KR101409764B1 - Comprising lactic acid fermentation product of Gelidium amansil extract as an Active Ingredient for Improving Skin Wrinkle - Google Patents
Comprising lactic acid fermentation product of Gelidium amansil extract as an Active Ingredient for Improving Skin Wrinkle Download PDFInfo
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- KR101409764B1 KR101409764B1 KR1020130018628A KR20130018628A KR101409764B1 KR 101409764 B1 KR101409764 B1 KR 101409764B1 KR 1020130018628 A KR1020130018628 A KR 1020130018628A KR 20130018628 A KR20130018628 A KR 20130018628A KR 101409764 B1 KR101409764 B1 KR 101409764B1
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- extract
- fermented
- lactic acid
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- lactobacillus
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Abstract
Description
본 발명은 피부 주름 개선 효능을 가지는 우뭇가사리 주정추출 유산균 발효물 및 이를 유효성분으로 함유하는 제품에 관한 것으로, 보다 상세하게는 우뭇가사리 주정추출물을 유산균을 이용하여 발효시킨 피부 주름 개선 효능을 가지는 우뭇가사리 주정추출 유산균 발효물 및 이를 유효성분으로 함유하는 제품에 관한 것이다
The present invention relates to a fermented product of Lactobacillus acidophilus extract having the effect of improving skin wrinkles and a product containing the same as an active ingredient. More particularly, the present invention relates to a fermented product of Lactobacillus sp. Extract having a skin wrinkle- Fermented products of lactic acid bacteria and products containing them as active ingredients
피부 노화(aging)는 나이가 듦에 따라 자연스럽게 신체 구조와 기능이 쇠퇴되는 현상으로 피부표면 전체에서 나이와 관련된 피부 기능, 구조 또는 모양의 변화를 나타내는 생리적 노화와 자외선에 의한 광노화로 구분된다. 평균수명이 길어짐에 따라 젊고 건강한 삶에 대한 욕구가 증대되면서 전 세계적으로 건강이나 피부 미용에 대한 관심이 고조되고 있다. 또한, 늘어난 여가시간 또는 바깥 활동으로 자외선에 노출되는 빈도가 늘어남에 따라 색소 침착, 거칠음, 탄력소실 등과 같은 피부 손상이 유발되어 피부 노화 현상이 가속화 된다. 이러한 태양광선에 의한 노화를 광노화(photoaging)라고 하며, 특히 자외선은 피부 세포내 신호전달체계를 교란하여 콜라겐, 탄력섬유 등의 합성 감소, 콜라겐 분해효소 활성화 등을 통해 주름을 생성시킨다. 또한 자외선은 물분자와의 결합을 통해 피부 수분유지 기능을 담당하는 히알루론산 합성을 저해하여 피부 건조를 유발한다. 이러한 피부 구성 성분들의 자외선에 의한 파괴는 굵고 깊은 주름을 생성시키며 피부 노화를 더욱 촉진하는 것으로 보고되고 있다. 최근 피부노화를 예방 또는 개선 할 수 있는 다양한 기능성 소재 탐색이 활발히 이루어지고 있다.
Skin aging is a phenomenon in which body structure and function decline naturally with age. It is divided into physiological aging and ultraviolet ray photoaging that show changes in age, skin function, structure or shape throughout the skin surface. As life expectancy increases, the desire for a younger, healthier life has increased and interest in health and skin care has increased worldwide. In addition, as the frequency of exposure to ultraviolet rays increases due to increased leisure time or outdoor activities, skin aging phenomenon is accelerated due to skin damage such as pigmentation, roughness, loss of elasticity and the like. Such aging by sunlight is called photoaging, and ultraviolet rays disturb the signal transduction system in the skin cells, resulting in reduction of synthesis of collagen and elastic fibers, and activation of collagenase. Ultraviolet rays inhibit the synthesis of hyaluronic acid, which is responsible for maintaining skin moisture through binding with water molecules, and cause skin dryness. Ultraviolet destruction of these skin constituents has been reported to produce thick and deep wrinkles and further promote skin aging. Recently, various functional materials capable of preventing or improving skin aging have been actively searched.
우뭇가사리는 우뭇가사리과에 속하는 홍조류의 해조류로 학명은 Gelidium amansii 이다. 바닷말의 일종으로 주로 한천의 주원료로 이용되며, 여러해살이 해조류로서 여름의 번식기가 지나면 본체의 상부는 녹아 없어지고 하부만 남아 있다가 다음해 봄에 다시 새싹이 자라난다. 동해안·남해안과 황해의 바깥 도서에 분포하며 동해 남부 연안에서 가장 많이 생산된다. 바닷속 20~30m 깊이의 바위에 붙어 자라는데, 바깥바다에 면하고 바닥이 모래로 되어 있으며, 해수의 소통이 잘되는 곳에 산다. 우뭇가사리를 민물에 깨끗이 씻어 햇볕에 말린 것을 고아서 찌꺼기를 걸러내고 식히면 우무가 된다. 이 우무는 예로부터 채쳐서 콩국에 띄워 청량음료로 사용하여 왔다. 우뭇가사리에는 3가지의 몸체가 있는데, 즉 유성 세대인 수배우체와 암배우체 및 무성 세대인 포자체이다. 이것들은 그 생김새가 매우 비슷하므로, 생식 기관이나 핵상을 조사하지 않으면 서로 구별할 수가 없다. 수배우체는 작은 가지의 표면 부근 세포가 분열하여 정자를 만든다. 이에 반해서 암배우체는 작은 가지를 이루는 세포열로부터 '조과기'라고 하는 특수한 세포열이 뻗어 나와서 그 끝 세포가 조과기가 되고 그 속에 난자가 생긴다. 그 후 조과기로부터 수정모가 체표를 뚫고 나오면 방출된 정자가 수정모 끝에 붙고, 이어서 정핵이 수정모 내부를 지나 조과기 안의 난핵과 합쳐지게 된다. 핵융합이 일어난 후에는 조과기로부터 특수한 연락사가 뻗어 나와, 가까운 영양 조직으로부터 자라는 데 필요한 영양분을 받아들인다. 수정핵은 이 연락사를 통해 이동하면서, 군데군데에서 '조포사'라는 세포사를 내어 그 곳에 과포자를 만든다. 이때 과포자 및 그것을 만드는 조포사나 연락사 등은 2n의 핵상을 가진다. 즉, 2n인 몸이 핵상이 n인 암배우체에 기생하는 것이다. 이러한 2n의 몸체를 '과포자체'라고 하는데, 얼마 후에는 이로부터 과포자가 방출되며, 이것이 바위 등에 붙어 발아하면 포자체가 되는 것이다. 포자체는 작은 가지 끝에 수많은 포자낭을 만드는데, 포자낭이 생긴 작은 가지는 주걱 모양으로 변화한다. 이 포자낭은 감수 분열을 하여 사분 포자를 만들어 방출하며, 방출된 사분 포자는 바위 등에 붙어서 발아하여, 4개 중 2개는 수배우체로, 다른 2개는 암배우체로 자라게 된다. 즉, 우뭇가사리에는 암수의 배우체 세대, 암배우체에 기생하는 작은 과포자 세대 및 사분 포자체 세대의 3가지 세대가 있으며, 생활사는 이 3세대가 순환하면서 이루어지게된다.
