KR102184119B1 - Novel strain Saccharomyces cerevisiae EY2-27 having skin whitening and use of the same - Google Patents
Novel strain Saccharomyces cerevisiae EY2-27 having skin whitening and use of the same Download PDFInfo
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- KR102184119B1 KR102184119B1 KR1020190061320A KR20190061320A KR102184119B1 KR 102184119 B1 KR102184119 B1 KR 102184119B1 KR 1020190061320 A KR1020190061320 A KR 1020190061320A KR 20190061320 A KR20190061320 A KR 20190061320A KR 102184119 B1 KR102184119 B1 KR 102184119B1
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- saccharomyces cerevisiae
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- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
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Abstract
Description
본 발명은 피부 미백 활성을 갖는 신균주 사카로마이세스 세레비지애 EY2-27 및 이의 용도에 관한 것이다.The present invention relates to a new strain of Saccharomyces cerevisiae EY2-27 having skin whitening activity and use thereof.
성형, 미용, 화장에 대한 일반 대중의 관심이 급속도로 높아지고 있다. 특히 동안 열풍 및 하얀 얼굴에 대한 선호 현상으로 인하여 미백화장품 및 주름개선화장품에 대한 관심이 지속적이면서도 폭발적으로 증가하고 있다.The general public's interest in plastic surgery, beauty and makeup is rapidly increasing. In particular, the interest in whitening cosmetics and wrinkle-improving cosmetics is constantly and explosively increasing due to the hot air and the preference for white faces.
특히 미백화장품에 대한 관심은 미백 증진 소재에 대한 수요의 증가와 효소, 유산균 등의 미생물을 이용한 화장품의 개발로 이어지고 있고, 최근에는 피부에 안정성이 높으며 피부 미백활성이 우수한 소재를 천연소재로부터 찾고자 하는 연구가 활발히 진행되고 있다.In particular, interest in whitening cosmetics is leading to an increase in demand for whitening enhancing materials and the development of cosmetics using microorganisms such as enzymes and lactic acid bacteria, and recently, a desire to find materials with high skin stability and excellent skin whitening activity from natural materials. Research is actively underway.
한편, 피부색은 주로 피부 속에 존재하는 멜라닌(melanin)이라는 색소의 함량에 의해 결정되는데, 멜라닌은 피부의 기저층에 존재하는 멜라닌 생성세포(멜라노사이트, melanocyte)에 의해서 생합성되고 세포질 돌기를 통하여 각질형성세포(keratinocyte)의 각화과정에 의해서 표피의 기저층에서 각질층으로 이동하게 된다. 이 멜라닌을 형성하는 주효소인 티로시나아제(tyrosinase)는 속도결정단계(rate-limiting step)인 타이로신(tyrosine)에서 디하이드록시페닐알라닌(3,4-dihydroxyphenylalanine, DOPA, 도파)으로의 전환단계와 이후 단계인 도파(DOPA)에서 도파퀴논(DOPA quinone)으로의 전환단계를 매개한다. 따라서 티로시나아제 효소의 활성을 저해하고 멜라닌 생성을 억제시켜 피부 미백 효과를 얻을 수 있다.On the other hand, skin color is mainly determined by the content of a pigment called melanin in the skin, which is biosynthesized by melanocytes (melanocytes) in the base layer of the skin, and keratinocytes through cytoplasmic processes (keratinocyte) is transferred from the basal layer of the epidermis to the stratum corneum by the keratinization process. This melanin-forming main enzyme, tyrosinase, is a conversion step from tyrosine, a rate-limiting step, to dihydroxyphenylalanine (3,4-dihydroxyphenylalanine, DOPA, dopa), and A subsequent step, a step of conversion from DOPA to DOPA quinone, is mediated. Therefore, it is possible to obtain a skin whitening effect by inhibiting the activity of the tyrosinase enzyme and suppressing the production of melanin.
현재 미백제로 가장 많이 사용되는 알부틴은 L-타이로신과 경쟁적으로 작용하는 저해제이다. 또한 미백제로 많이 사용되는 코직산(Kojic acid)는 타이로시나아제 활성 부위의 구리를 킬레이팅(chelating)하여 타이로신에서 도파로, 그리고 도파에서 도파퀴논으로 진행되는 과정을 저해한다. 그러나 이러한 미백제들은 피부 부작용을 초래할 수 있는 문제점이 있어 부작용이 없으면서도 피부 미백 활성을 높일 수 있는 새로운 소재의 개발이 필요한 시점이다.Arbutin, which is currently most commonly used as a whitening agent, is an inhibitor that acts competitively with L-tyrosine. Also, Kojic acid, which is often used as a whitening agent, inhibits the process of tyrosine to waveguide and waveguide to dopaquinone by chelating copper at the tyrosinase active site. However, since these whitening agents have a problem that can cause skin side effects, it is time to develop a new material capable of enhancing skin whitening activity without side effects.
이에 본 발명자들은 피부의 미백 활성을 증진시킬 수 있는 방법으로, 피부에 바르는 화장품 뿐만 아니라 미백 활성을 증진시킬 수 있는 기능성 식품의 원료로도 사용할 수 있는 소재를 미생물로부터 찾기 위해 연구하던 중, 미백 활성을 증진시킬 수 있는 새로운 균주를 동정하고 이의 우수한 미백활성을 확인함으로써 본 발명을 완성하였다.Accordingly, the inventors of the present invention were researching from microorganisms to find a material that can be used as a raw material for functional foods that can enhance whitening activity as well as cosmetics applied to the skin as a way to increase the whitening activity of the skin. The present invention was completed by identifying a new strain capable of enhancing and confirming its excellent whitening activity.
따라서 본 발명의 목적은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주를 제공하는 것이다.Accordingly, an object of the present invention is to provide a strain of Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P).
본 발명의 다른 목적은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물을 유효성분으로 포함하는 미백 활성을 갖는 식품 조성물 또는 화장료 조성물을 제공하는 것이다.Another object of the present invention is a food composition or cosmetic composition having a whitening activity comprising Saccharomyces cerevisiae EY2-27 (accession number: KCCM 12439P) strain, its lysate, its culture or its fermentation as an active ingredient Is to provide.
본 발명의 다른 목적은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주를 이용하여 전분질 원료를 발효시키는 단계를 포함하는, 미백 활성을 갖는 발효물의 제조방법을 제공하는 것이다.Another object of the present invention is to provide a method for producing a fermented product having whitening activity, comprising fermenting a starchy raw material using Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P) strain.
상기와 같은 본 발명의 목적을 달성하기 위해서, 본 발명은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주를 제공한다. In order to achieve the object of the present invention as described above, the present invention provides a Saccharomyces cerevisiae EY2-27 (accession number: KCCM 12439P) strain.
본 발명의 일실시예에 있어서, 상기 균주는 티로시나제(tyrosinase) 억제 활성 및 멜라닌(melanin) 생성 억제를 통해 미백 활성을 갖는 것일 수 있다.In one embodiment of the present invention, the strain may have a whitening activity through inhibition of tyrosinase inhibition and melanin production.
또한 본 발명은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물을 유효성분으로 포함하는 미백 활성을 갖는 식품 조성물을 제공한다. In addition, the present invention provides a food composition having a whitening activity comprising the Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P) strain, its lysate, its culture, or its fermentation as an active ingredient.
또한 본 발명은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물을 유효성분으로 포함하는 미백 활성을 갖는 화장료 조성물을 제공한다. In addition, the present invention provides a cosmetic composition having a whitening activity comprising Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P) strain, its lysate, its culture, or its fermentation as an active ingredient.
또한 본 발명은 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주를 이용하여 전분질 원료를 발효시키는 단계를 포함하는, 미백 활성을 갖는 발효물의 제조방법을 제공한다. In addition, the present invention provides a method for producing a fermented product having whitening activity, comprising fermenting a starchy raw material using Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P) strain.
