KR20230109833A - Composition for improving skin condition with anti-wrinkle, anti-oxidant and moisturizing of UVB-damaged skin from extract of Citrus junos seed shell as active ingredient - Google Patents
Composition for improving skin condition with anti-wrinkle, anti-oxidant and moisturizing of UVB-damaged skin from extract of Citrus junos seed shell as active ingredient Download PDFInfo
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- KR20230109833A KR20230109833A KR1020220005575A KR20220005575A KR20230109833A KR 20230109833 A KR20230109833 A KR 20230109833A KR 1020220005575 A KR1020220005575 A KR 1020220005575A KR 20220005575 A KR20220005575 A KR 20220005575A KR 20230109833 A KR20230109833 A KR 20230109833A
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- composition
- citron
- skin
- improving skin
- derived
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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
- A61K8/18—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
- A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
- A61K8/9783—Angiosperms [Magnoliophyta]
- A61K8/9789—Magnoliopsida [dicotyledons]
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/75—Rutaceae (Rue family)
- A61K36/752—Citrus, e.g. lime, orange or lemon
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Abstract
유자박 유래 분획물을 포함하는 피부개선용 조성물이 제공된다. A composition for improving skin comprising a fraction derived from citron peel is provided.
Description
본 발명은 유자박 유래 분획물의 추출물을 포함하는 자외선에 의해 손상된 피부의 주름 개선, 보습 및 미백 기능의 피부개선용 조성물에 관한 것으로, 보다 상세하게는 부작용을 일으키지 않고, 자외선(UVB)에 의해 유도되는 피부 주름 억제 효과, 피부 손상에 대한 복구 효과, 보습 효과, 미백 효과가 우수하여 노화방지 및 피부 복구를 위한 화장품 조성물에 유용하게 사용될 수 있는 유자박 유래 분획물을 포함하는 피부주름개선, 피부보습 및 미백 기능을 갖는 피부개선용 조성물에 관한 것이다.The present invention relates to a composition for improving wrinkles, moisturizing and whitening of skin damaged by ultraviolet rays, containing an extract of a fraction derived from citron peel, for skin improvement, and more particularly, to a composition for improving skin with wrinkle improvement, moisturizing and whitening functions, including a fraction derived from citron peel, which does not cause side effects and has excellent skin wrinkle suppression effect, repair effect for skin damage, moisturizing effect, and whitening effect, which can be usefully used in cosmetic compositions for anti-aging and skin restoration. It's about the composition.
피부는 인체의 가장 첫 번째 보호막으로서 기능을 하고 있다. 온도, 습도의 변화, 자외선, 박테리아와 다양한 환경 자극으로부터 인체를 보호해 주며 항상성 유지에도 중요한 기능을 하고 있다. 그러나 자외선, 박테리아 감염과 같은 환경 자극을 받은 피부는 정상적인 기능을 수행 할 수 없으며 주름 생성, 탄력 손실 등의 피부노화를 촉진하게 된다. 이러한 피부 노화를 방지하고 보다 건강한 피부를 유지하기 위해 다양한 식물들로부터 얻은 생리활성물질들이 포함된 화장품을 사용함으로써 피부의 기능과 건강을 유지하고 있다. The skin functions as the first protective layer of the human body. It protects the human body from changes in temperature and humidity, ultraviolet rays, bacteria and various environmental stimuli and plays an important role in maintaining homeostasis. However, skin exposed to environmental stimuli such as ultraviolet rays and bacterial infections cannot perform normal functions and accelerates skin aging such as wrinkle formation and loss of elasticity. In order to prevent such skin aging and maintain healthier skin, the function and health of the skin are maintained by using cosmetics containing physiologically active substances obtained from various plants.
자외선은 파장에 따라 자외선A (UVA), 자외선B (UVB) 자외선C (UVC) 3 가지로 분류 되는데, 이 중 자외선C는 오존층에 완전히 흡수된다. 따라서 자외선A (UVA) 와 자외선B (UVB)가 피부 노화와 손상을 일으키는 주범이고 특히 자외선B (UVB)의 경우, 피부를 뚫고 들어가 피부암을 일으키는 원인이기도 하다.Ultraviolet rays are classified into three types according to the wavelength: ultraviolet A (UVA), ultraviolet B (UVB), and ultraviolet C (UVC). Among them, ultraviolet C is completely absorbed by the ozone layer. Therefore, ultraviolet A (UVA) and ultraviolet B (UVB) are the main culprits of skin aging and damage, and in the case of ultraviolet B (UVB) in particular, it is also the cause of skin cancer by penetrating the skin.
활성산소 (Reactive Oxygen Species, ROS)는 산소로 전자가 전달되면서 순차적으로 생성되는 수퍼옥사이드(Superoxide), 과산화수소(H2O2), 하이드록시래디칼(Hydroxyl radical) 등으로 세포 내 다양한 효소와 지질, DNA를 손상시킨다. 특히 자외선이 피부 세포에 자극을 주어 세포 노화가 진행 될수록 산화 손상으로 인해 세포 내 활성산소가 증가 하는데 이는 심각한 피부 손상을 야기 할 수 있다.Reactive Oxygen Species (ROS) are superoxide, hydrogen peroxide (H 2 O 2 ), and hydroxyl radicals, which are sequentially generated as electrons are transferred to oxygen, and damage various enzymes, lipids, and DNA in cells. In particular, as UV rays stimulate skin cells and cell aging progresses, intracellular active oxygen increases due to oxidative damage, which can cause serious skin damage.