It is a seaweed of red algae belonging to the mugwort family. Its scientific name is Gelidium amansii. It is a kind of pale endosymbionts. It is mainly used for agar. It is a perennial seaweed. After the breeding season of summer, the upper part of the body melts away and remains only the lower part. It is distributed in the East Sea, South Sea and Yellow Sea outbreaks, and it is produced the most in the southern coast of the East Sea. It grows on
과거 오랫동안 유산균은 장관 면역 조절에 중요한 역할을 담당하며 장내 균총을 개선하여 몸에 유익한 효능을 나타내는 것으로 알려져 왔다. 최근에는 피부 장벽 기능 향상 등 피부건강 조절기능을 가진 유산균이 보고되고 있으며, 유산균 섭취가 장 내 면역력 조절뿐만 아니라, 전신의 면역을 조절하여 피부 면역에 영향을 준다는 보고가 있다. 또한 최근 보고에 따르면 임상 및 동물실험에서 급성적인 자외선 조사에 의해 유도된 피부의 과도한 면역반응이 유산균 경구 섭취에 의해 완화 되었다. 그러나, 만성적인 자외선 조사에 의한 피부의 광노화 및 광손상에 대한 유산균의 보호 효과는 아직 연구된 바가 없는 실정이다.
Lactobacillus has long been known to play an important role in the regulation of intestinal immunity and to improve the intestinal microflora and to show beneficial effects on the body. Recently, lactic acid bacteria having skin health controlling function such as improvement of skin barrier function have been reported. It has been reported that lactic acid bacterium intake affects skin immunity by controlling not only intestinal immunity but also whole body immunity. Recent reports also show that excessive immune responses to skin induced by acute ultraviolet irradiation in clinical and animal studies have been alleviated by oral ingestion of lactic acid bacteria. However, the protective effect of lactic acid bacteria against photoaging and photodamage of skin caused by chronic ultraviolet irradiation has not been studied yet.
이에 본 발명자들은 피부 주름 개선 효과가 우수한 천연물을 찾기 위해 연구를 제주도 자생식물 및 해조류를 대상으로 실시하여 다양한 천연물 중 본 발명의 우뭇가사리 주정 추출물을 유산균을 이용하여 발효한 우뭇가사리 주정추출물의 유산균 발효물의 피부 주름 개선 효과를 확인한 결과 상당히 우수한 효능을 나타냄을 확인하여 본 발명을 완성하였다.
Therefore, the present inventors conducted a study on native plants and algae of Jeju Island in order to find natural products having excellent skin wrinkle-reducing effect. The inventors of the present invention have found that extracts of the extracts of the present invention are obtained by extracting the extracts of fermented Lactobacillus sp. The present inventors confirmed that the wrinkle-improving effect was remarkably excellent as a result of confirming the wrinkle-improving effect.
상기와 같은 문제점을 해결하기 위하여, 본 발명은 우뭇가사리 주정추출물을 유산균을 이용하여 발효시킴으로써 인체에는 안전하면서 피부 주름 개선 효과가 우수한 우뭇가사리 주정추출물의 유산균 발효물 및 이를 유효성분으로 함유하는 약학조성물, 식품조성물 및 화장료 조성물을 제공하는 것을 목적으로 한다.
In order to solve the above-mentioned problems, the present invention provides a fermented product of Lactobacillus acidus extract of Staphylococcus aureus, which is safe for human body and has excellent effect of improving skin wrinkles by fermenting the extract of Staphylococcus aureus with lactic acid bacteria, It is an object of the present invention to provide a composition and a cosmetic composition.
상기와 같은 목적을 달성하기 위하여, 본 발명은 우뭇가사리 주정추출물을 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB)(기탁번호: KCTC 12371BP)로 발효시킨 우뭇가사리 주정추출 유산균 발효물을 제공하는 것을 특징으로 한다.In order to achieve the above object, the present invention relates to a method for producing Pepsiococcus sp. pentosaceus BB) (Accession No .: KCTC 12371BP). The fermented product of Lactobacillus sp.
또한, 본 발명은 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선용 약학 조성물, 식품 조성물 및 화장료 조성물을 제공하는 것을 특징으로 한다.
The present invention also provides a pharmaceutical composition for improving skin wrinkles, a food composition, and a cosmetic composition, which comprises a fermented product of Lactobacillus acidus extract of Staphylococcus aureus as an active ingredient.
이하, 본 발명을 상세히 설명한다.Hereinafter, the present invention will be described in detail.
본 발명의 우뭇가사리 주정추출물을 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB) 균주로 발효시킨 유산균 배양물을 '우뭇가사리 주정추출 유산균 발효물'이라 정의하여 사용하기로 한다.
The cultivated lactic acid bacteria obtained by fermenting the extract of Phellodendron spp. Of the present invention with a strain of Pediococcus pentosaceus BB will be used as "fermented product of Phellodendron spp. Extract".
본 발명에 따른 균주를 분리하기 위하여 출원인이 분리하여 보유 중인 유산균 라이브러리를 대상으로 가장 높은 프로콜라겐 생성능을 가진 균주를 Procollagen Type I C-peptide(PIP) EIA kit(Takara, MK101)를 이용하여 프로콜라겐(procollagen) 타입 I C-펩타이드의 생성량을 정량하여 분리하였고, 연속 계대 배양을 통하여 순수 분리하였다. 상기 분리된 균주로부터 염색체 DNA를 추출하여 DNA 정제 키트(purification kit)를 이용하여 상기 균주로부터 DNA를 정제하였고, 이를 16S rRNA 분자의 양끝에 존재하는 보존성이 높은 염기서열 영역을 표적으로 삼아 PCR primer[27F (5'-agagtttgatcctggctcag-3')와 1492R (5'-ggctaccttgttacgactt-3')]를 이용하여 중합효소연쇄반응(PCR)을 실시하여 약 1.5kb 크기의 PCR 산물을 획득하였다. 상기 획득한 중합효소 연쇄반응 산물을 DNA 정제 키트를 이용하여 정제한 후, 염기서열 분석을 실시하였고, 온라인상에서 상동성 비교 프로그램인 BLAST를 사용하여 상동성을 검색하였다. 검색 결과, 상기 분리한 균주는 Pediococcus pentosaceus strain CTSPL1의 16s rRNA 유전자와 100％ 동일함(identities)을 확인하였다. 따라서, 상기 균주를 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB)로 명명하고, 한국생명공학연구원 생명자원센타에 2013년 2월 15일자(기탁번호: KCTC 12371BP)로 기탁하였다.
In order to isolate the strain according to the present invention, the strain having the highest pro-collagen production ability was isolated from the lactic acid bacteria library which was separated and retained by the applicant. Procollagen Type I C-peptide (PIP) EIA kit (Takara, MK101) The amount of procollagen type I C-peptide was quantified and separated, and purified by serial passaging. The chromosomal DNA was extracted from the isolated strain and the DNA was purified from the strain using a DNA purification kit. The DNA was purified using a PCR primer [ The PCR product was amplified by PCR using 5'-aggtttgatcctggctcag-3 ') and 1492R (5'-ggctaccttgttacgactt-3'). The obtained polymerase chain reaction product was purified using a DNA purification kit, followed by sequencing, and homology was searched using BLAST, a homology comparison program on-line. As a result of the search, the isolated strains were found to have 100% identities with the 16s rRNA gene of Pediococcus pentosaceus strain CTSPL1. Therefore, the strain was named Pediococcus pentosaceus BB and deposited with the Korea Life Science Research Institute, Bio-Resource Center on Feb. 15, 2013 (Accession No .: KCTC 12371BP).
이와 같은 방법으로 분리된 본 발명에 따른 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB) 균주의 균학적 성질과 미생물의 분류학적 동정은 Hammes 등[The prokaryotes, 1563-1578, 2nd Edition, pringer-Verlag Co. (1992)]의 방법에 따라 수행하였으며, API 50 CHL bacterial isolation kit(Biomerieux, France)을 이용하여 결과를 확인하였고, 이를 표 1[페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB) 균주의 생리적 및 생화학적 검사결과]에 나타내었다. 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB)균주는 그람양성 구균이며, 혐기적인 조건에서도 성장하며, 카탈라아제는 양성을 나타내었다.
