KR101099665B1 - A cosmetic composition comprising the aplysia kurodai extract for skin moisturizing effect and improving skin wrinkles - Google Patents
A cosmetic composition comprising the aplysia kurodai extract for skin moisturizing effect and improving skin wrinkles Download PDFInfo
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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
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/98—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin
- A61K8/987—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin of species other than mammals or birds
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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
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/98—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution of animal origin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
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- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Dermatology (AREA)
- Gerontology & Geriatric Medicine (AREA)
- Zoology (AREA)
- Birds (AREA)
- Epidemiology (AREA)
- Cosmetics (AREA)
Abstract
Description
본 발명은 군소 추출물을 함유하여 피부 보습 및 주름개선 효과가 뛰어난 화장용 조성물에 관한 것이다.
The present invention relates to a cosmetic composition containing a small amount extract excellent in skin moisturizing and wrinkle improvement effect.
피부는 나이가 들거나 외부의 자극 등에 지속적으로 노출되게 되면 피부가 손상되고 생리 기능이 저하되는 등 피부 노화가 빠르게 진행된다. 피부 노화는 나이가 들어감에 따라 필연적으로 발생하는 자연스런 현상이다. 그러나, 생활수준의 향상으로 삶의 질에 대한 요구가 증대되고 있고 여러 가지 환경요인에 의해 피부 노화의 발생 요인이 증가한 최근에 들어서 피부 노화에 대한 대책이 절실히 요구된다. As the skin ages or is continuously exposed to external stimuli, skin aging progresses rapidly. Skin aging is a natural phenomenon that inevitably occurs with age. However, in recent years, the demand for quality of life has increased due to the improvement of living standards, and in recent years, the occurrence factor of skin aging has increased due to various environmental factors.
피부 수분량은 진피에서 70% 정도이나 표피로 가면서 감소하여 최외각층인 각질층에서는 약 10~30%가 된다. 진피에서 공급된 물은 각질층 상부로 확산되어 외부로 손실된다. 이것을 경피수분손실(TEWL: Trans Epidermal Water Loss)이라 하며 이 경피수분손실을 막아주는 것 또한 각질층이 지질막 성분인 피지성분과 표피 지질이라고 알려져 있다. 이러한 각질층의 수분보유능은 생체 방어라는 관점에서 중요할 뿐 아니라, 매끄럽고 유연한 피부 표면을 위한 미용상의 관점에서도 대단히 중요한 기능이다. Skin moisture level is about 70% in the dermis, but decreases to the epidermis, about 10-30% in the stratum corneum, the outermost layer. Water supplied from the dermis diffuses up the stratum corneum and is lost to the outside. This is called Trans Epidermal Water Loss (TEWL), and it is also known that the epidermal moisture is prevented from the sebaceous and epidermal lipids, which are the stratum corneum. Moisture retention of these stratum corneum is not only important from the point of view of biological defense, but also from the cosmetic point of view for smooth and flexible skin surface.
그러나 이러한 기능에도 불구하고, 피부는 혹독한 외부환경 즉, 바람, 날씨, 햇빛 등과 세안, 면도 등과 더불어 노화라는 인자가 더해져서 쉽게 수분을 잃게 되어 건조하고 거친 피부가 되기 쉽다. However, in spite of these functions, the skin is easily exposed to harsh external environment, such as wind, weather, sunlight, face wash, shaving, etc., and aging, and thus easily loses moisture, resulting in dry and rough skin.
나이가 들어가면서 피부는 점차 그 고유의 기능이 저하되면서 노화 현상이 일어나고 그 대표적인 예가 주름 생성이다. 이러한 노화의 주된 원인인 태양광의 자외선에 의해 생성된 프리라디칼은 피부의 결합 조직인 콜라겐과 엘라스틴의 산화적 절단을 촉진시키며 결과적으로는 결합조직의 비정상적인 교차결함에 의해 주름을 형성시킨다. 또한 피부의 수분 보유능력에 가장 큰 역할을 하고 있는 히아루론산의 절단 및 저분자화를 촉진시켜 각질층의 수분 보유능력을 감소시키는 한 원인이 되고 있다. 일반적으로 각질층의 수분이 10% 이하로 되는 경우에는 피부의 생리작용이 저하되어 피부가 본래의 기능을 잃게 되어 거칠어지게 된다. As the skin ages, its skin gradually deteriorates, and aging occurs. A typical example is wrinkle formation. The free radicals produced by the ultraviolet rays of sunlight, the main cause of aging, promote the oxidative cleavage of collagen and elastin, the connective tissues of the skin, and as a result, wrinkles are formed by abnormal cross defects of connective tissues. In addition, it promotes the cleavage and low molecular weight of hyaluronic acid, which plays a major role in the skin's water retention ability, thereby reducing the water retention capacity of the stratum corneum. In general, when the moisture of the stratum corneum is less than 10%, the physiological action of the skin is lowered and the skin loses its original function and becomes rough.
