KR102139000B1 - Antioxidant, anti-wrinkle, whitening and antibiotic cosmetic composition comprising fermented Methyl Sulfonyl methane extract and the preparation method of the same - Google Patents

Antioxidant, anti-wrinkle, whitening and antibiotic cosmetic composition comprising fermented Methyl Sulfonyl methane extract and the preparation method of the same Download PDF

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KR102139000B1
KR102139000B1 KR1020190019833A KR20190019833A KR102139000B1 KR 102139000 B1 KR102139000 B1 KR 102139000B1 KR 1020190019833 A KR1020190019833 A KR 1020190019833A KR 20190019833 A KR20190019833 A KR 20190019833A KR 102139000 B1 KR102139000 B1 KR 102139000B1
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dietary sulfur
extract
fermented
msm
sulfur
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KR1020190019833A
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Korean (ko)
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이지은
김진구
신용산
노낙경
장문식
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주식회사 리딩프로스
주식회사 리더스코스메틱
신용산
주식회사 더가든오브내추럴솔루션
노낙경
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/46—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing sulfur
    • A61K8/466—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing sulfur containing sulfonic acid derivatives; Salts
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q17/00—Barrier preparations; Preparations brought into direct contact with the skin for affording protection against external influences, e.g. sunlight, X-rays or other harmful rays, corrosive materials, bacteria or insect stings
    • A61Q17/005—Antimicrobial preparations
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/08—Anti-ageing preparations
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
    • A61K2800/52—Stabilizers
    • A61K2800/522—Antioxidants; Radical scavengers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/80—Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
    • A61K2800/85—Products or compounds obtained by fermentation, e.g. yoghurt, beer, wine

Abstract

The present application relates to a fermented dietary sulfur extract (MSM-F10) having excellent anti-wrinkle, antioxidant, whitening, and antibacterial effects and a manufacturing method thereof. The manufacturing method of the fermented dietary sulfur extract (MSM-F10) includes the following steps of: obtaining a sterilized dietary sulfur lysate; obtaining a fermented dietary sulfur; and obtaining the fermented dietary sulfur extract.

Description

발효 식이유황 추출물을 함유하는 항산화, 항주름, 미백 및 항균 기능성 화장료 조성물 및 그 제조방법{Antioxidant, anti-wrinkle, whitening and antibiotic cosmetic composition comprising fermented Methyl Sulfonyl methane extract and the preparation method of the same}Antioxidant, anti-wrinkle, whitening and antibiotic cosmetic composition comprising fermented Methyl Sulfonyl methane extract and the preparation method of the same}

본 출원은 발효 식이유황 추출물(MSM-F10) 및 그 제조방법과 상기 추출물을 함유하는 화장료 조성물에 관한 것이다.The present application relates to a fermented dietary sulfur extract (MSM-F10), a method for manufacturing the same, and a cosmetic composition containing the extract.

천연 식이유황 (MSM, methyl sulfonyl methane)은 모든 살아있는 유기체에서 발견되는 유기 유황의 자연 형태로서 34% 이상의 황을 함유하며 식물에서 추출해 낸 무독성 천연 물질로서 이 화학물의 분자식은 (CH3)2SO2로 표시된다. 무독성의 백색 결정인 식이유황은 인체의 피부, 뼈, 머리카락에 다량 존재하는데 머리카락이 윤기가 없고, 피부가 주름이 지고, 손톱과 발톱이 딱딱하게 각질화되는 이러한 현상들은 모두 유황 성분이 현격하게 결핍되어 가기 때문이며, 그만큼 유황성분이 머리카락, 뼈, 피부조직에 깊이 관여하고 있다.Natural dietary sulfur (MSM, methyl sulfonyl methane) is a natural form of organic sulfur found in all living organisms. It contains more than 34% sulfur and is a non-toxic natural substance extracted from plants. Its molecular formula is (CH 3 ) 2 SO 2 It is indicated by. Dietary sulfur, a non-toxic white crystal, is present in human skin, bones, and hair in large quantities, but all of these phenomena in which the hair is not shiny, the skin is wrinkled, and the nails and toenails are hardly keratinized are all lacking sulfur components. Because it goes, the sulfur component is deeply involved in hair, bone, and skin tissue.

유황은 크게 광물성, 동물성, 식물성 유황으로 분류된다. 광물성 유황은 우리가 흔이 아는 유황으로 화공약품으로 대표적인 것이 황토이다. 주로 유황오리, 유황온천, 비료, 성냥이나 화약의 주원료이다. 동물성 유황은 웅담, 우황, 녹용, 사향 등 모든 동물들의 쓸개가 동물성 유황이다. 마지막으로 식물성 유황은 Natural organinc sulfur로 소나무, 은행나무, 오가피, 뽕나무, 헛개나무, 해초류 등에서 추출 정제하여 수득한다.Sulfur is largely classified into mineral, animal, and vegetable sulfur. Mineral sulfur is a sulfur that we commonly know, and representative of chemicals is ocher. Mainly raw materials for sulfur duck, sulfur hot spring, fertilizer, matches or gunpowder. Animal sulfur is animal gallbladder of all animals, including ungdam, wu-wang, antler, musk. Finally, vegetable sulfur is natural organinc sulfur, which is obtained by extracting and purifying from pine, ginkgo, oak, mulberry, hut, and seaweed.

유황의 이점으로 과거에 국부자극제, 피부질환, 신경마비 등을 치료하는 약제로 사용되었음이 알려져 있고, 관절염, 류마티즘 등에 도움이 되는 건강보조식품으로 사용되고 있다.It is known that it has been used as a medicine to treat local irritants, skin diseases, and nerve paralysis in the past as an advantage of sulfur, and is used as a health supplement to help arthritis and rheumatism.

