운 피부층으로 대체됨에 따라 건강한 장벽기능을 유지할 수 있다. 또한, 세포간 지질은 각질형성세포인 케라티노사이트(keratinocyte)의 효소에 의해 분해되어 세라마이드, 지방산, 콜레스테롤 등을 생성하는데, 이에 따라 피부의 지질 장벽을 형성한다고 알려져 있다. 즉, 피부 장벽이 약화될 경우 피부 세포의 재생 속도가 늦어져 표피 노화가 가속화되며, 외부 자극에 피부가 노출되거나 피부 면역력이 약해지고, 피부 수분이 빠져나가거나 부족해지는 등의 문제가 발생할 수 있다.Keratin is produced from keratinocytes. Keratinocytes are formed in the basal layer (lowest layer of the epidermis), undergo cell division, move to the upper layer of the epidermis, and fall off from the skin surface. Keratin within cells is a major protein that makes up the cytoskeleton, and plays a role in protecting cells from external damage, and regulates the signal transduction system through interactions with other proteins involved in cell survival. Perform. Keratin is the most abundant intercellular fibrous connecting protein in the epidermis. Keratin has a large molecular weight and is connected by cystine disulfide bonds. As a result, old dead skin cells are normally shed and replaced with a new skin layer, thereby maintaining a healthy barrier function. In addition, intercellular lipids are decomposed by enzymes of keratinocytes, which are keratinocytes, to produce ceramides, fatty acids, cholesterol, etc., which are known to form the lipid barrier of the skin. In other words, when the skin barrier is weakened, the regeneration rate of skin cells slows down, which accelerates epidermal aging, and problems such as exposure of the skin to external stimuli, weakened skin immunity, and loss or lack of skin moisture may occur.
한편, 사이클로덱스트린(Cyclodextrin; CD)은 전분을 합성요소에 작용하는 효소(Cycloamylose Gucanotransferase: CGT-Aae)로 분해하여 얻은 말토올리고당(Cyclic Oligosaccharides)으로 Glucopyranose 분자가 α-1,4-Glycoside 결합을 한 구조물이다. CD는 glucose 분자가 6개로 결합된 α-CD, 7개인 β-CD, 8개인 γ-CD가 있으며 현재 공업적으로 생산되고 있다. 그 중에서도 β-CD은 구조적으로 안정성이 높고 가격도 저렴하여 화장품, 식품 및 의약 산업에 다양하게 응용되고 있다.Meanwhile, cyclodextrin (CD) is a maltooligosaccharide (Cyclic Oligosaccharides) obtained by decomposing starch with an enzyme (Cycloamylose Gucanotransferase: CGT-Aae) that acts on synthetic urea. Glucopyranose molecules have an α-1,4-Glycoside bond. It is a structure. CDs include α-CD, which consists of six glucose molecules, β-CD, which contains seven glucose molecules, and γ-CD, which contains eight glucose molecules, and is currently being produced industrially. Among them, β-CD is structurally stable and inexpensive, so it is widely used in the cosmetics, food, and pharmaceutical industries.
사이클로덱스트린은 소수성의 내부와 친수성의 외부로 이루어진 3차원 구조를 지녀서 물에 녹으면서도 다양한 지용성 물질을 포접할 수 있는 기능을 가지고 있다. 사이클로덱스트린의 이러한 기능은 난용성 물질의 용해도를 증가시켜 줄 뿐만 아니라 냄새나 맛의 차폐, 조해성 물질의 안정화, 자극성 감소 등을 돕는 역할을 할 수 있으며, 사이클로덱스트린은 구성단위가 글루코즈(glucose)이므로 독성이 없는 인체에 매우 안전한 물질로서 화장품 원료를 수용성 제형으로 녹이는데 큰 강점이 있다. 또 특이한 컵 모양의 구조를 지녀 일정한 분자량 범위(약 300이하)의 분자량들을 컵 내부에 포획할 수 있는 가능성이 있다. 이러한 컵과 같은 공간을 형성하는 특성은 분자 간 상호작용에 의해 형성된다. 사이클로덱스트린의 구조적특성은 소수성의 공동내에 불안정한 구조를 가지고 있는 각종 소수성 화합물인 게스트(guest)물질로 포접시켜 포접 복합체를 형성할 수 있으며, 이를 호스트-게스트 복합체(host-guest complex)라고 한다. 사이클로덱스트린의 소수성 내부로 분자의 크기 및 성질에 따라 유입되는 guest 분자는 사이클로덱스트린 분자에 의해 포접되어 외부적인 요인으로부터 보호 받을 수 있을 뿐 아니라, 산소, 열, 빛에 대한 안정성이 강화된다. 뿐만 아니라 사이클로덱스트린의 host-guest complex 특성을 이용하면 소수성 공간내에 있는 소수성 분자를 생체 속으로 전달해 주는 역할을 하게 된다. Cyclodextrin has a three-dimensional structure consisting of a hydrophobic interior and a hydrophilic exterior, so it has the ability to encapsulate various fat-soluble substances while being soluble in water. This function of cyclodextrin not only increases the solubility of poorly soluble substances, but also helps mask odor or taste, stabilize deliquescent substances, and reduce irritation. Since cyclodextrin's constituent unit is glucose, It is a non-toxic material that is very safe for the human body and has great strengths in dissolving cosmetic raw materials into a water-soluble formulation. In addition, it has a unique cup-shaped structure, so there is the possibility of capturing molecular weights within a certain molecular weight range (about 300 or less) inside the cup. The characteristic of forming this cup-like space is formed by interactions between molecules. The structural characteristics of cyclodextrin allow it to form an inclusion complex by incorporating guest materials, which are various hydrophobic compounds with an unstable structure, into a hydrophobic cavity, and this is called a host-guest complex. Guest molecules that flow into the hydrophobic interior of cyclodextrin, depending on the size and nature of the molecule, are enclosed by the cyclodextrin molecule and are not only protected from external factors, but also their stability against oxygen, heat, and light is enhanced. In addition, by using the host-guest complex characteristics of cyclodextrin, it plays the role of delivering hydrophobic molecules within the hydrophobic space into the living body.
이에 사이클로덱스트린의 host-guest 상호작용을 이용하여 소수성 화합물을 공동에 포접한 물질의 용해성 및 안정성을 증진시키고, 서방형성질(sustained-release, slow-release)을 향상시키는 등의 생체 이용률을 증가시키는 다양한 연구가 진행되고 있다.Accordingly, the host-guest interaction of cyclodextrin is used to improve the solubility and stability of substances containing hydrophobic compounds in the cavity and to increase bioavailability, such as improving sustained-release (slow-release). Various research is underway.
본 발명은 캡슐레이션(capsulation) 기술을 적용한 캡슐화케라틴을 제공하고자 한다.The present invention seeks to provide encapsulated keratin using encapsulation technology.
또한, 본 발명은 상기 캡슐화케라틴 조성물의 제조방법을 제공하는 것을 다른 목적으로 한다.Another object of the present invention is to provide a method for producing the encapsulated keratin composition.