또한 피부의 수분 보유 능력에 가장 큰 역할을 하고 있는 히아루론산의 절단 및 저분자화를 촉진시켜 각질층의 수분 보유능력을 감소시키는 한 원인이 되고 있다. 일반적으로 각질층의 수분이 10% 이하로 되는 경우에는 피부의 생리작용이 저하되어 피부가 본래의 기능을 잃게 되어 거칠어지게 된다. In addition, it promotes the cleavage and low molecular weight of hyaluronic acid, which plays a major role in the skin's water retention ability, thereby reducing the water retention capacity of the stratum corneum. In general, when the moisture of the stratum corneum is less than 10%, the physiological action of the skin is lowered and the skin loses its original function and becomes rough.
피부 노화가 진행되면 피부 세포들의 증식 및 재생 능력이 저하된다. 표피에서는 표피 세포의 증식이 저하되고 신진대사가 저하된다. 표피와 마찬가지로 진피에서는 콜라겐이나 엘라스틴과 같은 결합 조직 뿐만 아니라 세포간 지질 성분인 글리코스아미노글리칸과 글리코프로테인 등의 합성 및 분해를 담당하는 섬유아세포의 증식 활성이 저하되고 그 대사 기능도 저하된다. As the skin ages, skin cells proliferate and regenerate. In the epidermis, proliferation of epidermal cells is reduced and metabolism is reduced. Like the epidermis, the dermis decreases the proliferative activity of the fibroblasts responsible for the synthesis and degradation of not only connective tissues such as collagen and elastin but also glycosaminoglycans and glycoproteins, which are intercellular lipid components, and their metabolic functions are also reduced.
따라서, 이러한 현상을 방지하여 매끄럽고 촉촉한 피부를 유지시켜 주는 것이 화장품을 사용하는 가장 중요한 목적이다. 피부에 화장품을 바르게 되면 피부에 인공막을 만들어주어 외부의 자극을 막아주고 피부 내로부터의 수분 손실을 막아줄 뿐 아니라 어느 정도의 수분을 피부에 공급하게 된다. Therefore, it is the most important purpose of using cosmetics to prevent such a phenomenon to maintain a smooth and moist skin. Applying cosmetics to the skin creates an artificial membrane on the skin that prevents external irritation, prevents the loss of moisture from the skin, and supplies some amount of moisture to the skin.
그러나 일반적인 화장품으로서는 매끄럽고 촉촉하며, 피부에 부담감이 없는 피부 관리를 하는 것이 쉬운 일이 아니다. 즉, 종래 보습목적의 크림 또는 로션에 사용되어온 수용성 다가 알코올류, 피로리돈카르복실산 나트륨, 요소, 아미노산, 엘라스틴 등과 같은 보습제들은 각각 문제점 등을 갖고 있다. However, as a general cosmetic, it is not easy to take care of skin that is smooth and moist and has no burden on the skin. That is, moisturizing agents such as water-soluble polyhydric alcohols, sodium pyridonecarboxylic acid sodium, urea, amino acids, elastin, and the like, which have been conventionally used for moisturizing creams or lotions, have problems.
종래 화장품에 보습제로 가장 많이 쓰고 있는 수용성 다가알코올류의 경우 - OH기가 3개 이상인 글리세린, 솔비톨 등은 뛰어난 보습력에 반해 끈적임이 심하여 사용시 불쾌감을 느끼게 하며, -OH기가 2개인 프로필렌 글리콜, 1,3-부틸렌글리콜 등은 다량 사용 시 피부 부작용이 있게 된다. In the case of water-soluble polyhydric alcohols, which are most commonly used as moisturizers in conventional cosmetics-Glycerine and sorbitol having three or more OH groups are highly sticky in contrast to excellent moisturizing properties, and make them feel unpleasant when used. Butylene glycol, etc. will have side effects when used in large amounts.
이에 피부 보습 및 주름 개선 용도로 달팽이 추출물을 개발하는 연구 개발이 활발하게 진행되고 있으나, 달팽이 추출물로부터 피부 보습 등의 효과를 얻기에는 한계가 있어 왔다.