The mycological properties of the Pediococcus pentosaceus BB strain according to the present invention and the taxonomic identification of the microorganisms according to the present invention are described in Hammes et al. [The prokaryotes, 1563-1578, 2nd Edition, Co. (1992)]. The results were confirmed using an API 50 CHL bacterial isolation kit (Biomerieux, France), and the results are shown in Table 1. The physiological and physiological characteristics of the Pediococcus pentosaceus BB strain Biochemical test results]. The Pediococcus pentosaceus BB strain was a Gram-positive staphylococcus, growing under anaerobic conditions, and the catalase was positive.
한편, 본 발명의 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부주름 개선효능을 갖는 약학적 조성물은 단독 또는 약제학적으로 사용되는 부형제들과 함께 약제학적으로 통상으로 사용되는 방법에 따라 정제, 캡슐제 등과 같은 제재형태로 제제화하여 사용될 수 있다.
On the other hand, the pharmaceutical composition having skin wrinkle-reducing effect containing the fermented product of the extract of L. acidophilus extract of the present invention having the skin wrinkle-reducing effect of the present invention as an active ingredient can be used as a pharmaceutical, usually in combination with excipients used alone or in pharmaceutical preparations And may be formulated in the form of tablets, capsules and the like.
사람의 경우, 통상적인 1일 투여량은 1~30mg/kg 체중의 범위일 수 있고, 1회 또는 수회로 나누어 투여할 수 있다. 그러나, 실제 투여량은 투여경로, 환자의 연령, 성별 및 체중, 건강상태 및 질환의 중증도 등의 여러 관련 인자에 비추어 결정되어야 한다.For humans, a typical daily dose may range from 1 to 30 mg / kg body weight, and may be administered in single or divided doses. However, the actual dosage should be determined in light of various relevant factors such as the route of administration, the age, sex and weight of the patient, the health condition, and the severity of the disease.
물론, 본 발명의 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선 효능을 갖는 약학조성물은 독성 및 부작용은 거의 없으므로 예방 목적으로 장기간 복용 시에도 안심하고 사용할 수 있는 약제이다.
Of course, the pharmaceutical composition having the skin wrinkle-reducing effect containing the fermented extract of L. acidophilus extract of the present invention having the skin wrinkle-reducing effect of the present invention has little toxicity and side effects, so that it can be safely used for long- It is medicine.
또한, 본 발명의 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 조성물은 식품, 식품첨가제, 음료, 음료첨가제, 발효유, 건강기능식품 등으로 사용될 수 있다. 식품, 식품첨가제, 음료, 음료첨가제, 또는 건강기능식품으로 사용되는 경우, 각종 식품류, 발효유, 육류, 음료수, 초콜렛, 스넥류, 과자류, 피자, 라면, 기타 면류, 껌류, 아이스크림류, 알코올 음료, 비타민 복합제, 주류 및 그 밖의 건강기능식품일 수 있으나, 이에 한정되는 것은 아니다.In addition, the composition containing the fermented extract of L. acidophilus extract having the skin wrinkle-reducing effect of the present invention as an active ingredient can be used as a food, a food additive, a beverage, a beverage additive, a fermented milk, a health functional food, When used as a food, a food additive, a beverage, a beverage additive, or a health functional food, it is possible to use various foods, fermented milk, meat, beverage, chocolate, snack, confectionery, pizza, ramen, other noodles, gums, ice cream, But may be, but not limited to, a combination, a mainstream and other health functional food.
특히, 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선 효능을 갖는 발효유는 우뭇가사리 주정추출 유산균 발효물, 유산균 배양액 및 혼합과즙시럽을 일정비율로 조합하여 150bar에서 균질한 후 10℃이하로 냉각한 후 용기에 포장하여 발효유를 제조한다.
Particularly, the fermented milk having the skin wrinkle-reducing effect containing the fermented product of the extract of L. acidophilus extract having the effect of improving the skin wrinkles is obtained by mixing the fermented product of L. acidophilus extracted with L. acidophilus, the culture of L. acidophilus and the mixed fruit syrup at a certain ratio, After cooling to below 10 ° C, it is packaged in a container to make fermented milk.
또한, 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선 효능을 갖는 기능성 음료는 혼합과즙시럽, 우뭇가사리 주정추출 유산균 발효물 및 물을 일정한 비율로 조합하여 150bar에서 균질한 후 10℃ 이하로 냉각한 후 유리병, 패트병 등 소포장 용기에 포장하여 기능성 음료를 제조한다.
In addition, functional beverages having skin wrinkle-reducing effect containing the fermented extract of L. acidophilus extract having the effect of improving the skin wrinkle are mixed at a constant ratio, After cooling to below 10 ° C, the beverage is packaged in small containers such as glass bottles and plastic bottles to produce functional beverages.
또한, 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선 효능을 갖는 건강기능식품은 우뭇가사리 주정추출 유산균 발효물을 포함하는 것 이외에 영양보조 성분으로 비타민 B1, B2, B5, B6, E 및 초산에스테르, 니코틴산 아미드, 올리고당 등이 첨가될 수 있으며 여타의 식품 첨가물이 첨가되어도 무방하다.
Further, in addition to health food with a wrinkle effect containing agar alcoholic extract of lactic acid bacteria fermented with a wrinkle effect as an active ingredient comprises an agar alcoholic extract lactic acid fermentation as a nutritional adjunct ingredients vitamin B 1, B 2 , B 5 , B 6 , E and acetic acid esters, nicotinic acid amides, oligosaccharides and the like may be added, and other food additives may be added.
또한, 본 발명의 피부 주름 개선 효능을 갖는 우뭇가사리 주정추출 유산균 발효물은 피부 주름 개선 효능을 갖는 화장품 조성물로 사용할 수 있다. 상기 화장료로 사용 시 본 발명에 따른 우뭇가사리 주정추출 유산균 발효물 외에 기타 화장료의 제형 또는 사용목적 등에 맞게 임의로 선정한 물질을 첨가할 수 있다. 예를 들어, 정제수, 유분, 계면활성제, 보습제, 고급 알코올, 증저점제, 킬레이트제, 색소, 지방산, 산화방지제, 방부제, 왁스, pH 조절제, 향료 등이 첨가될 수 있다. 또한 본 발명의 조성물로 제조되는 화장료의 제형은 특별히 제한되는 바가 없으며, 예를 들면, 영양 크림, 수렴 화장수, 유연 화장수, 로션, 에센스, 영양젤, 마사지 크림, 마스크 팩 등의 제형을 가질 수 있다.
In addition, the fermented extract of L. acidophilus extract of the present invention having the skin wrinkle-reducing effect of the present invention can be used as a cosmetic composition having an effect of improving skin wrinkles. When used as the above-mentioned cosmetics, in addition to the fermented product of Lactobacillus acidus extract extract of Staphylococcus aureus according to the present invention, a substance arbitrarily selected according to the purpose of formulation or use of other cosmetics may be added. For example, purified water, an oil, a surfactant, a moisturizer, a high alcohol, an evaporation agent, a chelating agent, a pigment, a fatty acid, an antioxidant, an antiseptic, a wax, a pH adjusting agent and a perfume may be added. The formulation of the cosmetic composition of the present invention is not particularly limited and may have formulations such as a nutritional cream, a convergent lotion, a soft lotion, a lotion, an essence, a nutritional gel, a massage cream, .
본 발명의 우뭇가사리 주정추출 유산균 발효물은 추출 과정이 간단하여 단시간에 대량으로 추출 가능할 뿐만 아니라 피부주름 개선 효능이 우수하여 피부주름 개선용 약학 조성물, 식품 조성물, 화장료 조성물 등으로 활용이 가능하다.
The fermented extract of L. acidophilus extract of the present invention of the present invention can be extracted as a large amount in a short time due to its simple extraction process, and can be used as a pharmaceutical composition for improving skin wrinkles, a food composition, a cosmetic composition and the like.