본 발명은 케라틴을 캡슐화함에 따라 피부장벽기능 강화 효과가 우수한 화장료 조성물을 제공하는 것을 또 다른 목적으로 한다.Another object of the present invention is to provide a cosmetic composition that has an excellent effect of strengthening the skin barrier function by encapsulating keratin.
본 발명은 정제수, 하이드록시프로필사이클로덱스트린, 카페인, 폴리덱스트로오스, 하이드롤라이드즈케라틴, 케라틴, 말토덱스트린, 사이클로덱스트린, 아시아티코사이드, 페녹시에탄올, 잔탄검 및 하이드록시에틸셀룰로오스를 혼합한 후, 균질화하여 제조한 캡슐화케라틴을 제공한다.The present invention is a mixture of purified water, hydroxypropylcyclodextrin, caffeine, polydextrose, hydrolides keratin, keratin, maltodextrin, cyclodextrin, asiaticoside, phenoxyethanol, xanthan gum and hydroxyethylcellulose. Afterwards, the encapsulated keratin prepared by homogenization is provided.
본 발명에 있어서, 상기 캡슐화케라틴은, 바람직하게는 정제수 69.25~75.35 중량%, 하이드록시프로필사이클로덱스트린 12.09~16.09 중량%, 카페인 2.20~2.40 중량%, 폴리덱스트로오스 1.80~2.20 중량%, 하이드롤라이드즈케라틴 1.68~2.08 중량%, 케라틴 1.68~2.08 중량%, 말토덱스트린 1.56~1.96 중량%, 사이클로덱스트린 0.8~1 중량%, 아시아티코사이드 0.71~1.11 중량%, 페녹시에탄올 0.7~0.9 중량%, 잔탄검 0.50~0.70 중량% 및 하이드록시에틸셀룰로오스 0.13~0.23 중량%로 조성되는 것이 좋다.In the present invention, the encapsulated keratin is preferably 69.25 to 75.35% by weight of purified water, 12.09 to 16.09% by weight of hydroxypropylcyclodextrin, 2.20 to 2.40% by weight of caffeine, 1.80 to 2.20% by weight of polydextrose, and hydrol. Rydeskeratin 1.68~2.08% by weight, keratin 1.68~2.08% by weight, maltodextrin 1.56~1.96% by weight, cyclodextrin 0.8~1% by weight, asiaticoside 0.71~1.11% by weight, phenoxyethanol 0.7~0.9% by weight, It is preferably composed of 0.50 to 0.70% by weight of xanthan gum and 0.13 to 0.23% by weight of hydroxyethylcellulose.
또한, 본 발명은 하이드록시프로필사이클로덱스트린, 말토덱스트린, 사이클로덱스트린, 잔탄검, 하이드록시에틸셀룰로오스를 혼합하는 단계 (a); 카페인, 폴리덱스트로오스, 하이드롤라이즈드케라틴, 케라틴, 아시아티코사이드, 페녹시에탄올, 잔탄검을 첨가하는 단계 (b); 상기 단계 (b) 후, 정제수를 용매로 하여 40~80℃에서 완전히 용해시키는 단계 (c); 를 포함하는 것을 특징으로 하는 케라틴의 캡슐화방법을 제공한다.In addition, the present invention includes the step (a) of mixing hydroxypropylcyclodextrin, maltodextrin, cyclodextrin, xanthan gum, and hydroxyethylcellulose; Step (b) adding caffeine, polydextrose, hydrolyzed keratin, keratin, asiaticoside, phenoxyethanol, and xanthan gum; After step (b), (c) completely dissolving at 40-80°C using purified water as a solvent; It provides a method for encapsulating keratin, comprising:
본 발명에 있어서, 상기 케라틴의 캡슐화방법은, 바람직하게는 정제수 69.25~75.35 중량%, 하이드록시프로필사이클로덱스트린 12.09~16.09 중량%, 카페인 2.20~2.40 중량%, 폴리덱스트로오스 1.80~2.20 중량%, 하이드롤라이드즈케라틴 1.68~2.08 중량%, 케라틴 1.68~2.08 중량%, 말토덱스트린 1.56~1.96 중량%, 사이클로덱스트린 0.8~1 중량%, 아시아티코사이드 0.71~1.11 중량%, 페녹시에탄올 0.7~0.9 중량%, 잔탄검 0.50~0.70 중량% 및 하이드록시에틸셀룰로오스 0.13~0.23 중량%를 사용하는 것이 좋다.In the present invention, the method for encapsulating keratin is preferably 69.25 to 75.35% by weight of purified water, 12.09 to 16.09% by weight of hydroxypropylcyclodextrin, 2.20 to 2.40% by weight of caffeine, 1.80 to 2.20% by weight of polydextrose, Hydrolide keratin 1.68~2.08% by weight, keratin 1.68~2.08% by weight, maltodextrin 1.56~1.96% by weight, cyclodextrin 0.8~1% by weight, asiaticoside 0.71~1.11% by weight, phenoxyethanol 0.7~0.9% by weight %, it is recommended to use 0.50~0.70% by weight of xanthan gum and 0.13~0.23% by weight of hydroxyethylcellulose.
또한, 본 발명은 상기 캡슐화케라틴을 포함하는 화장료 조성물을 제공한다.Additionally, the present invention provides a cosmetic composition containing the encapsulated keratin.
본 발명에 있어서, 상기 화장료 조성물은, 바람직하게는 각질형성세포 분화 유도에 따른 피부 장벽 강화 효과가 있는 것이 좋다.In the present invention, the cosmetic composition preferably has a skin barrier strengthening effect by inducing keratinocyte differentiation.
본 발명은 케라틴에 캡슐화(capsulation) 기술을 적용하여, 케라틴의 안정성을 더욱 증진시키고, 피부에 효과적으로 전달할 수 있는 케라틴의 캡슐화방법을 제공한다. 또한, 본 발명은 캡슐화케라틴을 포함하여 피부장벽기능 강화 효과가 우수한 화장료 조성물을 제공한다. 더욱 구체적으로 본 발명에서는 캡슐화를 통하여 더욱 우수한 세포독성, 항염증효과, 세포재생효과를 확인하였는데, 케라틴캡슐화를 통하여 세포독성은 나타나지 않았으며, 염증반응에 수반되는 Nitric Oxide (NO), Tumor necrosis factor-α (TNF-α), Iinterleukin-6 (IL-6), 프로스타글란딘 E2 등이 감소하였으며, UV 손상에 대한 피부세포의 재생효과가 있음을 확인하였다.The present invention applies encapsulation technology to keratin to further improve the stability of keratin and provides a method of encapsulating keratin that can be effectively delivered to the skin. In addition, the present invention provides a cosmetic composition that contains encapsulated keratin and has an excellent skin barrier function strengthening effect. More specifically, in the present invention, superior cytotoxicity, anti-inflammatory effect, and cell regeneration effect were confirmed through encapsulation. Cytotoxicity was not observed through keratin encapsulation, and Nitric Oxide (NO) and Tumor necrosis factor accompanying inflammatory response were confirmed. -α (TNF-α), Iinterleukin-6 (IL-6), and prostaglandin E 2 were decreased, and it was confirmed that there was a regenerative effect on skin cells against UV damage.