The research and development to develop a snail extract for the purpose of skin moisturizing and wrinkle improvement has been actively progressed, but there has been a limit to obtain the effect of skin moisturizing from snail extract.
본 발명의 목적은 달팽이 추출물 대비 피부 보습 및 주름개선 효과가 뛰어난 피부 보습 및 주름개선용 화장용 조성물을 제공하는 것이다. It is an object of the present invention to provide a skin moisturizing and anti-wrinkle cosmetic composition having excellent skin moisturizing and anti-wrinkle effect compared to snail extract.
본 발명의 다른 목적은 세포 독성이 없고 부작용이 문제되지 않는 천연물 유래의 피부 보습 및 주름개선용 화장용 조성물을 제공하는 것이다.
Another object of the present invention is to provide a cosmetic composition for moisturizing and improving wrinkles derived from natural products, which is not cytotoxic and does not cause side effects.
본 발명의 하나의 관점은 군소(Aplysia kurodai) 육질을 단백분해효소 처리하고, 에탄올 불용성 물질을 침전시켜 얻어지는 추출물을 유효성분으로 함유하는 피부 보습 및 주름개선용 화장용 조성물에 관한 것이다.One aspect of the present invention relates to a cosmetic composition for skin moisturizing and wrinkle improvement comprising an extract obtained by proteolytic treatment of Aplysia kurodai meat and precipitating ethanol insoluble substances.
일 구체예에서, 상기 추출물은 화장용 조성물 중 0.001~50중량%로 함유될 수 있다.
본 발명의 다른 관점은 군소 육질을 분리하고 동결건조하는 단계; 상기 동결건조물에 단백분해효소를 처리하고 원심분리하여 제1상등액을 얻는 단계; 상기 제1상등액에 산을 처리한 후 원심분리하여 제2상등액을 얻는 단계; 및 상기 제2상등액에 에탄올을 첨가하여 에탄올 불용성 물질을 침전시키고 원심분리하여 침전물을 회수하는 단계;를 포함하는 제1항의 화장용 조성물 제조방법에 관한 것이다.
In one embodiment, the extract may be contained in 0.001 to 50% by weight of the cosmetic composition.
Another aspect of the invention is the step of separating and lyophilizing small meat; Treating the lyophilized protease and centrifuging to obtain a first supernatant; Treating the first supernatant with acid and then centrifuging to obtain a second supernatant; And a step of recovering a precipitate by adding ethanol to the second supernatant to precipitate an ethanol insoluble substance and centrifuging the same.
본 발명은 달팽이 추출물 대비 피부 보습 및 주름 개선 효과가 뛰어나고 천연물로부터 유래된 군소 추출물을 함유하는 피부 보습 및 주름개선용 화장용 조성물을 제공하였다.
The present invention provides a cosmetic composition for skin moisturizing and anti-wrinkle, which has a superior skin moisturizing and wrinkle improvement effect compared to the snail extract and contains elemental extracts derived from natural products.
도 1은 군소 추출물 및 달팽이 추출물의 수분 보유능을 나타낸 것으로, X축은 추출물 처리 후 경과시간(시간)을 나타내고, Y축은 수분 보유능(%)을 나타낸다.
도 2는 군소 추출물의 농도에 따른 인간 각질형성세포의 세포활성도를 나타낸 것으로, X축은 군소 추출물의 농도(%)를 나타내고, Y축은 세포활성도(%)를 나타낸다.
도 3은 군소 추출물, 달팽이 추출물 및 레티놀의 농도에 따른 콜라겐의 합성능을 나타낸 것으로, X축은 추출물의 농도(%)를 나타내고, Y축은 코라겐의 합성능(%)을 나타낸다. Figure 1 shows the water retention capacity of the extract and snail extract, the X-axis shows the elapsed time (hours) after the extract treatment, the Y-axis shows the water retention (%).
Figure 2 shows the cell activity of human keratinocytes according to the concentration of the extract, X-axis shows the concentration (%) of the extract, Y-axis shows the cell activity (%).
Figure 3 shows the collagen synthesis capacity according to the concentration of the extract, snail extract and retinol, the X-axis shows the concentration (%) of the extract, the Y-axis shows the synthesis capacity (%) of the collagen.
본 발명의 피부 보습 및 주름개선용 화장용 조성물은 군소 추출물을 유효성분으로 함유할 수 있다.The cosmetic composition for skin moisturizing and wrinkle improvement of the present invention may contain a small extract as an active ingredient.