피부는 또한 신체 내부의 수분 증발을 감소시키는 역할도 한다. 피부의 각질층은 표피를 통한 수분 손실을 최소화하고 병원체, 알레르겐 및 독성 화학물질의 유입을 방지하는데 필수적이다. 히알루론산 (hyaluronic acid, HA)은 글리코사미노글리칸 (glycosaminoglycan, GAG)의 한 종류로, 주로 피부 조직의 세포 외 매트릭스 (ECM)에 분포되어 있다. 히알루론산은 수분과 강하게 결합하여 피부 수분 유지에 중요한 역할을 한다. 필라그린 (filaggrin)은 각질층의 형성, 표피의 말단 분화를 촉진하는 주요 구조 단백질이다. 세라마이드는 피부 표피의 각질층에 우세한 지질로 존재하며 수분 유지 및 장벽기능에서 중요한 역할을 한다. 표피의 세라마이드는 세린 팔미토일트랜스퍼라제 (serinpalmitoyltransferase, SPT)와 같은 효소에 의해 합성된다.The skin also serves to reduce the evaporation of water inside the body. The stratum corneum of the skin is essential for minimizing water loss through the epidermis and preventing the entry of pathogens, allergens and toxic chemicals. Hyaluronic acid (HA) is a type of glycosaminoglycan (GAG), which is mainly distributed in the extracellular matrix (ECM) of skin tissue. Hyaluronic acid binds strongly to moisture and plays an important role in maintaining skin moisture. Filaggrin is a major structural protein that promotes the formation of the stratum corneum and terminal differentiation of the epidermis. Ceramide is present as a predominant lipid in the stratum corneum of the skin epidermis and plays an important role in water retention and barrier function. Ceramides in the epidermis are synthesized by enzymes such as serine palmitoyltransferase (SPT).
그리고 최근 화학성분이나 합성 성분의 유해성에 대한 인식이 높아지면서 천연 화장품에 대한 주목과 소비가 늘고 있으며, 천연 화장품들은 주로 우리가 섭취하는 식물에서 추출한 유래 성분이 포함된 화장품으로 효능과 안정성이 입증되어 인체에 자극이 적고 안전하다는 인식이다. 따라서, 이러한 흐름에 따라 질병에 효과가 있다고 알려진 다양한 식물의 추출물에 대해 효능을 입증하는 연구가 필요한 실정이다. In addition, as awareness of the harmfulness of chemical or synthetic ingredients has recently increased, attention and consumption of natural cosmetics are increasing. Natural cosmetics are mainly cosmetics containing ingredients extracted from plants that we consume. Therefore, in accordance with this trend, studies proving the efficacy of extracts of various plants known to be effective against diseases are required.
유자(Citrus junos)는 운향과 귤속에 속하는 교목성으로 중국의 사천성, 호북성, 운남성 및 티베트 등지에 야생하며 우리나라에는 신라시대에 중국으로부터 전래되어 현재 남해안에 걸쳐 재배되어 오고 있다. 유자는 주로 설탕으로 당 절임 한 청의 형태로 제조되어 차, 드레싱 소스, 식초 등으로 사용된다. 유자의 알려진 성분으로는 caffeic acid, tannic acid 등의 페놀성 화합물과 rutin, hesperidin, naringenin과 같은 플라보노이드류, limonoid류 및 vitamin C가 함유되어 있는 것으로 알려졌다. 특히 함유량이 비교적 많운 limonoid의 경우 위암, 유방암, 간암 등에 있어서 항암효과가 있다고 보고되고 있다. Yuzu ( Citrus junos ) is an arboreal tree belonging to the genus Rutaceae and is wild in China's Sichuan, Hubei, Yunnan and Tibet. Yuzu is mainly prepared in the form of sugar-pickled cheong and used as tea, dressing sauce, and vinegar. Citron is known to contain phenolic compounds such as caffeic acid and tannic acid, flavonoids such as rutin, hesperidin, and naringenin, limonoids, and vitamin C. In particular, limonoid, which has a relatively high content, has been reported to have anticancer effects in stomach cancer, breast cancer, and liver cancer.
하지만, 유자의 총 중량 중 15%에 이르는 씨는 대부분 폐기 처분되는 실정이며, 특히 이들 유자씨나 유자씨박을 활용하여 식품 및 화장품에서 응용 연구나 생리활성 연구는 국내외적으로 보고된 바가 거의 없다.However, most of the seeds, which account for 15% of the total weight of citron, are discarded, and in particular, application studies or physiological activity studies in food and cosmetics using these citron seeds or citron seed meal are rarely reported domestically and internationally.
따라서, 본 발명이 해결하고자 하는 과제는 유자씨나 유자씨박을 활용한 피부개선용 조성물과, 그 제조방법을 제공하는 것이다. Therefore, the problem to be solved by the present invention is to provide a composition for improving skin using citron seeds or citron seed meal, and a manufacturing method thereof.
상기 과제를 해결하기 위하여, 본 발명은 유자박 유래 분획물을 포함하는 피부개선용 조성물을 제공한다. In order to solve the above problems, the present invention provides a composition for improving skin comprising a fraction derived from citron peel.