본 발명의 일실시예에 있어서, 상기 발효물은, (1) 입국에 물 및 사카로마이세스 세레비지애 EY2-27(기탁번호: KCCM 12439P) 균주, 이의 파쇄물 또는 이의 배양물을 혼합하고 발효시켜 1차 발효물을 제조하는 단계; (2) 상기 1차 발효물에 증자된 전분질 원료 및 물을 혼합한 후 발효시켜 2차 발효물을 제조하는 단계; 및 (3) 상기 2차 발효물을 증류시키고 남은 잔사물을 수득하는 단계를 포함하는 과정으로 수득된 것일 수 있다.In one embodiment of the present invention, the fermented product is (1) mixed with water and Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P) strain, its lysate or its culture, and fermented To prepare a first fermented product; (2) preparing a second fermented product by fermenting after mixing the starchy raw material and water steamed in the first fermented product; And (3) distilling the secondary fermented product and obtaining the remaining residue.
본 발명의 일실시예에 있어서, 상기 전분질 원료는 서류 또는 곡류일 수 있다.In one embodiment of the present invention, the starchy raw material may be documents or grains.
본 발명의 일실시예에 있어서, 상기 서류는 고구마 또는 감자이고, 상기 곡류는 쌀, 찹쌀, 보리, 겉보기, 옥수수, 메밀, 귀리 또는 조일 수 있다.In one embodiment of the present invention, the document may be sweet potato or potato, and the grain may be rice, glutinous rice, barley, apparent, corn, buckwheat, oats, or crushed rice.
본 발명의 일실시예에 있어서, 상기 (1) 단계에서 상기 발효는 20~25℃의 온도에서 4~6일간 발효시키는 것일 수 있다.In one embodiment of the present invention, the fermentation in step (1) may be fermentation for 4 to 6 days at a temperature of 20 to 25°C.
본 발명의 일실시예에 있어서, 상기 (2) 단계에서 전분질 원료는 물로 세척 후, 2~4시간 동안 상온의 물에서 침지한 후, 40~60분 동안 스팀으로 증자하고, 20~30분 동안 상온에서 냉각시킨 것일 수 있다.In an embodiment of the present invention, in the step (2), the starchy raw material is washed with water, immersed in water at room temperature for 2 to 4 hours, then increased with steam for 40 to 60 minutes, and then for 20 to 30 minutes. It may be cooled at room temperature.
본 발명의 일실시예에 있어서, 상기 (2) 단계에서 2차 발효는 20~25℃의 온도에서 8~12일간 발효시키는 것일 수 있다.In one embodiment of the present invention, the second fermentation in step (2) may be fermentation for 8 to 12 days at a temperature of 20 to 25°C.
본 발명의 일실시예에 있어서, 상기 (3) 단계에서 증류는 상기 2차 발효물에 함유된 알코올을 증류기를 이용하여 제거하되, 증류 후 남은 잔사물에 함유된 알코올 성분이 1%(v/v) 미만이 되도록 수행하는 것일 수 있다.In one embodiment of the present invention, in the distillation in step (3), the alcohol contained in the secondary fermentation is removed using a still, but the alcohol component contained in the residue remaining after distillation is 1% (v/ v) It may be performed to be less than.
본 발명은 미백 활성을 갖는 신규한 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주에 관한 것이다. 본 발명에 따른 상기 균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물은 티로시나제 억제 활성이 우수하고 멜라닌 생성 저해 활성이 우수하여 피부 미백 활성을 갖는 화장료 조성물 및 식품 조성물의 용도로 유용하게 사용될 수 있다. The present invention relates to a novel Saccharomyces cerevisiae EY2-27 (KCCM12439P) strain having whitening activity. The strain, its lysate, its culture, or its fermented product according to the present invention has excellent tyrosinase inhibitory activity and excellent melanin production inhibitory activity, so it can be usefully used for cosmetic compositions and food compositions having skin whitening activity.
도 1은 본 발명의 일실시예에서 본 발명의 신균주인 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주를 이용하여 수득한 고구마 발효물의 증류 잔사물에 대한 티로시나제 억제 활성을 분석한 결과를 나타낸 것이다.
도 2는 본 발명의 일실시예에서 본 발명의 신균주인 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주를 이용하여 수득한 고구마 발효물의 증류 잔사물에 대한 멜라닌 생성 억제 활성을 분석한 결과를 나타낸 것이다. 1 is a result of analyzing tyrosinase inhibitory activity against distillation residues of fermented sweet potato obtained using Saccharomyces cerevisiae EY2-27 (KCCM12439P) strain, a new strain of the present invention in an embodiment of the present invention Is shown.
Figure 2 is an analysis of the melanin production inhibitory activity on the distillation residue of the fermented sweet potato obtained using the Saccharomyces cerevisiae EY2-27 (KCCM12439P) strain, a new strain of the present invention in an embodiment of the present invention It shows the results.
본 발명은 미백 활성을 갖는 신규한 균주를 제공함에 특징이 있으며, 구체적으로 본 발명은 피부 미백 활성을 갖는 신균주인 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주를 제공함에 특징이 있다. The present invention is characterized by providing a novel strain having whitening activity, and in particular, the present invention is characterized by providing a new strain of Saccharomyces cerevisiae EY2-27 (KCCM12439P) having skin whitening activity. .
본 발명자들은 체내 부작용을 유발하지 않으면서 효과적으로 피부 미백 활성을 증진시킬 수 있는 새로운 소재를 천연에서 찾기 위해 연구하던 중, 미백 활성을 갖는 신균주를 분리 및 동정하였고, 본 발명에서 분리한 신규 미생물을 16s rDNA 염기서열에 기초한 분자계통분류학 분석을 통해 분리된 상기 균주가 종래 존재하지 않은 신규한 균주임을 확인하였으며, 사카로마이세스 세레비지애 EY2-27 균주를 한국미생물보존센터에 2019.02.19.일자로 기탁하여 기탁번호 KCCM 12439P를 부여받았다.The inventors of the present invention isolated and identified a new strain having whitening activity while studying to find a new material in nature that can effectively enhance skin whitening activity without causing side effects in the body, and the novel microorganisms isolated in the present invention Through molecular phylogenetic analysis based on 16s rDNA sequence, it was confirmed that the isolated strain was a novel strain that did not exist before, and the Saccharomyces cerevisiae EY2-27 strain was transferred to the Korea Microbiological Conservation Center on February 19, 2019. It was deposited with and was assigned the deposit number KCCM 12439P.
이에 본 발명자들은 본 발명에서 분리 및 동정한 사카로마이세스 세레비지애 EY2-27 균주에 대해 피부 미백활성이 있는지 확인하기 위하여, 상기 EY2-27 균주 및 상기 EY2-27 균주를 이용하여 제조한 발효물에 대한 멜라닌 생성 저해활성 및 티로시나제 억제 활성을 분석하였다.Accordingly, in order to confirm whether the present inventors have skin whitening activity against the Saccharomyces cerevisiae EY2-27 strain isolated and identified in the present invention, fermentation prepared using the EY2-27 strain and the EY2-27 strain The melanin production inhibitory activity and tyrosinase inhibitory activity in water were analyzed.
그 결과, 본 발명의 사카로마이세스 세레비지애 EY2-27 균주의 배양물이 우수한 미백 활성이 있는 것으로 나타났고, 또한 상기 균주를 이용하여 수득한 발효물이 멜라닌 생성 저해 활성 및 티로시나제 억제 활성이 있는 것으로 나타났으며, 이러한 미백 활성은 다른 종류의 사카로마이세스 세레비지애 균주들에 비해 더 우수한 것으로 나타났다.As a result, it was found that the culture of the Saccharomyces cerevisiae EY2-27 strain of the present invention has excellent whitening activity, and the fermentation product obtained using the strain has melanin production inhibitory activity and tyrosinase inhibitory activity. This whitening activity was found to be superior to other types of Saccharomyces cerevisiae strains.