종래 공지된 선행기술문헌으로는 대한민국 등록특허 제 10-1454874에서 식이유황을 함유하는 항균 화장품 조성물이 개시되었고, 대한민국 등록특허 제 10-1866065에는 식물성 식이유황 추출물 제조방법과 이를 함유하는 식초음료가 그리고 등록특허 제10-1874830호에는 규산염광물 호마노석과 식이유황을 함유하는 항균 화장료 조성물이 각각 개시되었으나, 식이유황을 바실러스속 세균으로 발효시켜 추출한 발효 식이유황 추출물(MSM-F10)의 항산화, 항균, 항주름 및 미백 효과에 대하여는 상기 문헌 어디에도 암시되거나 개시된 바가 없다. 이러한 배경 하에, 본 발명자들은 식이유황만을 원료로 하여 항산화, 항균, 항주름 및 미백 효과가 뛰어난 발효 추출물을 함유하는 화장료 조성물을 개발하기 위하여 예의 연구 노력한 결과, 식이유황의 바실러스 발효 추출물(MSM-F10)을 함유하는 화장료 조성물을 제조하고 효과를 확인함으로써 본 출원을 완성하였다.As a prior art document known in the prior art, an antibacterial cosmetic composition containing dietary sulfur was disclosed in Korean Patent Registration No. 10-1454874, and Korean Patent Registration No. 10-1866065 discloses a method for preparing vegetable dietary sulfur extract and a vinegar beverage containing the same. Patent registration No. 10-1874830 discloses an antibacterial cosmetic composition containing silicate minerals, homanite and dietary sulfur, respectively, but anti-oxidation, antibacterial, and anti-bacterial properties of fermented dietary sulfur extract (MSM-F10) extracted by fermenting dietary sulfur as Bacillus bacteria. No anti-wrinkle and whitening effects have been implied or disclosed anywhere in the literature. Under these backgrounds, the present inventors conducted a thorough research to develop a cosmetic composition containing fermented extract having excellent antioxidant, antibacterial, anti-wrinkle and whitening effects using only dietary sulfur as a raw material, and Bacillus fermented extract of dietary sulfur (MSM-F10 ) And completed the present application by preparing a cosmetic composition and confirming the effect.

본 출원의 목적은 발효 식이유황 추출물(MSM-F10)의 제조방법 및 상기 방법으로 제조된 발효 식이유황 추출물(MSM-F10)을 제공하는 데에 있다.The purpose of the present application is to provide a method for producing a fermented dietary sulfur extract (MSM-F10) and a fermented dietary sulfur extract (MSM-F10) prepared by the above method.

본 출원의 다른 목적은 발효 식이유황 추출물(MSM-F10)을 유효성분으로 포함하는 항주름, 항산화, 미백, 항균 기능성 화장료 조성물을 제공하는 데에 있다.Another object of the present application is to provide an anti-wrinkle, antioxidant, whitening, antibacterial functional cosmetic composition comprising fermented dietary sulfur extract (MSM-F10) as an active ingredient.

본 출원의 다른 목적 및 이점은 첨부한 청구범위 및 도면과 함께 하기의 상세한 설명에 의해 보다 명확해질 것이다. 본 명세서에 기재되지 않은 내용은 본 출원의 기술 분야 또는 유사 분야에서 숙련된 자이면 충분히 인식하고 유추할 수 있는 것이므로 그 설명을 생략한다.Other objects and advantages of the present application will become more apparent by the following detailed description in conjunction with the appended claims and drawings. The contents not described in this specification can be sufficiently recognized and inferred by those skilled in the technical field or similar field of the present application, and thus the description thereof will be omitted.

이하, 본 출원 내용에 대하여 구체적으로 설명하면 다음과 같다. 한편, 본 출원에서 개시한 일 실시 양태의 설명 및 실시 형태는 공통된 사항에 대하여다른 양태의 설명 및 실시 형태에도 적용될 수 있다. 또한, 본 출원에서 개시된다양한 요소들의 모든 조합이 본 출원의 범주에 속한다. 더불어, 하기 기술된 구체적인 서술에 의하여 본 출원의 범주가 제한된다고 볼 수 없다.Hereinafter, the contents of the present application will be described in detail as follows. On the other hand, the description and embodiments of one embodiment disclosed in the present application may also be applied to the description and embodiments of other embodiments with respect to common matters. In addition, all combinations of the various elements disclosed in this application are within the scope of this application. In addition, the scope of the present application is not considered to be limited by the specific descriptions described below.