본 발명의 일 실시예에서, 상기 유자박 유래 분획물은 유자씨로부터 유래된 분획물이며, 상기 피부개선용 조성물은 자외선에 의해 유도되는 주름 개선, 보습, 또는 미백 효과를 갖는다. In one embodiment of the present invention, the citron peel-derived fraction is a fraction derived from citron seeds, and the skin improving composition has an ultraviolet-induced wrinkle improvement, moisturizing, or whitening effect.
본 발명의 일 실시예에서, 상기 분획물은 상기 유자씨 박 추출물을 에틸아세테이트(Ethyl acetate, EA), 헥산(Hex), 부탄올(BuOH)을 용매로 사용하여 분획된 것이다. In one embodiment of the present invention, the fraction is obtained by fractionating the yuja seed gourd extract using ethyl acetate (EA), hexane (Hex), and butanol (BuOH) as solvents.
본 발명의 일 실시예에서, 상기 조성물은 자외선에 의해 유도되는 주름 개선은 MMP-1/9/13 단백질, Collagen-1 단백질 또는 이를 코딩하는 mRNA의 발현을 억제 또는 증가시켜 주름 개선 효과를 갖는다. In one embodiment of the present invention, the composition has an anti-wrinkle effect by suppressing or increasing the expression of MMP-1/9/13 protein, Collagen-1 protein or mRNA encoding them for wrinkle improvement induced by ultraviolet rays.
본 발명의 일 실시예에서, 상기 추출물은 자외선에 의해 손상된 보습 관련 인자인 히알루론산, 필라그린 및 SPT 중 적어도 어느 하나의 발현을 증가시켜 보습효과를 갖는다. In one embodiment of the present invention, the extract has a moisturizing effect by increasing the expression of at least one of hyaluronic acid, filaggrin, and SPT, which are moisturizing-related factors damaged by ultraviolet rays.
본 발명의 일 실시예에서, 상기 조성물은 자극에 의해 생성된 멜라닌 생성 촉진인자 TRP-1, TRP-2, 티로시나제(Tyrosinase) 및 Mitf 중 적어도 어느 하나의 발현을 감소시킴으로써 미백효과를 갖는다. In one embodiment of the present invention, the composition has a whitening effect by reducing the expression of at least one of TRP-1, TRP-2, tyrosinase, and Mitf, which are melanin production promoters produced by stimulation.
본 발명은 또한 상술한 피부개선용 조성물을 포함하는 화장료를 제공한다. The present invention also provides a cosmetic comprising the above-described composition for improving skin.
본 발명은 또한 유자박 유래 분획물을 포함하는 피부개선용 약학 조성물을 제공하며, 상기 약학 조성물은 자외선에 의해 유도되는 주름 개선 또는 미백용 조성물이다. The present invention also provides a pharmaceutical composition for skin improvement comprising a fraction derived from citron peel, wherein the pharmaceutical composition is a composition for improving wrinkles or whitening induced by ultraviolet rays.
본 발명에 따르면, 유자박 유래 분획물을 포함하는 피부 노화 방지용 약학조성물은 천연물로부터 얻어진 물질을 함유하기 때문에 부작용을 일으키지 않고, 자외선에 의해 유도되는 피부 주름 억제 효과, 피부 손상에 대한 복구 효과, 보습 효과, 미백 효과가 우수하여 노화방지 및 피부 복구를 위한 화장료와 자외선 손상에 따른 피부개선과 치료용의 약학 조성물(예를 들면 크림 등의 외용제)로 사용될 수 있다. According to the present invention, the pharmaceutical composition for preventing skin aging, including a fraction derived from citron peel, does not cause side effects because it contains substances obtained from natural products, and has an excellent effect of inhibiting skin wrinkles induced by ultraviolet rays, repairing skin damage, moisturizing effect, and whitening effect, so that it can be used as a cosmetic for anti-aging and skin recovery, and a pharmaceutical composition for improving and treating skin damage caused by ultraviolet rays (for example, external preparations such as creams).
도 1은 본 발명의 실시예에 따른 유자 씨박으로부터 추출물과, 용매 특성에 따른 분획물을 제조하는 단계를 설명하는 도면이다.
도 2는 인간각질세포(HaCaT)에서 유자박 유래 분획물의 세포독성 분석 결과 및 UVB 조사로 유도되는 주름 인자와 손상된 콜라겐이 유자박 유래 분획물 처리에 따라 생성량이 변화하는 결과를 나타낸 도면이다.
도 3은 인간섬유아세포(HDF)에서 유자박 유래 분획물의 세포독성 분석 결과 및 UVB 조사로 유도되는 주름 인자와 손상된 콜라겐이 유자박 유래 분획물 처리에 따라 생성량이 변화하는 결과를 나타낸 도면이다.
도 4는 UVB 조사로 손상된 히알루론산 (hyaluronic acid) 생성량과 피부 표피의 구성에 필요한 필라그린 (filaggrin)과 SPT 효소가 유자박 유래 분획물 처리에 따라 생성량이 회복되는 변화를 나타낸 결과이다.
도 5는 α-MSH 자극에 의한 흑색종세포(B16F10) 멜라닌 생성에 유자박 유래 분획물 (EA, Hex, BuOH)처리에 따른 변화를 분석한 그래프와 그림이다.
도 6은 각 세포에 에탄올 100%와 50% 유자박유래추출물과 분획물 (EA, Hex, BuOH)을 (50 νg/ml) 24시간 처리한 후, Enzyme linked immunoassay (ELISA assay)를 통해 히알루론산 (hyaluronic acid) 생성량을 측정한 결과이다.