도 1의 (a)는 캡슐화 전의 케라틴 사진을 육안으로 관찰한 도면이고, (b)는 캡슐화케라틴을 육안으로 관찰한 도면이다.
도 2는 본 발명에서 제조한 캡슐화케라틴의 세포독성을 나타낸 그래프이다.
도 3은 본 발명에서 제조한 캡슐화케라틴의 NO 생성 억제능을 나타낸 그래프이다.
도 4는 본 발명에서 제조한 캡슐화케라틴의 IL-6 발현 억제능을 나타낸 그래프이다.
도 5는 본 발명에서 제조한 캡슐화케라틴의 TNF-α 발현 억제능을 나타낸 그래프이다.
도 6은 본 발명에서 제조한 캡슐화케라틴의 비색을 육안으로 관찰한 결과이다.
도 7은 본 발명에서 제조한 캡슐화케라틴의 활성산소 소거 효과를 나타낸 그래프이다.
도 8은 본 발명에서 제조한 캡슐화케라틴의 UV 조사 후 세포 독성을 나타낸 그래프이다.
도 9는 본 발명에서 제조한 캡슐화케라틴의 UV 조사 후 프로스타글란딘 E2 발현 억제 효과를 나타낸 그래프이다.
도 10은 본 발명에서 제조한 캡슐화케라틴의 각질형성세포 증식 효과를 나타낸 그래프이다.Figure 1 (a) is a picture of keratin before encapsulation observed with the naked eye, and (b) is a picture observed with the naked eye of encapsulated keratin.
Figure 2 is a graph showing the cytotoxicity of the encapsulated keratin prepared in the present invention.
Figure 3 is a graph showing the NO production inhibition ability of the encapsulated keratin prepared in the present invention.
Figure 4 is a graph showing the ability of the encapsulated keratin prepared in the present invention to inhibit IL-6 expression.
Figure 5 is a graph showing the ability of the encapsulated keratin prepared in the present invention to inhibit TNF-α expression.
Figure 6 shows the results of visual observation of the colorimetric color of the encapsulated keratin prepared in the present invention.
Figure 7 is a graph showing the active oxygen scavenging effect of the encapsulated keratin prepared in the present invention.
Figure 8 is a graph showing the cytotoxicity of the encapsulated keratin prepared in the present invention after UV irradiation.
Figure 9 is a graph showing the effect of inhibiting prostaglandin E 2 expression after UV irradiation of the encapsulated keratin prepared in the present invention.
Figure 10 is a graph showing the keratinocyte proliferation effect of the encapsulated keratin prepared in the present invention.
본 발명은 캡슐화케라틴의 제조방법 및 이의 방법으로 제조된 캡술화케라틴을 포함하는 피부장벽 개선 또는 강화용 화장료 조성물을 제공한다. 각질은 일반적으로 오래된 각질 세포가 정상적으로 탈락하고 새로운 피부층으로 대체되는 턴 오버(turn over)가 이루어지는데, 노화 등의 이유로 턴 오버 주기가 길어짐에 따라 오래된 각질세포의 탈락이 늦어지게 된다. 이러한 현상에 의해 각질층의 피부 장벽 기능이 약화되어 피부 수분 유지력이 감소하고 면역기능이 약해지는 등의 증상이 나타난다.The present invention provides a method for producing encapsulated keratin and a cosmetic composition for improving or strengthening the skin barrier containing encapsulated keratin produced by the method. Keratin generally undergoes turnover, in which old keratinocytes are normally shed and replaced by a new skin layer. As the turnover cycle becomes longer due to aging, etc., the shedding of old keratinocytes is delayed. Due to this phenomenon, the skin barrier function of the stratum corneum is weakened, causing symptoms such as reduced skin moisture retention and weakened immune function.
일반적으로 피부장벽 개선용 화장료 조성물은 각질층의 접착제 역할을 하는 지질인 세라마이드를 중심으로 연구개발되어 보습효과를 중심으로 보고 있다. 그러나 본 발명은 각질층의 주성분인 각질형성세포를 분화시킴에 따라 피부장벽 자체의 기능을 개선 또는 강화하고, 피부장벽 개선으로부터 피부 보습효과를 증진시킬 수 있는 화장료 조성물을 제공하고자 한다.In general, cosmetic compositions for improving the skin barrier are researched and developed focusing on ceramide, a lipid that acts as an adhesive for the stratum corneum, and are focused on the moisturizing effect. However, the present invention seeks to provide a cosmetic composition that can improve or strengthen the function of the skin barrier itself by differentiating keratinocytes, the main component of the stratum corneum, and enhance the skin moisturizing effect by improving the skin barrier.
한편, 세포 내의 케라틴은 세포골격(cytoskeleton)을 이루는 주요단백질로 외부에서 가해지는 손상으로부터 세포를 보호해 주는 역할을 하며, 세포생존에 관여하는 다른 단백질들과 상호작용을 통하여 신호전달체계의 조절기능을 수행한다. 케라틴은 표피에 가장 풍부하게 존재하는 세포간 섬유상 연결 단백질로서 케라틴은 분자량이 크며 시스틴 이중황화결합(cystine disulfide bonds)에 의해서 연결되어 있기에 불용성이며 잘 분해되지 않고 분자량이 큰 이유로 쉽게 용해되지 않는다.Meanwhile, keratin within cells is a major protein that makes up the cytoskeleton and plays a role in protecting cells from external damage, and regulates the signal transduction system through interactions with other proteins involved in cell survival. Perform. Keratin is an intercellular fibrous connecting protein that is most abundant in the epidermis. Keratin has a large molecular weight and is connected by cystine disulfide bonds, so it is insoluble, does not decompose easily, and does not dissolve easily due to its large molecular weight.
본 발명은 마이셀(micelle) 형태로 제조되는 캡슐 내부에 지용성 물질은 케라틴을 포집시켜 제조되는데, 마이셀 캡슐레이션(capsulation) 기술을 적용하여 케라틴을 안정적으로 보존 할 수 있을 뿐만 아니라, 케라틴의 효능을 피부에 효과적으로 전달할 수 있도록 하여, 케라틴에 특화된 캡슐화 방법을 제공함에 그 목적이 크다 하겠다. 본 발명에서 캡슐은 마이셀 형태의 캡슐을 의미하는 것으로 정의한다. In the present invention, the oil-soluble substance is manufactured by trapping keratin inside a capsule manufactured in the form of micelles. By applying micelle encapsulation technology, not only can keratin be stably preserved, but the efficacy of keratin can be extended to the skin. The main purpose is to provide an encapsulation method specialized for keratin so that it can be effectively delivered to the body. In the present invention, capsule is defined to mean a micelle-type capsule.
구체적으로, 본 발명은 정제수, 하이드록시프로필사이클로덱스트린, 카페인, 폴리덱스트로오스, 하이드롤라이드즈케라틴, 케라틴, 말토덱스트린, 사이클로덱스트린, 아시아티코사이드, 페녹시에탄올, 잔탄검 및 하이드록시에틸셀룰로오스를 혼합한 후, 균질화하여 제조한 캡슐화케라틴을 제공한다.Specifically, the present invention relates to purified water, hydroxypropylcyclodextrin, caffeine, polydextrose, hydrolide keratin, keratin, maltodextrin, cyclodextrin, asiaticoside, phenoxyethanol, xanthan gum, and hydroxyethylcellulose. Encapsulated keratin prepared by mixing and homogenizing is provided.