군소(Aplysia kurodai)는 대한민국 전 연안에 분포하며 연체동물문 복족강 군소목 군소과에 속하는 연체동물이다. Aplysia kurodai is a mollusk that is distributed all over the coast of Korea and belongs to the mollusk moon Bok-gang group.
상기 군소 추출물은 군소에 단백질 분해효소를 처리하고 에탄올 불용성 물질을 침전시켜 얻어질 수 있다. 이때 추출을 위한 군소 부위는 내장을 제거하고 육질을 이용하는 것이 바람직하다.
본 발명의 상기 화장용 조성물은 군소 육질을 분리하고 동결건조하는 단계; 상기 동결건조물에 단백분해효소를 처리하고 원심분리하여 제1상등액을 얻는 단계; 상기 제1상등액에 산을 처리한 후 원심분리하여 제2상등액을 얻는 단계; 및 상기 제2상등액에 에탄올을 첨가하여 에탄올 불용성 물질을 침전시키고 원심분리하여 침전물을 회수하는 단계;를 포함하여 제조될 수 있다.
상기 군소의 추출 방법은 구체적으로 통상의 단백질 분해 효소 예를 들면 플라보자임(Flavourzyme)을 군소에 첨가하여 군소의 육질을 가수분해한다. 얻은 가수 분해물을 원심분리한다. 얻은 상등액(제1상등액)에 산 예를 들면 트리클로로아세트산을 첨가하여 가수분해된 단백질 분해물을 침전시켜 제거한다. 여기서 얻은 상등액(제2상등액)에 알코올성 용매(예를 들면 탄소수 1-5의 알코올 또는 물과의 혼합물, 특히 에탄올)를 첨가하여 알코올 불용성 물질(침전물)을 얻은 다음, 침전물을 원심분리하여 회수함으로써 이를 군소 추출물로 사용할 수 있다.The elemental extract can be obtained by treating proteolytic enzymes in the element and precipitating ethanol insoluble material. At this time, it is preferable to remove the viscera and use meat quality for the small portions for extraction.
The cosmetic composition of the present invention comprises the steps of separating and lyophilizing small meat; Treating the lyophilized protease and centrifuging to obtain a first supernatant; Treating the first supernatant with acid and then centrifuging to obtain a second supernatant; And ethanol is added to the second supernatant to precipitate the ethanol insoluble substance and to recover the precipitate by centrifugation.
Specifically, the method of extracting a small amount of hydrolysates the small amount of the small amount by adding a conventional proteolytic enzyme, for example, Flavorzyme to the small amount. Centrifuge the obtained hydrolyzate. An acid such as trichloroacetic acid is added to the obtained supernatant (first supernatant) to precipitate and remove the hydrolyzed proteolytic products. The supernatant (second supernatant) obtained here is added an alcoholic solvent (e.g., a mixture of alcohols having 1 to 5 carbon atoms or water, in particular ethanol) to obtain an alcohol insoluble substance (precipitate), and then the precipitate is recovered by centrifugation. It can be used as a component extract.
본 발명의 실시예에 따르면, 군소 추출물은 달팽이 추출물 대비 피부 보습 및 주름 개선 효과를 비교한 결과, 동일 추출물 농도에서 현저하게 향상된 수분보유능과 콜라겐 합성능을 갖고 있음을 알 수 있다.According to an embodiment of the present invention, the result of comparing the moisturizing and wrinkle improvement effect compared to the extract snail extract, it can be seen that it has a significantly improved water retention and collagen synthesis ability at the same extract concentration.
상기 군소 추출물은 화장용 조성물 중 0.001~50중량%로 함유될 수 있다. 상기 범위 내에서, 피부 보습 및 주름 개선 효과가 뛰어날 수 있고, 함유량 증가 대비 개선 효과가 클 수 있다. 바람직하게는 화장용 조성물 중 0.05~20.0중량%로 함유될 수 있다.The ingredient extract may be contained in 0.001 to 50% by weight of the cosmetic composition. Within this range, the skin moisturizing and wrinkle improving effect can be excellent, and the improvement effect compared to the content increase can be large. Preferably it may be contained in 0.05 to 20.0% by weight of the cosmetic composition.