도 1은 인간 피부 섬유아세포에서 우뭇가사리 주정추출 유산균 발효물의 프로콜라겐 생성 촉진 효과를 측정한 그래프이다.
도 2는 우뭇가사리 주정추출 유산균 발효물의 자외선 조사에 의해 억제된 프로콜라겐 발현 회복효과를 측정한 그래프이다.
도 3은 우뭇가사리 주정추출 유산균 발효물의 자외선 조사에 의해 유도된 MMP-1 생성 억제 효과를 측정한 그래프이다.FIG. 1 is a graph showing the effect of promoting the production of procollagen by fermented lactic acid bacteria extracted from the mugwort extract of human skin fibroblasts.
2 is a graph showing the effect of restoring the expression of procollagen inhibited by ultraviolet irradiation of a fermented product of Lactobacillus sp.
FIG. 3 is a graph showing the effect of inhibiting MMP-1 production induced by ultraviolet irradiation of fermented product of Lactobacillus sp.
이하 실시예를 통하여 본 발명을 보다 상세하게 설명한다. 그러나 다음의 실시예는 본 발명의 범위를 한정하는 것은 아니며, 본 발명의 기술적 사상의 범위 내에서 당업자에 의한 통상적인 변화가 가능하다.
Hereinafter, the present invention will be described in more detail with reference to examples. However, the following embodiments are not intended to limit the scope of the present invention, and ordinary variations by those skilled in the art within the scope of the technical idea of the present invention are possible.
<실시예 1>≪ Example 1 >
우뭇가사리 주정추출물의 제조Preparation of Extracts of Ugukgasari
우뭇가사리 건조분말을 70%의 농도를 가진 주정(酒精)을 이용하여 침지시킨 후, 3시간 동안 50℃에서 반응시켜 추출을 실시하였다. 필터 백(filter bag)을 이용하여 우뭇가사리를 제거한 후, 여과된 추출물을 55℃에서 감압농축하였고, 이를 동결건조를 통하여 우뭇가사리 주정추출물을 제조하였다.
The dried powder was immersed in alcohol with a concentration of 70% and then subjected to extraction at 50 ° C for 3 hours. The extracts were filtered off with a filter bag, and the filtered extracts were concentrated under reduced pressure at 55 ° C.
<실시예 2>≪ Example 2 >
우뭇가사리 주정추출 유산균 발효물의 제조Preparation of Fermented Lactobacillus extract
상기 실시예 1의 우뭇가사리 주정추출물에 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB) 균주를 1.0 Ⅹ 106cfu/㎖로 접종하여 37℃에서 48시간 동안 배양하여 우뭇가사리 주정추출 유산균 발효물을 제조하였다.
The Pepsiococcus pentosaceus BB strain was inoculated at 1.0 × 10 6 cfu / ml to the extract of the mugwort extract of Example 1 and cultured at 37 ° C. for 48 hours to prepare a fermented product of Mugwort grape extract .
<실시예 3>≪ Example 3 >
우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 약학적 조성물의 제조Preparation of Pharmaceutical Composition Containing Fermented Lactic Acid Bacteria Extracted from Sugarcane Extract as Active Ingredient
본 발명의 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 하는 약학조성물의 제제 예를 설명하나, 본 발명은 이를 한정하고자 함이 아닌 단지 구체적으로 설명하고자 함이다.
The pharmaceutical composition of the present invention comprising the fermented product of Lactobacillus extract of Lactobacillus japonica extract of the present invention is described, but the present invention is not intended to be limited thereto but is specifically explained.
정제의 제조Manufacture of tablets
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물의 동결건조 분말 100㎎, 옥수수전분 100㎎, 유당 100㎎ 및 폴리비닐피롤리돈 97㎎을 균질하게 혼합하여 습식과립법으로 과립화하고 스테아린산 마그네슘 2㎎을 가하여 혼합한 후 1정이 400㎎이 되도록 타정하였다.
100 mg of the lyophilized powder of fermented extract of Lactobacillus acidophilus extract of Example 2, 100 mg of corn starch, 100 mg of lactose and 97 mg of polyvinylpyrrolidone were homogeneously mixed and granulated by the wet granulation method, and 2 mg of magnesium stearate After mixing, the mixture was compressed to 400 mg per tablet.
캡슐제의 제조Preparation of capsules
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물의 동결건조 분말 100㎎, 옥수수 전분 100㎎, 유당 100㎎, 스테아린산 마그네슘 2㎎을 완전히 혼합한 후 통상의 캡슐제의 제조방법에 따라서 경질 젤라틴 캡슐에 충전하여 캡슐제를 제조하였다.
100 mg of the lyophilized powder of fermented product of the extract of L. acidophilus extract of Example 2, 100 mg of corn starch, 100 mg of lactose and 2 mg of magnesium stearate were thoroughly mixed and filled into hard gelatin capsules according to the conventional preparation method of capsules Capsules were prepared.
<실시예 4><Example 4>
우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 발효유의 제조Preparation of Fermented Milk Containing Fermented Lactic Acid Bacteria as an Active Ingredient
유산균 배양액과 본 발명의 우뭇가사리 주정추출 유산균 발효물 및 혼합과즙시럽으로 구성된 발효유를 제조하는 방법은 다음과 같다.A method for preparing a fermented milk composed of a culture medium of lactic acid bacteria and fermented lactic acid bacteria of the present invention and a mixed fruit juice syrup is as follows.
먼저, 유산균 배양액은 원유 95.36중량%와 탈지분유(또는 혼합분유) 4.6중량%를 교반하여 15℃에서의 비중은 1.0473~1.0475, 적정산도는 0.200~0.220%, pH는 6.65~6.70, 20℃에서의 브릭스(Brixo)는 16.3~16.5% 정도가 되도록 혼합하였다. 혼합 후에 이를 UHT 열처리(135℃에서 2초간 살균)하고 40℃로 냉각한 뒤, 페디오코커스 펜토사시우스(Pediococcus pentosaceus)균과 유당분해효소(Valley laboratory, USA)를 각기 0.02중량%씩 첨가하고 6시간 동안 배양하여 BCP 배지에서의 총 유산균수가 1.0 Ⅹ 109 cfu/㎖ 이상, 적정산도가 0.89~0.91%, pH는 4.55~4.65가 되도록 하여 제조하였다.The lactic acid bacteria culture solution was prepared by stirring 95.36 wt% of crude oil and 4.6 wt% of skim milk powder (or mixed powdered milk) and measuring specific gravity at 15 ° C from 1.0473 to 1.0475, a titratable acidity of 0.200 to 0.220%, pH of 6.65 to 6.70, of Brix (Brix o) were mixed such that the degree of 16.3 ~ 16.5%. After mixing, the mixture was heat-treated with UHT (sterilized at 135 ° C for 2 seconds), cooled to 40 ° C, added with 0.02% by weight of Pediococcus pentosaceus and a lyotropic enzyme (Valley laboratory, USA) And cultured for 6 hours to obtain a total amount of lactic acid bacteria in the BCP medium of 1.0 × 10 9 cfu / ml or more, a titratable acidity of 0.89 to 0.91%, and a pH of 4.55 to 4.65.
그 다음, 혼합과즙시럽은 액상과당 13중량%, 백설탕 5중량%, 혼합과즙농축액 56Brixo 10.9중량%, 펙틴 1.0중량%, 후레쉬 후르츠 믹스 에센스 0.1중량% 및 정제수 70중량%를 30~35℃에서 교반하여 혼합한 후 UHT 열처리(135℃에서 2초간 살균)한 후 냉각하여 제조하였다.Next, the mixed fruit juice syrup was prepared by mixing 13 wt% of liquid fructose, 5 wt% of white sugar, 10.9 wt% of mixed juice concentrate 56Brix o , 1.0 wt% of pectin, 0.1 wt% of fresh fruit mix essence and 70 wt% of purified water at 30-35 캜 Mixed with stirring, heat-treated with UHT (sterilized at 135 ° C for 2 seconds), and cooled.