본 발명에서는 통상적으로 피부자극을 유발하는 유화제 성분을 사용하지 않고도 캡슐화케라틴을 유화물로 제조할 수 있었다. 구체적으로, 하이드록시프로필사이클로덱스트린, 말토덱스트린, 사이클로덱스트린, 잔탄검, 하이드록시에틸셀룰로오스를 본 발명에서 착안한 배합비로 혼합하여 유화제 대용으로 사용함으로써, 피부자극을 유발하는 유화제를 전혀 사용하지 않고도 케라틴 함유 유화물을 제조할수 있었던 것이다.In the present invention, encapsulated keratin could be manufactured as an emulsion without using emulsifier ingredients that typically cause skin irritation. Specifically, by mixing hydroxypropylcyclodextrin, maltodextrin, cyclodextrin, xanthan gum, and hydroxyethylcellulose in a mixing ratio inspired by the present invention and using it as a substitute for an emulsifier, keratin can be produced without using any emulsifiers that cause skin irritation. It was possible to produce a containing emulsion.
이를 통해 본 발명의 캡슐화케라틴은, 정제수, 하이드록시프로필사이클로덱스트린, 카페인, 폴리덱스트로오스, 하이드롤라이드즈케라틴, 케라틴, 말토덱스트린, 사이클로덱스트린, 아시아티코사이드, 페녹시에탄올, 잔탄검 및 하이드록시에틸셀룰로오스를 포함하여 구성된다. 이때, 상기 캡슐화케라틴은 바람직하게는, 정제수 65.35~75.35 중량%, 하이드록시프로필사이클로덱스트린 12.09~16.09 중량%, 카페인 2.20~2.40 중량%, 폴리덱스트로오스 1.80~2.20 중량%, 하이드롤라이드즈케라틴 1.68~2.08 중량%, 케라틴 1.68~2.08 중량%, 말토덱스트린 1.56~1.96 중량%, 사이클로덱스트린 0.8~1 중량%, 아시아티코사이드 0.71~1.11 중량%, 페녹시에탄올 0.7~0.9 중량%, 잔탄검 0.50~0.70 중량% 및 하이드록시에틸셀룰로오스 0.13~0.23 중량%로 조성되는 것이 좋다.Through this, the encapsulated keratin of the present invention contains purified water, hydroxypropylcyclodextrin, caffeine, polydextrose, hydrolides keratin, keratin, maltodextrin, cyclodextrin, asiaticoside, phenoxyethanol, xanthan gum, and hydrolyzed keratin. It is composed of oxyethylcellulose. At this time, the encapsulated keratin is preferably 65.35 to 75.35% by weight of purified water, 12.09 to 16.09% by weight of hydroxypropylcyclodextrin, 2.20 to 2.40% by weight of caffeine, 1.80 to 2.20% by weight of polydextrose, and hydrolide keratin. 1.68~2.08% by weight, keratin 1.68~2.08% by weight, maltodextrin 1.56~1.96% by weight, cyclodextrin 0.8~1% by weight, asiaticoside 0.71~1.11% by weight, phenoxyethanol 0.7~0.9% by weight, xanthan gum 0.50 It is recommended that it be composed of ~0.70% by weight and 0.13~0.23% by weight of hydroxyethylcellulose.
본 발명에서 사용되는 카페인은 피부에 흡수될 경우 흡수된 국소 부위의 피부 보호 효과를 발휘한다고 알려져 있으며, 착향제, 피부컨디셔닝제로 화장품 원료로 사용한다. 또한, 폴리덱스트로오스는 D-글루코오스를 축합하여 얻은 폴리머로 고체성분을 희석하는 기능을 한다. 또한, 하이드롤라이즈드케라틴은 케라틴단백가수분해물이며, 케라틴은 피부 표면이나 모발 표면에 피막을 형성하여 수분 증발을 차단함에 따라 보습 효과를 발휘한다. 또한, 아시아티코사이드는 병풀의 유효성분 중 하나로 콜라겐 합성을 촉진하며, 주름 개선, 피부 탄력 유지 등의 효과가 있다고 알려져 있다. 또한, 페녹시에탄올은 방부효과를 위해 화장품에 첨가된다. Caffeine used in the present invention is known to exert a skin protection effect in the local area where it is absorbed when absorbed into the skin, and is used as a cosmetic ingredient as a flavoring agent and skin conditioning agent. In addition, polydextrose is a polymer obtained by condensing D-glucose and functions to dilute solid components. In addition, hydrolyzed keratin is a hydrolyzed product of keratin protein, and keratin exerts a moisturizing effect by forming a film on the surface of the skin or hair to block moisture evaporation. Additionally, asiaticoside is one of the active ingredients of Centella asiatica and is known to promote collagen synthesis and has effects such as improving wrinkles and maintaining skin elasticity. Additionally, phenoxyethanol is added to cosmetics for its preservative effect.
한편, 본 발명의 캡슐화케라틴은, 바람직하게, 하이드록시프로필사이클로덱스트린, 말토덱스트린, 사이클로덱스트린, 잔탄검, 하이드록시에틸셀룰로오스를 혼합한 무유화제 유화물과, 카페인, 폴리덱스트로오스, 케라틴, 하이드롤라이즈드케라틴, 아시아티코사이드, 페녹시에탄올을 정제수와 혼합하고, 이들이 정제수에 완전히 용해되어 맑은 용액이 되도록 40~80℃에서 10분~60분 교반하여 제조할 수 있다. 더욱 바람직하게는 60℃에서 30분 교반하여 제조하는 것이 좋다.Meanwhile, the encapsulated keratin of the present invention is preferably a non-emulsifier emulsion of hydroxypropylcyclodextrin, maltodextrin, cyclodextrin, xanthan gum, and hydroxyethylcellulose, caffeine, polydextrose, keratin, and hydrol. It can be prepared by mixing raised keratin, asiaticoside, and phenoxyethanol with purified water and stirring at 40 to 80°C for 10 to 60 minutes so that they are completely dissolved in purified water to form a clear solution. More preferably, it is prepared by stirring at 60°C for 30 minutes.
한편, 본 발명의 캡슐화케라틴은 NO 생성 억제 효과가 우수하며, IL-6 발현 억제, TNF-α 생성 억제, 프로스타글란딘 E2 발현 억제 효과가 있음을 하기 실험에서 확인하였다.Meanwhile, it was confirmed in the following experiment that the encapsulated keratin of the present invention has an excellent effect of suppressing NO production, and has the effect of suppressing IL-6 expression, inhibiting TNF-α production, and prostaglandin E 2 expression.