도 7은 aCaT 세포를 배양하여 UVB (30 mJ/cm2)를 조사 한 후, 각 세포에 에탄올 100%와 50% 유자박유래추출물과 분획물 (EA, Hex, BuOH)을 (50 νg/ml) 24시간 처리한 후, Enzyme linked immunoassay (ELISA assay)를 통해 통해 Collagen-1의 전구체인 Pro-collagen-1을 측정한 결과이다.
도 8은 흑색종세포(B16F10)에 α-MSH(200 nM)과 에탄올 100%와 50% 유자박유래추출물, 분획물 (EA, Hex, BuOH)을 (50 νg/ml) 24시간 처리한 후, α-MSH (200 nM)의 처리로 인해 증가한 멜라닌 함유량 (Melanin contents)을 측정한 결과이다. 1 is a view explaining steps for preparing an extract from citron seed meal and a fraction according to solvent characteristics according to an embodiment of the present invention.
2 is a diagram showing the results of cytotoxicity analysis of the citron peel-derived fraction in human keratinocytes (HaCaT) and the change in the amount of wrinkle factors and damaged collagen induced by UVB irradiation according to the treatment of the citron peel-derived fraction.
3 is a view showing the results of cytotoxicity analysis of the citron peel-derived fraction in human fibroblasts (HDF) and the change in the amount of wrinkle factors and damaged collagen induced by UVB irradiation according to treatment with the citron peel-derived fraction.
4 is a result showing the change in the production amount of hyaluronic acid damaged by UVB irradiation and the recovery of filaggrin and SPT enzymes required for the construction of the skin epidermis according to the treatment of the citron peel-derived fraction.
5 is a graph and a picture analyzing changes according to treatment with citron peel-derived fractions (EA, Hex, BuOH) in melanin production of melanoma cells (B16F10) by α-MSH stimulation.
6 is a result of measuring the amount of hyaluronic acid produced by Enzyme linked immunoassay (ELISA assay) after each cell was treated with 100% ethanol and 50% citron peel-derived extracts and fractions (EA, Hex, BuOH) (50 νg/ml) for 24 hours.
7 is a result of measuring Pro-collagen-1, a precursor of Collagen-1, through Enzyme linked immunoassay (ELISA assay) after culturing aCaT cells and irradiating UVB (30 mJ/cm 2 ), and then treating each cell with 100% ethanol and 50% citron peel-derived extracts and fractions (EA, Hex, BuOH) (50 νg/ml) for 24 hours.
8 shows melanoma cells (B16F10) treated with α-MSH (200 nM) and 100% and 50% ethanol-derived extracts and fractions (EA, Hex, BuOH) (50 νg/ml) for 24 hours, and then measured the increased melanin contents (Melanin contents) due to treatment with α-MSH (200 nM).
이하, 도면과 실시예를 참조하여 본 발명의 구체적인 실시형태를 설명하기로 한다. 그러나 이는 예시에 불과하며 본 발명은 이에 제한되지 않는다. Hereinafter, specific embodiments of the present invention will be described with reference to drawings and examples. However, this is only an example and the present invention is not limited thereto.
본 발명을 설명함에 있어서, 본 발명과 관련된 공지기술에 대한 구체적인 설명이 본 발명의 요지를 불필요하게 흐릴 수 있다고 판단되는 경우에는 그 상세한 설명을 생략하기로 한다. 그리고, 후술되는 용어들은 본 발명에서의 기능을 고려하여 정의된 용어들로서 이는 사용자, 운용자의 의도 또는 관례 등에 따라 달라질 수 있다. 그러므로 그 정의는 본 명세서 전반에 걸친 내용을 토대로 내려져야 할 것이다. 또한 본 발명의 기술적 사상은 청구범위에 의해 결정되며, 이하의 실시예는 본 발명의 기술적 사상을 본 발명이 속하는 기술분야에서 통상의 지식을 가진 자에게 효율적으로 설명하기 위한 일 수단일 뿐이다. In describing the present invention, if it is determined that a detailed description of the known technology related to the present invention may unnecessarily obscure the subject matter of the present invention, the detailed description will be omitted. In addition, terms to be described later are terms defined in consideration of functions in the present invention, which may vary according to the intention or custom of a user or operator. Therefore, the definition should be made based on the contents throughout this specification. In addition, the technical spirit of the present invention is determined by the claims, and the following examples are only one means for efficiently explaining the technical spirit of the present invention to those skilled in the art to which the present invention belongs.
실시예 Example
유자박유래 추출물 및 분획물 제조 Manufacture of citron peel-derived extracts and fractions
도 1은 본 발명의 실시예에 따른 유자 씨박으로부터 추출물과, 용매 특성에 따른 분획물을 제조하는 단계를 설명하는 도면이다. 1 is a view explaining steps for preparing an extract from citron seed meal and a fraction according to solvent characteristics according to an embodiment of the present invention.
도 1을 참조하면, 먼저 유자 씨에서 오일을 추출(분리)하고 남은 부산물인 유자 씨 박 (각 100 g)에 50% 및 100% 에탄올을 가하여 2시간 1회 초음파추출한 후 여과한 후 감압농축 및 동결건조를 진행하여 유자박 50% 에탄올 추출물(4.79 g) 및 100% 에탄올 추출물(4.51 g)을 얻었고 그 수율은 각각 4.51%, 4.79%이다.Referring to FIG. 1, first, oil was extracted (separated) from citron seeds, and 50% and 100% ethanol were added to the remaining by-products, citron seed gourd (each 100 g), ultrasonically extracted once for 2 hours, filtered, concentrated under reduced pressure and freeze-dried to obtain citron peel 50% ethanol extract (4.79 g) and 100% ethanol extract (4.51 g), and the yields were 4.51%, respectively. It is 4.79%.