따라서 본 발명의 사카로마이세스 세레비지애 EY2-27 균주는 피부 미백 증진을 위한 화장품 및 기능성 식품의 소재로 사용할 수 있음을 알 수 있었다.Therefore, it was found that the Saccharomyces cerevisiae EY2-27 strain of the present invention can be used as a material for cosmetics and functional foods for enhancing skin whitening.
그러므로 본 발명은 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물을 유효성분으로 포함하는 피부 미백 활성을 갖는 화장료 조성물을 제공할 수 있다.Therefore, the present invention can provide a cosmetic composition having skin whitening activity comprising the Saccharomyces cerevisiae EY2-27 (KCCM12439P) strain, its lysate, its culture, or its fermentation as an active ingredient.
본 발명의 피부 미백 활성을 갖는 화장료 조성물은 본 발명에 따른 균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물을 유효성분으로 함유할 수 있으며, 적합한 부형제 또는 담체를 추가로 포함할 수 있다. The cosmetic composition having skin whitening activity of the present invention may contain a strain according to the present invention, a lysate thereof, a culture product thereof, or a fermented product thereof as an active ingredient, and may further include a suitable excipient or carrier.
본 발명에서 상기 배양물은 본 발명에 따른 신균주를 적합한 액체 배지에서 배양한 배양액 자체, 상기 배양액을 여과 또는 원심분리하여 균주를 제거한 여액(여과액 또는 원심분리한 상등액), 상기 배양액을 초음파 처리하거나 상기 배양액에 용해효소(lysozyme)를 처리하여 수득한 세포 파쇄액(파쇄물) 등을 포함하나, 이에 한정되는 것은 아니다.In the present invention, the culture is a culture solution obtained by culturing a new strain according to the present invention in a suitable liquid medium, a filtrate from which the strain is removed by filtration or centrifugation (filtrate or centrifugal supernatant), and ultrasonic treatment of the culture solution Or a cell lysate (lysate) obtained by treating the culture medium with a lysozyme, but is not limited thereto.
또한 본 발명에서 상기 발효물은 본 발명에 따른 신균주를 이용하여 발효시킨 발효물이라면 모두 포함할 수 있으며, 바람직하게는 상기 발효물은 증자된 전분질 원료에 본 발명의 신균주를 처리한 후, 발효시켜 수득한 발효물을 증류 처리한 다음, 알코올이 제거되고 남은 잔사물일 수 있다.In addition, in the present invention, the fermented product may include any fermented product fermented using the new strain according to the present invention, and preferably, the fermented product is treated with the new strain of the present invention in the increased starchy raw material, The fermented product obtained by fermentation may be distilled, and then the alcohol may be removed and a remaining residue.
본 발명의 상기 피부 미백 활성을 갖는 화장료 조성물은 화장수류, 크림류, 로션류, 팩류, 파운데이션류, 메이크업베이스류 등과 같은 다양한 제형으로 제조될 수 있으며, 구체적으로 액상, 크림상, 페이스트상, 고체상 등 다양한 성상으로 적용가능하다. 또한 통상적인 화장료 또는 피부연고 제조법을 사용하여 본 발명의 피부 미백 활성을 갖는 화장료 조성물을 제조할 수 있다.The cosmetic composition having the skin whitening activity of the present invention can be prepared in various formulations such as lotions, creams, lotions, packs, foundations, makeup bases, etc., specifically liquid, cream, paste, solid, etc. It can be applied in a variety of properties. In addition, a cosmetic composition having skin whitening activity of the present invention can be prepared using a conventional cosmetic or skin ointment preparation method.
한편, 본 발명의 피부 미백 활성을 갖는 화장료 조성물에 있어 본 발명의 유효성분의 함량은 조성물 총 중량 대비 0.0001 내지 20 중량%인 것이 바람직하다. 유효성분의 함량이 0.0001 중량% 미만인 경우에는 본래 목적하는 피부의 미백 효과를 충분하게 달성할 수 없어 바람직하지 못하며, 유효성분의 함량이 20 중량%를 초과하는 경우에는 조성물의 안정성에 문제를 발생시킬 수 있고, 이로 인해 조성물의 외관에 문제를 일으킬 우려가 있다.On the other hand, in the cosmetic composition having skin whitening activity of the present invention, the content of the active ingredient of the present invention is preferably 0.0001 to 20% by weight based on the total weight of the composition. If the content of the active ingredient is less than 0.0001% by weight, it is not desirable because the intended skin whitening effect cannot be sufficiently achieved.If the content of the active ingredient exceeds 20% by weight, it may cause problems in the stability of the composition. There is a risk of causing a problem in the appearance of the composition due to this.
또한 본 발명은 본 발명에 따른 신균주, 이의 파쇄물, 이의 배양물 또는 이의 발효물을 유효성분으로 포함하는 미백 활성을 갖는 식품 조성물을 제공할 수 있다. In addition, the present invention can provide a food composition having a whitening activity comprising a new strain according to the present invention, a lysate thereof, a culture product thereof, or a fermented product thereof as an active ingredient.
본 발명의 식품 조성물은 기능성 식품(functional food), 영양 보조제(nutritional supplement), 건강식품(health food) 및 식품 첨가제(food additives) 등의 모든 형태를 포함한다. 상기 유형의 식품 조성물은 당 업계에 공지된 통상적인 방법에 따라 다양한 형태로 제조할 수 있다.The food composition of the present invention includes all forms such as functional food, nutritional supplement, health food and food additives. Food compositions of this type can be prepared in various forms according to conventional methods known in the art.
예를 들면, 건강식품으로는 본 발명의 신균주들을 이용하여 차, 쥬스 및 드링크의 형태로 제조하여 음용하도록 하거나, 과립화, 캡슐화 및 분말화 하여 섭취할 수 있다. 또한, 미백 활성 또는 미백 활성의 증진 효과가 있다고 알려진 공지의 물질 또는 활성 성분과 함께 혼합하여 조성물의 형태로 제조할 수 있다.For example, as a health food, the new strains of the present invention may be prepared in the form of tea, juice, and drinks to be consumed, or granulated, encapsulated, and powdered to be consumed. In addition, it may be prepared in the form of a composition by mixing with a known substance or active ingredient known to have a whitening activity or a whitening activity enhancing effect.
또한, 기능성 식품으로는 음료(알코올성 음료 포함), 과실 및 그의 가공식품(예: 과일통조림, 병조림, 잼, 마아말레이드 등), 어류, 육류 및 그 가공식품(예: 햄, 소시지콘비이프 등), 빵류 및 면류(예: 우동, 메밀국수, 라면, 스파게티, 마카로니 등), 과즙, 각종 드링크, 쿠키, 엿, 유제품(예: 버터, 치이즈 등), 식용식물유지, 마아가린, 식물성 단백질, 레토르트 식품, 냉동식품, 각종 조미료(예: 된장, 간장, 소스 등) 등에 본 발명의 신균주, 이의 배양물, 이의 파쇄물 또는 이의 발효물을 첨가하여 제조할 수 있다.In addition, functional foods include beverages (including alcoholic beverages), fruits and processed foods thereof (eg, canned fruit, canned food, jam, marmalade, etc.), fish, meat and processed foods thereof (eg, ham, sausage corn beef, etc.) ), breads and noodles (e.g. udon, soba, ramen, spaghetti, macaroni, etc.), fruit juice, various drinks, cookies, syrup, dairy products (e.g. butter, cheese, etc.), edible vegetable oil, margarine, vegetable protein, Retort foods, frozen foods, various seasonings (eg, miso, soy sauce, sauce, etc.) can be prepared by adding a new strain of the present invention, a culture product thereof, a crushed product thereof, or a fermented product thereof.