산화질소는 세포독성이나 각종 염증반응을 일으키는 것으로 알려져 있으며, TNF-α, IL-6와 같은 사이토카인(cytokine)은 염증성 질환과 관련이 있다고 알려져 있는데, 이러한 TNF-α, IL-6의 발현을 저해시키거나 COX-2 활성저해에 기인하는 프로스타글란딘 E2의 생성 억제를 통해 초기 염증성 분자의 증가를 수반하는 병변 과정을 조절할 수 있다고 알려진 바 있다. 이에 본 발명의 캡슐화케라틴은 염증성 사이토카인 억제 효과, 염증 완화 효과가 있어 민감성 피부를 완화할 수 있음을 확인하였다.Nitric oxide is known to cause cytotoxicity and various inflammatory reactions, and cytokines such as TNF-α and IL-6 are known to be related to inflammatory diseases. The expression of TNF-α and IL-6 It is known that the lesion process accompanied by an increase in early inflammatory molecules can be controlled by inhibiting the production of prostaglandin E 2 due to inhibition of COX-2 activity. Accordingly, it was confirmed that the encapsulated keratin of the present invention has the effect of suppressing inflammatory cytokines and alleviating inflammation and can alleviate sensitive skin.
또한, 피부는 자외선에 의해 히스타민, 프로스타글란딘 등의 염증 물질을 생성하는데, 본 발명의 캡슐화케라틴은 UV 조사 후, 프로스타글란딘 E2의 발현 정도를 효과적으로 억제함에 따라, UV 손상에 대한 피부세포의 재생 기능이 있는 것으로 판단되었다.In addition, the skin produces inflammatory substances such as histamine and prostaglandin by ultraviolet rays, and the encapsulated keratin of the present invention effectively suppresses the expression level of prostaglandin E 2 after UV irradiation, thereby improving the regenerative function of skin cells against UV damage. It was judged that there was.
또한, 본 발명의 캡슐화케라틴은 활성산소 소거 효과가 우수하여 항산화 효능이 있는 것으로 나타났다. 따라서, 본 발명의 캡슐화케라틴을 화장료 조성물에 적용할 경우 항산화 효능을 기대할 수 있을 것으로 판단되며, 지속적인 사용은 피부 환경의 산화 스트레스를 감소하는데 영향을 미칠 것으로 예상된다.In addition, the encapsulated keratin of the present invention was found to have an antioxidant effect due to its excellent oxygen radical scavenging effect. Therefore, it is expected that antioxidant efficacy can be expected when the encapsulated keratin of the present invention is applied to a cosmetic composition, and continuous use is expected to have an effect in reducing oxidative stress in the skin environment.
또한, 본 발명의 캡슐화케라틴은 각질형성세포를 증식시키는 효과가 있는 것으로 나타났다. 정상 각질형성세포의 증식 효과는 상처 치유나 피부 세포 재생과 관련이 있다고 알려져 있으며, 표피의 대부분을 차지하는 각질형성세포에 대한 증식을 보이는 본 발명의 캡슐화케라틴은 표피의 재생 또는 회복에 도움을 줌으로써 피부 장벽 개선 또는 강화 효과가 나타나는 것으로 판단되었다. 이에 본 발명의 캡슐화케라틴은 피부 각질형성세포 분화 유도 효과, 표피 재생 또는 회복 효과가 있는 것으로 판단되었으며, 이에 따라 피부 장벽기능 개선 또는 강화 효과가 있음을 확인하였다.In addition, the encapsulated keratin of the present invention was shown to be effective in proliferating keratinocytes. It is known that the proliferation effect of normal keratinocytes is related to wound healing and skin cell regeneration, and the encapsulated keratin of the present invention, which shows proliferation of keratinocytes, which make up most of the epidermis, helps in regeneration or recovery of the epidermis, thereby improving skin It was judged that there was an effect of improving or strengthening the barrier. Accordingly, it was determined that the encapsulated keratin of the present invention had an effect of inducing skin keratinocyte differentiation and an effect of epidermal regeneration or recovery, and accordingly, it was confirmed that it had an effect of improving or strengthening the skin barrier function.
이하, 본 발명에 대해 하기 실시예 및 실험예에서 더욱 상세히 설명하고자 한다. 다만, 본 발명의 권리범위가 하기 실시예 및 실험예에만 한정되는 것은 아니고, 이와 등가의 기술적 사상의 변형까지를 모두 포함한다.Hereinafter, the present invention will be described in more detail in the following examples and experimental examples. However, the scope of the present invention is not limited to the following examples and experimental examples, and includes all modifications of the technical idea equivalent thereto.
[실시예 1 : 캡슐화케라틴의 제조][Example 1: Preparation of encapsulated keratin]
정제수, 하이드록시프로필사이클로덱스트린, 카페인, 폴리덱스트로오스, 하이드롤라이즈드케라틴, 케라틴, 말토덱스트린, 사이클로덱스트린, 아시아티코사이드, 페녹시에탄올, 잔탄검, 하이드록시에틸셀룰로오스를 아래 표 1과 같은 배합비로 혼합하여 마이셀 형태의 캡슐 내부에 케라틴이 포집된 케라틴 용액(이하, 캡슐화케라틴)을 제조하였다. 아래 표 1의 함량비를 이용하여 정제수에 완전히 용해시켜 맑은 용액이 되도록 40~80℃에서 30분간 아지믹서와 호모믹서를 이용하여 교반하여, 마이셀(micelle) 수용액을 제조하였다.Purified water, hydroxypropylcyclodextrin, caffeine, polydextrose, hydrolyzed keratin, keratin, maltodextrin, cyclodextrin, asiaticoside, phenoxyethanol, xanthan gum, and hydroxyethylcellulose as shown in Table 1 below. By mixing in the mixing ratio, a keratin solution (hereinafter referred to as encapsulated keratin) in which keratin was captured inside a micelle-shaped capsule was prepared. Using the content ratio in Table 1 below, a micelle aqueous solution was prepared by completely dissolving in purified water and stirring at 40-80°C for 30 minutes using an azimixer and homomixer to form a clear solution.
도 1에 캡슐화 전 케라틴과 캡슐화 후 케라틴을 나타내었다. 도 1의 (a)는 캡슐화 전 케라틴으로 상기 표 1의 배합비에 따라 혼합한 형태를 육안으로 관찰한 결과이며, 도 1의 (b)는 캡슐화케라틴을 육안으로 관찰한 결과이다.Figure 1 shows keratin before encapsulation and keratin after encapsulation. Figure 1 (a) is the result of visual observation of the keratin mixed according to the mixing ratio in Table 1 before encapsulation, and Figure 1 (b) is the result of visual observation of the encapsulated keratin.