상기 군소 추출물은 약학적 조성물 또는 화장용 조성물로 사용될 수 있다. 약학적 조성물로 사용될 경우, 사용시에 연고제, 크림제, 로숀제, 액제, 스프레이제 등의 국소 적용제 등의 다양한 제제로 이용할 수 있다. 바람직하게는, 연고제, 로숀제, 스프레이제 등의 국소 적용 제제로 제형화시켜 피부 일부분에 직접 적용시키는 것이 효과적이다. 또한, 화장용 조성물로 사용될 경우, 상기 화장용 조성물은 그 제형이 화장수, 로션, 영양크림, 에센스, 마사지크림 및 팩으로 이루어진 군으로부터 선택되는 제형으로 사용될 수 있다.The group extract may be used as a pharmaceutical composition or a cosmetic composition. When used as a pharmaceutical composition, it can be used in various formulations such as topical application such as ointment, cream, lotion, liquid, spray, etc. in use. Preferably, it is effective to formulate into topical application preparations, such as ointments, lotions, sprays, etc., and apply them directly to a portion of the skin. In addition, when used as a cosmetic composition, the cosmetic composition may be used in a formulation selected from the group consisting of a lotion, lotion, nutrition cream, essence, massage cream and pack.
이하, 본 발명의 바람직한 실시예를 통해 본 발명의 구성 및 작용을 더욱 상세히 설명하기로 한다. 다만, 이는 본 발명의 바람직한 예시로 제시된 것이며 어떠한 의미로도 이에 의해 본 발명이 제한되는 것으로 해석될 수는 없다.Hereinafter, the configuration and operation of the present invention through the preferred embodiment of the present invention will be described in more detail. However, this is presented as a preferred example of the present invention and in no sense can be construed as limiting the present invention.
여기에 기재되지 않은 내용은 이 기술 분야에서 숙련된 자이면 충분히 기술적으로 유추할 수 있는 것이므로 그 설명을 생략하기로 한다.
Details that are not described herein will be omitted since those skilled in the art can sufficiently infer technically.
실시예: 군소 추출물의 제조Example: Preparation of Bovine Extract
군소는 끓는 물에서 5분 동안 자숙하고, 육질과 내장을 분리한 다음, 육질 부분만을 동결건조시켜 -20℃의 냉동고에서 보관하면서 추출물 원재료로 사용하였다. 군소 동결건조 시료 1g에 대해 50mM sodium phosphate 완충액(pH 6.0)을 10배량으로 첨가하였다. 동결건조 시료에 대하여 단백질 농도비 1/50이 되도록 단백질 분해 효소(Flavourzyme)를 첨가하고 60℃의 항온 수조에서 흔들어주면서 15시간 동안 반응시켰다. 얻은 가수분해물을 원심 분리(3,000g, 30분)하고 상등액을 취하였다. 상등액에 대해 최종 농도가 0.5%가 되도록 트리클로로아세트산을 첨가하고 25℃에서 1시간 동안 방치하였다. 그런 다음 얻은 반응물을 원심분리(3,000g, 30분)하여 침전물을 제거하고 상등액을 취하였다. 상등액에 대해 10배량의 에탄올을 첨가하여 에탄올 불용성 물질을 침전시키고, 원심분리(3,000g, 30분)하여 침전물을 분리함으로써 군소 추출물을 얻었다.
Sauce was boiled in boiling water for 5 minutes, the meat and the intestines were separated, and only the meat part was lyophilized and stored in a freezer at -20 ° C and used as an extract raw material. Ten milliliters of 50 mM sodium phosphate buffer (pH 6.0) was added to 1 g of small lyophilized sample. Proteolytic enzyme (Flavourzyme) was added to the lyophilized sample so as to have a protein concentration ratio of 1/50 and reacted for 15 hours while shaking in a constant temperature bath at 60 ° C. The obtained hydrolyzate was centrifuged (3,000 g, 30 minutes) and the supernatant was taken. Trichloroacetic acid was added to a final concentration of 0.5% relative to the supernatant and left at 25 ° C. for 1 hour. Then, the obtained reaction was centrifuged (3,000 g, 30 minutes) to remove the precipitate and the supernatant was taken. 10 times the amount of ethanol was added to the supernatant to precipitate the ethanol insoluble substance, and centrifuged (3,000 g, 30 minutes) to separate the precipitate to obtain a small extract.
비교예:달팽이 추출물의 제조Comparative Example: Preparation of Snail Extract
상기 실시예에서 군소 대신에 달팽이(Acusta despecta sieboldiana)를 사용한 것을 제외하고는 동일한 방법을 실시하여 달팽이 추출물을 제조하였다.
Except for using a snail ( Acusta despecta sieboldiana ) in place of the above embodiment in the same manner to prepare a snail extract.