본 발명의 식품 조성물 중 상기 본 발명의 유효성분에 대한 바람직한 함유량으로는 이에 한정되지 않지만 바람직하게는 최종적으로 제조된 식품 중 고형물 함량 0.01 내지 50 중량%이다. 또한 식품 첨가제의 형태로 사용하기 위해서는 본 발명의 조성물을 분말 또는 농축액 형태로 제조하여 사용할 수 있다.The preferred content of the active ingredient of the present invention in the food composition of the present invention is not limited thereto, but is preferably 0.01 to 50% by weight of the solid content in the finally prepared food. In addition, for use in the form of a food additive, the composition of the present invention may be prepared and used in the form of a powder or concentrate.
나아가 본 발명자들은 본 발명의 신균주를 이용한 미백 활성이 우수한 발효물을 제조하기 위한 연구를 진행하였는데, 그 결과, 전분질 원료를 기질로 하고 본 발명의 신균주를 이용하여 발효시킨 발효물이 다른 종류의 균주를 이용하여 발효시킨 발효물에 비해 더 우수한 미백 활성이 있는 것을 확인하였다.Further, the present inventors conducted a study to prepare a fermented product having excellent whitening activity using the new strain of the present invention. As a result, the fermented product fermented using the new strain of the present invention using a starchy raw material as a substrate is different. It was confirmed that it has better whitening activity than the fermented product fermented using the strain of.
따라서 본 발명은 본 발명에서 분리 및 동정한 상기 신균주인 사카로마이세스 세레비지애 EY2-27(KCCM12439P)를 이용하여 전분질 원료를 발효시키는 단계를 포함하는, 미백 활성을 갖는 발효물의 제조방법을 제공할 수 있다. Accordingly, the present invention provides a method for producing a fermented product having whitening activity, comprising fermenting a starchy raw material using Saccharomyces cerevisiae EY2-27 (KCCM12439P), the new strain isolated and identified in the present invention. Can provide.
구체적으로 상기 방법은, (1) 입국에 물 및 본 발명의 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주, 이의 파쇄물 또는 이의 배양물을 혼합하고 발효시켜 1차 발효물을 제조하는 단계; (2) 상기 1차 발효물에 증자된 전분질 원료 및 물을 혼합한 후 발효시켜 2차 발효물을 제조하는 단계; 및 (3) 상기 2차 발효물을 증류시키고 남은 잔사물을 수득하는 단계를 포함할 수 있다.Specifically, the method comprises the steps of: (1) mixing and fermenting water and Saccharomyces cerevisiae EY2-27 (KCCM12439P) strain of the present invention, a lysate or a culture thereof, to prepare a primary fermentation product ; (2) preparing a second fermented product by fermenting after mixing the starchy raw material and water steamed in the first fermented product; And (3) distilling the secondary fermented product and obtaining the remaining residue.
본 발명의 상기 방법을 보다 구체적으로 설명하면 다음과 같다.The method of the present invention will be described in more detail as follows.
먼저 (1) 입국에 물 및 본 발명의 사카로마이세스 세레비지애 EY2-27(KCCM12439P) 균주, 이의 파쇄물 또는 이의 배양물을 혼합하고 발효시켜 1차 발효물을 제조한다.First, (1) water and Saccharomyces cerevisiae EY2-27 (KCCM12439P) strain of the present invention, a lysate or a culture thereof are mixed and fermented to prepare a primary fermented product.
여기서 상기 입국은 증자된 쌀을 이용하여 통상적인 방법으로 제조한 입국을 사용할 수 있으며, 본 발명의 일실시예에서는 증자한 쌀에 균을 접종하고 항온항습기 내에서 33℃의 온도 및 습도 95%로 유지하면서 48시간 동안 균을 번식시켜 입국을 제조하여 사용하였다.Here, the entry may use an entry manufactured by a conventional method using increased rice, and in one embodiment of the present invention, inoculation of the bacteria in the increased rice and at a temperature of 33°C and a humidity of 95% in a thermo-hygrostat. While maintaining, the bacteria were propagated for 48 hours to prepare and use the immigration.
상기 1차 발효물은 입국에 물과 본 발명의 신균주, 이의 파쇄물 또는 이의 배양물을 첨가하고 발효시켜 수득할 수 있는데, 바람직하게는 상기 입국에 물과 본 발명의 신균주의 배양물을 첨가하고 20~25℃의 온도에서 4~6일간 발효시켜 수득할 수 있다.The primary fermentation product can be obtained by fermentation by adding water and a new strain of the present invention, a lysate thereof, or a culture thereof to entry into the country, preferably, adding water and a culture of the new strain of the present invention to the entry And can be obtained by fermentation for 4-6 days at a temperature of 20-25°C.
1차 발효물의 제조가 완료되면, 그 다음으로 1차 발효물에 증자된 전분질 원료 및 물을 첨가하고 혼합한 후 발효시켜 2차 발효물을 제조한다.When the production of the first fermented product is completed, the starchy raw material and water are then added to the first fermented product, mixed, and fermented to prepare a second fermented product.
이때 상기 전분질 원료로는 서류 또는 곡류를 사용할 수 있으며, 상기 서류로는 이에 제한되지는 않으나, 고구마 또는 감자를 사용할 수 있으며, 바람직하게는 고구마를 사용할 수 있다.In this case, documents or grains may be used as the starchy raw material, and the documents are not limited thereto, but sweet potatoes or potatoes may be used, and sweet potatoes may be preferably used.
상기 곡류로는 이에 제한되지는 않으나, 쌀, 찹쌀, 보리, 겉보기, 옥수수, 메밀, 귀리 또는 조를 사용할 수 있다.The grains are not limited thereto, but rice, glutinous rice, barley, apparent, corn, buckwheat, oats, or millet may be used.
상기 전분질 원료는 물로 세척 후, 2~4시간 동안 상온의 물에서 침지한 후, 40~60분 동안 스팀으로 증자하고, 20~30분 동안 상온에서 냉각시킨 것을 사용한다.The starchy raw material is washed with water, immersed in water at room temperature for 2 to 4 hours, steamed for 40 to 60 minutes, and cooled at room temperature for 20 to 30 minutes.
상기 2차 발효물의 제조를 위한 발효는 20~25℃의 온도에서 8~12일간 발효시킨다.Fermentation for the production of the secondary fermented product is fermented for 8 to 12 days at a temperature of 20 to 25°C.
이렇게 2차 발효물의 제조가 완료되면, 이후 상기 2차 발효물을 증류시키고 남은 잔사물을 수득하며, 이때 상기 증류는 2차 발효물에 함유된 알코올을 증류기를 이용하여 제거함으로써 남은 잔사물에는 알코올 성분이 1%(v/v) 미만이 되도록 한다.When the production of the secondary fermentation product is completed in this way, the secondary fermentation product is then distilled and the remaining residue is obtained. In this case, the distillation is performed by removing alcohol contained in the secondary fermentation product using a distiller. Make sure the component is less than 1% (v/v).
이상의 과정을 통해 제조된 본 발명의 신균주를 이용하여 수득한 발효물, 바람직하게는 발효물의 증류 잔사물은 멜라닌 생성 억제 활성이 우수하고 티로시나제 억제 활성이 우수하여 피부 미백을 효과적으로 증진시킬 수 있다.The fermented product obtained using the new strain of the present invention prepared through the above process, preferably the distilled residue of the fermented product, has excellent melanin production inhibitory activity and tyrosinase inhibitory activity, so that skin whitening can be effectively promoted.