[실험예 1 : 캡슐화케라틴의 세포독성평가][Experimental Example 1: Cytotoxicity evaluation of encapsulated keratin]
본 실험에서는 상기 실시예 1에서 제조한 캡슐화케라틴의 세포독성을 평가하였다. 캡슐화케라틴 100%를 배지로 하였으며, 캡슐화케라틴은 0.25, 0.5, 1, 2%(v/v)로 각각 희석하여 사용하였다. 96 well-plate에 Raw 264.7 세포를 4×104 cell/well 농도로 분주하고, 37℃, 5% CO2 배양기에서 24시간 배양하였으며, 24시간 후에 캡슐화케라틴을 농도별로 처리하고 1시간 후 LPS를 처리하여 37℃, 5% CO2 배양기에서 24시간 배양한 후 배지를 모두 제거하고, MTT kit 프로토콜에 따라 MTT 시약을 처리하고 590㎚에서 흡광도를 측정하여 세포독성을 확인하였다. 대조군으로는 무처리군을 이용하였다.In this experiment, the cytotoxicity of the encapsulated keratin prepared in Example 1 was evaluated. 100% encapsulated keratin was used as the medium, and the encapsulated keratin was diluted to 0.25, 0.5, 1, and 2% (v/v), respectively. Raw 264.7 cells were distributed in a 96 well- plate at a concentration of 4 After treatment and culturing in a 37°C, 5% CO 2 incubator for 24 hours, all medium was removed, MTT reagent was treated according to the MTT kit protocol, and cytotoxicity was confirmed by measuring absorbance at 590 nm. An untreated group was used as the control group.
도 2에서 보듯이, 본 발명의 캡슐화케라틴은 세포 생존율이 우수한 것을 확인하였다.As shown in Figure 2, it was confirmed that the encapsulated keratin of the present invention had excellent cell viability.
[실험예 2 : 캡슐화케라틴의 NO 생성 억제 효과 확인] [Experimental Example 2: Confirmation of NO production inhibition effect of encapsulated keratin]
본 실험에서는 상기 실시예 1에서 제조한 캡슐화케라틴의 NO 생성 억제 효과를 확인하였다. 캡슐화케라틴은 0.25, 0.5, 1, 2%(v/v)로 각각 희석하여 사용하였다. 96 well-plate에 Raw 264.7 세포를 4×104 cell/well 농도로 분주하고, 37℃, 5% CO2 배양기에서 24시간 배양하였으며, 24시간 후에 양성 대조물질(덱사메타손 10㎍/㎖)과 캡슐화케라틴을 농도별로 처리하고 1시간 후 LPS를 처리하여 37℃, 5% CO2 배양기에서 24시간 배양한 후 세포배양액과 Griess 시약을 1:1로 혼합하고 10분 후 550㎚에서 흡광도를 측정하였다. NO 생성량은 표준곡선을 이용하여 산출하였으며, NO 생성 억제능(%)을 아래와 같이 계산하였다.In this experiment, the NO production inhibition effect of the encapsulated keratin prepared in Example 1 was confirmed. Encapsulated keratin was used diluted to 0.25, 0.5, 1, and 2% (v/v), respectively. Raw 264.7 cells were distributed in a 96 well-plate at a concentration of 4 × 10 4 cells/well, cultured for 24 hours in a 5% CO 2 incubator at 37°C, and encapsulated with a positive control substance (dexamethasone 10 μg/ml) after 24 hours. Keratin was treated at different concentrations, and after 1 hour, LPS was treated and cultured in an incubator at 37°C and 5% CO 2 for 24 hours. Cell culture medium and Griess reagent were mixed 1:1, and absorbance was measured at 550 nm after 10 minutes. The amount of NO production was calculated using a standard curve, and the NO production inhibition ability (%) was calculated as follows.
▶ NO 생성 억제능 (%) = [(N-S)/(N-C)]×100▶ NO production inhibition ability (%) = [(N-S)/(N-C)]×100
N : Negative coltrol(LPS 처리군)N: Negative control (LPS treatment group)
C : Control(무처리군)C: Control (untreated group)
S : Sample(시료 처리군)S: Sample (sample treatment group)
도 3에서 보듯이, 본 발명의 캡슐화케라틴은 NO 생성을 효과적으로 억제하는 점을 확인하였다.As shown in Figure 3, it was confirmed that the encapsulated keratin of the present invention effectively inhibits NO production.
[실험예 3 : 캡슐화케라틴의 IL-6 발현 억제 효과 확인] [Experimental Example 3: Confirmation of the inhibitory effect of encapsulated keratin on IL-6 expression]
본 실험에서는 상기 실시예 1에서 제조한 캡슐화케라틴의 IL-6 발현 억제 효과를 확인하였다. 캡슐화케라틴 100%를 배지로 하였으며, 캡슐화케라틴은 0.5, 1, 2%(v/v)로 각각 희석하여 사용하였다. 6 well-plate에 Raw 264.7 세포를 2×105 cell/well 농도로 분주하고, 37℃, 5% CO2 배양기에서 24시간 배양하였으며, 24시간 후에 양성 대조물질(덱사메타손 10㎍/㎖)과 캡슐화케라틴을 농도별로 처리하고 1시간 후 LPS를 처리하여 37℃, 5% CO2 배양기에서 24시간 배양한 후 IL-6 ELISA kit에 제시된 방법에 따라 상층액을 이용하여 450㎚에서 흡광도를 측정하였다. IL-6 발현량은 표준곡선을 이용하여 산출하였다.In this experiment, the inhibitory effect of the encapsulated keratin prepared in Example 1 on IL-6 expression was confirmed. 100% encapsulated keratin was used as a medium, and the encapsulated keratin was diluted to 0.5, 1, and 2% (v/v), respectively. Raw 264.7 cells were dispensed into a 6 well-plate at a concentration of 2×10 5 cells/well, cultured at 37°C in a 5% CO 2 incubator for 24 hours, and encapsulated with a positive control substance (dexamethasone 10 μg/ml) after 24 hours. Keratin was treated at different concentrations, and 1 hour later, LPS was treated and cultured in an incubator at 37°C and 5% CO 2 for 24 hours. Then, the absorbance was measured at 450 nm using the supernatant according to the method suggested in the IL-6 ELISA kit. IL-6 expression level was calculated using a standard curve.
도 4에서 보듯이, 본 발명의 캡슐화케라틴은 IL-6 발현을 효과적으로 억제하는 점을 확인하였다.As shown in Figure 4, it was confirmed that the encapsulated keratin of the present invention effectively inhibits IL-6 expression.
[실험예 4 : 캡슐화케라틴의 TNF-α 생성 억제 효과 확인] [Experimental Example 4: Confirmation of the inhibitory effect of encapsulated keratin on TNF-α production]
본 실험에서는 상기 실시예 1에서 제조한 캡슐화케라틴의 TNF-α 생성 억제 효과를 확인하였다. 캡슐화케라틴 100%를 배지로 하였으며, 캡슐화케라틴은 0.5, 1, 2%(v/v)로 각각 희석하여 사용하였다. 6 well-plate에 Raw 264.7 세포를 2×105 cell/well 농도로 분주하고, 37℃, 5% CO2 배양기에서 24시간 배양하였으며, 24시간 후에 양성 대조물질(덱사메타손 10㎍/㎖)과 캡슐화케라틴을 농도별로 처리하고 1시간 후 LPS를 처리하여 37℃, 5% CO2 배양기에서 24시간 배양한 후 TNF-α ELISA kit에 제시된 방법에 따라 상층액을 이용하여 450㎚에서 흡광도를 측정하였다. TNF-α 생성량은 표준곡선을 이용하여 산출하였다.In this experiment, the inhibitory effect of the encapsulated keratin prepared in Example 1 on TNF-α production was confirmed. 100% encapsulated keratin was used as a medium, and the encapsulated keratin was diluted to 0.5, 1, and 2% (v/v), respectively. Raw 264.7 cells were dispensed into a 6 well-plate at a concentration of 2×10 5 cells/well, cultured at 37°C in a 5% CO 2 incubator for 24 hours, and encapsulated with a positive control substance (dexamethasone 10 μg/ml) after 24 hours. Keratin was treated at different concentrations, and 1 hour later, LPS was treated and cultured in an incubator at 37°C and 5% CO 2 for 24 hours. Then, the absorbance was measured at 450 nm using the supernatant according to the method suggested in the TNF-α ELISA kit. The amount of TNF-α production was calculated using a standard curve.
