KR20180017952A - Natural Preservative Composition Containing Plant Extract Complex as Active Ingredient - Google Patents
Natural Preservative Composition Containing Plant Extract Complex as Active Ingredient Download PDFInfo
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- KR20180017952A KR20180017952A KR1020160102613A KR20160102613A KR20180017952A KR 20180017952 A KR20180017952 A KR 20180017952A KR 1020160102613 A KR1020160102613 A KR 1020160102613A KR 20160102613 A KR20160102613 A KR 20160102613A KR 20180017952 A KR20180017952 A KR 20180017952A
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Abstract
Description
본 발명은 식물 복합 추출물을 유효성분으로 함유하는 천연 방부제 조성물에 관한 것으로, 더욱 구체적으로 정향 추출물, 단삼 추출물, 황백 추출물, 및 황련 추출물을 유효성분으로 함유하는 천연 방부제 조성물에 관한 것이다.The present invention relates to a natural preservative composition containing a plant complex extract as an active ingredient, and more particularly, to a natural preservative composition containing a clove extract, a rosemary extract, a yellowish extract, and a Rhodiola extract as an active ingredient.
생물체의 항상성(homeostasis)을 유지하기 위한 과정에서 중요한 역할을 담당하고 있는 물질들 중 일부가 각종 생물체 유래의 생리 활성 물질이다. 지금까지 수많은 생리 활성 물질에 대해 많은 연구가 진행되고 있으며, 그 중, 미생물, 식물, 또는 동물체 등에서 분리된 항균물질에 대한 연구는 생명과학 및 의학 분야에서 매우 중요한 영역이다.Some of the substances that play an important role in maintaining the homeostasis of organisms are physiologically active substances derived from various organisms. Numerous researches on physiologically active substances have been carried out so far. Among them, studies on antimicrobial substances isolated from microorganisms, plants, or animals are very important fields in life sciences and medicine.
모든 생명체는 생존을 위하여 항균 및 항진균 물질을 생산하는 것으로 알려져 있는데, 항균 및 항진균 물질을 생산하는 미생물에 대해서 많은 연구가 이루어지고 있으며, 항균 및 항진균 물질을 생산하는 미생물 또는 이들에게서 분리된 물질을 이용해 항균제 또는 항진균제를 개발하려는 연구들이 오래전부터 시도되고 있다. All living organisms are known to produce antimicrobial and antifungal substances for survival. Many studies have been conducted on microorganisms that produce antimicrobial and antifungal substances. Using microorganisms that produce antimicrobial and antifungal substances, or materials isolated from them Studies to develop antibacterial or antifungal agents have long been attempted.
항균물질은 미생물을 죽이면서 인체 또는 동물에게는 독성이 낮고 체내의 효소 등에 의해 비활성화되지 않는 선택적 독성작용(selective toxicity)을 갖는 물질로서, 이는 주로 DNA의 복제, 유전정보의 전사 및 해독, 전자에너지의 수송, 세포벽의 생합성 등을 저해함으로써 미생물의 증식을 억제하는 기전을 통해 효과를 나타낸다.The antimicrobial substance is a substance having a selective toxicity which is low in toxicity to a human body or animal and is not inactivated by an enzyme in the body while killing a microorganism. This is mainly a DNA replication, a transcription and decryption of genetic information, Transport, and biosynthesis of the cell wall, thereby exhibiting an effect through a mechanism of inhibiting the proliferation of microorganisms.
현재까지 개발된 주된 항균물질들은 미생물, 식물 등 천연에서 유래한 것과 화학적으로 합성된 것으로 나뉠 수 있는데, 천연 유래 또는 화학적으로 합성된 많은 항균물질이 유해균에 의해 유발된 질병의 치료에 사용되고 있으나, 최근에는 화학적으로 합성된 항균물질에 대한 내성 증가로 인해 천연에서 유래한 항균물질을 이용한 항균제 개발에 관심이 증가하고 있는 추세이고, 최근 삶의 질이 향상됨에 따라, 여성용 질염의 치료 또는 예방에 효과가 있는 천연유래 물질을 탐색하고자 하는 연구가 활발하게 진행되고 있다. The main antimicrobial substances developed so far can be divided into microorganisms, plants and the like, which are chemically synthesized with natural ones. Many natural or chemically synthesized antimicrobial substances are used for the treatment of diseases caused by harmful bacteria, There is an increasing interest in the development of antimicrobial agents using natural antimicrobial substances due to the increase in the resistance to chemically synthesized antimicrobial substances. Recently, as the quality of life is improved, it is effective in the treatment or prevention of female vaginitis Researches are being actively pursued to search for naturally occurring substances.
한편, 방부제는 화장품, 의약품, 생활용품, 식품 등 다양한 제품군의 품질을 장기간 유지하기 위해 필수적으로 사용되는 물질로, 특히 유통기간이 비교적 길고 기름이나 물이 주성분으로 구성되고, 미생물의 생장에 필요한 탄소원, 질소원 및 단백질원 등으로 조성된 화장품의 경우에는 필수적으로 사용되는 물질이다. 화장품은 일반적으로 상온 1년~ 3년간의 유통기간을 가지고 있으며, 개봉 후 지속적으로 손과의 접촉으로 인하여 미생물에 매우 취약하다. 따라서 이러한 미생물로부터의 오염을 차단하며 제품의 안전과 품질을 보장하기 위하여 방부력이 높은 방부제를 사용해야 한다. 그러나, 기존 화장품에 사용되는 대부분의 방부제는 화학적 방법에 의해서 합성된 화합물이 대부분이고, 이러한 화합물을 인체 피부에 장기적으로 사용하는 경우, 피부 노화 촉진, 피부 자극, 발암 및 내분비계의 교란시킬 수 있는 문제가 있고, 생분해성 결여로 피부에 잔존하는 등의 문제가 있다. 따라서, 최근에는 인체에 안전하며, 환경 친화적인 천연물 유래의 방부제 소재에 대한 기술개발의 필요성이 증대되고 있는 실정이다. On the other hand, preservatives are essential substances for long-term maintenance of various product groups such as cosmetics, medicines, household products, foods, etc. Especially, they have a relatively long distribution period and are composed mainly of oil or water, , A nitrogen source, a protein source, and the like. Cosmetics generally have a shelf life of 1 year ~ 3 years at room temperature and are very vulnerable to microorganisms due to continuous contact with hands after opening. Therefore, in order to prevent contamination from these microorganisms and to ensure the safety and quality of the product, a preservative with high flotation power should be used. However, most of the preservatives used in conventional cosmetics are compounds synthesized by chemical methods, and when these compounds are used for human skin for a long time, the skin aging promotion, skin irritation, carcinogenesis and endocrine disruption There is a problem, and there is a problem such as remaining on the skin due to lack of biodegradability. Therefore, in recent years, there is a growing need for the development of a preservative material derived from natural materials that is safe for humans and environmentally friendly.
또한, 대표적인 방부제인 파라벤(paraben, p-hydroxybenzoic acid ester), 포름알데하이드 도너(formaldehyde-donor) 및 이소티아졸리논(isothiazolinone) 등은 피부 알러지, 환경 호르몬으로서의 작용 가능성(Andera Countti et al., Contact Dermatitis, 37:35, 1997; Edwin et al.,Toxicology and Applied Pharmacology, 153:12, 1998)과 지속적인 체내 축척으로 인한 급, 만성 독성 및 돌연변이 유발 등의 유해성이 널리 인식되고 있기 때문에 일부 생산기업들은 소비자의 요구에 대응하여 대체 방부제 개발에 노력을 하고 있다.In addition, representative preservatives such as paraben, p-hydroxybenzoic acid ester, formaldehyde-donor and isothiazolinone are known to act as skin allergies and environmental hormones (Andera Countti et al., Contact Because toxicity, such as rapid, chronic toxicity and mutagenesis, due to persistent systemic accumulation is widely recognized, some producers (eg, Dermatitis, 37:35, 1997; Edwin et al., Toxicology and Applied Pharmacology, In response to consumer needs, we are making efforts to develop alternative preservatives.
이러한 문제점들로 인하여 제품의 안전성과 경제성이 우수한 천연방부제에 대한 연구가 지속되고 있으며, 선행특허로는 향신료, 정유, 한약재 등 동·식물기원의 추출물(한국등록특허 제10-09862490호, 제10-07876340호, 제10-07825990호, 제10-09814070호, 제10-07298310호, 제10-0822416호, 제10-09428300호, 한국공개특허 제2009-0025654호, 제2007-0080707호 및 제 2009-0075282호)과 박테리오신 같은 미생물 기원의 천연 항균성 물질이 연구개발 된 바 있다.Because of these problems, studies on natural preservatives that are superior in safety and economical efficiency of products have been continuing. Prior art patents include extracts of Dong-yi and plant origin such as spices, essential oils and herb medicines (Korean Patent No. 10-09862490, No. 10 -07876340, 10-07825990, 10-09814070, 10-07298310, 10-0822416, 10-09428300, Korean Patent Publication Nos. 2009-0025654, 2007-0080707, 2009-0075282) and bacteriocins have been researched and developed.