본 출원의 목적을 달성하기 위하여 본 출원은 발효 식이유황 추출물(MSM-F10)의 제조방법 및 상기 제조방법에 따라 제조된 발효 식이유황 추출물(MSM-F10)을 제공한다(도 1). 이는 식이유황(Methyl Sulfonyl Methane)을 50배의 정제수에 혼합한 후 멸균하여 멸균된 식이유황 용해물을 수득하는 단계와; 상기 멸균된 식이유황 용해물에 Bacillus Subtilis 균주를 접종 후 30℃, 120rpm에서 1일 배양하여 발효시킨 다음 식이유황 발효물을 수득하는 단계와; 상기에서 수득한 식이유황 발효물을 정제수, 메탄올, 에탄올, 글리세린, 에틸아세테이트, 부틸렌글리콜, 프로필렌글리콜, 디클로로메탄, 클로로포름, 에틸에테르, 부틸렌글리콜, 헥산 및 이의 혼합물로 이루어지는 군으로부터 선택되는 용매를 식이유황 발효물의 50배(V/V) 사용하여 추출함으로써 발효 식이유황 추출물을 수득하는 단계로 이루어지는 것이 특징인 발효 식이유황 추출물(MSM-F10)의 제조방법으로 제조되는 것이 특징이다. In order to achieve the object of the present application, the present application provides a method for preparing fermented dietary sulfur extract (MSM-F10) and a fermented dietary sulfur extract (MSM-F10) prepared according to the method (FIG. 1). This is a step of mixing the dietary sulfur (Methyl Sulfonyl Methane) with 50 times purified water and sterilizing to obtain a sterilized dietary sulfur lysate; Bacillus Subtilis strain inoculated into the sterilized dietary sulfur lysate and cultured for 1 day at 30°C and 120 rpm, followed by fermentation to obtain dietary sulfur fermentation product; The dietary sulfur fermentation product obtained above is a solvent selected from the group consisting of purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane, chloroform, ethyl ether, butylene glycol, hexane and mixtures thereof. It is characterized in that it is produced by the production method of fermented dietary sulfur extract (MSM-F10) characterized in that it consists of the step of obtaining a fermented dietary sulfur extract by extracting using 50 times (V/V) of the dietary sulfur fermentation.

상기 식이유황(Methyl Sulfonyl methane)의 멸균은 121 ℃에서 5분 내지 25분, 바람직하게는 10분 내지 20분, 더 바람직하게는 15분 멸균하고 외부 균의 접촉이 없도록 유지하여 멸균된 식이유황 용해물을 수득하는 단계와; 상기 멸균된 식이유황 용해물에 104 내지 109, 바람직하게는 105 내지 108, 더욱 바람직하게는 106 내지 107 cell/ml의 일정 농도의 Bacillus Subtilis 균주를 1%(V/V) 접종 후, 30℃, 120rpm에서 1일 내지 3일, 바람직하게는 1일 배양하여 발효시켜 발효 식이유황 용해물을 수득하는 단계와; 상기 발효 식이유황 용해물을 정제수, 메탄올, 에탄올, 글리세린, 에틸아세테이트, 부틸렌글리콜, 프로필렌글리콜, 디클로로메탄, 클로로포름, 에틸에테르, 부틸렌글리콜, 헥산 및 이의 혼합물로 이루어지는 군으로부터 선택되는 용매를 발효 식이유황 용해물의 20 내지 80배, 바람직하게는 40 내지 60배, 더욱 바람직하게는 50배(V/V) 사용하여 추출함으로써 발효 식이유황 추출물을 수득하는 단계로 이루어지는 제조방법으로 제조되는 것이 특징이다. 상기 용매의 종류 및 추출시 필요한 용량은 추출물의 농도, 제형 및 용도에 따라 선택하여 추출할 수 있다.Sterilization of the dietary sulfur (Methyl Sulfonyl methane) is 5 minutes to 25 minutes at 121 ℃, preferably 10 minutes to 20 minutes, more preferably 15 minutes to sterilize and maintain the absence of contact of external bacteria for sterilized dietary sulfur Obtaining a seafood; 10 4 to 10 9, preferably 10 5 to 10 8, more preferably from 10 6 to 10 1% of the Bacillus Subtilis strain in a concentration of 7 cell / ml in the lysate the sterilized diet sulfur (V / V) After inoculation, a step of incubating at 30°C and 120 rpm for 1 to 3 days, preferably 1 day, to obtain fermented dietary sulfur lysate; The fermented dietary sulfur lysate is fermented with a solvent selected from the group consisting of purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane, chloroform, ethyl ether, butylene glycol, hexane and mixtures thereof. It is characterized by being produced by a production method comprising the step of obtaining a fermented dietary sulfur extract by extracting using 20 to 80 times, preferably 40 to 60 times, more preferably 50 times (V/V) of the dietary sulfur lysate. to be. The type of solvent and the capacity required for extraction can be selected and extracted according to the concentration, formulation and use of the extract.

본 출원의 목적을 달성하기 위하여 본 출원은 발효 식이유황 추출물(MSM-F10)을 유효성분으로 함유하는 항산화, 항주름, 미백 및 항균 기능성 화장료 조성물을 제공한다. 본 발명의 화장료 조성물에서 발효 식이유황 추출물(MSM-F10)을 액상의 중량으로서 0.1~20 중량%의 양을 화장료에 첨가할 수 있고, 바람직하게는 0.1~15 중량%, 더욱 바람직하게는 10중량%를 첨가할 수 있다. 본 발명의 항산화, 항주름, 미백 및 항균 기능성 화장료 조성물은 그 제형에 있어서 특별히 한정되는 바가 없으며, 예를 들면, 화장수, 영양로션, 영양크림, 마사지크림, 에센스, 팩 등의 제형을 가질 수 있다. 또한, 각 제형의 상기 화장료 조성물에 있어서, 상기의 해란초 추출물 외에 다른 성분들을 기타 화장료의 제형 또는 사용목적 등에 따라 임의로 선정하여 배합할 수 있다.In order to achieve the object of the present application, the present application provides an antioxidant, anti-wrinkle, whitening and antibacterial functional cosmetic composition containing fermented dietary sulfur extract (MSM-F10) as an active ingredient. In the cosmetic composition of the present invention, fermented dietary sulfur extract (MSM-F10) may be added in an amount of 0.1 to 20% by weight as a liquid weight, preferably 0.1 to 15% by weight, more preferably 10% by weight % Can be added. The antioxidant, anti-wrinkle, whitening and antibacterial functional cosmetic composition of the present invention is not particularly limited in its formulation, and may have, for example, a lotion, nutrition lotion, nutrition cream, massage cream, essence, pack, etc. . In addition, in the cosmetic composition of each formulation, other ingredients in addition to the above-mentioned seaweed extract may be arbitrarily selected and blended according to other cosmetic formulations or purpose of use.