실험예 1: 수분 보유능 평가Experimental Example 1: Evaluation of Water Retention Capacity
추출물의 수분 보유능을 검증하였다. 페트리 디쉬에 상기 실시예와 비교예에서 제조한 군소 추출물과 달팽이 추출물을 각각 첨가하고 적당한 크기의 종이를 적신 다음 종이의 무게를 재었다. 각 추출물을 함유하는 종이를 빈 페트리 디쉬에 넣고 시간이 지남에 따라 종이의 무게를 측정하여 수분 보유능을 확인하였다. The water retention capacity of the extract was verified. The Petri dishes were added with the extracts of the small and snail extracts prepared in Examples and Comparative Examples, respectively, and wetted with paper of a suitable size, and then weighed. The paper containing each extract was placed in an empty Petri dish to determine the moisture retention by weighing the paper over time.
음성 대조군으로 추출물 대신에 정제수를 첨가하고 동일한 방법을 실시하였다. 수분 보유능은 하기 식 1에 의해 평가할 수 있고, 그 결과를 하기 표 1 및 도 1에 나타내었다.As a negative control, purified water was added instead of the extract and the same method was performed. Moisture retention capacity can be evaluated by the following
<식 1><
수분 보유능(%)=(경과시간 후 무게-종이 무게)X100/(최초 무게 -종이무게)Moisture Retention Capacity (%) = (Weight-Paper Weight After Elapsed Time) X100 / (Initial Weight-Paper Weight)
Cow extract
상기 표 1 및 도 1에서 살핀 바와 같이, 군소 추출물은 동일 시간에 대해 달팽이 추출물보다 수분보유능이 현저하게 뛰어난 것으로 나타났다.
As shown in Table 1 and Figure 1, the salin extract was found to be significantly better water retention than the snail extract for the same time.
실험예Experimental Example 2: 세포 독성 및 활성도 실험 2: Cytotoxicity and Activity Experiment
추출물이 피부에 대한 자극과 세포 독성이 없음을 확인한다. 인간 각질형성 세포 HaCat(ACTT, CLS 300493, USA)를 96웰 플레이트에 5 x 103 셀/웰씩 동일하게 헤마사이토미터(hemacytometer)를 이용하여 계수한 후, 분주하고, 37℃, 5%의 이산화탄소 조건에서 24시간 동안 배양하였다. 24시간 배양한 후 얻은 세포 배양액을 제거하고, 상기 실시예와 비교예에서 얻은 군소 추출물과 달팽이 추출물을 0, 0.05, 0.1, 0.5, 1 및 2(중량/부피)%로 각 웰에 100㎕씩 넣고 DMEM 배지에서 37℃, 5% 이산화탄소 조건에서 24시간 동안 배양하였다. 그 후 웰당 WST를 10㎕씩 첨가하고 동일한 조건에서 4시간 동안 더 배양하였다. 그 후 ELISA reader(Thermo, Multiskan EX)로 420nm에서 흡광도를 측정하였다.Confirm that the extract is non-irritating and cytotoxic to the skin. Human keratinocytes HaCat (ACTT, CLS 300493, USA) were counted using a hemacytometer equally at 5 x 10 3 cells / well in 96-well plates, aliquoted, and then aliquoted at 37 ° C., 5% Incubated for 24 hours in carbon dioxide conditions. Cell culture solution obtained after culturing for 24 hours was removed, and 100 µl of each of the small and snail extracts obtained in Examples and Comparative Examples was 0, 0.05, 0.1, 0.5, 1 and 2 (weight / volume) in each well. Incubated for 24 hours at 37 ℃, 5% carbon dioxide conditions in DMEM medium. Thereafter, 10 μl of WST was added per well, and the cells were further incubated for 4 hours under the same conditions. Thereafter, the absorbance was measured at 420 nm with an ELISA reader (Thermo, Multiskan EX).
상기에서, 추출물 대신에 동일한 양의 정제수를 사용한 것을 제외하고는 동일한 방법을 실시하여 흡광도를 측정하였다. 양성 대조군으로 추출물 대신에 동일한 양의 DMEM(10% FBS 함유)를 사용하고, 음성 대조군으로 추출물 대신에 동일한 양의 DMSO(10% 농도)를 사용하여 흡광도를 측정하였다.In the above, absorbance was measured by the same method except that the same amount of purified water was used instead of the extract. Absorbance was measured using the same amount of DMEM (containing 10% FBS) instead of the extract as a positive control and the same amount of DMSO (10% concentration) instead of the extract as the negative control.