그러나, 상기 선행특허들은 단순히 향신료, 정유, 한약재 등 동·식물기원의 추출물의 항균 효과만 검증하고 있을 뿐, 추출물이 화장료 조성물의 방부제로 사용되기에 적합한지에 대한 검증이 이루어지지 않아 그 활용도가 매우 미미하였다. 또한, 화장품 방부제로서의 고유 기능 이외에 피부주름 개선, 미백 활성 등 방부제 자체가 보유한 기능성에 대한 연구는 이루어진바 없다. However, the above-mentioned patents merely verify the antimicrobial effect of extracts of plant and plant origin such as spices, essential oils and herbal medicines, and it is not verified whether the extract is suitable for use as a preservative in cosmetic compositions. Lt; / RTI > In addition, there have been no studies on the functional properties of preservatives themselves, such as skin wrinkles and whitening activity, in addition to their unique functions as cosmetic preservatives.
이에 본 발명자들은 넓은 항균 스펙트럼 및 우수한 항균 효과를 갖는 소재로, 자체적으로 피부주름 개선, 미백 활성 등의 기능성을 보유한 천연 추출물의 조합을 찾기 위해 예의 연구노력한 결과, 정향, 단삼, 황백, 및 황련의 복합 추출물을 방부제 조성물로 활용하는 경우, 화장품의 부패를 일으키는 다양한 세균 및 진균을 효과적으로 억제함과 동시에, 피부 주름개선 및 미백의 기능성을 보유하여, 화장료 조성물의 방부제로 유용하게 사용할 수 있음을 확인하고, 본 발명을 완성하게 되었다.Accordingly, the present inventors have made extensive efforts to find a combination of natural extracts having a broad spectrum of antibacterial activity and excellent antimicrobial activity and having functions such as skin wrinkle improvement, whitening activity, etc. As a result, When the combined extract is used as a preservative composition, it can be effectively used as a preservative for cosmetic compositions, effectively inhibiting various bacteria and fungi causing the decay of cosmetics, and having skin wrinkle improving and whitening function , Thereby completing the present invention.
따라서, 본 발명의 주된 목적은 천연 방부 및 항균 효과를 보유함과 동시에 피부 주름개선, 미백, 및 피부염증 개선의 기능성을 보유한 식물 유래의 천연 방부제 조성물을 제공하는 데 있다.Accordingly, it is a main object of the present invention to provide a natural antiseptic composition derived from plants having natural antiseptic and antimicrobial effects, and having the function of improving skin wrinkles, whitening, and skin inflammation.
본 발명의 다른 목적 및 이점은 하기의 발명의 상세한 설명, 청구범위 및 도면에 의해 보다 명확하게 된다.Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.
본 발명의 한 양태에 따르면, 본 발명은 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물을 유효성분으로 함유하는 천연 복합 방부제 조성물을 제공한다.According to one aspect of the present invention, there is provided a natural combination preservative composition comprising clove extract, ginsenosides extract, Huang Baek extract and Rhodiola extract as an active ingredient.
본 발명자들은 화장료에 기능성을 부여할 수 있는 천연 방부 소재를 찾고자 수백여 종의 식물 추출물들의 항균활성을 1차적으로 검증하였고, 우수한 항균활성을 갖는 복합 식물추출물의 피부 기능성을 2차적으로 검증한 바 있다. 그 결과, 정향, 단삼, 황백, 및 황련의 복합 식물 추출물이 넓은 항균 스펙트럼과 우수한 항균 활성을 갖는 동시에 피부 주름개선, 미백, 피부 염증 개선 등의 효과를 갖는 것을 발견하였고, 또한 피부에 적용시 안전성에도 문제가 없음을 확인하였다.The inventors of the present invention firstly examined the antibacterial activity of hundreds of plant extracts in order to find a natural preservative material capable of imparting functionality to cosmetics, and secondly verified the skin function of a complex plant extract having excellent antibacterial activity have. As a result, it has been found that the combined plant extracts of clodshell, danshang, hwangbyeong, and chrysanthemum have a broad antibacterial spectrum and excellent antimicrobial activity, and also have effects such as improvement of skin wrinkles, whitening, improvement of skin inflammation, It is confirmed that there is no problem.
본 발명에서 사용되는 용어 "방부"는 미생물의 작용에 의해 식품, 화장품, 의약품 등의 물질이 부패하는 것을 막는 것을 의미하며, 항생, 항균을 총칭하는 의미로 사용된다. 또한, 용어 "방부제"는 식품, 화장품, 의약품 등의 물질을 보존하기 위한 목적으로 방부하기 위해서 첨가되는 약제를 의미한다. 따라서 인체에 해가 없어야 하며, 또 그 첨가로 인해 식품, 화장품, 의약품 등의 물질의 품질을 손상시키지 않아야 한다. The term " antiseptic " used in the present invention means preventing substances such as foods, cosmetics, and pharmaceuticals from being decomposed by the action of microorganisms, and is used generically to refer to antibiotics and antibacterials. In addition, the term "preservative" means a medicament added to preserve a substance such as food, cosmetics, medicines or the like in order to preserve it. Therefore, there should be no harm to the human body, and the addition thereof should not impair the quality of materials such as food, cosmetics, and medicines.
본 발명의 "정향"은 학명 유제니아 카리오필라타(Eugenia caryophyllatum) 또는 학명 시지기움 아로마티카(Syzygium aromaticum)로 분류된 식물을 의미한다. 본 발명의 실시예에서는 건조된 정향 100g 분쇄한 후 물과 에탄올의 혼합 수용액 으로 12시간 가온 환류 추출하여 냉침시키고, 1.2㎛ 여과지를 이용, 여과하여 여과액 얻고, 감압농축 하여 12g을 수득하였다. The term " clove "of the present invention means a plant classified as scientific name Eugenia caryophyllatum or Syzygium aromaticum. In the example of the present invention, 100 g of the dried clove was pulverized, and the mixture was heated and refluxed for 12 hours by heating in a mixed aqueous solution of water and ethanol, followed by cooling and filtration using 1.2 탆 filter paper to obtain a filtrate, which was then concentrated under reduced pressure to obtain 12 g.
본 발명의 "단삼"은 꿀풀과에 속하는 다년생 초본식물로 학명 살비아 밀티오리자(Salvia miltiorrhiza BUNGE)로 분류된 식물의 뿌리를 건조한 약재를 의미한다. 본 발명의 실시예에서는 건조된 단삼 100g 분쇄한 후 물과 에탄올의 혼합 수용액으로 12시간 가온 환류 추출하여 냉침시키고, 1.2㎛ 여과지를 이용, 여과하여 여과액 얻고, 감압농축 하여 8.2g을 수득하였다. "Danshang" of the present invention means a herbaceous perennial herbaceous plant belonging to the family Lamiaceae, which has dried root of a plant classified as Salvia miltiorrhiza BUNGE. In the example of the present invention, 100 g of dried ginseng was pulverized, and the mixture was heated and refluxed for 12 hours in a mixed aqueous solution of water and ethanol. The mixture was cooled down and filtered through 1.2 쨉 m filter paper to obtain a filtrate. The filtrate was concentrated under reduced pressure to obtain 8.2 g.
본 발명의 "황백(phellodendron bark)"은 황백나무(Phellodendron amurense Rupr.) 수피를 건조한 건조품을 의미한다. 본 발명의 실시예에서는 건조된 황백 100g 분쇄한 후 물과 에탄올의 혼합 수용액으로 12시간 가온 환류 추출하여 냉침시키고, 1.2㎛ 여과지를 이용, 여과하여 여과액 얻고, 감압농축 하여 10.5g을 수득하였다. The "phellodendron bark" of the present invention means a dried product of bark of Phellodendron amurense Rupr. In the example of the present invention, 100 g of dried yellowish white was pulverized, and the mixture was heated and refluxed for 12 hours with heating in a mixed aqueous solution of water and ethanol. The mixture was cooled and filtered through 1.2 탆 filter paper to obtain a filtrate, which was concentrated under reduced pressure to obtain 10.5 g.
본 발명의 "황련"은 쌍떡잎식물 미나리아재비목 미나리아재비과의 상록 여러해살이풀로 학명 콥티스 치네시스(coptis Chinensis)로 분류된 식물을 의미한다. 본 발명의 실시예에서는 건조된 황련 100g 분쇄한 후 물과 에탄올의 혼합 수용액으로 12시간 가온 환류 추출하여 냉침시키고, 1.2㎛ 여과지를 이용, 여과하여 여과액 얻고, 감압농축 하여 11g을 수득하였다. The term " Rhodiola "of the present invention refers to a plant classified as coptis Chinensis, an evergreen perennial herb that belongs to the family Ranunculaceae. In the example of the present invention, 100 g of dried persimmon was crushed, and the mixture was heated and refluxed for 12 hours in a mixed aqueous solution of water and ethanol. The mixture was cooled down and filtered using 1.2 쨉 m filter paper to obtain a filtrate, which was concentrated under reduced pressure to obtain 11 g.