이후 활성이 뛰어난 유자박 50% 에탄올 추출물(4.79 g)을 증류수에 현탁 시킨 후, 도 1에 제시된 바와 같이 용매의 극성에 따라 n-hexane 분획물 (1.55 g), EtOAc 분획물 (0.12 g), n-butanol 분획물 (0.19 g), 물 분획물 (2.88 g)을 얻었다. 이하 실험예를 통하여 본 발명을 보다 상세히 설명하며, 하기에서 EA, Hex, BuOH는 용매 종류에 따른 분획물을, 50%와 100%는 에탄올 농도별 추출물을 나타낸다.Thereafter, the citron peel 50% ethanol extract (4.79 g) having excellent activity was suspended in distilled water, and as shown in FIG. 1, n -hexane fraction (1.55 g), EtOAc fraction (0.12 g), n -butanol fraction (0.19 g), and water fraction (2.88 g) were obtained according to the polarity of the solvent. The present invention will be described in more detail through the following experimental examples. In the following, EA, Hex, and BuOH represent fractions according to solvent types, and 50% and 100% represent extracts according to ethanol concentrations.
실험예 1Experimental Example 1
도 2는 인간각질세포(HaCaT)에서 유자박 유래 분획물의 세포독성 분석 결과 및 UVB 조사로 유도되는 주름 인자와 손상된 콜라겐이 유자박 유래 분획물 처리에 따라 생성량이 변화하는 결과를 나타낸 도면이다. 도 2를 참조한 결과는 다음과 같다. 2 is a diagram showing the results of cytotoxicity analysis of the citron peel-derived fraction in human keratinocytes (HaCaT) and the change in the amount of wrinkle factors and damaged collagen induced by UVB irradiation according to the treatment of the citron peel-derived fraction. The results with reference to FIG. 2 are as follows.
패널 A: 용매별 유자박 유래 분획물 (EA, Hex, BuOH)을 다양한 농도 (0-5-10-50 νg/ml)로 인간각질세포(HaCaT)에 24 시간 처리하여 유자박 유래 분획물 (YJP-EA, Hex, BuOH)에 대한 세포의 생존율을 MTT 분석법으로 측정하였다. Panel A: Citron peel-derived fractions (EA, Hex, BuOH) for each solvent were treated with human keratinocytes (HaCaT) at various concentrations (0-5-10-50 νg/ml) for 24 hours, and cell viability for the citron peel-derived fractions (YJP-EA, Hex, BuOH) was measured by MTT assay.
패널 B와 C: HaCaT세포에 UVB (30mJ/cm2)를 조사 한 후, 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml)을 24 시간 동안 처리하여 웨스턴 블롯과 Reverse transcription polymerase chain reaction (RT-PCR)을 수행하였다. Panels B and C: HaCaT cells were irradiated with UVB (30mJ/cm 2 ), then treated with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours, followed by Western blotting and reverse transcription polymerase chain reaction (RT-PCR).
UVB로 유도된 주름 인자 (MMP-1/9/13)가 유자박 유래 분획물 처리 후 효과적으로 감소되었고, 반대로 피부 탄력과 관련된 Collagen-1은 UVB 조사로 손상 되었다가 유자박 유래 분획물 처리 후 다시 복구 되는 것을 확인하였다. It was confirmed that UVB-induced wrinkle factors (MMP-1/9/13) were effectively reduced after treatment with the citron peel-derived fraction, and on the contrary, Collagen-1, which is related to skin elasticity, was damaged by UVB irradiation and then restored after treatment with the citron peel-derived fraction.
패널 D: HaCaT세포에 UVB (30mJ/cm2)를 조사 한 후, 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml)을 24 시간 동안 처리하여 Enzyme linked immunoassay (ELISA assay)를 통해 Collagen-1의 전구체인 Pro-collagen-1이 증가하는 것을 측정하였다.Panel D: HaCaT cells were irradiated with UVB (30mJ/cm 2 ) and then treated with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours, and an increase in Pro-collagen-1, a precursor of collagen-1, was measured through an enzyme linked immunoassay (ELISA assay).
도 3은 인간섬유아세포(HDF)에서 유자박 유래 분획물의 세포독성 분석 결과 및 UVB 조사로 유도되는 주름 인자와 손상된 콜라겐이 유자박 유래 분획물 처리에 따라 생성량이 변화하는 결과를 나타낸 도면이다. 도 3을 참조한 결과는 다음과 같다. 3 is a view showing the results of cytotoxicity analysis of the citron peel-derived fraction in human fibroblasts (HDF) and the change in the amount of wrinkle factors and damaged collagen induced by UVB irradiation according to treatment with the citron peel-derived fraction. The results referring to FIG. 3 are as follows.