세포 활성 정도는 하기 식 2에 의해 나타낼 수 있고, 그 결과를 하기 표 2 및 도 2에 나타내었다.The degree of cell activity can be represented by the following
<식 2><
세포 활성도(%) = (실험군의 흡광도 X 100)/양성 대조군의 흡광도% Cell activity = (
세포 독성 물질인 DMSO를 넣은 음성 대조군의 경우 세포 활성도는 22%로 나왔다. 그러나, 상기 표 2에서 나타난 바와 같이, 군소 추출물은 높은 농도에서도 세포 독성을 보이지 않았고, 오히려 인간 각질세포 생장을 촉진시켜 세포 재생효과가 있는 것을 확인하였다. 또한, 군소 추출물은 달팽이 추출물보다 인간 각질세포 생장 촉진 효과가 높았음을 알 수 있다.
In the negative control group containing the cytotoxic substance DMSO, the cell activity was 22%. However, as shown in Table 2, the extract showed no cytotoxicity at high concentrations, rather it was confirmed that there is a cell regeneration effect by promoting the growth of human keratinocytes. In addition, it can be seen that the extract is higher in human keratinocyte growth promoting effect than the snail extract.
실험예 3: Collagen 합성 실험Experimental Example 3: Collagen Synthesis Experiment
군소 추출물이 주름 개선능이 있는지 여부를 확인한다. 인간 섬유아세포(human fibroblast cell)를 이용하여 추출물의 주름개선능 여부를 확인한다. Check if the extract has anti-wrinkle ability. Human fibroblast cells (human fibroblast cells) to determine whether the wrinkle improvement of the extract.
인간 섬유아세포(ACTT, PCS-201-012, USA)를 24웰 플레이트에 4 x 104셀/웰씩 동일하게 헤마사이토미터(hemacytometer)를 이용하여 계수한 후, 분주하고, 37℃, 5%의 이산화탄소 조건에서 24시간 동안 배양하였다. 24시간 배양한 후 얻은 세포 배양액을 제거하고, 상기 실시예에서 얻은 추출물을 0.05, 0.1 및 0.2(중량/부피)%로 각 웰에 처리하고 DMEM 배지에서 37℃, 5% 이산화탄소 조건에서 24시간 동안 배양하였다. 그 후 배양액을 회수하여 96웰 플레이트에 100㎕씩 넣고 4℃에서 overnight하여 코팅한다. PBS_T solution으로 3반복 washing을 하고 5% Skim milk를 100㎕씩 각 웰에 넣고 90분 동안 흔들어주었다. Collagen antibody(COL1A1(L-19), Santa Cruz Biotechnology, Inc)(1:500)를 100㎕씩 각 웰에 넣고 1시간 동안 흔들어주었다. PBS-T solution으로 3반복 washing 한 후 Secondary antibody(Anti Goat IgG Antibody(Alkaline Phosphatase, Novus Biologicals)(1:10,000)를 100ul씩 각 웰에 넣고 1시간 동안 흔들어주었다. 각 웰 당 AP 100㎕씩 넣고 30분간 암 조건에서 흔들어주었다. 그 후 ELISA reader(Thermo, Multiskan EX)로 405nm에서 흡광도를 측정하였다.Human fibroblasts (ACTT, PCS-201-012, USA) were counted using a hemacytometer equally at 4 x 10 4 cells / well in a 24-well plate, aliquoted, 37 ° C., 5% Incubated for 24 hours under carbon dioxide conditions. Cell cultures obtained after incubation for 24 hours were removed, and the extracts obtained in the above examples were treated in each well at 0.05, 0.1, and 0.2 (weight / volume)% in DMEM medium at 37 ° C., 5% carbon dioxide conditions for 24 hours. Incubated. After that, the culture solution is collected, and 100 µl each is placed in a 96-well plate and coated overnight at 4 ° C. After washing three times with PBS_T solution and put the 5% Skim milk in each well 100μl was shaken for 90 minutes. Collagen antibody (COL1A1 (L-19), Santa Cruz Biotechnology, Inc) (1: 500) was added to each well 100μl and shaken for 1 hour. After washing three times with PBS-T solution, 100 μl of secondary antibody (Anti Goat IgG Antibody (Alkaline Phosphatase, Novus Biologicals) (1: 10,000) was added to each well and shaken for 1 hour. After shaking for 30 minutes under dark conditions, absorbance was measured at 405 nm with an ELISA reader (Thermo, Multiskan EX).