본 발명에서 사용되는 용어, "추출물"은 식물의 추출 처리에 의하여 얻어지는 추출액, 상기 추출액의 희석액이나 농축액, 상기 추출액을 건조하여 얻어지는 건조물, 상기 추출액의 조정제물이나 정제물, 또는 이들의 혼합물 등, 추출액 자체 및 추출액을 이용하여 형성 가능한 모든 제형의 추출물을 포함한다. 본 발명의 상기 추출물은 바람직하게 추출 후 건조 분말 형태로 제조되어 사용될 수 있다. The term "extract" used in the present invention refers to an extract obtained by extracting a plant, a diluted solution or concentrate of the extract, a dried product obtained by drying the extract, a controlled preparation or a purified product of the extracted solution, Extracts themselves and extracts of all formulations which can be formed using extracts. The extract of the present invention can be preferably prepared in the form of a dry powder after extraction.
본 발명의 상기 정향, 단삼, 황백, 및 황련 추출물은, 천연, 잡종 또는 변종 식물로부터 추출될 수 있고, 식물 조직 배양물에서도 추출이 가능하며, 잎, 줄기, 뿌리, 꽃, 열매 등의 부위에서 추출한 추출물을 포함한다. The clove extracts of the present invention can be extracted from natural, hybrid, or variant plants and can be extracted from plant tissue cultures. The extracts of leaves, stems, roots, flowers, Includes extracted extracts.
본 발명의 천연 복합 방부제 조성물에서, 상기 정향 추출물은 0.01 내지 10 중량부, 상기 단삼 추출물은 0.01 내지 10 중량부, 상기 황백 추출물은 0.01 내지 5 중량부, 및 상기 황련 추출물은 0.01 내지 5 중량부로 전체 조성물 중량대비 포함될 수 있다. In the natural combination preservative composition of the present invention, the clove extract is 0.01 to 10 parts by weight, the raw ginseng extract is 0.01 to 10 parts by weight, the yellowish white extract is 0.01 to 5 parts by weight, May be included relative to the weight of the composition.
본 발명의 천연 복합 방부제 조성물은 넓은 항균 스펙트럼을 갖는데, 구체적으로 그람양성균, 그람음성균, 효모, 및 진균에 항균 활성을 갖는다.The natural complex antiseptic composition of the present invention has a broad antibacterial spectrum, and specifically has antimicrobial activity against Gram-positive bacteria, Gram-negative bacteria, yeast, and fungi.
본 발명의 조성물은 구체적으로 바실러스(Bacillus) 속 균주, 및 스타필로코커스(Staphylococcus) 속의 그람양성 균주에 대해 항균 활성을 갖고, 보다 구체적으로 스타필로코커스 아우레우스(Staphylococcus aureus), 바실러스 서브틸리스(Bacillus subtilis) 등의 세균에 대하여 항균 활성이 있으나, 이에 한정되는 것은 아니다.The compositions of the present invention specifically, Bacillus (Bacillus) spp, and Staphylococcus (Staphylococcus) in the Gram have antibacterial activity against a positive strains, and more particularly Staphylococcus aureus (Staphylococcus aureus), Bacillus subtilis ( Bacillus subtilis ), but the present invention is not limited thereto.
또한, 본 발명의 조성물은 구체적으로 에스체리치아(Escherichia) 속 균주, 슈도모나스(Pseudomonas) 속의 그람음성 균주에 대해 항균활성을 갖고, 보다 구체적으로 에스체리치아 콜라이(Escherichia coli), 슈도모나스 에루지노사(Pseudomonas aeruginosa) 균주에 대해 항균 활성이 있으나, 이에 한정되는 것은 아니다.In addition, the compositions of the present invention specifically has the S cherry teeth (Escherichia) spp, Pseudomonas (Pseudomonas) in the gram-negative antibacterial activity against strains, more particularly S. Cherry Escherichia coli (Escherichia coli , and Pseudomonas aeruginosa , but the present invention is not limited thereto.
또한, 본 발명의 조성물은 구체적으로는 칸디다 속 진균 및 아스퍼질러스 속 진균에 대해 항진균 활성을 갖고, 보다 구체적으로는 칸디다 알비칸스(Candida albicans), 아스퍼질러스 니가(Aspergillus niger)에 대해 항진균 활성이 있으나, 이에 한정되는 것은 아니다. In addition, the compositions of the present invention specifically has a Candida genus fungi and Aspergillus antifungal activity against fungi in, more specifically from Candida albicans (Candida albicans , and Aspergillus niger , but are not limited thereto.
본 발명의 실시예에서는 정향, 단삼, 황백, 및 황련의 복합추출물의 항균 효과를 검정한 결과, 이들 복합 추출물이 단일 추출물에 비해 매우 우수한 항균 활성을 갖는 것을 확인하였다.In the examples of the present invention, it was confirmed that the combined extracts of clodshell, danshang, hwangbyeong, and chrysanthemum were highly antimicrobial against the single extracts.
본 발명의 천연 복합 방부제 조성물에서, 상기 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물은 4:3:2:3의 중량비로 혼합되는 것이 가장 바람직하다. In the natural combination preservative composition of the present invention, it is most preferable that the clove extract, the ginseng extract, the yellowish white extract and the chrysanthemum extract are mixed at a weight ratio of 4: 3: 2: 3.
본 발명의 실시예에서 상기 추출물들을 다양한 비율로 혼합하여 천연 복합 방부제 조성물(시험예 1 내지11)을 제조하였으며, 제조된 천연 복합 방부제 조성물의 방부력을 확인하기 위해 원형여과지법(paper disc method) 및 미생물 생장 최소억제 농도 평가를 수행하였다. 그 결과, 시험예 1 내지 11에서 전반적으로 우수한 항균활성을 나타내는 것을 확인하였으며, 특히 대조군으로 사용한 메틸파라벤과 자몽종자 추출물과 비교하였을 때, 시험예 8의 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물이 4:3:2:3의 비율로 혼합된 천연 복합방부제 조성물이 가장 우수한 조합임을 확인할 수 있었다.In the examples of the present invention, the natural preservatives compositions (Test Examples 1 to 11) were prepared by mixing the extracts in various ratios, and a paper disc method was used to confirm the repelling force of the prepared natural combination preservative compositions. And minimum inhibitory concentration of microorganisms. As a result, it was confirmed that the antibacterial activity was excellent overall in Test Examples 1 to 11. In particular, when compared with the methylparaben and grapefruit seed extracts used as the control group, the clove extracts, roots extracts, It was confirmed that the natural combination preservative composition mixed at a ratio of 4: 3: 2: 3 is the best combination.
본 발명의 천연 복합 방부제 조성물에서, 상기 조성물은 멜라닌 색소의 생성을 억제하여 피부미백 활성을 갖는 것을 특징으로 한다. 본 발명에 따르면, 멜란 A 세포에 본 발명의 정향, 단삼, 황백 및 황련의 복합 추출물을 처리하고 배양시킨 후 멜라닌 생성량을 비교한 결과, 나이아신-펩타이드를 처리한 경우 대조군에 비해 멜라닌 생성이 최대 70% 수준으로 억제되는 것을 확인할 수 있었다(도 1).In the natural combination preservative composition of the present invention, the composition is characterized by having a skin whitening activity by inhibiting the production of melanin pigment. According to the present invention, melanin A cells were treated with a combination extract of cloves, dansam, yellow whites, and yellow radishes of the present invention and melanin production was measured. As a result, melanin production was 70 % (Fig. 1).
한편, 피부섬유아세포의 콜라겐은 피부 재생, 피부 탄력, 피부 주름 형성 및 피부 손상 시 조직의 수복 또는 재생과 직접적인 관련이 있다고 볼 수 있다.On the other hand, collagen of skin fibroblast is directly related to restoration or regeneration of tissue in skin regeneration, skin elasticity, wrinkle formation and skin damage.
즉, 피부 섬유아세포의 콜라겐 또는 프로콜라겐의 합성이 촉진되거나, 콜라겐을 분해하는 기질 단백질 분해효소(matrix metallo-proteinase, 이하 'MMP'라 함)의 합성이 저해되면, 피부 탄력 개선, 피부 재생, 피부 주름 개선, 상처 치유, 손상된 피부 조직의 수복 및 재생; 및 피부 노화 방지 등의 효과를 얻을 수 있다.That is, when the synthesis of collagen or procollagen of skin fibroblast is promoted or the synthesis of matrix metallo-proteinase (hereinafter, referred to as 'MMP'), which degrades collagen, is inhibited, skin elasticity improvement, skin regeneration, Improvement of skin wrinkles, wound healing, restoration and regeneration of damaged skin tissue; And prevention of skin aging can be obtained.