패널 A: 유자박 유래 분획물 (EA, Hex, BuOH)을 다양한 농도 (0-5-10-50 νg/ml)로 인간섬유아세포(HDF)에 24 시간 처리하여 유자박 유래 분획물 (YJP-EA, Hex, BuOH)에 대한 세포의 생존율을 MTT 분석법으로 측정하였다. Panel A: Human fibroblasts (HDF) were treated with the citron peel-derived fractions (EA, Hex, BuOH) at various concentrations (0-5-10-50 νg/ml) for 24 hours, and cell viability for the citron peel-derived fractions (YJP-EA, Hex, BuOH) was measured by MTT assay.
패널 B와 C: HDF세포에 UVB (100mJ/cm2)를 조사 한 후, 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml)을 24 시간 동안 처리하여 웨스턴 블롯과 Reverse transcription polymerase chain reaction (RT-PCR)을 수행하였다. UVB로 유도된 주름 인자 (MMP-1/9/13)가 유자박 유래 분획물 처리 후 효과적으로 감소되었고, 반대로 피부 탄력과 관련된 Collagen-1은 UVB 조사로 손상 되었다가 유자박 유래 분획물 처리 후 다시 복구 되는 것을 확인하였다. Panels B and C: HDF cells were irradiated with UVB (100mJ/cm 2 ), then treated with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours, followed by Western blotting and reverse transcription polymerase chain reaction (RT-PCR). It was confirmed that UVB-induced wrinkle factors (MMP-1/9/13) were effectively reduced after treatment with the citron peel-derived fraction, and on the contrary, Collagen-1, which is related to skin elasticity, was damaged by UVB irradiation and then restored after treatment with the citron peel-derived fraction.
패널 D: HDF세포에 UVB (100mJ/cm2)를 조사 한 후, 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml)을 24 시간 동안 처리하여 Enzyme linked immunoassay (ELISA assay)를 통해 Collagen-1의 전구체인 Pro-collagen-1이 증가하는 것을 측정하였다.Panel D: HDF cells were irradiated with UVB (100mJ/cm 2 ), then treated with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours, and an increase in Pro-collagen-1, a precursor of collagen-1, was measured through an enzyme linked immunoassay (ELISA assay).
도 4는 UVB 조사로 손상된 히알루론산 (hyaluronic acid) 생성량과 피부 표피의 구성에 필요한 필라그린 (filaggrin)과 SPT 효소가 유자박 유래 분획물 처리에 따라 생성량이 회복되는 변화를 나타낸 결과이다. 도 4를 참조한 결과는 다음과 같다. 4 is a result showing the change in the production amount of hyaluronic acid damaged by UVB irradiation and the recovery of filaggrin and SPT enzymes required for the construction of the skin epidermis according to the treatment of the citron peel-derived fraction. The results with reference to FIG. 4 are as follows.
패널 A: HaCaT 세포에 UVB (30mJ/cm2) 조사로 인해 감소한 히알루론산 (hyaluronic acid) 생성량이 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml) 24 시간 동안 처리함에 따라 생성량이 회복되는 것을 Enzyme linked immunoassay (ELISA assay)를 통해 확인하였다. Panel A: It was confirmed through Enzyme linked immunoassay (ELISA assay) that the amount of hyaluronic acid production, which was reduced by UVB (30mJ/cm 2 ) irradiation in HaCaT cells, was restored by treatment with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours.
패널 B: HaCaT세포에 UVB (30mJ/cm2)를 조사 후, 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml)을 24 시간 동안 처리하여 웨스턴 블롯을 수행하였다. UVB 조사로 인해 손상되었던 표피를 구성하는데 필요한 필라그린 (filaggrin)과 SPT가 유자박 유래 분획물 처리에 따라 다시 증가함을 확인하였다. Panel B: HaCaT cells were irradiated with UVB (30mJ/cm 2 ) and then treated with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours, and western blotting was performed. It was confirmed that filaggrin and SPT required to construct the epidermis damaged by UVB irradiation increased again according to the treatment of the citron peel-derived fraction.
패널 C: HDF세포에 UVB (100mJ/cm2) 조사로 인해 감소한 히알루론산 (hyaluronic acid) 생성량이 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml) 24 시간 동안 처리함에 따라 생성량이 회복되는 것을 Enzyme linked immunoassay (ELISA assay)를 통해 확인하였다. Panel C: It was confirmed through Enzyme linked immunoassay (ELISA assay) that the amount of hyaluronic acid production, which was reduced by UVB (100mJ/cm 2 ) irradiation in HDF cells, was restored by treatment with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours.
패널 D: HDF세포에 UVB (100mJ/cm2)를 조사 후, 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml)을 24 시간 동안 처리하여 웨스턴 블롯을 수행하였다. UVB 조사로 인해 손상되었던 표피를 구성하는데 필요한 필라그린 (filaggrin)과 SPT가 유자박 유래 분획물 처리에 따라 다시 증가함을 확인하였다.Panel D: HDF cells were irradiated with UVB (100 mJ/cm 2 ) and then treated with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours, and Western blotting was performed. It was confirmed that filaggrin and SPT required to construct the epidermis damaged by UVB irradiation increased again according to the treatment of the citron peel-derived fraction.
도 5는 α-MSH 자극에 의한 흑색종세포(B16F10) 멜라닌 생성에 유자박 유래 분획물 (EA, Hex, BuOH)처리에 따른 변화를 분석한 그래프와 그림이다. 도 5를 참조한 결과는 다음과 같다. 5 is a graph and a picture analyzing changes according to treatment with citron peel-derived fractions (EA, Hex, BuOH) in melanin production of melanoma cells (B16F10) by α-MSH stimulation. The results referring to FIG. 5 are as follows.