본 출원의 발효 식이유황 추출물은 뛰어난 항주름, 항산화, 미백 및 항균 효과가 있어 화장품에 적용시 주름, 항산화, 미백 및 항균 기능성 화장료 조성물을 제공하는 장점이 있다.The fermented dietary sulfur extract of the present application has an excellent anti-wrinkle, antioxidant, whitening and antibacterial effect, and thus has an advantage of providing a wrinkle, antioxidant, whitening and antibacterial functional cosmetic composition when applied to cosmetics.

도 1은 본 발명에 따른 식이유황 발효 추출물(MSM-F10)의 제조방법을 도시한 다이어그램이며, 도 2는 그에 따라 제조된 식이유황 발효 추출물(MSM-F10)과 발효 전 식이유황 용해물 항주름 효과 실험 결과를 나타내는 그래프이다.
도 3은 본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 항산화 효과 실험 결과를 나타내는 그래프이다.
도 4는 본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 미백 효과 실험 결과를 나타내는 그래프이다.1 is a diagram showing a method of preparing a dietary sulfur fermentation extract (MSM-F10) according to the present invention, and FIG. 2 is an anti-wrinkle dietary sulfur fermentation extract (MSM-F10) and dietary sulfur lysate prepared before fermentation. It is a graph showing the results of the effect experiment.
Figure 3 is a graph showing the antioxidant effect of the present invention fermented dietary sulfur extract (MSM-F10) and the antioxidant effect of dietary sulfur lysate before fermentation.
Figure 4 is a graph showing the experimental results of the whitening effect of the present invention fermented dietary sulfur extract (MSM-F10) and dietary sulfur lysate before fermentation.

이하, 본 출원에 따른 실시예를 통하여 본 발명을 보다 상세히 설명하고자 한다. 다만 본 출원의 하기 실시예는 본 출원의 일 예시에 불과하다. 이들 실시예는 본 발명을 보다 구체적으로 설명하기 위한 것으로서, 첨부된 청구항에 제시된 본 발명의 범위가 이들 실시예에 의해 제한되지 않는다는 것은 본 발명이 속하는 기술분야에서 통상의 지식을 가진 자에게 있어 명백할 것이다.Hereinafter, the present invention will be described in more detail through examples according to the present application. However, the following examples of the present application are only examples of the present application. These examples are intended to illustrate the present invention more specifically, and it is apparent to those skilled in the art to which the present invention pertains that the scope of the present invention set forth in the appended claims is not limited by these examples. something to do.

실시예 1. 발효 식이유황 추출물(MSM-F10)의 제조Example 1. Preparation of fermented dietary sulfur extract (MSM-F10)

본 발명 발효 식이유황 추출물은, 식이유황을 50배의 정제수에 혼합한 후 121 ℃에서 15분 멸균하고 외부 균의 접촉이 없도록 유지하여 상온에서 온도를 내려 멸균된 식이유황 용해물을 수득하였다. 다음, 상기 멸균된 식이유황 용해물에 106 내지 107 cell/ml의 일정 농도의 Bacillus subtilis 균주를 1%(v/v) 접종 후 30℃, 120rpm에서 1일 배양하여 발효시켜 식이유황 발효물을 수득하였다. 다음, 상기 식이유황 발효물을 정제수, 메탄올, 에탄올, 글리세린, 에틸아세테이트, 부틸렌글리콜, 프로필렌글리콜, 디클로로메탄, 클로로포름, 에틸에테르, 부틸렌글리콜, 헥산 및 이의 혼합물로 이루어지는 군으로부터 선택되는 용매를 식이유황 발효물의 50배(V/V) 사용하여 추출함으로써 발효 식이유황 추출물을 제조하였고, 이를 MSM-F10으로 명명하였다.The fermented dietary sulfur extract of the present invention, after mixing the dietary sulfur with 50 times purified water, sterilized at 121° C. for 15 minutes and kept free from contact with external bacteria to lower the temperature at room temperature to obtain a sterilized dietary sulfur lysate. Next, the Bacillus subtilis strain having a constant concentration of 10 6 to 10 7 cell/ml in the sterilized dietary sulfur lysate was inoculated with 1% (v/v) and incubated for 1 day at 30°C and 120 rpm to ferment the dietary sulfur fermentation product. Was obtained. Next, the dietary sulfur fermentation product is a solvent selected from the group consisting of purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane, chloroform, ethyl ether, butylene glycol, hexane, and mixtures thereof. A fermented dietary sulfur extract was prepared by extraction using 50 times (V/V) of dietary sulfur fermentation, which was named MSM-F10.

실험예 1. 발효 식이유황 추출물(MSM-F10)의 항주름 효과Experimental Example 1. Anti-wrinkle effect of fermented dietary sulfur extract (MSM-F10)