본 발명의 천연 복합 방부제 조성물은 MMP-1 단백질의 발현을 억제하는 것을 특징으로 한다.The natural combination preservative composition of the present invention is characterized by inhibiting the expression of MMP-1 protein.
구체적으로, 본 발명의 천연 복합 방부제 조성물은 자외선 조사된 정상 인간 진피 섬유아세포(NHDF)에서 세포 독성을 저감시키고, 콜라겐 분해를 촉진시키는 기질 단백질 분해효소-1(matrix metallo-proteinase)의 발현을 억제시키는바, 피부 탄력 증가, 피부 재생, 피부 주름 개선, 상처 치유, 손상된 피부 조직의 수복 및 재생; 및 피부 노화 방지 등의 효과 등을 가질 수 있다. 구체적으로는 피부주름의 예방 또는 개선의 효과를 가질 수 있다(도 2 참조).Specifically, the natural combination preservative composition of the present invention reduces cytotoxicity in normal human dermal fibroblast (NHDF) irradiated with ultraviolet rays and inhibits the expression of matrix metalloproteinase-1, which promotes collagen degradation Increasing skin elasticity, regenerating skin, improving skin wrinkles, healing wounds, restoring and regenerating damaged skin tissue; And prevention of skin aging and the like. Specifically, it can have an effect of preventing or improving wrinkles of the skin (see Fig. 2).
또한, 본 발명의 천연 복합 방부제 조성물은 IL-6 단백질의 발현을 억제하는 것을 특징으로 한다(도 3 참조). 손상된 피부는 염증반응을 통해 더욱 확대되며, 피부 발적, 피부 과민반응, 또는 광과민 반응 등을 통해 임상적으로 피부가 더욱 악화되는 경향을 보인다. 따라서, 손상된 피부 상태의 효과적인 개선을 위해서 염증반응의 조기 차단은 매우 중요하다.In addition, the natural combination preservative composition of the present invention is characterized by inhibiting the expression of IL-6 protein (see FIG. 3). Damaged skin is further enlarged through the inflammatory reaction, and the skin tends to get worse clinically through skin eruption, skin irritation, or photosensitization. Therefore, early interception of the inflammatory response is very important for effective improvement of the damaged skin condition.
본 발명의 바람직한 실시예에서는, 상기 천연 복합 방부제 조성물을 처리시 염증 매개 물질인 IL-6 단백질의 생성이 억제되는 것으로 나타났으며, 이러한 IL-6 단백질 발현에 대한 억제능은 농도 의존적인 것으로 나타났다. 따라서, 본 발명의 조성물은 염증매개 물질인 IL-6 단백질의 생성을 효과적으로 억제하고, 피부 주변의 염증반응을 효과적으로 차단하여 손상된 피부의 상태개선에 효과적으로 작용한다.In a preferred embodiment of the present invention, the production of IL-6 protein, which is an inflammation mediator, is inhibited during the treatment of the natural combination preservative composition, and the inhibitory effect on IL-6 protein expression is concentration-dependent. Accordingly, the composition of the present invention effectively inhibits the production of IL-6 protein, which is an inflammation mediator, effectively blocks the inflammation reaction around the skin and effectively works to improve the condition of damaged skin.
본 발명의 다른 양태에 따르면, 본 발명은 상기 천연 복합 방부제 조성물을 유효성분으로 함유하는 화장료 조성물을 제공한다. According to another aspect of the present invention, there is provided a cosmetic composition comprising the natural combination preservative composition as an active ingredient.
본 발명의 상기 천연 복합 방부제 조성물은 천연유래 식물 소재에서 추출되어 인체에 해가 없으며, 화장품이 유해 미생물에 의해 부패하는 것을 효과적으로 막아주고, 그 첨가로 인해 화장품의 물질의 품질을 손상시키지 않으므로 화장료 조성물에 포함되어 사용되는데 적합하고, 특히 피부미백, 피부주름개선 및 피부염증 완화와 관련된 화장료 조성물에 포함되어 사용되는 것이 바람직하다.Since the natural combination preservative composition of the present invention is extracted from natural plant material and is free from harm to the human body, it effectively prevents the decay of the cosmetic by harmful microorganisms and does not impair the quality of the cosmetic material, And it is particularly preferable to be used in a cosmetic composition related to skin whitening, skin wrinkle improvement and skin inflammation relief.
본 발명의 화장료 조성물에서, 상기 천연 복합 방부제 조성물의 함량은 0.001 내지 10 중량%로 함유될 수 있다. In the cosmetic composition of the present invention, the content of the natural complex preservative composition may be 0.001 to 10% by weight.
본 발명의 실시예에서 상기 천연 복합 방부제 조성물을 이용하여 크림 제형을 제조하였으며, 천연 복합 방부제를 포함한 제형의 방부력을 확인한 결과, 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물을 혼합한 천연 복합 방부제 조성물을 이용한 화장료는 우수한 방부 효능을 갖는 것을 확인할 수 있었다.In the examples of the present invention, a cream formulation was prepared by using the natural combination preservative composition. As a result of confirming the repellency of the formulation containing the natural combination preservative, the natural combination preservative mixed with clove extract, ginseng extract, It was confirmed that cosmetic compositions using the composition had excellent preservative efficacy.
또한, 본 발명의 천연 복합 방부제를 포함한 제형의 피부자극 정도를 확인하기 위해 상기 조합의 천연 복합 방부제를 포함한 제형을 이용하여 건강한 성인 남녀 20명을 대상으로 피부자극 실험을 진행한 결과, 대부분 자극이 없거나 경미한 정도였으며, 화학성분의 방부제가 포함된 크림보다 평균 자극지수가 낮은 것을 확인할 수 있었다.In order to confirm the degree of skin irritation of the formulation containing the natural combination preservative of the present invention, skin irritation test was conducted on 20 healthy adult men and women using a formulation containing the natural combination preservative of the combination. As a result, The average irritation index was lower than that of the cream containing chemical preservatives.
또한, 다양한 비율로 혼합된 천연 복합 방부제 조성물을 이용하여 제조된 크림 제형의 안정성을 확인한 결과, 로션이 분리되는 과정인 크리밍, 응집과 같은 현상은 관찰되지 않았으며, 로션 성분의 산화에 의한 특이취도 나타나지 않는 것을 확인하였다.Further, as a result of confirming the stability of the cream formulation prepared using the natural combination preservative composition mixed at various ratios, no phenomenon such as creaming and flocculation, which is a separation process of lotion, was observed, It was confirmed that there were no symptoms.
본 발명의 천연 복합 방부제 조성물을 첨가할 수 있는 화장품의 제형으로는, 예를 들어, 스킨로션, 스킨 소프너, 스킨토너, 아스트린젠트, 로션, 밀크로션, 모이스처 로션, 영양로션, 맛사지크림, 영양크림, 모이스처크림, 핸드크림, 에센스, 영양에센스, 팩, 비누, 샴푸, 클렌징폼, 클렌징로션, 클렌징크림, 바디로션, 바디클렌저, 유액, 프레스파우더, 루스파우더, 아이섀도 등의 제형으로 제조될 수 있고, 통상적인 화장료 제조법에 적용시킬 수 있다. 구제척으로 액상, 크림상, 페이스트상 및 고체상등 다양한 성상으로 적용이 가능하며, 이들 각 제형에 적합하고 당업계에 주지된 각종의 통상적인 보조제와 담체를 포함할 수 있다.Examples of the formulations of cosmetics to which the natural combination preservative composition of the present invention can be added include skin lotions, skin softeners, skin toners, astringents, lotions, milk lotions, moisturizing lotions, nutrition lotions, massage creams, It can be made into formulations such as moisturizing cream, hand cream, essence, nutritional essence, pack, soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, latex, press powder, loose powder, eye shadow , And can be applied to a conventional cosmetic preparation method. It may be applied in various forms such as liquid, cream, paste, and solid forms, and may include various conventional auxiliary agents and carriers suitable for each formulation and known in the art.
또한, 본 발명의 다른 양태에 따르면, 본 발명은 상기 천연 복합 방부제 조성물을 유효성분으로 함유하는 약학적 조성물을 제공한다.According to another aspect of the present invention, there is also provided a pharmaceutical composition comprising the natural combination preservative composition as an active ingredient.