패널 A: α-MSH(200 nM)과 유자박 유래 분획물 (EA, Hex, BuOH)을 다양한 농도 (0-5-10-50 νg/ml)로 흑색종세포(B16F10)에 24 시간 처리하여 유자박 유래 분획물 (YJP-EA, Hex, BuOH)에 대한 세포의 생존율을 MTT 분석법으로 측정하였다. Panel A: α-MSH (200 nM) and citron peel-derived fractions (EA, Hex, BuOH) were treated with melanoma cells (B16F10) at various concentrations (0-5-10-50 νg/ml) for 24 hours.
패널 B: B16F10세포에 α-MSH(200 nM)로 인해 증가한 멜라닌 함유량 (Melanin contents)이 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml) 24 시간 동안 처리함에 따라 함유량이 감소되는 것을 흡광도 측정를 통해 확인하였다. 패널 C: B16F10세포에 α-MSH(200 nM)로 인해 증가한 Tyrosinase 활성이 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml) 24 시간 동안 처리함에 따라 감소되는 것을 흡광도 측정을 통해 확인하였다. Panel B: The melanin contents increased by α-MSH (200 nM) in B16F10 cells were reduced by treatment with the citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours. It was confirmed through absorbance measurement. Panel C: It was confirmed through absorbance measurement that the increased Tyrosinase activity of B16F10 cells by α-MSH (200 nM) was decreased by treatment with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours.
패널 D: B16F10세포에 α-MSH(200 nM)로 인해 증가한 Mushroom tyrosinase 활성이 유자박 유래 분획물 EA, Hex, BuOH (0-5-10-50 νg/ml) 24 시간 동안 처리함에 따라 감소되는 것을 흡광도 측정을 통해 확인하였다. 패널 E: B16F10세포에 α-MSH(200 nM)로 유도된 멜라닌 생성 촉진인자 (TRP-1, TRP-2, Tyrosinase, Mitf)가 유자박 유래 분획물 처리 후 효과적으로 감소되었음을 웨스턴 블롯으로 확인하였다.Panel D: It was confirmed through absorbance measurement that the Mushroom tyrosinase activity increased by α-MSH (200 nM) in B16F10 cells was decreased by treatment with citron peel-derived fractions EA, Hex, and BuOH (0-5-10-50 νg/ml) for 24 hours. Panel E: It was confirmed by Western blot that α-MSH (200 nM)-induced melanogenesis promoters (TRP-1, TRP-2, Tyrosinase, Mitf) in B16F10 cells were effectively reduced after treatment with the citron peel-derived fraction.
실험예 2Experimental Example 2
도 6은 각 세포에 에탄올 100%와 50% 유자박유래추출물과 분획물 (EA, Hex, BuOH)을 (50 νg/ml) 24시간 처리한 후, Enzyme linked immunoassay (ELISA assay)를 통해 히알루론산 (hyaluronic acid) 생성량을 측정한 결과이다. 6 is a result of measuring the amount of hyaluronic acid produced by Enzyme linked immunoassay (ELISA assay) after each cell was treated with 100% ethanol and 50% citron peel-derived extracts and fractions (EA, Hex, BuOH) (50 νg/ml) for 24 hours.
도 6을 참조하면, UVB 조사로 손상된 히알루론산 (hyaluronic acid) 생성량은 기존 에탄올 100%와 50% 유자박유래추출물 보다 분획물 (EA, Hex, BuOH)의 처리에서 최대 60% 증가된 회복량을 확인할 수 있었다.Referring to FIG. 6, it was confirmed that the amount of hyaluronic acid damaged by UVB irradiation was increased by up to 60% in the treatment of the fractions (EA, Hex, BuOH) compared to the existing 100% ethanol and 50% citron peel-derived extracts.
도 7은 aCaT 세포를 배양하여 UVB (30 mJ/cm2)를 조사 한 후, 각 세포에 에탄올 100%와 50% 유자박유래추출물과 분획물 (EA, Hex, BuOH)을 (50 νg/ml) 24시간 처리한 후, Enzyme linked immunoassay (ELISA assay)를 통해 통해 Collagen-1의 전구체인 Pro-collagen-1을 측정한 결과이다. 7 is a result of measuring Pro-collagen-1, a precursor of Collagen-1, through Enzyme linked immunoassay (ELISA assay) after culturing aCaT cells and irradiating UVB (30 mJ/cm 2 ), and then treating each cell with 100% ethanol and 50% citron peel-derived extracts and fractions (EA, Hex, BuOH) (50 νg/ml) for 24 hours.
도 7을 참조하면, 피부 탄력과 관련된 Collagen-1은 UVB 조사로 손상 되었다가 에탄올 100%와 50% 유자박유래추출물 보다 분획물 (EA, Hex, BuOH)의 처리에서 최대 43% 증가된 회복량을 확인할 수 있었다.Referring to FIG. 7, Collagen-1 related to skin elasticity was damaged by UVB irradiation, and it was confirmed that the amount of recovery increased by up to 43% in the treatment of fractions (EA, Hex, BuOH) compared to 100% ethanol and 50% citron peel-derived extract.