그런 다음, 상기 유산균배양액 69.5중량%와 상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물의 동결건조 분말 0.1중량% 및 상기 혼합과즙시럽 30.4중량%를 조합하여 150bar에서 균질한 후 10℃ 이하로 냉각하여 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유한 발효유를 제조하였다.
Then, 69.5% by weight of the culture liquid of lactic acid bacteria, 0.1% by weight of the lyophilized powder of the fermented extract of the extract of L. acidum of Example 2, and 30.4% by weight of the mixed fruit juice syrup were combined and homogenized at 150 bar, Fermented milk containing fermented extract of lactic acid fermented with alcohol as an active ingredient was prepared.
<실시예 5>≪ Example 5 >
우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 기능성 음료의 제조Preparation of Functional Beverage Containing Fermented Lactic Acid Bacteria Extracted with Sugar Extract as an Active Ingredient
본 발명의 우뭇가사리 주정추출 유산균 발효물 및 혼합과즙시럽으로 구성된 기능성 음료를 제조하는 방법은 다음과 같다.The method for producing a functional beverage comprising the fermented product of the extract of L. acidophilus extract of the present invention and the mixed fruit syrup of the present invention is as follows.
먼저, 혼합과즙시럽은 액상과당 13중량%, 백설탕 2.5중량%, 갈색설탕 2.5중량%, 혼합과즙농축액 56Brixo 10.9중량%, 펙틴 1.0중량%, 후레쉬 후르츠 믹스 에센스 0.1중량% 및 정제수 70중량%를 30~35℃에서 교반하여 혼합한 후 UHT 열처리(135℃에서 2초간 살균)한 후 냉각하여 제조하였다.
First, a mixed juice syrup was 13% by weight of liquid fructose, white sugar, 2.5 wt%, brown sugar 2.5% by weight, mixed juice concentrate 56Brix o 10.9% by weight of pectin, 1.0% by weight, fresh 0.1% by weight fruit mix essence and purified water 70% The mixture was stirred at 30 to 35 ° C, mixed, and heat-treated by UHT (sterilized at 135 ° C for 2 seconds) and cooled.
그리고, 상기의 방법으로 제조된 혼합과즙시럽 30.4중량%와 상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물의 동결건조분말 0.1중량% 및 나머지 정제수 69.5중량%를 조합하여 150bar에서 균질한 후 10℃ 이하로 냉각한 후 이를 유리병, 패트병 등 소포장 용기에 포장하여 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 기능성 음료를 제조하였다.
Then, 30.4% by weight of the mixed fruit juice syrup prepared by the above method was combined with 0.1% by weight of the lyophilized powder of the fermented product of the extract of L. acidophilus extract of Example 2 and 69.5% by weight of the remaining purified water, homogenized at 150 bar, After cooling, the product was packed in a small bottle container such as a glass bottle or a plastic bottle to produce a functional beverage containing the fermented product of L. acidophilus extracted from Aspergillus oryzae.
<실시예 6>≪ Example 6 >
우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 건강기능식품의 제조Preparation of Health Functional Foods containing Fermented Lactobacillus fermented Extracts as Active Ingredients
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물의 동결건조 분말 0.1중량%에 영양보조성분(비타민 B1, B2, B5, B6, E 및 초산에스테르, 니코틴산 아미드) 및 올리고당을 상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물의 동결건조 분말 100중량부에 대하여 10중량부가 되도록 첨가하여 고속회전 혼합기에서 혼합하였다. 상기 혼합물 100중량부에 대하여 멸균 정제수 10중량부를 첨가, 혼합하고 직경 1~2mm의 과립상으로 성형하였다. 상기 성형된 과립은 40~50℃의 진공건조기에서 건조시킨 후 12~14 메쉬(mesh)를 통과시켜 균일하게 과립을 제조하였다. 상기와 같이 제조된 과립은 적당량씩 압출 성형되어 정제 또는 분말로 되거나 경질캡슐에 충전되어 경질캡슐제품으로 제조하였다.
(Vitamin B 1 , B 2 , B 5 , B 6 , E and acetic acid ester, nicotinic acid amide) and oligosaccharide were added to 0.1% by weight of the lyophilized powder of the fermented product of the extract of L. acidophilus of Example 2, Was added in an amount of 10 parts by weight based on 100 parts by weight of the lyophilized powder of the fermented extract of L. acidophilus extract of L. acidus, and the mixture was mixed in a high-speed rotary mixer. To 100 parts by weight of the mixture, 10 parts by weight of sterilized purified water was added and mixed to form granules having a diameter of 1 to 2 mm. The molded granules were dried in a vacuum drier at 40 to 50 DEG C and then passed through 12 to 14 mesh to prepare uniform granules. The granules thus prepared were extruded in suitable amounts to be purified or powdered or filled into hard capsules to prepare hard capsule products.
<실시예 7>≪ Example 7 >
우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 화장료의 제조Preparation of Cosmetic Ingredients Containing Fermented Lactic Acid Bacteria Extract Extracted from Sugarcane Extract as an Active Ingredient
유연화장수Softening longevity
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 0.01중량%, 글리세린 3중량%, 부틸렌 글리콜 2중량%, 프로필렌 글리콜 2중량%, 카복시비닐폴리머 0.1중량%, 에탄올 10중량%, 트리에탄올아민 0.1중량% 및 잔량의 방부제, 미량색소, 미량향료 및 미량정제수를 혼합하여 통상의 유연화장수 제조방법에 따라 제조하였다.
Example 2 was carried out in the same manner as in Example 2, except that the fermented extract of L. acidophilus extract of L. rudder extract, 3 wt% of glycerin, 2 wt% of butylene glycol, 2 wt% of propylene glycol, 0.1 wt% of carboxyvinyl polymer, 10 wt% of ethanol, And minor amounts of preservatives, trace pigments, trace flavors and trace amounts of purified water were mixed and prepared according to a conventional method for preparing softened water.
영양화장수Nutritional lotion
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 0.01중량%, 밀납 4중량%, 폴리소르베이트 60 1.5중량%, 소르비탄세스퀴올레이트 0.5중량%, 유동파라핀 5중량%, 스쿠알란 5중량%, 카프릴릭/카프릭 트리글리세라이드 5중량%, 글리세린 3중량%, 부틸렌 글리콜 3중량%, 프로필렌 글리콜 3중량%, 카복시비닐폴리머 0.1중량%, 트리에탄올아민 0.2중량%, 잔량의 방부제, 미량색소, 미량향료 및 미량정제수를 혼합하여 통상의 영양화장수 제조방법에 따라 제조하였다.
The fermented product of the extract of L. acidophilus extract of Example 2, a fermented product of 0.01% by weight of waxy fermented product, 0.5% by weight of sorbitan sesquioleate, 5% by weight of liquid paraffin, 5% by weight of squalane, 5% by weight of glycerin / capric triglyceride, 3% by weight of glycerin, 3% by weight of butylene glycol, 3% by weight of propylene glycol, 0.1% by weight of carboxyvinyl polymer, 0.2% by weight of triethanolamine, And a trace amount of purified water were mixed and prepared according to a conventional method for producing a nutritional lotion.
영양크림Nourishing cream
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 0.01중량%, 밀납 10중량%, 폴리소르베이트 60 1.5중량%, 소르비탄세스퀴올레이트 0.5중량%, 유동파라핀 10중량%, 스쿠알란 5중량%, 카프릴릭/카프릭 트리글리세라이드 5중량%, 글리세린 5중량%, 부틸렌 글리콜 3중량%, 프로필렌 글리콜 3중량%, 트리에탄올아민 0.2중량% 및 잔량의 방부제, 미량색소, 미량향료 및 미량정제수를 혼합하여 통상의 영양크림 제조방법에 따라 제조하였다.