또한, 본 발명의 미백 활성을 갖는 발효물의 증류 잔사물은 본 발명의 신균주 대신 다른 종류의 미생물(사카로마이세스속 균주, 락토바실러스속 균주)을 이용하여 제조된 잔사물에 비해 더 우수한 미백 활성이 있는 것을 확인할 수 있었다. 나아가 본 발명의 사카로마이세스 세레비지애 EY2-27 균주, 상기 균주를 이용하여 수득한 발효물 또는 상기 발효물의 증류 잔사물은 미백 활성을 갖는 주류의 제조에도 사용될 수 있으며, 상기 주류는 이에 제한되지는 않으나, 희석식 소주, 증류식 소주, 일반 증류주, 약주, 탁주, 맥주, 인삼주, 매실주 또는 과실주일 수 있다. In addition, the distilled residue of the fermentation having the whitening activity of the present invention is better than the residue prepared using other types of microorganisms (Saccharomyces strain, Lactobacillus strain) instead of the new strain of the present invention. It was confirmed that there is activity. Furthermore, the Saccharomyces cerevisiae EY2-27 strain of the present invention, the fermented product obtained by using the strain, or the distilled residue of the fermentation product may be used in the manufacture of alcoholic beverages having whitening activity, and the alcoholic beverage is limited thereto. Although it is not, it may be diluted soju, distilled soju, general spirits, yakju, takju, beer, ginseng wine, plum wine, or fruit wine.
이하, 실시예를 통하여 본 발명을 보다 상세히 설명하고자 한다. 이들 실시예는 본 발명을 보다 구체적으로 설명하기 위한 것으로, 본 발명의 범위가 이들 실시예에 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail through examples. These examples are for explaining the present invention more specifically, and the scope of the present invention is not limited to these examples.
<< 실시예Example 1> 1>
신균주의Neophyte 분리 및 동정 Separation and sympathy
본 발명자는 미백 활성이 우수한 신규한 균주를 분리 및 동정하기 위해 다음과 같은 실험을 수행하였다. 먼저 신균주의 분리 및 동정을 위한 과정으로, 곰팡이 균사 발육 억제와 세균 생육 억제를 위해 chloramphenicol 0.01%, sodium propionate 0.3%를 첨가하여 만든 PDA(Potato dextrose agar)배지에 희석한 고구마소주 술덧 0.1ml을 도말 한 후 30℃에서 24hr 배양하였다. 배양 후, 형성된 콜로니를 취하여 다시 획선 배양하여 오염되지 않은 순수한 단일 콜로니를 확보하였고, 확보된 균주로부터 18s rDNA의 분석을 통해 종래 존재하지 않았던 신규 균주를 분리하였다. The present inventors performed the following experiments to isolate and identify a novel strain having excellent whitening activity. First, as a process for the isolation and identification of new strains, 0.1 ml of sweet potato soju diluted in PDA (Potato dextrose agar) medium made by adding 0.01% chloramphenicol and 0.3% sodium propionate to inhibit the growth of fungal mycelium and inhibit the growth of bacteria. After smearing, it was incubated at 30° C. for 24 hours. After cultivation, the formed colonies were taken and cultivated again to obtain a pure single colony without contamination, and a new strain that did not exist before was isolated from the obtained strain through 18s rDNA analysis.
이때, 균주의 동정을 위한 18s rRNA 시퀀싱은 전문 기관인 ‘마크로젠’에 의뢰하여 진행하였으며 진행방법은 순수분리가 확인된 균주의 whole genomic DNA를 분리한 후 PCR을 이용하여 증폭하였다. PCR증폭에 사용된 primer는 ITS1 (TCCGTAGGTGAACCTGCGG)과 ITS4 (TCCTCCGCTTATTGATATGC)를 이용하였다. PCR 조건은 95℃에서 5분, 94℃에서 45초, 55℃에서 1분, 72℃에서 1분, 72℃에서 10초의 사이클을 총 35회 반복하였다. PCR prodect의 분석은 ABI 3730XL sequencing 기계를 이용하였다. 분석이 된 염기서열을 BioEdit seqencing 프로그램을 이용하여 alignment시킨 후 그 결과를 NCBI의 data base와 비교하여 동정하였다. At this time, 18s rRNA sequencing for strain identification was conducted by requesting a specialized agency, “Macrogen,” and the method was amplified using PCR after separating the whole genomic DNA of the strain confirmed to be pure. The primers used for PCR amplification were ITS1 (TCCGTAGGTGAACCTGCGG) and ITS4 (TCCTCCGCTTATTGATATGC). PCR conditions were 95°C for 5 minutes, 94°C for 45 seconds, 55°C for 1 minute, 72°C for 1 minute, 72°C for 10 seconds, a total of 35 cycles were repeated. PCR prodect analysis was performed using an ABI 3730XL sequencing machine. After the analyzed nucleotide sequence was aligned using the BioEdit seqencing program, the result was identified by comparing it with the NCBI data base.
분석 결과, 종래 규명되지 않았던 새로운 균주를 확인하였는데, 이를 본 발명자들은 Saccharomyces cerevisiae EY2-27로 명명하였고, 상기 균주의 18s rRNA에 대한 ITS1의 영역 및 ITS4 영역의 염기서열을 분석하여 이를 서열번호 1(ITS1) 및 2(ITS4)에 기재하였으며, 국제미생물기탁기관인 한국미생물보존센터(KCCM)에 2019년 02월 19일자로 기탁하여 기탁번호 KCCM12439P를 부여받았다.As a result of the analysis, a new strain, which was not previously identified, was identified, which the present inventors named Saccharomyces cerevisiae EY2-27, and analyzed the nucleotide sequences of the ITS1 region and the ITS4 region for the 18s rRNA of the strain, and this was obtained as SEQ ID NO: 1 ( It was described in ITS1) and 2 (ITS4), and was deposited with the Korea Microbial Conservation Center (KCCM), an international microbial deposit organization, on February 19, 2019, and was given the deposit number KCCM12439P.
여기서 상기 ITS (Internal Transcribed Spacer)는 염색체의 small-subunit ribosomal RNA(SSU rRNA)와 large-subunit rRNA(LSU rRNA) 유전자 사이에 위치한 DNA 영역을 의미하는 것으로, 진핵생물은 18S rRNA에 대한 특정 ITS1, ITS4 영역에 대한 염기서열 분석결과를 얻을 수 있고, 종래 알려진 균주와의 염기서열 유사성 분석을 통해 신균주 여부를 파악할 수 있다. Here, the ITS (Internal Transcribed Spacer) refers to a DNA region located between the small-subunit ribosomal RNA (SSU rRNA) and large-subunit rRNA (LSU rRNA) genes of the chromosome, and in eukaryotes, a specific ITS1 for 18S rRNA, The result of nucleotide sequence analysis for the ITS4 region can be obtained, and whether a new strain can be identified through nucleotide sequence similarity analysis with a conventionally known strain.
<< 실시예Example 2> 2>
본 발명의 Of the present invention 신균주를New strain 이용한 우수한 미백 활성을 갖는 Having excellent whitening activity 발효물의Fermented 제조 Produce
본 발명자들은 상기 실시예 1에서 분리 및 동정한 신균주인 Saccharomyces cerevisiae EY2-27를 이용하여 미백 활성이 우수한 발효물을 다음과 같은 방법으로 제조하였다.The present inventors prepared a fermented product having excellent whitening activity by using Saccharomyces cerevisiae EY2-27, a new strain isolated and identified in Example 1, as follows.