도 5에서 보듯이, 본 발명의 캡슐화케라틴은 TNF-α 생성량을 효과적으로 억제하는 점을 확인하였다.As shown in Figure 5, it was confirmed that the encapsulated keratin of the present invention effectively suppresses the amount of TNF-α production.
[실험예 5 : 캡슐화케라틴의 활성산소 소거 효과 확인] [Experimental Example 5: Confirmation of active oxygen scavenging effect of encapsulated keratin]
본 실험에서는 상기 실시예 1에서 제조한 캡슐화케라틴의 활성산소 소거 효과를 확인하였다. 캡슐화케라틴은 1, 1.5, 2, 2.5 %(v/v)로 각각 희석하여 사용하였다. 양성대조군으로는 강력한 항산화제인 비타민 C와 글루타티온(glutathione)을 각각 0.01%(v/v)를 이용하였다. 70%(v/v) 에탄올을 96 well에 115㎕ 씩 분주하고, 캡슐화케라틴과 양성대조군을 5㎕ 씩 분주한 뒤 517㎚에서 흡광도를 측정하여 반응 전 흡광도값을 측정하였다. 이후, DPPH 용액을 80㎕ 씩 분주하고 517㎚에서 흡광도를 측정하여 반응 후 흡광도를 측정하였다. 반응 전후 흡광도값을 이용하여 결과를 분석하였다. 모든 결과는 유의함을 통계처리를 통해 검증하였다.In this experiment, the active oxygen scavenging effect of the encapsulated keratin prepared in Example 1 was confirmed. Encapsulated keratin was used by diluting it to 1, 1.5, 2, and 2.5% (v/v), respectively. As a positive control group, 0.01% (v/v) each of vitamin C and glutathione, which are powerful antioxidants, were used. 115 ㎕ of 70% (v/v) ethanol was dispensed into 96 wells, 5 ㎕ of encapsulated keratin and positive control were dispensed, and the absorbance was measured at 517 nm to measure the absorbance before reaction. Afterwards, 80 ㎕ of DPPH solution was dispensed and the absorbance was measured at 517 nm to measure the absorbance after reaction. The results were analyzed using the absorbance values before and after the reaction. The significance of all results was verified through statistical processing.
도 6에 본 발명의 캡슐화케라틴을 농도별로 처리함에 따른 비색을 육안으로 관찰한 결과를 나타내었다. 도 6에서 보듯이, 본 발명의 캡슐화케라틴은 육안으로도 충분히 관찰 가능한 수준의 항산화 효능을 보이는 것을 확인하였다. 한편, 도 7에서 보듯이, 캡슐화케라틴의 항산화 효능을 분석한 결과 모든 농도에서 항산화 효능이 있는 것으로 나타났다. 특히, 양성 대조군인 글루타티온(glutathione)과 비교할 경우에도 유사한 항산화 효능을 보이는 점을 확인하였다.Figure 6 shows the results of visual observation of colorimetry according to the treatment of the encapsulated keratin of the present invention at different concentrations. As shown in Figure 6, it was confirmed that the encapsulated keratin of the present invention exhibits antioxidant efficacy at a level that can be sufficiently observed with the naked eye. Meanwhile, as shown in Figure 7, as a result of analyzing the antioxidant effect of encapsulated keratin, it was found that it had antioxidant effect at all concentrations. In particular, it was confirmed that it showed similar antioxidant efficacy when compared to glutathione, a positive control.
[실험예 6 : 캡슐화케라틴의 프로스타글란딘 E[Experimental Example 6: Prostaglandin E of encapsulated keratin 22 발현 억제 효과 확인] Confirmation of expression inhibition effect]
본 실험에서는 상기 실시예 1에서 제조한 캡슐화케라틴의 프로스타글란딘 E2 발현 억제 효과를 확인하였다.In this experiment, the inhibitory effect of prostaglandin E 2 expression of the encapsulated keratin prepared in Example 1 was confirmed.
먼저 캡슐화케라틴의 처리 농도를 결정하기 위해 각질형성세포에서 세포독성 시험을 진행하였다. 세포 독성의 판정은 무처리군 대비 20% 이상의 세포 활성이 떨어질 때의 기준으로 하였다. 캡슐화케라틴은 증류수를 이용하여 희석하였으며, 양성대조군으로는 NS-398을 이용하였다.First, a cytotoxicity test was conducted on keratinocytes to determine the treatment concentration of encapsulated keratin. Determination of cytotoxicity was made based on when cell activity decreased by more than 20% compared to the untreated group. Encapsulated keratin was diluted using distilled water, and NS-398 was used as a positive control.
각질형성세포(keratinocyte)를 5×104 cell/6well 1일간 배양하였으며, Starvation 상태로 1일간 배양한 후 캡슐화케라틴을 처리하고, 양성대조군인 NS-398은 0.0002%(v/v)을 처리하여 1일간 배양하였으며, 배지를 제거한 후 DPBS로 교환하고 UVB를 조사하였다. 배양 배지를 원심 분리 후 프로스타글란딘 E2 assay kit를 이용하여 프로스타글란딘 E2 발현 양을 측정하였으며, MTT 방법을 이용하여 세포 활성을 측정하여 프로스타글란딘 E2 값을 보정 및 분석하였다. 도 9에서 보듯이, 본 발명의 캡슐화케라틴을 처리한 경우 프로스타글란딘 E2 발현량을 효과적으로 억제하는 점을 확인하였다.Keratinocytes were cultured at 5 Cultured for 1 day, the medium was removed, replaced with DPBS, and UVB irradiated. After centrifuging the culture medium, the amount of prostaglandin E 2 expression was measured using a prostaglandin E 2 assay kit, and cell activity was measured using the MTT method to correct and analyze the prostaglandin E 2 value. As shown in Figure 9, it was confirmed that treatment with the encapsulated keratin of the present invention effectively suppresses the expression level of prostaglandin E 2 .
즉, 본 발명의 캡슐화케라틴은 UV 조사 후, 프로스타글란딘 E2의 발현 정도를 효과적으로 억제함에 따라, UV 손상에 대한 피부세포의 재생 기능이 있는 것으로 판단되었다.That is, the encapsulated keratin of the present invention was determined to have a regenerative function in skin cells against UV damage as it effectively suppressed the expression level of prostaglandin E 2 after UV irradiation.