상기 실시예1에서 수득한 본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 항주름 효과를 비교하고 확인하기 위하여, 피부 섬유아세포(HDFn)를 48 well 플레이트에 5×104 cells/well의 농도로 배양 배지 0.3 mL로 접종하여 37℃, 5% CO2에서 배양하였다. 24시간 지난 이후 각 추출물 및 용해물을 농도별(0.5%(v/v), 1%(v/v), 2%(v/v))로 포함한 새로운 배지로 24시간 동안 배양하였다. 24시간 배양 이후 배양액을 모아서 enzymes-linked immunoassay kit (Takara사)를 이용하여 콜라겐 전구체의 C-말단의 양을 측정하였다. 또한, 콜라겐 용액 640 ng/mL을 이용하여 콜라겐 농도 0 ng-640 ng 사이의 standard curve를 얻어 배지에 있는 콜라겐 양을 계산하였다. 모든 실험은 3번 반복하여 실험하였다.To compare and confirm the anti-wrinkle effect of the present invention fermented dietary sulfur extract (MSM-F10) obtained in Example 1 and dietary sulfur lysate prior to fermentation, skin fibroblasts (HDFn) were 5×10 in 48 well plates. Inoculated with 0.3 mL of culture medium at a concentration of 4 cells/well and cultured at 37°C and 5% CO 2 . After 24 hours, each extract and lysate was cultured for 24 hours with fresh medium containing concentrations (0.5% (v/v), 1% (v/v), 2% (v/v)). After incubation for 24 hours, the culture solution was collected and the amount of the C-terminus of the collagen precursor was measured using an enzymes-linked immunoassay kit (Takara). In addition, a standard curve between 0 ng-640 ng of collagen concentration was obtained using a collagen solution of 640 ng/mL, and the amount of collagen in the medium was calculated. All experiments were repeated three times.

본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 항주름 효과를 비교한 결과, 발효 식이유황 추출물(MSM-F10)이 발효 전 식이유황 용해물보다 콜라겐 합성 효과가 뛰어나고, 특히 2%(v/v)의 농도에서 20%의 더 증가된 콜라겐 합성 효과가 있음을 확인하였다(도 2).As a result of comparing the anti-wrinkle effect of the fermented dietary sulfur extract (MSM-F10) and the dietary sulfur lysate before fermentation, the fermented dietary sulfur extract (MSM-F10) has a better collagen synthesis effect than the dietary sulfur lysate before fermentation, In particular, it was confirmed that there is an increased collagen synthesis effect of 20% at a concentration of 2% (v/v) (FIG. 2).

실험예 2. 발효 식이유황 추출물(MSM-F10)의 항산화 효과Experimental Example 2. Antioxidative effect of fermented dietary sulfur extract (MSM-F10)

상기 실시예1에서 수득한 본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 항산화 효과를 비교하고 확인하기 위하여, DCFH-DA(2＇7＇-dichloro flurescin0diacetate)가 사용되었다. 우선, 피부 섬유아세포(HDFn)를 DMEM(10% 혈청 및 1% 항생제 첨가) 배지에 현탁하여 37℃, 5% CO2 배양기에서 well당 1x105 세포수가 되도록 24시간 배양하였다. 다음 상기 피부 섬유아세포에 추출물 및 용해물 농도별(0.5%(v/v), 1%(v/v), 2%(v/v))로 처리하여 37℃, 5% CO2 배양기에서 1시간 배양한 후 산화스트레스 유발 물질인 H2O2 1mM 및 DCFDA 20M을 첨가하고, excitation 485nm, emission 535nm 조건에서 spectrofluorophotomer로 측정하였다. 시료를 넣지 않고 항산화 활성을 측정한 처리군을 대조군으로 이용하였고 항산화 활성은 하기 수학식1의 방법으로 계산하였다. 모든 실험은 3번 반복하여 실험하였다.To compare and confirm the antioxidant effects of the present invention fermented dietary sulfur extract (MSM-F10) obtained in Example 1 and dietary sulfur lysate before fermentation, DCFH-DA (2＇7＇-dichloro flurescin0diacetate) was used. . First, skin fibroblasts (HDFn) were suspended in DMEM (10% serum and 1% antibiotic added) medium and cultured for 24 hours at 37° C. in a 5% CO 2 incubator to be 1×10 5 cells per well. Next, the skin fibroblasts were treated with extract and lysate concentrations (0.5% (v/v), 1% (v/v), 2% (v/v)) at 37°C, 5% CO 2 incubator 1 After incubation with time, H 2 O 2 1mM and DCFDA 20M, which are oxidative stress-inducing substances, were added, and measured with a spectrofluorophotomer at excitation 485nm and emission 535nm. A treatment group in which antioxidant activity was measured without a sample was used as a control, and antioxidant activity was calculated by the method of Equation 1 below. All experiments were repeated three times.

Figure 112019017970053-pat00001
Figure 112019017970053-pat00001

본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 항산화 효과를 비교한 결과, 발효 식이유황 추출물(MSM-F10)이 발효 전 식이유황 용해물보다 ROS 생성 감소 효과가 뛰어나고, 특히 2%(v/v)의 발효 식이유황 추출물(MSM-F10) 농도에서 25%의 ROS 생성을 감소시키는 효과가 있음을 확인하였다(도 3).As a result of comparing the antioxidant effect of the fermented dietary sulfur extract (MSM-F10) and the dietary sulfur lysate before fermentation, the fermented dietary sulfur extract (MSM-F10) has a superior ROS production reduction effect than the dietary sulfur lysate before fermentation, In particular, it was confirmed that there is an effect of reducing the production of 25% ROS at a concentration of 2% (v/v) fermented dietary sulfur extract (MSM-F10) (FIG. 3).