먼저 쌀에 균을 혼합 및 배양하여 입국을 제조하였다. 이때 입국은 증자된 쌀(10 kg)에 균(백국균 2g)을 접종하여 통상적인 방법에 따라 제조하였고, 증자한 쌀에 균을 접종하고 항온 항습기 내에서 33℃의 온도 및 습도 95%로 유지하면서 48시간 동안 균을 번식시켜 입국을 제조하였다. 이후 상기 제조된 입국에 본 발명에서 동정한 신균주의 각 배양액을 물과 혼합하고 발효시켜 1차 발효액을 제조하였다. 여1차 발효를 위해 상기에서 제조한 입국 513 g에 물 718 g을 첨가하여 혼합한 후, 입국 당화액에서 배양하여 108/ml 이상의 균주가 됨을 확인한 본 발명의 신균주 배양액 10 ml을 접종하였다. 본 발명의 효모 배양액인 입국 당화액은 상기 방법으로 제조된 입국 200g과 물 600 g을 섞은 후 65℃에서 3 시간 30 분 동안 교반하여 당화시켰다. 당화 후, 4,700 rpm에서 30 분간 원심분리한 후 여과포로 여과한 뒤 물로 10˚brix로 희석하여 제조하였다. 상기 혼합액을 25℃에서 5 일간 발효시켜 1차 발효액을 제조하였다.First, the rice was mixed and cultured to prepare an immigration. At this time, the entry was made according to the usual method by inoculating the increased rice (10 kg) with a fungus (2g of Bacillus bacteria), and inoculating the increased rice with the fungus and maintaining the temperature and humidity of 95% at 33℃ in a thermo-hygrostat. While carrying out the breeding for 48 hours, the immigration was prepared. Thereafter, each culture solution of the new strain identified in the present invention was mixed with water and fermented to prepare a first fermentation broth at the entry of the prepared country. For the primary fermentation, 718 g of water was added to 513 g of the above-prepared country for fermentation and mixed, and then 10 ml of the culture solution of the new strain of the present invention was inoculated, which was confirmed to be a strain of 10 8 /ml or more by culturing in the saccharification solution of the country . The immigration saccharification solution, which is the yeast culture solution of the present invention, was saccharified by mixing 200 g of immigration prepared by the above method and 600 g of water and stirring at 65° C. for 3 hours and 30 minutes. After saccharification, the mixture was centrifuged at 4,700 rpm for 30 minutes, filtered through a filter cloth, and diluted with water to 10˚ brix. The mixed solution was fermented at 25° C. for 5 days to prepare a first fermented solution.
상기 제조된 1차 발효액은 2차 발효를 수행하였는데, 즉, 상기 1차 발효액에 증자된 고구마 및 물을 첨가하여 혼합하고 발효시켜 2차 발효액을 제조하였다.The prepared primary fermentation broth was subjected to secondary fermentation, that is, steamed sweet potatoes and water were added to the primary fermentation broth, mixed, and fermented to prepare a secondary fermentation broth.
구체적으로, 2차 발효를 위해 먼저 고구마를 준비하는데, 이물질이나 오염원을 제거하기 위하여 물로 깨끗이 세척하고 수분의 충분한 흡수를 위해 3 시간 동안 상온의 물에 침지한 후 상압에서 60 분간 스팀으로 고구마를 증자하였다. 증자 후 20 분간 상온에서 냉각시켰고, 이후 상기에서 제조된 1차 발효액에 물 1,743 g 및 증자된 고구마 1,026 g를 첨가하고 혼합하여, 25℃에서 10 일간 발효시켜 본 발명의 신균주를 이용한 2차 발효액(고구마 술덧) 4,000 g을 제조하였다.Specifically, sweet potatoes are first prepared for the secondary fermentation. To remove foreign substances or contaminants, wash them clean with water, immerse them in water at room temperature for 3 hours for sufficient absorption of moisture, and increase the sweet potatoes with steam for 60 minutes at normal pressure. I did. After steaming, it was cooled at room temperature for 20 minutes, and then 1,743 g of water and 1,026 g of steamed sweet potatoes were added to the first fermentation broth prepared above, mixed, and fermented at 25° C. for 10 days to use the new strain (Sweet potato mash) 4,000 g was prepared.
이후, 2차 발효액을 증류시키고 남은 잔사물을 수득하였다. 구체적으로 상기 단계에서 수득한 2차 발효액을 단식 증류기를 이용하여 알코올을 제거하였는데, 내부온도는 40℃에서 110 mmHg로 감압 증류하였다. 감압 펌프는 수봉식 펌프를 이용하였다. 알코올을 제거하기 위한 증류기는 유리재 5 L용량의 라운드형 플라스크와 포말방지 증류 컬럼, 횡형 냉각사관, 감압 형성벨브가 달린 눈금있는 사이트 글래스, 유출액을 포집하는 리시버를 구비하는 증류기를 사용하였다. 알코올 제거를 위한 초기 발효액량은 2 L, 초기 발효액 온도는 25℃, 유출 증기의 냉각을 위한 냉각기의 설정온도는 5℃가 되게 하였고, 15 L/분의 속도로 순환하게 하였다. 알코올 제거기에 투입되는 열로써 알코올 제거 속도를 조절하였다. 알코올을 제거하고 수득한 잔사물 중의 알코올 성분은 1% v/v 미만이 되게 하였다. 이와 같은 방법으로 본 발명의 신균주를 이용한 미백 활성을 갖는 고구마 발효물로서, 고구마 발효물의 증류 잔사물을 수득하였다.Thereafter, the secondary fermentation broth was distilled to obtain the remaining residue. Specifically, alcohol was removed from the secondary fermentation broth obtained in the above step using a single-type still, and the internal temperature was distilled under reduced pressure to 110 mmHg at 40°C. The pressure reducing pump used a water ring pump. The distiller for removing alcohol was a distillation machine equipped with a 5 L glass round flask, an anti-foam distillation column, a horizontal cooling tube, a graduated sight glass with a pressure reducing valve, and a receiver for collecting the effluent. The initial fermentation broth amount for alcohol removal was 2 L, the initial fermentation broth temperature was 25°C, and the set temperature of the cooler for cooling the effluent steam was 5°C, and circulated at a rate of 15 L/min. The alcohol removal rate was controlled by heat input to the alcohol remover. The alcohol was removed and the alcohol component in the obtained residue was made to be less than 1% v/v. In this way, as a fermented sweet potato having whitening activity using the new strain of the present invention, a distilled residue of the fermented sweet potato was obtained.
<< 실시예Example 3> 3>
미백 활성 분석Whitening activity assay
상기 실시예 2를 통해 제조한 본 발명의 신균주를 이용한 고구마 발효물의 증류 잔사물에 대한 미백 활성 분석을 수행하였다. 미백 활성 분석은 티로시나제 활성 억제 효능분석과 멜라닌 생성 억제 효능 분석으로 수행하였다. 이때 대조군으로는 본 발명의 신균주를 첨가하지 않은 군을 사용하였고, 비교군으로는 본 발명의 신균주가 아닌 다른 사카로마이세스 세레비지애 균주를 이용하여 제조한 고구마 발효물의 증류 잔사물군을 사용하였다. 실험에 사용한 각 실험군은 하기 표에 나타내었다.A whitening activity analysis was performed on the distilled residue of the fermented sweet potato product using the new strain of the present invention prepared in Example 2. The whitening activity analysis was performed by the tyrosinase activity inhibition efficacy analysis and the melanin production inhibition efficacy analysis. At this time, a group to which the new strain of the present invention was not added was used as a control group, and a group of distilled residues of sweet potato fermentation prepared using a Saccharomyces cerevisiae strain other than the new strain of the present invention was used as a control group. Was used. Each experimental group used in the experiment is shown in the table below.
<3-1> <3-1> 티로시나제Tyrosinase 활성 억제효능 분석 Activity inhibitory effect analysis
티로시나제 활성 억제 효능 분석은 다음과 같은 방법으로 수행하였다. 먼저 튜브에 20 ml의 0.1 M potassium phosphate 버퍼를 넣고, 여기에 0.02 mg/ml mushroom tyrosinase를 20 ml 넣고 첨가한 후, 본 발명의 증류 잔사물을 농도별(250ug/ml, 500ug/ml, 1000ug/ml)로 40 ml 첨가하여 25℃에서 10분간 반응시켰으며, 기질로 1 mM L-3,4-dihydroxyphenylalanine (L-DOPA)를 40ml 넣고 25℃ 에서 30분간 반응시켰다. 이후 475 nm 파장영역에서 흡광도를 측정하였으며, 이때 상기 과정을 총 3회 반복하였고 평균값을 측정하여 티로시나제 활성 억제 정도를 분석하였다.The analysis of the tyrosinase activity inhibition efficacy was performed in the following manner. First, 20 ml of 0.1 M potassium phosphate buffer was added to the tube, and 20 ml of 0.02 mg/ml mushroom tyrosinase was added thereto, and then the distillation residue of the present invention was added by concentration (250ug/ml, 500ug/ml, 1000ug/ ml) was added and reacted at 25° C. for 10 minutes, and 40 ml of 1 mM L-3,4-dihydroxyphenylalanine (L-DOPA) was added as a substrate and reacted at 25° C. for 30 minutes. Thereafter, absorbance was measured in a wavelength region of 475 nm. At this time, the above process was repeated three times, and the average value was measured to analyze the degree of inhibition of tyrosinase activity.