본 발명의 상기 천연 복합 방부제 조성물은 천연유래 식물 소재에서 추출되어 인체에 해가 없으며, 약품이 유해 미생물에 의해 부패하는 것을 효과적으로 막아주고, 그 첨가로 인해 화장품의 물질의 품질을 손상시키지 않으므로 약학적 조성물에 포함되어 사용되는데 적합하며, 특히 피부미백, 피부주름개선 및 피부염증 완화와 관련된 약학적 조성물에 포함되어 사용되는 것이 바람직하다.The natural combination preservative composition of the present invention is extracted from a naturally occurring plant material and has no harmful effect on the human body. It effectively prevents the decay of the drug by the harmful microorganisms and does not impair the quality of the substance of the cosmetic due to the addition thereof, It is preferable to be used in a pharmaceutical composition related to skin whitening, skin wrinkle improvement and skin irritation mitigation.
또한, 본 발명의 다른 양태에 따르면, 본 발명은 상기 천연 복합 방부제 조성물을 유효성분으로 함유하는 식품 조성물을 제공한다.According to another aspect of the present invention, there is also provided a food composition comprising the natural combination preservative composition as an active ingredient.
본 발명의 상기 천연 복합 방부제 조성물은 천연유래 식물 소재에서 추출되어 인체에 해가 없으며, 식품이 유해 미생물에 의해 부패하는 것을 효과적으로 막아주고, 그 첨가로 인해 식품의 물질의 품질을 손상시키지 않으므로 식품 조성물에 포함되어 사용되는데 적합하며, 특히 피부미백, 피부주름개선 및 피부염증 완화와 관련된 기능성 식품 조성물에 포함되어 사용되는 것이 바람직하다.Since the natural combination preservative composition of the present invention is extracted from natural plant material and is free from harm to the human body, it effectively prevents the food from being decayed by harmful microorganisms and does not impair the quality of the food substance due to the addition thereof, , And it is particularly preferable to be used in a functional food composition related to skin whitening, skin wrinkle improvement and skin irritation mitigation.
본 발명의 천연 복합 방부제 조성물은 그람양성, 그람음성, 및 진균류에 대한 우수한 항균 효과를 갖는 동시에, 멜라닌 색소의 생성을 저해하고, 콜라겐 단백질의 파괴를 저해하며, 손상된 피부 조직에서 염증반응을 억제시키는 효과가 있으므로, 방부제 조성물의 활용을 보다 제고할 수 있을 것으로 기대된다.INDUSTRIAL APPLICABILITY The natural complex preservative composition of the present invention has an excellent antimicrobial effect on gram-positive, gram-negative and fungi, and inhibits the production of melanin pigment, inhibits collagen protein destruction, inhibits inflammatory reaction in damaged skin tissue It is expected that the use of the preservative composition can be further improved.
본 발명의 특징 및 이점을 요약하면 다음과 같다:The features and advantages of the present invention are summarized as follows:
(ⅰ) 본 발명은 정향, 단삼, 황백, 및 황련의 복합 추출물을 유효성분으로 포함하는 천연 복합 방부제 조성물을 제공한다.(I) The present invention provides a natural combination preservative composition comprising, as an active ingredient, a combined extract of cloves, persimmon, persimmon, and yellowtail.
(ⅱ) 본 발명의 천연 복합 방부제 조성물은 다양한 균들에 대해 폭넓은 항균 및 항진균 스펙트럼을 나타내고 있으며, 화장품, 약품, 및 식품 조성물 분야에서 해당 물질을 보존하기 위한 방부 목적으로 유용하게 이용가능하다.(Ii) The natural combination preservative composition of the present invention exhibits a wide spectrum of antimicrobial and antifungal spectrum against various fungi, and can be usefully used for preservation purpose for preserving the material in the field of cosmetics, medicine, and food composition.
(ⅲ) 또한, 본 발명의 천연 복합 방부제 조성물은 인간 진피 섬유 아세포에서 MMP-1의 발현을 억제하고, 피부 주름의 발생을 억제하며, 손상된 피부의 염증반응을 개선하는 효능을 나타낸다. 또한, 본 발명의 조성물은 인체에 매우 안전할 뿐만 아니라, 안정성도 매우 탁월하다.(Iii) In addition, the natural combination preservative composition of the present invention inhibits the expression of MMP-1 in human dermal fibroblasts, suppresses the occurrence of wrinkles of the skin, and improves the inflammatory reaction of damaged skin. In addition, the composition of the present invention is not only very safe for human body but also excellent in stability.
도 1은 본 발명의 천연 복합 방부제 조성물이 갖는 멜라닌 생성 억제 효과를 나타내는 그래프이다.
도 2는 본 발명의 천연 복합 방부제 조성물이 갖는 MMP-1 발현 억제 효과를 나타내는 그래프이다.
도 3은 본 발명의 천연 복합 방부제 조성물이 갖는 IL-6 억제 효과를 나타내는 그래프이다. BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph showing the melanin production inhibitory effect of the natural combination preservative composition of the present invention. FIG.
2 is a graph showing the inhibitory effect of MMP-1 on the natural combination preservative composition of the present invention.
3 is a graph showing the IL-6 inhibitory effect of the natural combination preservative composition of the present invention.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하기로 한다. 이들 실시예는 단지 본 발명을 예시하기 위한 것이므로, 본 발명의 범위가 이들 실시예에 의해 제한되는 것으로 해석되지는 않는다.Hereinafter, the present invention will be described in more detail with reference to Examples. These embodiments are only for illustrating the present invention, and thus the scope of the present invention is not construed as being limited by these embodiments.
실시예 1: 식물 추출물의 제조Example 1: Preparation of plant extracts
1-1. 정향 추출물의 제조1-1. Production of clove extract
건조 중량 100g의 분쇄한 정향을 70%(V/V) 에탄올 수용액으로 12시간 가온 환류추출하고 냉침한 후, 1.2㎛ 투과사이즈를 갖는 여과지(Glassfiber, sartorius, 미국)로 여과하였다. 이 여과된 추출액을 50℃ 이하에서 감압농축한 후 감압건조기(Eyela Rotary evaporator, Tokyo Rikakika Co.,LTD, 일본)를 이용해 완전히 건조시켜 정향 추출물 12g을 수득하였다.The pulverized cloves having a dry weight of 100 g were heated and refluxed with an aqueous 70% (v / v) ethanol solution for 12 hours, cooled, and then filtered with a filter paper (Glassfiber, sartorius, USA) having a throughput size of 1.2 mu m. The filtered extract was concentrated under reduced pressure at 50 ° C or less, and then completely dried using a vacuum dryer (Eyela Rotary evaporator, Tokyo Rikakika Co., LTD., Japan) to obtain 12 g of a clove extract.
1-2. 단삼 추출물의 제조1-2. Preparation of Radix Root Extract
건조 중량 100g의 분쇄한 단삼을 70%(V/V) 에탄올 수용액으로 12시간 가온 환류추출하고 냉침한 후, 1.2㎛ 투과사이즈를 갖는 여과지(Glassfiber, sartorius, 미국)로 여과하였다. 이 여과된 추출액을 50℃ 이하에서 감압농축한 후 감압건조기(Eyela Rotary evaporator, Tokyo Rikakika Co.,LTD, 일본)를 이용해 완전히 건조시켜 단삼 추출물 8.2g을 수득하였다.The pulverized raw ginseng having a dry weight of 100 g was heated and refluxed with an aqueous 70% (v / v) ethanol solution for 12 hours, cooled, and filtered through a filter paper (Glassfiber, sartorius, USA) having a throughput size of 1.2 mu m. The filtered extract was concentrated under reduced pressure at 50 ° C or less, and then completely dried using a vacuum dryer (Eyela Rotary evaporator, Tokyo Rikakika Co., LTD., Japan) to obtain 8.2 g of Radix Salviae Radix Extract.
1-3. 황백 추출물의 제조1-3. Production of yellowish white extract
건조 중량 100g의 분쇄한 황백을 70%(V/V) 에탄올 수용액으로 12시간 가온 환류추출하고 냉침한 후, 1.2㎛ 투과사이즈를 갖는 여과지(Glassfiber, sartorius, 미국)로 여과하였다. 이 여과된 추출액을 50℃ 이하에서 감압농축한 후 감압건조기(Eyela Rotary evaporator, Tokyo Rikakika Co.,LTD, 일본)를 이용해 완전히 건조시켜 황백 추출물 10.5g을 수득하였다.The pulverized yellowish white powder having a dry weight of 100 g was heated and refluxed with an aqueous 70% (v / v) ethanol solution for 12 hours, cooled and filtered with a filter paper (Glassfiber, sartorius, USA) having a 1.2 μm permeable size. The filtered extract was concentrated under reduced pressure at a temperature of 50 ° C or less, and then completely dried using a vacuum dryer (Eyela Rotary evaporator, Tokyo Rikakika Co., LTD., Japan) to obtain 10.5 g of a yellowish white extract.