[실험예 7 : 캡슐화케라틴의 각질형성세포 증식 효과 확인] [Experimental Example 7: Confirmation of keratinocyte proliferation effect of encapsulated keratin]
정상 각질형성세포에서 세포증식능은 세포 재생, 상처 회복과 관련이 있다. 본 실험에서는 본 발명의 캡슐화케라틴이 각질형성세포를 증식시키는 효과가 있는지 확인하였다.In normal keratinocytes, cell proliferation ability is related to cell regeneration and wound recovery. In this experiment, it was confirmed whether the encapsulated keratin of the present invention was effective in proliferating keratinocytes.
캡슐화케라틴의 처리 농도는 상기 실험예 6에서 확인한 바와 같이 결정하였다. 사람 유래 정상 각질형성세포(normal hUman keratinocyte)를 5×104 cell/24well 2일간 배양하고, starvation 상태로 1일간 배양한 뒤, 캡슐화케라틴을 농도별로 처리하고 72시간 배양하였으며, MTT 방법을 이용하여 세포 활성을 측정하여 분석하였다. 시험의 N=4 이며, 통계 기법 Student’s test를 이용하여 분석한 결과 모든 구간에서 유의성을 가짐을 확인하였다.The treatment concentration of encapsulated keratin was determined as confirmed in Experimental Example 6 above. Human-derived normal keratinocytes (normal hUman keratinocytes) were cultured for 2 days at 5 Cell activity was measured and analyzed. The number of tests was N = 4, and as a result of analysis using the statistical technique Student's test, it was confirmed that all sections had significance.
도 10에서 보듯이, 본 발명의 캡슐화케라틴은 적용 최저농도인 0.000001%(v/v) 처리 군부터 대조군 대비 7% 이상의 증식 효과를 나타내었으며, 증식 효과는 0.0001%(v/v)에서 대조군 대비 13% 이상의 증식 효능을 나타내는 것을 확인하였다. As shown in Figure 10, the encapsulated keratin of the present invention showed a proliferation effect of more than 7% compared to the control group from the lowest application concentration of 0.000001% (v/v) treatment group, and the proliferation effect was 0.0001% (v/v) compared to the control group. It was confirmed that it exhibited a proliferation efficacy of more than 13%.
정상 각질형성세포의 증식 효과는 상처 치유나 피부 세포 재생과 관련이 있다고 알려져 있으며, 표피의 대부분을 차지하는 각질형성세포에 대한 증식을 보이는 본 발명의 캡슐화케라틴은 표피의 재생 또는 회복에 도움을 줄 것으로 판단된다. It is known that the proliferation effect of normal keratinocytes is related to wound healing and skin cell regeneration, and the encapsulated keratin of the present invention, which shows proliferation of keratinocytes, which make up most of the epidermis, is expected to help in regeneration or recovery of the epidermis. It is judged.
Claims (6)
Mix purified water, hydroxypropylcyclodextrin, caffeine, polydextrose, hydrolide keratin, keratin, maltodextrin, cyclodextrin, asiaticoside, phenoxyethanol, xanthan gum, and hydroxyethylcellulose, then homogenize. Encapsulated keratin manufactured by
상기 캡슐화케라틴은,
정제수 69.25~75.35 중량%, 하이드록시프로필사이클로덱스트린 12.09~16.09 중량%, 카페인 2.20~2.40 중량%, 폴리덱스트로오스 1.80~2.20 중량%, 하이드롤라이드즈케라틴 1.68~2.08 중량%, 케라틴 1.68~2.08 중량%, 말토덱스트린 1.56~1.96 중량%, 사이클로덱스트린 0.8~1 중량%, 아시아티코사이드 0.71~1.11 중량%, 페녹시에탄올 0.7~0.9 중량%, 잔탄검 0.50~0.70 중량% 및 하이드록시에틸셀룰로오스 0.13~0.23 중량%로 조성되는 것을 특징으로 하는 캡슐화케라틴.
According to paragraph 1,
The encapsulated keratin is,
Purified water 69.25~75.35% by weight, hydroxypropylcyclodextrin 12.09~16.09% by weight, caffeine 2.20~2.40% by weight, polydextrose 1.80~2.20% by weight, hydrolides keratin 1.68~2.08% by weight, keratin 1.68~2.08 Weight%, maltodextrin 1.56~1.96% by weight, cyclodextrin 0.8~1% by weight, asiaticoside 0.71~1.11% by weight, phenoxyethanol 0.7~0.9% by weight, xanthan gum 0.50~0.70% by weight, and hydroxyethylcellulose 0.13 Encapsulated keratin, characterized in that it is composed of ~0.23% by weight.
카페인, 폴리덱스트로오스, 하이드롤라이즈드케라틴, 케라틴, 아시아티코사이드, 페녹시에탄올, 잔탄검을 첨가하는 단계 (b);
상기 단계 (b) 후, 정제수를 용매로 하여 40~80℃에서 완전히 용해시키는 단계 (c); 를 포함하는 것을 특징으로 하는 케라틴의 캡슐화방법.
Step (a) of mixing hydroxypropylcyclodextrin, maltodextrin, cyclodextrin, xanthan gum, and hydroxyethylcellulose;
Step (b) adding caffeine, polydextrose, hydrolyzed keratin, keratin, asiaticoside, phenoxyethanol, and xanthan gum;
After step (b), (c) completely dissolving at 40-80°C using purified water as a solvent; A method of encapsulating keratin, comprising:
상기 케라틴의 캡슐화방법은,
정제수 69.25~75.35 중량%, 하이드록시프로필사이클로덱스트린 12.09~16.09 중량%, 카페인 2.20~2.40 중량%, 폴리덱스트로오스 1.80~2.20 중량%, 하이드롤라이드즈케라틴 1.68~2.08 중량%, 케라틴 1.68~2.08 중량%, 말토덱스트린 1.56~1.96 중량%, 사이클로덱스트린 0.8~1 중량%, 아시아티코사이드 0.71~1.11 중량%, 페녹시에탄올 0.7~0.9 중량%, 잔탄검 0.50~0.70 중량% 및 하이드록시에틸셀룰로오스 0.13~0.23 중량%를 사용하는 것을 특징으로 하는 케라틴의 캡슐화방법.
According to paragraph 3,
The method of encapsulating the keratin is,
Purified water 69.25~75.35% by weight, hydroxypropylcyclodextrin 12.09~16.09% by weight, caffeine 2.20~2.40% by weight, polydextrose 1.80~2.20% by weight, hydrolides keratin 1.68~2.08% by weight, keratin 1.68~2.08 Weight%, maltodextrin 1.56~1.96% by weight, cyclodextrin 0.8~1% by weight, asiaticoside 0.71~1.11% by weight, phenoxyethanol 0.7~0.9% by weight, xanthan gum 0.50~0.70% by weight, and hydroxyethylcellulose 0.13 A method of encapsulating keratin, characterized by using ~0.23% by weight.
A cosmetic composition comprising the encapsulated keratin of claim 1.
상기 화장료 조성물은,
각질형성세포 분화 유도에 따른 피부 장벽 강화 효과가 있는 것을 특징으로 하는 화장료 조성물.According to clause 5,
The cosmetic composition is,
A cosmetic composition characterized by a skin barrier strengthening effect due to induction of keratinocyte differentiation.
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