<3-2> 멜라닌 생성 억제효능 분석<3-2> Analysis of melanin production inhibitory effect
멜라닌 생성 세포주 B16F10 세포를 6웰 플레이트 2 x 105 cells/well 농도로 배양하고, 배양 12시간 후 상기 본 발명의 증류 잔사물을 농도별(250ug/ml, 500ug/ml, 1000ug/ml)로 처리하였다. 증류 잔사물 처리 1시간 후 멜라닌 생성을 유도하는 a-melanocyte-stimulating hormone (a-MSH)를 100 nM 농도로 처리한 다음, 72시간 후 세포 배양액을 튜브에 옮기고 상온에서 48시간 반응시켰다. 반응완료 후 400 nm 파장영역에서 흡광도를 측정하였으며, 이때 상기 과정을 총 3회 반복하였고 평균값을 측정하여 멜라닌 생성 억제 정도를 분석하였다.The melanin-producing cell line B16F10 cells were cultured in a 6-well plate at a concentration of 2 x 10 5 cells/well, and after 12 hours of culture, the distillation residue of the present invention was treated at different concentrations (250 ug/ml, 500 ug/ml, 1000 ug/ml). I did. After 1 hour of treatment with the distillation residue, a-melanocyte-stimulating hormone (a-MSH), which induces melanogenesis, was treated at a concentration of 100 nM, and after 72 hours, the cell culture solution was transferred to a tube and reacted at room temperature for 48 hours. After completion of the reaction, the absorbance was measured in a wavelength region of 400 nm. At this time, the above process was repeated three times, and the average value was measured to analyze the degree of inhibition of melanin production.
분석 결과, 도 1에 나타낸 바와 같이, 티로시나제 활성 억제 효능은 본 발명의 신균주를 이용하여 수득한 증류 잔사물이 본 발명의 균주를 첨가하지 않고 수득한 대조군 잔사물에 비해 우수한 것으로 나타났고, 나아가 본 발명의 균주 처리 군은 비교군으로 사용한 다른 균주인 사카로마이세스 세레비지애 처리 군들에 비해서도 티로시나제 억제 활성이 더 우수한 것으로 나타났다.As a result of the analysis, as shown in Fig. 1, the distillation residue obtained using the new strain of the present invention was found to be superior to the control residue obtained without adding the strain of the present invention. The strain-treated group of the present invention was found to have more excellent tyrosinase inhibitory activity compared to the Saccharomyces cerevisiae-treated group, which is another strain used as a comparative group.
또한, 도 2에서는 멜라닌 생성 억제 분석 결과를 확인할 수 있었는데, 본 발명의 균주를 첨가하지 않은 대조군 잔사물에서는 멜라닌 생성량이 증가되어 있는 것으로 나타난 반면, 본 발명의 균주 처리 군에서는 멜라닌 생성량이 현저하게 감소되어 있는 것으로 나타났고, 이러한 멜라닌 감소 정도는 비교군으로 사용한 다른 균주인 사카로마이세스 세레비지애 처리 군에 비해서도 더 우수한 것으로 나타났다.In addition, in FIG. 2, it was possible to confirm the results of the analysis of inhibition of melanin production, whereas the amount of melanin production was increased in the control residue to which the strain of the present invention was not added, whereas the amount of melanin production was significantly reduced in the strain-treated group of the present invention. It was found that the degree of melanin reduction was better than that of the Saccharomyces cerevisiae treatment group, which is another strain used as a control group.
이상의 결과를 통해 본 발명에서 동정한 신균주 및 상기 신균주로 전분질 원료를 발효시켜 수득한 발효물의 증류 잔사물은 우수한 피부 미백 활성을 가지고 있는 바, 피부 미백용 화장품 및 기능성 식품의 제조에 유용하게 사용할 수 있음을 알 수 있었다.Through the above results, the new strains identified in the present invention and the distilled residues of the fermented product obtained by fermenting starchy raw materials with the new strains have excellent skin whitening activity, and are useful in the manufacture of cosmetics and functional foods for skin whitening. I could see that it could be used.
이제까지 본 발명에 대하여 그 바람직한 실시예들을 중심으로 살펴보았다. 본 발명이 속하는 기술 분야에서 통상의 지식을 가진 자는 본 발명이 본 발명의 본질적인 특성에서 벗어나지 않는 범위에서 변형된 형태로 구현될 수 있음을 이해할 수 있을 것이다. 그러므로 개시된 실시예들은 한정적인 관점이 아니라 설명적인 관점에서 고려되어야 한다. 본 발명의 범위는 전술한 설명이 아니라 특허청구범위에 나타나 있으며, 그와 동등한 범위 내에 있는 모든 차이점은 본 발명에 포함된 것으로 해석되어야 할 것이다.So far, the present invention has been looked at around its preferred embodiments. Those of ordinary skill in the art to which the present invention pertains will be able to understand that the present invention can be implemented in a modified form without departing from the essential characteristics of the present invention. Therefore, the disclosed embodiments should be considered from an illustrative point of view rather than a limiting point of view. The scope of the present invention is shown in the claims rather than the above description, and all differences within the scope equivalent thereto should be construed as being included in the present invention.