1-4. 황련 추출물의 제조1-4. Manufacture of HuangYin extract
건조 중량 100g의 분쇄한 황련을 70%(V/V) 에탄올 수용액으로 12시간 가온 환류추출하고 냉침한 후, 1.2㎛ 투과사이즈를 갖는 여과지(Glassfiber, sartorius, 미국)로 여과하였다. 이 여과된 추출액을 50℃ 이하에서 감압농축한 후 감압건조기(Eyela Rotary evaporator, Tokyo Rikakika Co.,LTD, 일본)를 이용해 완전히 건조시켜 단삼 추출물 11g을 수득하였다.The pulverized fermentation broth having a dry weight of 100 g was heated and refluxed with an aqueous 70% (v / v) ethanol solution for 12 hours, cooled, and filtered through a filter paper (Glassfiber, sartorius, USA) having a 1.2 μm permeation size. The filtered extract was concentrated under reduced pressure at 50 ° C or less, and then completely dried using a vacuum dryer (Eyela Rotary evaporator, Tokyo Rikakika Co., LTD., Japan) to obtain 11 g of Radix Salviae Radix extract.
실시예 2: 항균 활성 측정Example 2: Measurement of antimicrobial activity
2-1. 균주 및 배양조건2-1. Strain and culture conditions
상기 추출물들의 항균 효과를 측정하기 위하여 최소 저해농도(Minimum Inhibition Concentration, MIC)를 측정하여 항균활성을 조사하였다. 항균 효과를 측정하기 위하여 사용한 균주, 배지, 및 배양온도는 아래 표 1에 나타내었다. In order to measure the antibacterial effect of the extracts, the minimum inhibition concentration (MIC) was measured to investigate the antibacterial activity. The strains, media, and culture temperatures used to measure the antimicrobial effect are shown in Table 1 below.
[표 1][Table 1]
대장균(E coli CCARM0237), 슈도모나스 에루지노사(Pseudomonas aeruginosa KACC14021), 스타필로코커스 아우레우스(Staphylococcus aureus KCTC3881) 및 바실러스 서브틸리스(Bacillus subtilis KACC14394)의 균주와, 진균류인 칸디다 알비칸스 (Candida albicans KCTC7270), 아스퍼질러스 나이거 (Aspergillus niger KACC40280)를 사용하였다(CCARM 서울여대 항생제내성균주은행, KACC 농촌진흥청 농업미생물은행, KCTC 한국생명공학연구원 생물자원센터).A strain of Escherichia coli CCARM0237, Pseudomonas aeruginosa KACC14021, Staphylococcus aureus KCTC3881 and Bacillus subtilis KACC 14394, and a fungus Candida albicans, KCTC7270) and Aspergillus niger KACC40280 (CCARM Seoul National University College of Medicine), KACC Agricultural Microbiology Bank, KCTC Korea Research Institute of Bioscience and Biotechnology.
2-2. 디스크 확산법에 의한 항균력 검증2-2. Antimicrobial activity test by disk diffusion method
각각의 균주는 상기 표 1의 영양배지(Nutrient Broth) 용액을 제조하여 121℃에서 15분간 멸균한 후 각각의 배양된 균주 1 백금이를 영양배지 용액 (10㎖)에 접종하고, 진탕배양기(shaking incubator)에서 24시간 배양하여 항균실험을 위한 균주를 준비하였다.Each of the strains was prepared by preparing a nutrient broth solution of Table 1 and sterilized at 121 ° C for 15 minutes. Then, each of the cultured strains was inoculated into a nutrient broth (10 ml), shaking incubator for 24 hours to prepare a strain for the antibacterial experiment.
미생물 균주의 성장을 억제하는 항생물질의 최소억제농도를 측정하기 위해, 멸균된 배지에 배양된 균주를 세균은 1 X 108 cfu/㎖, 진균은 1 X 106 cfu/㎖이 되도록 각각을 접종하였다. 상기 실시예 1에서 제조된 4종의 단일 추출물은 2% 에탄올에 용해시켜 멸균한 페이퍼 디스크(직경 8mm)에 25㎕씩 두 번 점적한 후 충분히 건조시켰다. 추출물이 완전히 건조된 다음 평판 배지에 치상하여 28~37℃로 유지하여, 각각 1, 2, 3일(A. niger의 경우 7일까지 관찰) 후, 페이퍼 디스크 주변의 투명도(clear zone)를 측정하여 항균활성을 조사하였다. To measure the minimum inhibitory concentration of the antibiotic which inhibits growth of the microbial strain, inoculated with each of the strains cultured in a sterile culture medium bacteria is 1 X 10 8 cfu / ㎖, fungi such that a 1 X 10 6 cfu / ㎖ Respectively. Each of the four single extracts prepared in Example 1 was dissolved in 2% ethanol and dispensed in 25 μl onto a sterilized paper disk (diameter 8 mm) twice and dried sufficiently. After the extracts were completely dried, they were kept at 28 to 37 ° C for 1, 2, and 3 days (observed for 7 days in case of A. niger), and then the clear zone around the paper disk was measured To investigate the antimicrobial activity.
상기 추출물의 항균력을 비교 검증하기 위해, 시중에서 상용되는 메틸파라벤(Methyl Paraben)을 비교예 1로, 자몽종자추출물을 비교예 2로 사용하여 비교 실험하였다. In order to compare the antimicrobial activity of the extracts, Methyl Paraben (commercially available) and Comparative Example 2 were used as Comparative Example 1 and Comparative Example 2, respectively.
상기 정향, 단삼, 황백, 황련 추출물의 디스크 확산법에 의한 항균 활성을 아래 표 2에 나타내었다. 그 결과, 단일 추출물들의 항균 활성은 비교예 1 및 2와 비교하여 비교열위의 항균 활성을 갖는 것이 관찰되었다.The antimicrobial activity of the above-mentioned clove, Dansam, Hwangbuk, and Rhodiola extracts by the disk diffusion method is shown in Table 2 below. As a result, it was observed that the antimicrobial activity of the single extracts had antibacterial activities comparable to those of Comparative Examples 1 and 2.
[표 2][Table 2]
상기 정향, 단삼, 황백, 및 황련을 조합한 복합추출물의 항균 활성을 검정하기 위해 아래 표 3과 같이 조합하여 천연 복합 방부제 조성물을 제조하고, 상기와 같은 방법으로 각각의 시험예에 대한 디스크 확산법에 의한 항균 활성을 검정하여 표 4에 나타내었다. In order to test the antibacterial activity of the combined extracts of the combination of cloves, dansam, yellow whites, and chrysanthemums, a natural combination preservative composition was prepared as shown in Table 3 below, and the disc diffusion method for each test example And the results are shown in Table 4. < tb > < TABLE >
[표 3][Table 3]
[표 4][Table 4]
그 결과, 천연 복합 방부제는 시험예 7 내지 11은 모두 자몽종자추출물과 비교하여 동등 이상의 항균효과를 가지고 있는 것을 확인하였으며, 시험예 8과 11은 메틸파라벤 보다 우수한 항균효과고 있는 것을 확인하였고, 시험예 8은 가장 우수한 항균 활성을 갖는 것을 확인하였다.As a result, it was confirmed that all of the natural combination preservatives had the same antimicrobial effect as that of the grapefruit seed extracts in all of Test Examples 7 to 11, and Test Examples 8 and 11 had an antimicrobial effect superior to methylparaben Example 8 was confirmed to have the best antimicrobial activity.
2-3. 천연 복합 방부제의 미생물 생장 최소억제농도 평가2-3. Assessment of minimum inhibitory concentration of microorganism growth of natural compound preservatives
MIC 측정은 고형 한천 배지에서 각 세균과 진균을 배양한 후 1×106~8 CFU/mL에 준하여 5개의 집락을 얻고 배지에 희석하여 접종한 후 37℃에서 2~3시간 동안 유지하였다. 본 실험에 사용한 정향, 단삼, 황백, 및 황련의 단일 추출물들은 20.0 mg/mL 농도로 적정하여 각각 2배 계열로 희석하였다. 각 세균과 추출물이 함유된 배지를 96 well plate에 옮기고 37℃에서 16~24시간 동안 배양한 후, 600nm에서 Multilabel Counter(1420, Perkin Elmer)로 측정하여 단일 추출물들의 최소저해농도를 구하고, 이를 표 5에 나타내었다. MIC measurement was performed by incubating each bacterium and fungus in a solid agar culture medium and then obtaining 5 colonies according to 1 × 10 6 to 8 CFU / mL, diluted in the medium and inoculated, and maintained at 37 ° C. for 2 to 3 hours. Single extracts of clove, dansam, yellow whites, and chrysanthemums used in this experiment were titrated to 20.0 mg / mL and diluted to 2-fold series, respectively. The culture medium containing each bacterium and extract was transferred to a 96-well plate and cultured at 37 ° C for 16-24 hours. The minimum inhibitory concentration of single extracts was determined by measuring with a Multilabel Counter (1420, Perkin Elmer) at 600 nm Respectively.
또한, 표 3의 배합 비율에 따라 제조된 복합 추출물의 최소저해농도를 측정하여 표 6에 나타내었다. The minimum inhibitory concentrations of the compound extracts prepared according to the compounding ratios in Table 3 were measured and are shown in Table 6.