상기 군소 추출물 대신에 비교예의 달팽이 추출물을 사용한 것을 제외하고는 동일한 방법을 실시하여 흡광도를 측정하였다. 또한, 주름개선능이 있는 것으로 알려진 레티놀(0.5μM)에 대해 동일한 방법을 실시하여 흡광도를 측정하였다. 양성 대조군으로 추출물을 사용하지 않고 DMEM(10% FBS 함유)를 사용한 것을 제외하고는 동일한 방법을 실시하여 흡광도를 측정하였다. Absorbance was measured by the same method except that the snail extract of Comparative Example was used instead of the small extract. In addition, the absorbance was measured by the same method for retinol (0.5 µM) known to have wrinkle improvement. Absorbance was measured in the same manner, except that DMEM (containing 10% FBS) was used as the positive control and no extract was used.
Collagen 합성 정도는 하기 식 3에 의해 나타낼 수 있고, 그 결과를 하기 표 3 및 도 3에 나타내었다.Collagen synthesis degree can be represented by the following
<식 3><
Collagen 합성능(%) = (실험군의 흡광도 X 100)/양성 대조군의 흡광도Collagen synthetic ability (%) = (
상기 표 3에서 나타난 바와 같이, 군소 추출물은 동일한 농도에서 달팽이 추출물 대비 콜라겐 합성능이 현저히 높은 것으로 나타났다. 레티놀은 0.5μM로 사용시 콜라겐 합성능이 50%인 것으로 나타났다. 군소 추출물은 주름개선능이 있는 것으로 알려진 레티놀(0.5uM) 대비 0.2%에서 유사한 효과가 있음을 알 수 있다.
As shown in Table 3, the extract was significantly higher collagen synthesis capacity than the snail extract at the same concentration. Retinol was found to have a collagen synthesis capacity of 50% when used at 0.5 μM. It can be seen that the extract has a similar effect at 0.2% compared to retinol (0.5uM) known to have wrinkle improvement.
제제예Formulation example 1: 화장수의 제조 1: Preparation of the lotion
하기 표 4에 기재된 성분을 사용하여 통상의 방법으로 화장수를 제조하였다.To make up the lotion by the conventional method using the components shown in Table 4.
제제예Formulation example 2: 영양로션의 제조 2: Preparation of Nutritional Lotion
하기 표 5에 기재된 성분을 사용하여 통상의 방법을 사용하여 영양로션을 제조하였다. The nutrition lotion was prepared using the conventional method using the component of Table 5 below.
Claims (3)
A cosmetic composition for moisturizing skin and improving wrinkles comprising an extract obtained by treating proteolytic enzymes with Aplysia kurodai and precipitating ethanol insoluble substances.
상기 추출물은 상기 화장용 조성물 중 0.001-50중량%로 함유되는 것을 특징으로 하는 피부 보습 및 주름개선용 화장용 조성물.
The method of claim 1,
The extract is a cosmetic composition for skin moisturizing and wrinkle improvement, characterized in that it is contained in 0.001-50% by weight of the cosmetic composition.
상기 동결건조물에 단백분해효소를 처리하고 원심분리하여 제1상등액을 얻는 단계;
상기 제1상등액에 산을 처리한 후 원심분리하여 제2상등액을 얻는 단계; 및
상기 제2상등액에 에탄올을 첨가하여 에탄올 불용성 물질을 침전시키고 원심분리하여 침전물을 회수하는 단계;
를 포함하는 제1항의 화장용 조성물 제조방법.Separating and lyophilizing the beef meat;
Treating the lyophilized protease and centrifuging to obtain a first supernatant;
Treating the first supernatant with acid and then centrifuging to obtain a second supernatant; And
Adding ethanol to the second supernatant to precipitate the ethanol insoluble substance and centrifuging to recover the precipitate;
Cosmetic composition manufacturing method of claim 1 comprising a.
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KR20190067442A (en) * | 2017-12-07 | 2019-06-17 | 주식회사 선마린바이오테크 | Cosmetic composition comprising oysters, sea cucumber, sea squirt, Aplysia kurodai and buccinum striatissimum |
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KR101918773B1 (en) | 2012-05-03 | 2018-11-14 | 박시향 | Composition For Improving Hair Condition and Preventing Hair Loss Comprising an Extract of Aplysia kurodai |
KR20190067442A (en) * | 2017-12-07 | 2019-06-17 | 주식회사 선마린바이오테크 | Cosmetic composition comprising oysters, sea cucumber, sea squirt, Aplysia kurodai and buccinum striatissimum |
KR102011654B1 (en) * | 2017-12-07 | 2019-08-19 | 주식회사 선마린바이오테크 | Cosmetic composition comprising oysters, sea cucumber, sea squirt, Aplysia kurodai and buccinum striatissimum |
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