The fermented product of the extract of L. acidophilus extract of Example 2, the fermentation product of 0.01% by weight of waxy fermented alcohol extract, 10% by weight of wax, 1.5% by weight of polysorbate 60, 0.5% by weight of sorbitan sesquioleate, 10% by weight of liquid paraffin, 5% by weight of glycerin / capric triglyceride, 5% by weight of glycerin, 3% by weight of butylene glycol, 3% by weight of propylene glycol, 0.2% by weight of triethanolamine and the balance of preservatives, trace pigments, Lt; RTI ID = 0.0 > cream. ≪ / RTI >
맛사지 크림Massage Cream
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 0.01중량%, 밀납 10중량%, 폴리소르베이트 60 1.5중량%, 소르비탄세스퀴올레이트 0.8중량%, 유동파라핀 40중량%, 스쿠알란 5중량%, 카프릴릭/카프릭 트리글리세라이드 4중량%, 글리세린 5중량%, 부틸렌 글리콜 3중량%, 프로필렌 글리콜 3중량%, 트리에탄올아민 0.2중량% 및 잔량의 방부제, 미량색소, 미량향료 및 미량정제수를 혼합하여 통상의 맛사지크림 제조방법에 따라 제조하였다.
The fermented product obtained in Example 2 was prepared by mixing 0.01% by weight of fermented extract of L. acidum extract, 1.5% by weight of polysorbate 60, 0.8% by weight of sorbitan sesquioleate, 40% by weight of liquid paraffin, 5% by weight of squalane, A mixture of 4% by weight of ricin / capric triglyceride, 5% by weight of glycerin, 3% by weight of butylene glycol, 3% by weight of propylene glycol and 0.2% by weight of triethanolamine and the balance of preservatives, trace pigments, Of the present invention.
팩pack
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 0.01중량%, 폴리비닐알콜 13중량%, 소듐카복시메틸셀룰로스 0.2중량%, 알란토인 0.1중량%, 에탄올 5중량%, 노닐페닐에테르 0.3중량% 및 잔량의 방부제, 미량색소, 미량향료 및 미량정제수 를 혼합하여 통상의 팩 제조방법에 따라 제조하였다.
Except that 0.01% by weight of the fermented extract of L. acidophilus extract of Example 2, 13% by weight of polyvinyl alcohol, 0.2% by weight of sodium carboxymethyl cellulose, 0.1% by weight of allantoin, 5% by weight of ethanol, 0.3% by weight of nonylphenyl ether, , A trace pigment, a trace amount of flavor, and a trace amount of purified water were mixed and prepared according to a conventional pack preparation method.
<비교예 1>≪ Comparative Example 1 &
개우무 주정추출물의 제조Manufacture of Dogwood Extracts
대한민국 특허등록 제1096206호(발명의 명칭: 개우무 추출물을 유효성분으로 포함하는 피부 상태 개선용 조성물)의 방법에 따라 개우무 주정추출물을 제조하였다. Korean Patent Registration No. 1096206 (the title of the invention: composition for improving skin condition containing an extract of Asanovirus as an active ingredient) was prepared.
즉, 음건한 개우무 전체를 95% 에탄올 수용액으로 5시간씩 3회 환류추출하고 냉침한 후, 와트만(Whatman) 여과지로 여과하였다. 여과된 농축물을 50℃ 이하에서 감압농축 및 동결건조하여 개우무 주정추출물을 제조하였다.
That is, the whole shade of shade was refluxed for 3 hours with 95% aqueous ethanol solution for 5 hours, followed by cooling down and filtering with Whatman filter paper. The filtered concentrate was concentrated under reduced pressure at 50 DEG C or below and lyophilized to prepare a crude extract of the crude extract.
<비교예 2>≪ Comparative Example 2 &
우뭇가사리 주정추출물의 제조Preparation of Extracts of Ugukgasari
건조한 우뭇가사리를 70% 주정을 이용하여 4시간 동안 침지 후, 70℃에서 6시간 동안 환류추출하여 냉침한 후, 와트만(Whatman) No 1. 여과지로 여과하였다. 여과된 농축물을 50℃ 이하에서 감압농축 및 동결건조하였다 .
The dried mugwort was immersed in a 70% alcohol solution for 4 hours, refluxed at 70 ° C for 6 hours, cooled, and filtered through a Whatman No.1 filter paper. The filtered concentrate was concentrated under reduced pressure and freeze-dried at 50 DEG C or lower.
<시험예 1>≪ Test Example 1 >
인간 피부 섬유아세포에서 우뭇가사리 주정추출 유산균 발효물의 프로콜라겐 생성 촉진 효과 측정Measuring effect of promoting the production of pro-collagen in fermented lactic acid bacteria extracted from mugwort pulp extract from human skin fibroblast
인간 피부 섬유아세포인 Hs68 세포[입수처: 미국세포주은행(ATCC)]를 10% 우태아 혈청(fetal bovine serum; FBS)에 페니실린 7.5㎎/L, 스트렙토마이신 7.5㎎/L를 함유하고 있는 DMEM 배지를 사용하여 5% 농도의 CO2 배양기에서 37℃로 24시간 동안 배양하였다. 배지 조성물들은 모두 GIBCO BRL(Grand Island, NY, USA)사 제품을 사용하였다.
Human skin fibroblast Hs68 cells (obtained from American Cell Line Bank (ATCC)) were cultured in DMEM medium containing 7.5 mg / L of penicillin and 7.5 mg / L of streptomycin in 10% fetal bovine serum (FBS) Were cultured in a 5% CO 2 incubator at 37 ° C for 24 hours. All of the medium compositions were purchased from GIBCO BRL (Grand Island, NY, USA).
인간 피부 섬유아세포에서 프로콜라겐(procollagen) 생성량을 측정하기 위해서 Procollagen Type I C-peptide(PIP) EIA kit(Takara, MK101)를 이용하여 프로콜라겐(procollagen) 타입 I C-펩타이드의 생성량을 정량하였다.
Procollagen type I C-peptide (PIP) EIA kit (Takara, MK101) was used to quantify procollagen type I C-peptide production in human skin fibroblasts.
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물을 각각 1㎍/㎖, 10㎍/㎖씩 상기 배양된 인간 피부 섬유아세포인 Hs68 세포에 1시간 동안 전 처리한 후 UVB 50mJ/cm2를 조사하여 48시간 동안 배양하였다. 배양한 세포의 배지로부터 20㎕을 취하여 배지로 분비된 프로콜라겐의 양을 측정하였다.The second embodiment of the lactic acid bacteria fermentation extract agar alcohol each 1㎍ / ㎖, for 1 hour to 10㎍ / ㎖ by the cultured cells of Hs68 human skin fibroblast cells after pre-treatment by irradiating UVB 50mJ / cm 2 48 Lt; / RTI > The amount of procollagen secreted in the medium was measured by taking 20 占 퐇 from the medium of the cultured cells.
그 결과를 도 1에 나타내었다.The results are shown in Fig.
인간 피부 섬유아세포인 Hs68 세포에 우뭇가사리 주정추출 유산균 발효물을 무처리하고, UVB를 조사 하지 않은 군을 양성대조군(무처리군)으로 하였고, 인간 피부 섬유아세포인 Hs68 세포에 UVB 50mJ/cm2만을 조사한 군을 음성대조군(자외선)으로 하였으며, 우뭇가사리 주정추출 유산균 발효물 1㎍/㎖ 처리와 UVB 50mJ/cm2를 조사산 군을 실험군(1㎍/㎖) 및 우뭇가사리 주정추출 유산균 발효물 10㎍/㎖ 처리와 UVB 50mJ/cm2를 조사한 군을 실험군(10㎍/㎖)으로 하였다.