[표 5] 단일 추출물의 미생물 생장 최소 억제 농도(MIC) 측정(mg/ml)[Table 5] Minimum inhibitory concentration (MIC) of microorganism growth of single extract (mg / ml)
[표 6] 복합 추출물의 미생물 생장 최소 억제 농도(MIC) 측정(mg/ml)[Table 6] Microbial growth inhibition concentration (MIC) measurement (mg / ml)
그 결과, 시험예 1 내지 11에서 전반적으로 우수한 항균활성을 나타내는 것을 확인하였으며, 시험예 8이 대조군으로 사용한 메틸파라멘과 자몽종자추출물과 비슷하거나 우수한 항균활성을 보이는 것을 확인하였다.As a result, it was confirmed that the antibacterial activity in Test Examples 1 to 11 was excellent overall, and Test Example 8 showed similar or superior antimicrobial activity to methylparramen and grapefruit seed extract used as the control group.
또한, 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물 각각에 대한 미생물 생장최소억제 농도 평균값과 시험예 8를 비교한 결과, 시험예 8의 비율로 혼합하여 사용한 경우 각각의 평균값보다 60~80% 감소된 미생물 생장 최소억제 농도를 보여 뛰어난 미생물 생장 억제 효과를 확인할 수 있었다(표 7).In addition, when the average value of the minimum inhibitory concentration of microorganisms was compared with that of Test Example 8 for each of the clove extract, roots extract, Huang Baek extract and Rhodiola extract, it was 60 ~ 80% (Table 7). As shown in Table 7, the microbial growth inhibitory effect of the present invention was confirmed.
[표 7][Table 7]
즉, 각각의 추출물을 따로 사용하였을 때보다 본 발명에서와 같이 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물을 일정한 비율로 혼합하여 사용하였을 때 상승적인 미생물 생장 억제 효과(synergy effect)를 보이는 것을 확인하였다.That is, it was confirmed that the synergy effect of the clove extract, the ginseng extract, the yellowish white extract, and the chrysanthemum extract was mixed when they were mixed at a constant ratio as compared with the case where each extract was separately used, Respectively.
실시예 3. 천연 복합 방부제의 세포생존율 및 미백활성 측정Example 3. Measurement of cell survival rate and whitening activity of a natural combination preservative
본 발명의 복합물 에 대한 세포생존율을 측정하기 위해 MTT(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) 어세이를 실시하였다.(3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide) assay was performed to measure the cell viability of the composite of the present invention.
먼저, 0.5㎎/㎖의 MTT 시약 100㎕와 상기 시험예 8의 복합추출물이 처리된 멜란-A 세포 배양액을 600㎕로 줄인 뒤, 각 웰에 더하여 5% CO2, 95% O2, 37℃ 조건에서 2시간 동안 배양하였다.First, 0.5㎎ / ㎖ of MTT after the reduced
기질이 포함된 배지를 제거하고, 각 웰의 포르마잔 결정에 500㎕의 DMSO(dimethyl sulfoxide)를 분주하여 용해시켰다. 이후, 플레이트를 실온에서 30분 동안 교반한 다음, 100㎕씩 분주하여 마이크로플레이트 리더(microplate reader, Molecular Devices E09090, USA)로 570㎚에서 흡광도를 측정하였다(도 1 참조). The medium containing the substrate was removed, and 500 mu l of DMSO (dimethyl sulfoxide) was dissolved in the formazan crystals of each well. Thereafter, the plate was stirred at room temperature for 30 minutes, and 100 ㎕ aliquots were added thereto, and the absorbance was measured at 570 nm with a microplate reader (Molecular Devices E09090, USA) (see Fig. 1).
또한, 멜라닌 함량을 측정하기 위해, 선행문헌 [Hosoi et al., 1985]에 의해 보고된 방법을 수정하여 적용하였다. 먼저, 상기 멜란-A 세포에 상기 시험예 8의 복합추출물을 1, 10, 및 100 ug/ml 의 농도로 3일 동안 처리한 후, 배양된 배지를 제거하고, 각 웰을 PBS(phosphate buffer saline)로 두 번씩 세척하였다. 이후, 멜라닌을 용해하기 위해, 1N의 NaOH 400㎕를 각 웰에 더하고, 마이크로플레이트 리더를 사용하여 405㎚의 흡수파장에서 멜라닌을 측정하여 도 1에 나타내었다.In addition, to measure the melanin content, the method reported by the prior art [Hosoi et al., 1985] was modified and applied. First, the complex extract of Test Example 8 was treated for 3 days at a concentration of 1, 10, and 100 ug / ml in the melan-A cells. The cultured medium was removed and each well was washed with PBS ). ≪ / RTI > Then, in order to dissolve melanin, 400 1 of 1N NaOH was added to each well, and melanin was measured at an absorption wavelength of 405 nm using a microplate reader, and it is shown in Fig.
그 결과, 본 발명의 천연 복합추출물이 처리된 피부세포에서 농도 의존적으로 세포 생존율이 증가하고, 멜라닌 색소의 생성이 억제되는 것을 확인할 수 있었다. As a result, it was confirmed that the cell survival rate was increased in a concentration-dependent manner in the skin cells treated with the natural compound extract of the present invention and the production of melanin pigment was inhibited.
실시예 4. 천연 복합 방부제의 주름개선 효과 측정Example 4. Measurement of wrinkle-improving effect of natural compound preservatives
건강한 젊은 남성 지원자(SD 생명공학, 서울, 한국)를 선발하고, 피부 생검하여 정상 인간 진피 섬유아세포(NHDF)들을 획득하였다. 세포들은 100-㎜조직 배양 플레이트에서 플레이팅 되었고, 5% CO2를 함유하는 습기 환경, 37℃ 에서 10%의 열-비활성화된 FBS 및 1%의 페니실린-스트렙토마이신으로 보충된 DMEM 배지에서 배양되었다. 모든 실험은 오직 계대 6 내지 10 사이의 세포들을 사용하여 수행하였다.Healthy young male volunteers (SD Biotech, Seoul, Korea) were selected and skin biopsies were obtained to obtain normal human dermal fibroblasts (NHDF). Cells were plated in 100-mm tissue culture plates and cultured in DMEM medium supplemented with a humidified environment containing 5% CO 2 , 10% heat-inactivated FBS and 1% penicillin-streptomycin at 37 ° C . All experiments were performed using only cells between 6 and 10 passages.
상기 정상 인간 진피 섬유아세포들은 40㎜ 조직 배양 플레이트들(1.2×105 세포들)에 시드하였다. 정상 인간 진피 섬유아세포들이 80% 융합에 도달할 때, 그들을 인산 완충 식염수(PBS)로 두 차례 씻어내었고, 신선한 세럼-프리 배지 1980㎕와 샘플 20㎕는 각 웰에 첨가된 후, 정상 인간 진피 섬유아세포들은 따뜻한 PBS로 세 차례 세척하였다.The normal human dermal fibroblast cells were seeded in tissue culture plates 40㎜ (s 1.2 × 10 5 cells). When normal human dermal fibroblasts reached 80% confluence, they were washed twice with phosphate buffered saline (PBS), and 1980 신선한 fresh serum-free medium and 20 샘플 of sample were added to each well, Fibroblasts were washed three times with warm PBS.
MMP-1(매트릭스 메탈로프로테이나제) 활성은 UVB 조사 후 72시간 채취된 상층액에서 평가되었다. ELISA 리더기를 이용하여 채취된 상층액은 450nm에서 측정 되었다.MMP-1 (matrix metalloproteinase) activity was evaluated in the supernatant collected 72 hours after UVB irradiation. The supernatant collected using an ELISA reader was measured at 450 nm.
그 결과, 본 발명의 천연 복합추출물이 처리된 피부세포에서 농도 의존적으로 세포 생존율이 증가하고, MMP-1(매트릭스 메탈로프로테이나제) 활성이 억제되는 것을 확인할 수 있었다(도 2). As a result, it was confirmed that the cell survival rate was increased in a concentration-dependent manner in the skin cells treated with the natural compound extract of the present invention, and the activity of MMP-1 (matrix metalloproteinase) was inhibited (FIG. 2).
실시예 5. 천연 복합 방부제의 항염증 효과 측정Example 5. Measurement of anti-inflammatory effect of natural compound preservative
HaCaT cells (피부 표피 각질 세포주) 세포들은 100 ㎜ 조직 배양 플레이트에서 플레이팅 되었고, 5% CO2를 함유하는 습기 환경, 37℃ 에서 10%의 열-비활성화된 FBS 및 1%의 페니실린-스트렙토마이신으로 보충된 DMEM 배지에서 배양되었다. HaCaT cells were plated on 100 mm tissue culture plates and cultured in a humidified environment containing 5% CO 2 , 10% heat-inactivated FBS at 37 ° C and 1% penicillin-streptomycin ≪ / RTI > supplemented DMEM medium.