실험예 3. 발효 식이유황 추출물(MSM-F10)의 미백 효과Experimental Example 3. Whitening effect of fermented dietary sulfur extract (MSM-F10)

상기 실시예1에서 수득한 본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 미백 효과를 비교하고 확인하기 위하여, murine melanoma(B-16 F1) 세포를 적량의 FBS(fetal bovine serum)가 함유된 DMEM 배지로 6-well plate에 well당 1×105개로 접종한 후, 5% CO2, 37℃하에서 세포가 well 바닥에 약 80 % 이상 부착될 때까지 배양하였다. 배양 후 배지를 제거하고 추출물 및 용해물의 적당 농도(0.5%(v/v), 1%(v/v), 2%(v/v))로 희석된 배지로 교체한 후 5% CO2, 37℃하에서 일정시간 배양하였다. 배지를 제거한 세포를 PBS(phosphated buffer saline)로 세척하고 트립신으로 처리하여 세포를 회수하였다. 회수한 세포를 hematocytometer를 이용하여 세포수를 측정한 후 5,000∼10,000 rpm으로 10분간 원심분리한 다음 상등액을 제거하여 pellet을 얻었다. 상기 세포 pellet은 60℃에서 건조한 후 10% DMSO가 함유된 1M 수산화나트륨액 100 ㎕ 또는 적량의 cell lysis buffer를 넣어 60 ℃ 항온조에서 세포 내 멜라닌을 얻었다. 상기 액을 가지고 마이크로플레이트 측정기로 405nm에서 흡광도를 측정하여 세포 일정 수당 멜라닌 양 또는 일정 단백질당 멜라닌양을 구하였다.In order to compare and confirm the whitening effect of the present invention fermented dietary sulfur extract (MSM-F10) obtained in Example 1 and dietary sulfur lysate before fermentation, murine melanoma (B-16 F1) cells in an appropriate amount of FBS (fetal Bovine serum) was inoculated with DMEM medium containing 1×10 5 cells per 6-well plate, and cultured at 5% CO 2 and 37° C. until the cells adhered to the bottom of the wells by about 80% or more. After incubation, the medium is removed and replaced with a medium diluted to an appropriate concentration (0.5% (v/v), 1% (v/v), 2% (v/v)) of extracts and lysates, followed by 5% CO 2 , Incubated at 37 ℃ for a certain time. The cells from which the medium was removed were washed with PBS (phosphated buffer saline) and treated with trypsin to recover the cells. The recovered cells were counted using a hematocytometer, centrifuged at 5,000 to 10,000 rpm for 10 minutes, and the supernatant was removed to obtain pellets. The cell pellet was dried at 60° C., and then 100 μl of 1M sodium hydroxide solution containing 10% DMSO or an appropriate amount of cell lysis buffer was added to obtain intracellular melanin in a 60° C. thermostat. The absorbance was measured at 405 nm with a microplate measuring instrument to obtain the amount of melanin per cell or the amount of melanin per protein.

본 발명 발효 식이유황 추출물(MSM-F10)과 발효 전 식이유황 용해물의 미백 효과를 비교한 결과, 발효 식이유황 추출물(MSM-F10)이 발효 전 식이유황 용해물보다 미백 효과가 뛰어나고, 특히 2%(v/v)의 발효 식이유황 추출물(MSM-F10) 농도에서 20%의 멜라닌 생성을 감소시키는 효과가 있음을 확인하였다 (도 4).As a result of comparing the whitening effect of the fermented dietary sulfur extract (MSM-F10) and the dietary sulfur lysate before fermentation, the fermentation dietary sulfur extract (MSM-F10) has a better whitening effect than the dietary sulfur lysate before fermentation, especially 2 It was confirmed that there is an effect of reducing the production of melanin by 20% at a concentration of fermented dietary sulfur extract (MSM-F10) of% (v/v) (FIG. 4 ).

실험예 4. 발효 식이유황 추출물(MSM-F10)의 항균 효과Experimental Example 4. Antibacterial effect of fermented dietary sulfur extract (MSM-F10)

상기 실시예1에서 수득한 본 발명 발효 식이유황 추출물(MSM-F10)의 항균 효과를 확인하기 위하여 사용한 시험균주와 방법은 하기와 같다.The test strains and methods used to confirm the antibacterial effect of the fermented dietary sulfur extract (MSM-F10) of the present invention obtained in Example 1 are as follows.

1. 시험균주1. Test strain

1) Staphylococcus aureus : LB agar plate에서 37℃, 24시간 배양 후 single colony를 취하여 LB 액체배지에 접종 후 30℃ shaking incubator에서 18시간 배양 시킨 후 사용하였다(균수 109 cfu/ml정도).1) Staphylococcus aureus : After incubation at 37° C. for 24 hours in LB agar plate, single colony was inoculated into LB liquid medium, followed by incubation at 30° C. in shaking incubator for 18 hours (about 109 cfu/ml of bacteria).

2) Escherichia coli : LB agar plate에서 37℃, 24시간 배양 후 single colony를 취하여 LB 액체배지에 접종 후 30℃ shaking incubator에서 18시간 배양 시킨 후 사용하였다(균수 109 cfu/ml정도).2) Escherichia coli : After incubation at 37°C for 24 hours in LB agar plate, single colony was inoculated into LB liquid medium, followed by incubation at 30°C in shaking incubator for 18 hours (about 109 cfu/ml of bacteria).

3) Candida albicans : YM agar plate에서 30℃, 24시간 배양 후 single colony를 취하여 YM 액체배지에 접종 후 30℃ shaking incubator에서 18시간 배양 시킨 후 사용하였다(균수 109 cfu/ml정도).3) Candida albicans : After incubation at 30°C and 24 hours in YM agar plate, single colony was inoculated into YM liquid medium, followed by incubation at 30°C shaking incubator for 18 hours (about 109 cfu/ml of bacteria).