The human skin fibroblasts, Hs68 cells, were treated with no fermented Lactobacillus fermentation extract and the positive control group (untreated group) was not irradiated with UVB. Hs68 cells, which were human skin fibroblasts, were irradiated with UVB 50 mJ / cm 2 (1 ㎍ / ㎖) and UVB 50 mJ / cm 2 were irradiated with 1 ㎍ / ㎖ of lactic acid fermented product of Staphylococcus aureus extract and 10 ㎍ / ㎖ of fermented extract of Staphylococcus aureus extract. Ml and UVB 50 mJ / cm 2 were irradiated to the test group (10 μg / ml).
도 1에서 확인할 수 있는 바와 같이, 본 발명의 우뭇가사리 주정추출 유산균 발효물은 자외선 조사로 억제된 프로콜라겐(procollagen) 생성량을 효과적으로 증가시키는 것을 확인할 수 있었다.
As can be seen from FIG. 1, it was confirmed that the fermented product of L. acidophilus extract of the present invention effectively increases the amount of procollagen inhibited by ultraviolet irradiation.
<시험예 2> ≪ Test Example 2 &
우뭇가사리 주정추출 유산균 발효물의 자외선 조사에 의해 억제된 프로콜라겐 발현 회복효과 측정Measurement of the recovery effect of procollagen expression inhibited by ultraviolet irradiation of lactic acid fermented extract
인간 피부 섬유아세포인 Hs68 세포에서 우뭇가사리 주정추출 유산균 발효물이 자외선 조사에 의해 억제된 프로콜라겐 발현을 회복시키는지 측정하기 위해서 웨스턴 블랏팅 분석법(western blotting analysis)[Upham, B. L., Kang, K. S., Cho, H. Y., & Trosko, J. E. Carcinogenesis 18:37-42, 1997]을 이용하여 분석하였다.BL, Kang, KS, KS, and Cho, KS, in order to determine whether the fermented product of L. acidophilus extract from the human skin fibroblasts, Hs68 cells, restored the inhibition of the inhibition of procollagen by ultraviolet irradiation , HY, & Trosko, JE Carcinogenesis 18: 37-42, 1997].
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물을 각각 1㎍/㎖, 10㎍/㎖씩 상기 배양된 인간 피부 섬유아세포인 Hs68 세포에 처리하고 1시간 후 UVB 50mJ/cm2를 조사하여 48시간 동안 배양하였다. 배양된 세포로부터 단백질을 추출하기 위하여 RIPA buffer를 사용하였다. 단백질 함량은 DC assay kit[Bio-Rad Corp., Richmond, CA, USA]를 이용하여 결정하였다. 각각의 단백질 추출물들로부터 약 10㎍에 해당하는 단백질을 10% SDS-PAGE에 넣어 전기 영동하여 분리하였다. 프로콜라겐은 폴리클론 항체[Santacruz biotechnology, inc., Santa Cruz, CA, USA]를 이용하여 반응시킨 후, ECL 키트[Amersham, Life Science, Denver, USA]를 이용하여 감지하였다.The cultured human skin fibroblasts Hs68 cells were treated with 1 μg / ml and 10 μg / ml, respectively, of the fermented extract of L. acidophilus of Example 2 in the above Example 2, irradiated with UVB 50 mJ / cm 2 for 1 hour, Lt; / RTI > RIPA buffer was used to extract proteins from the cultured cells. Protein content was determined using a DC assay kit [Bio-Rad Corp., Richmond, CA, USA]. From each of the protein extracts, about 10 μg of the protein was separated by electrophoresis in 10% SDS-PAGE. Procollagen was reacted with a polyclonal antibody (Santacruz biotechnology, Inc., Santa Cruz, Calif., USA) and detected using an ECL kit (Amersham, Life Science, Denver, USA).
그 결과를 도 2에 나타내었다.The results are shown in Fig.
레인 1(Lane 1)은 무처리 대조군, Lane 1 (Lane 1) was an untreated control group,
레인 2(Lane 2)는 자외선(50mJ/cm2) 조사군,
레인 3(Lane 3)은 우뭇가사리 주정추출 유산균 발효물(1㎍/㎖)과 자외선(50 mJ/cm2) 처리군
레인 4(Lane 4)는 우뭇가사리 주정추출 유산균 발효물(10㎍/㎖)과 자외선(50 mJ/cm2) 처리군을 나타낸다.
베타액틴(β-actin)은 동일한 양의 세포로 실험하였음을 나타낸다.
Beta-actin indicates experimentation with the same amount of cells.
도 2에서 확인할 수 있는 바와 같이, 레인 2에서와 같이 자외선 조사에 의해 세포 내 프로콜라겐의 발현량이 감소한 반면에, 레인 3과 레인 4와 같이 우뭇가사리 주정추출 유산균 발효물을 처리한 군에서는 우뭇가사리 주정추출 유산균 발효물의 농도가 증가함에 따라 억제된 프로콜라겐의 발현이 점차 회복되는 것을 확인할 수 있었다.
As can be seen from FIG. 2, the expression level of intracellular procollagen was decreased by ultraviolet irradiation as in
<시험예 3>≪ Test Example 3 >
우뭇가사리 주정추출 유산균 발효물의 자외선 조사에 의해 유도된 MMP-1 생성 억제 효과 측정Measurement of Inhibitory Effect on MMP-1 Production Induced by Ultraviolet Irradiation of Fermented Lactic Acid Bacteria Extracted from Staphylococcus aureus
인간 피부 섬유아세포인 Hs68 세포에서 우뭇가사리 주정추출 유산균 발효물이 자외선 조사에 의해 유도된 MMP-1(Matrix metalloproteinase-1) 생성량을 억제시키는지 측정하기 위해서 Amersham Matrix metalloproteinase-1 ELISA System(GE Healthcare, RPN2610)을 사용하여 MMP-1의 생성량을 정량하였다.
(Amersham Matrix metalloproteinase-1 ELISA System (GE Healthcare, RPN2610, USA) was used to determine whether the fermented extract of L. acidophilus from human skin fibroblasts Hs68 cells inhibited the production of MMP-1 (Matrix metalloproteinase- ) Was used to quantify the amount of MMP-1 produced.
상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물을 각각 1㎍/㎖, 10㎍/㎖로 인간 피부 섬유아세포인 Hs68 세포에 1시간 동안 전 처리한 후 UVB 50mJ/cm2를 조사하여 48시간 동안 배양하였다. 배양한 세포의 배지로부터 100㎕을 취하여 배지로 분비된 MMP-1의 양을 측정하였다.
The second embodiment of the lactic acid bacteria fermentation
그 결과를 도 3에 나타내었다.The results are shown in Fig.
레인 1(Lane 1)은 무처리 대조군, Lane 1 (Lane 1) was an untreated control group,
레인 2(Lane 2)는 자외선(50mJ/cm2) 조사군,
레인 3(Lane 3)은 우뭇가사리 주정추출 유산균 발효물(1㎍/㎖)과 자외선(50 mJ/cm2) 처리군
레인 4(Lane 4)는 우뭇가사리 주정추출 유산균 발효물(10㎍/㎖)과 자외선(50 mJ/cm2) 처리군을 나타낸다.
도 3에서 확인할 수 있는 바와 같이, 레인 2의 자외선 처리군은 자외선 조사에 의해 MMP-1 생성량이 현저히 증가한 반면에, 레인 3 및 레인 4의 우뭇가사리 주정추출 유산균 발효물을 처리한 군에서는 우뭇가사리 주정추출 유산균 발효물의 농도가 증가함에 따라 자외선에 의해 증가한 MMP-1 생성을 효과적으로 억제하는 것을 확인할 수 있었다.