정상 인간 표피 각질 세포 (HaCaT cells)들은 40㎜ 조직 배양 플레이트들(1.0×105 세포들)에 시드하였다. 정상 인간 진피 섬유아세포들이 80% 융합에 도달할 때, 그들을 인산 완충 식염수(PBS)로 두 차례 씻어내었고, 신선한 세럼-프리 배지 1980㎕와 샘플 20㎕는 각 웰에 첨가된 후, HaCaT cells은 따뜻한 PBS로 세 차례 세척하였다.Normal human epidermal keratinocytes (HaCaT cells) were seeded (the 1.0 × 10 5 cells) in tissue culture plates 40㎜. When normal human dermal fibroblasts reached 80% confluence, they were washed twice with phosphate buffered saline (PBS), and 1980 신선한 fresh serum-free medium and 20 ㎕ of sample were added to each well and then HaCaT cells Washed three times with warm PBS.
IL-6 단백질은 UVB 조사 후 24시간 채취된 상층액에서 평가되었다. ELISA 리더기를 이용하여 채취된 상층액을 450nm에서 측정하였다.IL-6 protein was assessed in the supernatant collected 24 hours after UVB irradiation. The supernatant collected using an ELISA reader was measured at 450 nm.
그 결과, 본 발명의 천연 복합추출물이 처리된 피부세포에서 농도 의존적으로 세포 생존율이 증가하고, 염증매개 인자인 IL-6 단백질의 발현이 억제되는 것을 확인할 수 있었다(도 3). As a result, it was confirmed that the cell survival rate was increased in a concentration-dependent manner in the skin cells treated with the natural compound extract of the present invention and the expression of IL-6 protein as an inflammatory mediator was inhibited (FIG. 3).
실시예 6. 천연 복합 방부제를 함유하는 화장품의 방부력 시험Example 6. Flotation test of cosmetics containing natural complex preservatives
본 발명의 천연 복합 방부제 조성물을 함유하는 화장료의 방부력 시험을 행하기 위해 메틸파라벤이 0.15 중량%로 포함된 비교제조예 및 시험예 8의 천연 복합 방부제 조성물이 0.12 중량%로 함유된 제조예의 크림 제형을 제조하였다(표 8).In order to carry out the flotation test of the cosmetic composition containing the natural combination preservative composition of the present invention, the comparative preparation example containing methylparaben in an amount of 0.15% by weight and the cream of the production example containing 0.12% by weight of the natural combination preservative composition of Test Example 8 Formulations were prepared (Table 8).
[표 8][Table 8]
상기 비교제조예와 제조예의 크림 제형에 대한 방부 효과를 미국화장품공업협회(The Cosmetic, Toility and Fragrance Association, Inc., 이하 CTFA)의 미생물 가이드라인(microbiology Guidelines)의 시험법을 사용하여 시행하였다. 사용한 균주로는 대장균(E coli CCARM0237), 슈도모나스 에루지노사(Pseudomonas aeruginosa KACC14021), 및 스타필로코커스 아우레우스(Staphylococcus aureus KCTC3881)의 균주와, 진균류인 칸디다 알비칸스 (Candida albicans KCTC7270), 아스퍼질러스 나이거 (Aspergillus niger KACC40280)를 사용하였다. 비교예와 시험예들이 10g씩 담겨져 있는 병에 세균 3종(E.coli, P. aeruginosa, S. aureus)과 진균 2종(C. albicans, A. niger)을 혼합한 현탁액을 각각 1×106 cfu/g, 1×105 cfu/g이 되도록 접종한 다음, 25℃에서 보관 후 4시간, 1일, 2일, 3일, 7일 간격으로 무균 조작 하에서 1g의 샘플을 채취하여 희석액으로 적절히 희석하고 세균은 뉴트리언트 아가(Nutrient agar, Difco), 진균은 포테이토 덱스트로스 아가(potato dextrose agar, Difoc) 배지에 깔아 각각 35℃, 25℃에서 24 ~ 72시간 동안 배양 후 계수하였다.The preservative effects of the cream formulations of the comparative preparation examples and the preparation examples were tested using the microbiology guidelines of the CTFA. The strains used were Escherichia coli CCARM0237, Pseudomonas aeruginosa KACC14021, and Staphylococcus aureus KCTC3881, Candida albicans KCTC7270, Aspergillus species, Aspergillus niger KACC40280 was used. A suspension of three bacteria (E. coli, P. aeruginosa, S. aureus) and two fungi (C. albicans, A. niger) in a bottle containing 10 g of each of the comparative examples and the test examples was diluted 1 × 10 6 cfu / g, and 1 × 10 5 cfu / g. Then, 1 g of sample was collected under sterile conditions at 4 hours, 1 day, 2 days, 3 days and 7 days after storage at 25 ° C. The bacteria were diluted appropriately and the bacteria were counted on Nutrient agar (Difco) and the fungi were cultured on potato dextrose agar (Difoc) medium and incubated at 35 ° C and 25 ° C for 24 to 72 hours, respectively.
[표 9][Table 9]
그 결과, 시험제조예의 천연 복합 방부제를 함유한 크림의 방부 효과는 종래의 합성 방부제로 사용되는 비교제조예와 비슷한 효과를 보이므로(표 9 참조), 본 발명의 천연 복합 방부제 조성물을 매우 우수한 방부 효과를 가지는 것을 확인하였다.As a result, the antiseptic effect of the cream containing the natural combination preservative of the test production example showed similar effects to those of the comparative preparation used as a conventional synthetic preservative (see Table 9), so that the natural combination preservative composition of the present invention is excellent in preservation Respectively.
이상으로 본 발명의 특정한 부분을 상세히 기술하였는 바, 당업계의 통상의 지식을 가진 자에게 있어서 이러한 구체적인 기술은 단지 바람직한 구현예일 뿐이며, 이에 본 발명의 범위가 제한되는 것이 아닌 점은 명백하다. 따라서, 본 발명의 실질적인 범위는 첨부된 청구항과 그의 등가물에 의하여 정의된다고 할 것이다.While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.
Claims (9)
전체 조성물 중량대비 상기 정향 추출물은 0.01 내지 10 중량부, 상기 단삼 추출물은 0.01 내지 10 중량부, 상기 황백 추출물은 0.01 내지 5 중량부, 및 상기 황련 추출물은 0.01 내지 5 중량부인 것을 특징으로 하는 천연 복합 방부제 조성물.The method according to claim 1,
Characterized in that the amount of the clove extract is 0.01 to 10 parts by weight based on the weight of the whole composition, 0.01 to 10 parts by weight of the extract of Radix ginseng, 0.01 to 5 parts by weight of the yellowish white extract, and 0.01 to 5 parts by weight of the extract. Preservative composition.
상기 정향 추출물, 단삼 추출물, 황백 추출물 및 황련 추출물은 4:3:2:3의 중량비인 것을 특징으로 하는 천연 복합 방부제 조성물.The method according to claim 1,
Wherein the clove extract, ginsenoside R5 extract, Huang Baek extract and Rhodiola extract are in a weight ratio of 4: 3: 2: 3.
상기 조성물은 멜라닌 색소의 생성을 억제하여 피부미백 활성을 갖는 것을 특징으로 하는 천연 복합 방부제 조성물.The method according to claim 1,
Wherein the composition has skin whitening activity by inhibiting the production of melanin pigment.
상기 조성물은 MMP-1 단백질의 발현을 억제하여 피부 주름을 개선하는 것을 특징으로 하는 천연 복합 방부제 조성물.The method according to claim 1,
Wherein said composition inhibits the expression of MMP-1 protein to improve skin wrinkles.
상기 조성물은 IL-6 단백질의 발현을 억제하여 피부 염증 개선 활성을 갖는 것을 특징으로 하는 천연 복합 방부제 조성물.The method according to claim 1,
Wherein the composition inhibits the expression of IL-6 protein and has a skin inflammation-improving activity.
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| WO2019225786A1 (en) * | 2018-05-23 | 2019-11-28 | (주)에스디생명공학 | Antibacterial and antifungal composition comprising extract from clove, hibiscus, and coconut as effective ingredient |
| KR20200055623A (en) * | 2018-11-13 | 2020-05-21 | 주식회사 케이바이오랩 | Antibiotic composition containing extract of Syzygii Flos and extract of Phellodendron Cortex |
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| WO2019225786A1 (en) * | 2018-05-23 | 2019-11-28 | (주)에스디생명공학 | Antibacterial and antifungal composition comprising extract from clove, hibiscus, and coconut as effective ingredient |
| KR20200055623A (en) * | 2018-11-13 | 2020-05-21 | 주식회사 케이바이오랩 | Antibiotic composition containing extract of Syzygii Flos and extract of Phellodendron Cortex |
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