2. 본 발명 발효 식이유황 추출물(MSM-F10) 및 발효 전 식이유황 용해물을 각 100%, 50%, 25%, 12.5%, 6.2% 3.1%의 농도(v/v)와 positive control인 Tetracycline(50μg/ml, 12.5 μg/ml), Ketoconazole 0.25%(v/v) 그리고 각각의 균주를 104 만큼 well 에 넣어준 후 shaking incubator에서 30℃, 20시간 배양 후, 흡광도 600에서 측정하여 측정값을 저해활성으로 환산하여 계산하였다.2. The present invention fermented dietary sulfur extract (MSM-F10) and the dietary sulfur lysate before fermentation are 100%, 50%, 25%, 12.5%, 6.2%, concentration of 3.1% (v/v) and Tetracycline as positive control (50 μg/ml, 12.5 μg/ml), Ketoconazole 0.25% (v/v), and put each strain in a well for 10 4 , incubating at 30°C for 20 hours in a shaking incubator, and measuring at absorbance 600 Was calculated in terms of inhibitory activity.

본 발명 발효 식이유황 추출물(MSM-F10)은 E.coli를 농도 의존적으로 저해하는 항균 효과가 있음을 확인하였다 (표 1).It was confirmed that the fermented dietary sulfur extract (MSM-F10) of the present invention has an antimicrobial effect to inhibit E. coli in a concentration-dependent manner (Table 1).

Figure 112019017970053-pat00002
Figure 112019017970053-pat00002

이하, 본 발명을 사용하기에 바람직하고 다양한 화장품 제형을 개시하지만 본 발명의 권리범위는 개시된 제형에 제한되지 않음은 물론이다.Hereinafter, it is preferable to use the present invention and discloses various cosmetic formulations, but the scope of the present invention is not limited to the disclosed formulations.

제조예1. 화장수Preparation Example 1. toilet water

본 발명 상기 실시예1에서 얻은 본 발명 발효 식이유황 추출물(MSM-F10)이 함유된 화장수를 하기 표2와 같이 제조하였다.The present invention was prepared as shown in Table 2 below the cosmetic product containing the fermented dietary sulfur extract (MSM-F10) obtained in Example 1 above.

원료Raw material 원 료Raw material 함량(중량%)Content (% by weight) 1One 본 발명 발효 식이유황 추출물(MSM-F10)Fermented dietary sulfur extract of the present invention (MSM-F10) 10.010.0 22 글리세린glycerin 3.03.0 33 부틸렌 글리콜Butylene glycol 2.02.0 44 프로필렌 글리콜Propylene glycol 2.02.0 55 폴리옥시에칠렌 경화피마자유Polyoxyethylene cured castor oil 1.01.0 66 에탄올ethanol 10.010.0 77 트리에탄올아민Triethanolamine 0.10.1 88 방부제antiseptic 미량a very small amount 99 색소Coloring 미량a very small amount 1010 향료Spices 미량a very small amount 1111 정제수Purified water 잔량Balance

제조예2. 로션Preparation Example 2. Lotion

본 발명 상기 실시예1에서 얻은 본 발명 발효 식이유황 추출물(MSM-F10)이 함유된 로션을 하기 표3와 같이 제조하였다.Invention The lotion containing the present invention fermented dietary sulfur extract (MSM-F10) obtained in Example 1 was prepared as shown in Table 3 below.

원료Raw material 원 료Raw material 함량(중량%)Content (% by weight) 1One 본 발명 발효 식이유황 추출물(MSM-F10)Fermented dietary sulfur extract of the present invention (MSM-F10) 10.010.0 22 밀납Wax 1.01.0 33 폴리솔베이트 60Polysorbate 60 1.51.5 44 솔비단 세스퀴올레이트Solvidan Sesquioleate 0.50.5 55 유동 파라핀Floating paraffin 10.010.0 66 소르비단 스테아레이트Sorbidan stearate 1.01.0 77 친유형 모노스테아린신 글리세린Lipophilic monostearine glycerin 0.50.5 88 스테아린산Stearic acid 1.51.5 99 글리세릴스테아레이트/피이지-400 스테아레이트Glyceryl stearate/page-400 stearate 1.01.0 1010 프로필렌글리콜Propylene glycol 3.03.0 1111 카르복시폴리머Carboxy polymer 0.10.1 1212 트리에탄올아민Triethanolamine 0.20.2 1313 방부제antiseptic 미량a very small amount 1414 색소Coloring 미량a very small amount 1515 향료Spices 미량a very small amount 1616 정제수Purified water 잔량Balance

제조예3. 크림Preparation Example 3. cream

본 발명 상기 실시예1에서 얻은 본 발명 발효 식이유황 추출물(MSM-F10)이 함유된 크림을 하기 표4과 같이 제조하였다.Invention The cream containing the present invention fermented dietary sulfur extract (MSM-F10) obtained in Example 1 was prepared as shown in Table 4 below.

함량content 원 료Raw material 함량(중량%)Content (% by weight) 1One 본 발명 발효 식이유황 추출물(MSM-F10)Fermented dietary sulfur extract of the present invention (MSM-F10) 10.010.0 22 스테아린산Stearic acid 2.02.0 33 세틸알콜Cetyl alcohol 2.02.0 44 글리세릴모노스테아레이트Glyceryl monostearate 2.02.0 55 폴리옥시에틸렌소르비단모소스테아레이트Polyoxyethylene sorbitan monostearate 0.50.5 66 솔비단세스퀴올레이트Solvidan sesquioleate 0.50.5 77 글리세릴모노스테아레이트/글리세릴스테아레이트/폴리옥시에틸렌스테아레이트Glyceryl monostearate/glyceryl stearate/polyoxyethylene stearate 1.01.0 88 왁스Wax 1.01.0 99 유동파라핀스Floating paraffin 4.04.0 1010 스쿠알란Squalane 4.04.0 1111 카프릴릭/카프릭트리글리세라이드Caprylic/Capric Triglyceride 4.04.0 1212 카르복시비닐폴리머Carboxyvinyl polymer 0.30.3 1313 부틸렌글리콜Butylene glycol 5.05.0 1414 글리세린glycerin 3.03.0 1515 트리에탄올아민Triethanolamine 0.50.5 1616 정제수Purified water 잔량Balance

제조예4. 에센스 제조Preparation Example 4. Essence production

본 발명 상기 실시예1에서 얻은 본 발명 발효 식이유황 추출물(MSM-F10)이 함유된 에센스를 하기 표5과 같이 제조하였다.Invention The essence containing the fermented dietary sulfur extract (MSM-F10) of the present invention obtained in Example 1 was prepared as shown in Table 5 below.