<110> Korea Food Research Institute <120> Novel strain Saccharomyces cerevisiae EY2-27 having skin whitening and use of the same <130> NPDC78399 <160> 2 <170> KoPatentIn 3.0 <210> 1 <211> 828 <212> DNA <213> Artificial Sequence <220> <223> Saccharomyces cerevisiae(EY2-27) ITS1 sequence <400> 1 tggacaacta gcggaggact agcaatcgga cgtatggttt ggtttgggaa gagcatgaga 60 gcttttactg ggcaaaaaga caaaagaggg aaagtccagc cgggcctgcg cttaagtgcg 120 cggtcttgct aggcttgtaa gtttctttct tgctattcca aacggtgaaa gatttctgtg 180 cttttgttat aggacaatta aaaccgtttc aatacaacac actgtggagt tttcatatct 240 ttgcaacttt ttctttgggc attcaagcaa tcggggccca aaggtaacaa acacaaacaa 300 ttttatttat tcattaaatt tttgtcaaaa acaaaaattt tcgtaactgg aaattttaaa 360 atattaaaaa ctttcaacaa cggatctctt ggttctcgca tcgatgaaaa acgcagcgaa 420 atgcgatacg taatgtgaat tgcaaaattc cgtgaatcat caaatctttg aacgcacatt 480 gcgccccttg gtattccagg gggcatgcct gtttgagcgt catttccttc tcaaacattc 540 tgtttggtag tgagtgatac tctttgaagt taacttgaaa ttgctggcct tttcattgga 600 tgtttttttt tcaaagagag gtttctctgc gtgcttgagg taaatgcaat acggccgttt 660 taggtttaca actgcggcta tcttttttta ctgagcgtat tggaacgtat cgataaaaaa 720 gagcgctagg caacatgttc taaagttgac tcaatcagga ggataccgtg actattatta 780 atggggggga gaaaaaggcg cccccctttt ttcttattaa aaaagttt 828 <210> 2 <211> 862 <212> DNA <213> Artificial Sequence <220> <223> Saccharomyces cerevisiae(EY2-27) ITS4 sequence <400> 2 atgatggtac atacggcgtg gtttgaggcc tccttttata gcattgttcg cctagacgct 60 ctcttcttat cgataacgtt ccaatacgct cagtataaaa aagattagcc gcagttggta 120 aaacctaaaa cgaccgtact tgcattatac ctcaagcacg cagagaaacc tctctttgga 180 aaaaaaaaca tccaatgaaa aggccagcaa tttcaagtta actccaaaga gtatcactca 240 ctaccaaaca gaatgtttga gaaggaaatg acgctcaaac aggcatgccc cctggaatac 300 caaggggcgc aatgtgcgtt caaagattca atgattcacg gaattctgca attcacatta 360 cgtatcgcat ttcgctgcgt tcttcatcga tgcgagaacc aaaagatccg ttgttgaaag 420 tttttaatat tttaaaattt ccagttacga aaattcttgt ttttgacaaa aatttaatga 480 ataaataaaa ttgtttgtgt ttgttacctc tgggccccga ttgctcgaat gcccaaagaa 540 aaagttgcaa agatatgaaa actccacagt gtgttgtatt gaaacggttt taattgtcct 600 ataacaaaag cacagaaatc tctcaccgtt tggaatagca agaaagaaac ttacaagcct 660 agcaagaccg cgcacttaag cgcaggcccg gctggactct ccatctcttg tcttcttgcc 720 cagtaaaagc tctcatgctc ttgccaaaca aaaaaatcca ttttcaaaat ataaatttct 780 taatgacctc aggccccccc cccccaaaaa aggtgggggg gggggttaaa aaattttaaa 840 aagtttaaaa tgggattgga tt 862 <110> Korea Food Research Institute <120> Novel strain Saccharomyces cerevisiae EY2-27 having skin whitening and use of the same <130> NPDC78399 <160> 2 <170> KoPatentIn 3.0 <210> 1 <211> 828 <212> DNA <213> Artificial Sequence <220> <223> Saccharomyces cerevisiae (EY2-27) ITS1 sequence <400> 1 tggacaacta gcggaggact agcaatcgga cgtatggttt ggtttgggaa gagcatgaga 60 gcttttactg ggcaaaaaga caaaagaggg aaagtccagc cgggcctgcg cttaagtgcg 120 cggtcttgct aggcttgtaa gtttctttct tgctattcca aacggtgaaa gatttctgtg 180 cttttgttat aggacaatta aaaccgtttc aatacaacac actgtggagt tttcatatct 240 ttgcaacttt ttctttgggc attcaagcaa tcggggccca aaggtaacaa acacaaacaa 300 ttttatttat tcattaaatt tttgtcaaaa acaaaaattt tcgtaactgg aaattttaaa 360 atattaaaaa ctttcaacaa cggatctctt ggttctcgca tcgatgaaaa acgcagcgaa 420 atgcgatacg taatgtgaat tgcaaaattc cgtgaatcat caaatctttg aacgcacatt 480 gcgccccttg gtattccagg gggcatgcct gtttgagcgt catttccttc tcaaacattc 540 tgtttggtag tgagtgatac tctttgaagt taacttgaaa ttgctggcct tttcattgga 600 tgtttttttt tcaaagagag gtttctctgc gtgcttgagg taaatgcaat acggccgttt 660 taggtttaca actgcggcta tcttttttta ctgagcgtat tggaacgtat cgataaaaaa 720 gagcgctagg caacatgttc taaagttgac tcaatcagga ggataccgtg actattatta 780 atggggggga gaaaaaggcg cccccctttt ttcttattaa aaaagttt 828 <210> 2 <211> 862 <212> DNA <213> Artificial Sequence <220> <223> Saccharomyces cerevisiae (EY2-27) ITS4 sequence <400> 2 atgatggtac atacggcgtg gtttgaggcc tccttttata gcattgttcg cctagacgct 60 ctcttcttat cgataacgtt ccaatacgct cagtataaaa aagattagcc gcagttggta 120 aaacctaaaa cgaccgtact tgcattatac ctcaagcacg cagagaaacc tctctttgga 180 aaaaaaaaca tccaatgaaa aggccagcaa tttcaagtta actccaaaga gtatcactca 240 ctaccaaaca gaatgtttga gaaggaaatg acgctcaaac aggcatgccc cctggaatac 300 caaggggcgc aatgtgcgtt caaagattca atgattcacg gaattctgca attcacatta 360 cgtatcgcat ttcgctgcgt tcttcatcga tgcgagaacc aaaagatccg ttgttgaaag 420 tttttaatat tttaaaattt ccagttacga aaattcttgt ttttgacaaa aatttaatga 480 ataaataaaa ttgtttgtgt ttgttacctc tgggccccga ttgctcgaat gcccaaagaa 540 aaagttgcaa agatatgaaa actccacagt gtgttgtatt gaaacggttt taattgtcct 600 ataacaaaag cacagaaatc tctcaccgtt tggaatagca agaaagaaac ttacaagcct 660 agcaagaccg cgcacttaag cgcaggcccg gctggactct ccatctcttg tcttcttgcc 720 cagtaaaagc tctcatgctc ttgccaaaca aaaaaatcca ttttcaaaat ataaatttct 780 taatgacctc aggccccccc cccccaaaaa aggtgggggg gggggttaaa aaattttaaa 840 aagtttaaaa tgggattgga tt 862
Claims (12)
(2) 상기 1차 발효물에 증자된 고구마 원료 및 물을 혼합한 후 발효시켜 2차 발효물을 제조하는 단계; 및
(3) 상기 2차 발효물을 증류시키고 남은 잔사물을 수득하는 단계를 포함하는,
미백 활성을 갖는 고구마 발효물의 제조방법.(1) mixing and fermenting water and Saccharomyces cerevisiae EY2-27 (Accession No.: KCCM 12439P) strain, its lysate or its culture at entry to prepare a primary fermented product;
(2) mixing the steamed sweet potato raw material and water in the first fermented product and then fermenting it to prepare a second fermented product; And
(3) comprising the step of distilling the secondary fermented product and obtaining the remaining residue,
Method for producing fermented sweet potato products having whitening activity.
상기 (1) 단계에서 상기 발효는 20~25℃의 온도에서 4~6일간 발효시키는 것을 특징으로 하는, 미백 활성을 갖는 고구마 발효물의 제조방법.The method of claim 6,
In the step (1), the fermentation is characterized in that the fermentation is carried out for 4 to 6 days at a temperature of 20 to 25°C.
상기 (2) 단계에서 고구마는 물로 세척 후, 2~4시간 동안 상온의 물에서 침지한 후, 40~60분 동안 스팀으로 증자하고, 20~30분 동안 상온에서 냉각시킨 것을 특징으로 하는, 미백 활성을 갖는 고구마 발효물의 제조방법.The method of claim 6,
In the step (2), the sweet potato is washed with water, immersed in water at room temperature for 2 to 4 hours, then steamed for 40 to 60 minutes, and cooled at room temperature for 20 to 30 minutes, whitening A method for producing fermented sweet potato products having activity.
상기 (2) 단계에서 2차 발효는 20~25℃의 온도에서 8~12일간 발효시키는 것을 특징으로 하는, 미백 활성을 갖는 고구마 발효물의 제조방법.The method of claim 6,
The second fermentation in the step (2) is characterized in that the fermentation is performed for 8 to 12 days at a temperature of 20 to 25°C. A method of producing a fermented sweet potato having whitening activity.
상기 (3) 단계에서 증류는 상기 2차 발효물에 함유된 알코올을 증류기를 이용하여 제거하되, 증류 후 남은 잔사물에 함유된 알코올 성분이 1%(v/v) 미만이 되도록 수행하는 것을 특징으로 하는, 미백 활성을 갖는 고구마 발효물의 제조방법.The method of claim 6,
In the step (3), the distillation is performed so that the alcohol contained in the secondary fermentation product is removed using a still, but the alcohol component contained in the residue remaining after distillation is less than 1% (v/v). To, a method for producing a fermented sweet potato having a whitening activity.
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