As can be seen from FIG. 3, the amount of MMP-1 produced by ultraviolet irradiation was significantly increased in the ultraviolet treatment group of
<시험예 4><Test Example 4>
콜라겐 합성 증가율은 상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 10㎍/㎖, 상기 비교예 1의 개우무 주정추출물 10㎍/㎖, 상기 비교예 2의 우뭇가사리 주정 추출물 50㎍/㎖을 사용한 것을 제외하고는 상기 시험예 1과 동일한 방법으로 측정하여 정량하였다.
The collagen synthesis increase rate was the same as that of Example 2 except that 10 μg / ml of the fermented extract of the extract of the extract of the extract from the extract of Example 2, 10 μg / ml of the extract of the crude extract of Comparative Example 1 and 50 μg / Were measured and quantified in the same manner as in Test Example 1 above.
콜라게네이즈 억제율은 상기 실시예 2의 우뭇가사리 주정추출 유산균 발효물 10㎍/㎖, 상기 비교예 1의 개우무 주정추출물 10㎍/㎖, 상기 비교예 2의 우뭇가사리 주정 추출물 50㎍/㎖을 사용한 것을 제외하고는 상기 시험예 3과 동일한 방법으로 측정하여 정량하였다.The collagenase inhibition rate was determined by using 10 μg / ml of the fermented extract of Mugwort marjoram extract obtained from Example 2, 10 μg / ml of the crude mugwort alcohol extract of Comparative Example 1 and 50 μg / ml of the mugwort extract of Comparative Example 2 Except that the concentration was measured and quantified in the same manner as in Test Example 3 above.
그 결과를 표 2에 나타내었다.
The results are shown in Table 2.
상기 표 2에서 확인할 수 있는 바와 같이, 본 발명의 실시예 2(우뭇가사리 주정추출 유산균 발효물)는 콜라겐 합성을 증가시킴과 동시에 콜라겐 분해에 관여하는 콜라게네이즈(MMP1)의 활성을 억제한다는 것을 알 수 있었다.
As can be seen from the above Table 2, Example 2 (fermented product of Lactobacillus acidus extract extract of Staphylococcus aureus) of the present invention increased collagen synthesis and inhibited the activity of collagenase (MMP1) involved in collagen degradation I could.
또한, 본 발명의 실시예 2(우뭇가사리 주정추출 유산균 발효물)는 비교예 1(개우무 주정추출물)과 비교예 2(우뭇가사리 주정추출물)에 비해 월등히 높은 콜라겐합성 효능과 콜라케네이즈 억제효능을 가짐을 알 수 있었다.
In addition, Example 2 of the present invention (fermented product of Lactobacillus acidophilus extract extract of Staphylococcus aureus) has a significantly higher collagen synthesis effect and collagenase inhibitory effect than Comparative Example 1 (dogwood extract) and Comparative Example 2 (Staphylococcus aureus extract) And it was found.
이러한 사실로부터 본 발명의 우뭇가사리 주정추출 유산균 발효물(실시예 2)은 우뭇가사리 주정추출물을 페디오코커스 펜토사시우스 BB(Pediococcus pentosaceus BB) 균주로 발효시킴으로써 같은 속인 개우무 주정추출물(비교예 1)과 우뭇가사리 주정추출물(비교예 2)에 비하여 피부주름 개선 효능이 월등히 우수함을 알 수 있었으며, 이러한 주름 개선 효과는 농도 의존적임을 확인할 수 있었다.
From these facts, the fermented product of L. acidophilus extract of the present invention (Example 2) of the present invention can be obtained by fermenting the extract of L. acidophilus as a strain of Pediococcus pentosaceus BB, It was found that the effect of improving skin wrinkles was remarkably superior to that of the extract of mugwort gris (Comparative Example 2), and it was confirmed that the effect of improving wrinkles was concentration-dependent.
Claims (6)
A fermented product of Lactobacillus sp. Extract obtained by fermentation of Lactobacillus sp. Extract with Pediococcus pentosaceus BB (Accession No .: KCTC 12371BP).
A pharmaceutical composition for improving skin wrinkles containing the fermented product of extract of Lactobacillus sp.
A food composition for improving skin wrinkles containing the fermented product of Lactobacillus extract extract of L. japonica extract of claim 1 as an active ingredient.
A cosmetic composition for improving skin wrinkles containing the fermented product of extract of Lactobacillus acidus extract of the present invention as an active ingredient.
상기 식품 조성물은 발효유, 기능성 음료 및 건강기능식품 중에서 선택된 어느 하나의 제형을 가지는 것을 특징으로 하는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선용 식품 조성물.
The method of claim 3,
A food composition for improving skin wrinkles, comprising the fermented product of a fermented milk, a functional beverage, and a health functional food, wherein the food composition is one selected from the group consisting of fermented milk, functional beverage and health functional food.
상기 화장료 조성물은 영양 크림, 수렴 화장수, 유연 화장수, 로션, 에센스, 영양젤, 마스트 쉬트, 마사지 크림 및 마스크 팩 중에서 선택된 어느 하나의 제형을 가지는 것을 특징으로 하는 우뭇가사리 주정추출 유산균 발효물을 유효성분으로 함유하는 피부 주름 개선용 화장료 조성물.5. The method of claim 4,
The cosmetic composition according to any one of claims 1 to 3, wherein the cosmetic composition has any one of a nutritional cream, a convergent lotion, a soft lotion, a lotion, an essence, a nutritional gel, a mast sheet, a massage cream and a mask pack. A cosmetic composition for improving skin wrinkles.
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| WO2016162648A1 (en) | 2015-04-10 | 2016-10-13 | Societe D'exploitation De Produits Pour Les Industries Chimiques Seppic | Method for culturing cells of acrochaetium moniliforme red algae, method for obtaining an extract of the biomass thereof, and use of same in cosmetics |
| WO2018060567A1 (en) | 2016-09-30 | 2018-04-05 | Societe D'exploitation De Produits Pour Les Industries Chimiques Seppic | Process for preventing or slowing the appearance of unattractive signs, generated by pollutants present in the atmosphere, on the skin, the scalp, the hair or the mucous membranes |
| KR101901764B1 (en) * | 2016-11-30 | 2018-09-27 | 주영철 | Manufacturing method of pill-type Lactic acid bacteria complex |
| KR20190101110A (en) | 2018-02-22 | 2019-08-30 | 주식회사 휴모트메디컬 | Mask bead and method for manufacturing it |
| WO2020226470A1 (en) * | 2019-05-09 | 2020-11-12 | 한국 한의학 연구원 | Composition for ameliorating skin wrinkles and moisturizing skin, comprising pediococcus inopinatus as active ingredient |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2016162648A1 (en) | 2015-04-10 | 2016-10-13 | Societe D'exploitation De Produits Pour Les Industries Chimiques Seppic | Method for culturing cells of acrochaetium moniliforme red algae, method for obtaining an extract of the biomass thereof, and use of same in cosmetics |
| US11224627B2 (en) | 2015-04-10 | 2022-01-18 | Societe D'exploitation De Produits Pour Les Industries Chimiques Seppic | Method for culturing cells of Acrochaetium moniliforme red algae, method for obtaining an extract of the biomass thereof, and use of same in cosmetics |
| WO2018060567A1 (en) | 2016-09-30 | 2018-04-05 | Societe D'exploitation De Produits Pour Les Industries Chimiques Seppic | Process for preventing or slowing the appearance of unattractive signs, generated by pollutants present in the atmosphere, on the skin, the scalp, the hair or the mucous membranes |
| KR101901764B1 (en) * | 2016-11-30 | 2018-09-27 | 주영철 | Manufacturing method of pill-type Lactic acid bacteria complex |
| KR20190101110A (en) | 2018-02-22 | 2019-08-30 | 주식회사 휴모트메디컬 | Mask bead and method for manufacturing it |
| WO2020226470A1 (en) * | 2019-05-09 | 2020-11-12 | 한국 한의학 연구원 | Composition for ameliorating skin wrinkles and moisturizing skin, comprising pediococcus inopinatus as active ingredient |
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