함량content 원 료Raw material 함량(중량%)Content (% by weight) 1One 본 발명 발효 식이유황 추출물(MSM-F10)Fermented dietary sulfur extract of the present invention (MSM-F10) 10.010.0 22 시토 스테롤Citosterol 1.71.7 33 폴리글리세릴 2-올레이트Polyglyceryl 2-oleate 1.51.5 44 세라마이드Ceramide 0.70.7 55 스테아레스-4Steareth-4 1.21.2 66 콜레스테롤cholesterol 1.51.5 77 디세틸포스페이트Dicetyl phosphate 0.40.4 88 농글리세린Concentrated glycerin 5.05.0 99 마카다미아 오일Macadamia oil 15.015.0 1010 카르복시비닐폴리머Carboxyvinyl polymer 0.20.2 1111 산탄검Shotgun Sword 0.20.2 1212 방부제antiseptic 미량a very small amount 1313 향료Spices 미량a very small amount 1414 정제수Purified water 잔량Balance

이상의 설명으로부터, 본 출원이 속하는 기술분야의 당업자는 본 출원이 그 기술적 사상이나 필수적 특징을 변경하지 않고서 다른 구체적인 형태로 실시될 수 있다는 것을 이해할 수 있을 것이다. 이와 관련하여, 이상에서 기술한 실시예들은 모든 면에서 예시적인 것이며 한정적인 것이 아닌 것으로 이해해야만 한다. 본 출원의 범위는 상기 상세한 설명보다는 후술하는 특허 청구범위의 의미 및 범위 그리고 그 등가 개념으로부터 도출되는 모든 변경 또는 변형된 형태가 본 출원의 범위에 포함되는 것으로 해석되어야 한다.From the above description, those skilled in the art to which the present application pertains will appreciate that the present application may be implemented in other specific forms without changing the technical spirit or essential characteristics. In this regard, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. The scope of the present application should be construed to include all changes or modifications derived from the meaning and scope of the following claims rather than the above detailed description and equivalent concepts thereof.

Claims (5)

식이유황(Methyl Sulfonyl Methane)을 정제수에 혼합한 후 멸균하여 멸균된 식이유황 용해물을 수득하는 단계와;
상기에서 얻은 멸균된 식이유황 용해물에 Bacillus Subtilis 균주를 접종 후 30℃, 120rpm에서 1일 배양하여 발효시켜 식이유황 발효물을 수득하는 단계와;
상기에서 얻은 식이유황 발효물을 정제수, 메탄올, 에탄올, 글리세린, 에틸아세테이트, 부틸렌글리콜, 프로필렌글리콜, 디클로로메탄, 클로로포름, 에틸에테르, 부틸렌글리콜, 헥산 및 이의 혼합물로 이루어지는 군으로부터 선택되는 용매를 식이유황 발효물의 50배(V/V) 사용하여 추출함으로써 발효 식이유황 추출물을 수득하는 단계로 이루어지는 것이 특징인 발효 식이유황 추출물(MSM-F10)의 제조방법.Mixing dietary sulfur (Methyl Sulfonyl Methane) with purified water and sterilizing to obtain sterilized dietary sulfur lysate;
Inoculating Bacillus Subtilis strain into the sterilized dietary sulfur lysate obtained above, and then incubating at 30°C and 120 rpm for 1 day to ferment to obtain dietary sulfur fermentation products;
The fermented dietary sulfur obtained above is purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane, chloroform, ethyl ether, butylene glycol, hexane and a solvent selected from the group consisting of mixtures thereof Method for producing a fermented dietary sulfur extract (MSM-F10), characterized in that it consists of the step of obtaining a fermented dietary sulfur extract by extracting using 50 times (V/V) of the dietary sulfur fermentation. 제1항의 방법에 따라 제조된 발효 식이유황 추출물(MSM-F10).Fermented dietary sulfur extract prepared according to the method of claim 1 (MSM-F10). 제2항의 추출물을 유효성분으로 함유하는 것이 특징인 항산화 및 항균 기능성 화장료 조성물 Antioxidant and antibacterial functional cosmetic composition characterized by containing the extract of claim 2 as an active ingredient 제2항의 추출물을 유효성분으로 함유하는 것이 특징인 항주름 및 미백 기능성 화장료 조성물 Anti-wrinkle and whitening functional cosmetic composition characterized by containing the extract of claim 2 as an active ingredient 제 3 또는 제 4항에 있어서, 상기 기능성 화장료 조성물은 화장수, 로션, 크림 및 에센스 중에서 선택되는 어느 하나의 제형인 것이 특징인 화장료 조성물.
The cosmetic composition according to claim 3 or 4, wherein the functional cosmetic composition is any one selected from lotion, lotion, cream and essence.
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KR101874830B1 (en) * 2018-04-09 2018-07-06 전문식 Antibacterial Cosmetics Composition

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KR102367604B1 (en) * 2021-10-25 2022-02-28 주식회사 캘러스 Feed additive composition for companion animals with excellent anti-oxidant, anti-inflammatory and immune boostings effects

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