도 8은 흑색종세포(B16F10)에 α-MSH(200 nM)과 에탄올 100%와 50% 유자박유래추출물, 분획물 (EA, Hex, BuOH)을 (50 νg/ml) 24시간 처리한 후, α-MSH (200 nM)의 처리로 인해 증가한 멜라닌 함유량 (Melanin contents)을 측정한 결과이다. 8 shows melanoma cells (B16F10) treated with α-MSH (200 nM) and 100% and 50% ethanol extracts and fractions (EA, Hex, BuOH) (EA, Hex, BuOH) (50 νg/ml) for 24 hours, and then measured the increased melanin contents (Melanin contents) due to treatment with α-MSH (200 nM).
도 8을 참조하면, α-MSH (200 nM)의 처리로 인해 증가한 멜라닌 함유량 (Melanin contents)이 에탄올 100%와 50% 유자박유래추출물 보다 분획물 (EA, Hex, BuOH)의 처리에서 최대 35% 감소되는 결과를 확인할 수 있었다.Referring to FIG. 8, it was confirmed that the melanin contents (Melanin contents) increased by the treatment with α-MSH (200 nM) were reduced by up to 35% in the treatment of the fractions (EA, Hex, BuOH) compared to 100% ethanol and 50% citron peel-derived extracts.
이상의 결과는 유자박, 특히 유자 씨박의 추출물로부터 분획한 분획물이 추출물 대비 월등히 우수한 피부개선 효과를 가지는 것을 나타내며, 본 발명에 따른 분획물을 포함하는 조성물은 UVB에 의해 유도되는 피부 주름 억제 효과, 피부 손상에 대한 복구 효과, 보습 효과, 미백 효과가 우수하여 노화방지 및 피부 복구를 위한 화장품 조성물에 유용하게 사용될 수 있다.The above results show that the fraction obtained from the extract of citron peel, especially citron seed peel, has a far superior skin improvement effect compared to the extract, and the composition containing the fraction according to the present invention has an effect of suppressing skin wrinkles induced by UVB, a repair effect for skin damage, a moisturizing effect, and a whitening effect, so that it can be usefully used in a cosmetic composition for anti-aging and skin restoration.
Claims (10)
상기 유자박 유래 분획물은 유자씨로부터 유래된 분획물인 것을 특징으로 하는 피부개선용 조성물.According to claim 1,
The composition for improving skin, characterized in that the fraction derived from citron peel is a fraction derived from citron seeds.
상기 피부개선용 조성물은 자외선에 의해 유도되는 주름 개선, 보습, 또는 미백 효과를 갖는 것을 특징으로 하는 피부개선용 조성물.According to claim 1,
The composition for improving skin, characterized in that it has a wrinkle improvement, moisturizing, or whitening effect induced by ultraviolet rays.
상기 분획물은 상기 유자씨 박 추출물을 에틸아세테이트(Ethyl acetate, EA), 헥산(Hex), 부탄올(BuOH)을 용매로 사용하여 분획된 것을 특징으로 하는 피부개선용 조성물.According to claim 2,
The fraction is a composition for improving skin, characterized in that the fractionation of the citron seed gourd extract using ethyl acetate (EA), hexane (Hex), butanol (BuOH) as a solvent.
상기 조성물은 자외선에 의해 유도되는 주름 개선은 MMP-1/9/13 단백질, Collagen-1 단백질 또는 이를 코딩하는 mRNA의 발현을 억제 또는 증가시켜 주름 개선 효과를 갖는 것을 특징으로 하는 피부개선용 조성물.According to claim 1,
The composition is a composition for improving skin, characterized in that wrinkle improvement induced by ultraviolet rays has an effect of reducing wrinkles by inhibiting or increasing the expression of MMP-1/9/13 protein, collagen-1 protein or mRNA encoding them.
상기 추출물은 자외선에 의해 손상된 보습 관련 인자인 히알루론산, 필라그린 및 SPT 중 적어도 어느 하나의 발현을 증가시켜 보습효과를 갖는 것을 특징으로 하는 피부개선용 조성물.According to claim 1,
The extract is a composition for improving skin, characterized in that it has a moisturizing effect by increasing the expression of at least one of hyaluronic acid, filaggrin and SPT, which are moisturizing-related factors damaged by ultraviolet rays.
상기 조성물은 자극에 의해 생성된 멜라닌 생성 촉진인자 TRP-1, TRP-2, 티로시나제(Tyrosinase) 및 Mitf 중 적어도 어느 하나의 발현을 감소시킴으로써 미백효과를 갖는 것을 특징으로 하는 피부개선용 조성물.According to claim 1,
The composition has a whitening effect by reducing the expression of at least one of TRP-1, TRP-2, tyrosinase and Mitf, which are melanin production promoters produced by stimulation.
상기 약학 조성물은 자외선에 의해 유도되는 주름 개선 또는 미백용 조성물인 것을 특징으로 하는 약학 조성물. According to claim 8,
The pharmaceutical composition is a pharmaceutical composition, characterized in that the composition for improving wrinkles or whitening induced by ultraviolet rays.
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| KR1020220005575A KR20230109833A (en) | 2022-01-14 | 2022-01-14 | Composition for improving skin condition with anti-wrinkle, anti-oxidant and moisturizing of UVB-damaged skin from extract of Citrus junos seed shell as active ingredient |
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| KR1020220005575A KR20230109833A (en) | 2022-01-14 | 2022-01-14 | Composition for improving skin condition with anti-wrinkle, anti-oxidant and moisturizing of UVB-damaged skin from extract of Citrus junos seed shell as active ingredient |
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