KR101944013B1 - Antiobesitic composition comprising extract of Quercus aliena - Google Patents
Antiobesitic composition comprising extract of Quercus aliena Download PDFInfo
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- KR101944013B1 KR101944013B1 KR1020170082500A KR20170082500A KR101944013B1 KR 101944013 B1 KR101944013 B1 KR 101944013B1 KR 1020170082500 A KR1020170082500 A KR 1020170082500A KR 20170082500 A KR20170082500 A KR 20170082500A KR 101944013 B1 KR101944013 B1 KR 101944013B1
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/332—Promoters of weight control and weight loss
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- Coloring Foods And Improving Nutritive Qualities (AREA)
Abstract
본 발명은 갈참나무 추출물을 함유하는 항비만용 조성물에 관한 것으로서, 상기 갈참나무 추출물이 포함된 약학 조성물 또는 건강기능식품은 비만과 같은 대사질환의 치료용 조성물 또는 이들 질환의 예방과 개선을 위한 건강기능식품으로서 용이하게 이용될 수 있다.The present invention relates to an anti-obesity composition containing an alopecum oak extract, wherein the pharmaceutical composition or health functional food containing the alopecum oak extract comprises a composition for the treatment of metabolic diseases such as obesity, And can be easily used as a functional food.
Description
본 발명은 갈참나무 추출물을 함유하는 항비만용 조성물에 관한 것이다. 보다 자세하게는 본 발명은 지방세포의 분화억제 및 지방축적 억제 효과와 지방세포 분화 과정에 따른 산화 반응의 억제 효과가 있는 갈참나무의 잎의 알코올 추출물을 함유하는 항비만용 조성물에 관한 것이다. The present invention relates to an anti-obesity composition containing an oak extract. More specifically, the present invention relates to an anti-obesity composition containing an alcohol extract of a leaf of oak, which has an effect of suppressing differentiation of adipocytes and inhibiting lipid accumulation and inhibiting oxidation reaction according to an adipocyte differentiation process.
최근 경제 발전에 따른 생활 수준의 향상, 바쁜 생활 환경에 따른 운동 부족, 영양의 과잉 섭취 등으로 비만 인구가 급속히 늘고 있다. 비만은 에너지의 섭취와 소모의 불균형으로 인하여 체내에 에너지가 과잉으로 축적되어 지방조직이 비정상적으로 증가된 상태를 말한다. 세계보건기구(WHO)는 동양인 기준 BMI(Body Mass Index: 체질량 지수) 23~25를 과체중, 25~30을 비만, 30 이상을 고도비만으로 보고 있다. 비만의 원인으로는 고지방, 고열량의 식생활, 바쁜 사회적 환경에 따른 운동 부족, 내분비 이상 등 환경적 요인과 유전적 요인을 들 수 있는데, 이 중 비만의 50 내지 70% 정도가 환경적 요인에 의한 것으로 알려져 있고, 나머지가 유전적 요인에 의한 것으로 알려져 있다. 세계보건기구(WHO)의 통계에 따르면 비만은 발병률이 매년 증가하고 있다고 보고되며, 특히 OECD 국가의 경우, 성인 비만인구는 해마다 증가하는 추세이며, 미국의 경우 3명 중 1명은 비만인 것으로 조사되었다. 우리나라도 생활환경의 변화로 비만인구가 급증하고 있으며 체질량 지수(body mass index, BMI)가 25 이상인 한국 성인 비만인구는 매년 40여만 명씩 늘어 2005년 기준으로 10명 중 3명은 비만인 것으로 조사되었다. 또한, 질병관리본부의 자료에 따르면 1995년 비만인구는 전체 인구의 26.3%이었으나 2001년에는 전체 인구의 30.6%, 2005년에는 전체 인구의 31.5%로 급증하였고, 현재에도 지속적인 증가 추세를 나타내고 있다.In recent years, the population of obesity has been rapidly increasing due to improvements in living standards due to economic development, lack of exercise due to busy living environment, and excessive intake of nutrition. Obesity is a condition in which energy is excessively accumulated in the body due to unbalanced energy intake and consumption, resulting in an abnormally increased adipose tissue. The World Health Organization (WHO) considers Oriental BMI (body mass index) 23-25 as overweight, 25-30 obese, and over 30 as high obesity. The causes of obesity include environmental factors such as high fat, high calorie diet, lack of exercise due to busy social environment, endocrine abnormality, and genetic factors. Of these, about 50 to 70% of obesity is due to environmental factors And the rest is known to be caused by genetic factors. According to the statistics of the World Health Organization (WHO), the incidence of obesity is reported to increase year by year, especially in the OECD countries, the adult obesity population is increasing year by year. In Korea, the obesity population is increasing rapidly due to changes in the living environment, and the Korean adult obesity population with a body mass index (BMI) of 25 or more is increased by 40 million every year. As of 2005, 3 out of 10 were obese. According to data from the CDC, the obesity population in 1995 was 26.3% of the total population, but in 2001 it increased to 30.6% of the total population and to 31.5% of the total population in 2005, and is still showing a steady upward trend.
갈참나무(Quercus aliena Blume)는 재잘나무, 톱날갈참나무, 큰갈참나무 등으로도 불린다. 원산지는 한국, 중국, 일본, 인도 등이다. 산기슭에서 자란다. 높이는 25m, 지름은 1m 정도이고, 나무껍질은 그물처럼 얕게 갈라지며, 작은가지와 겨울눈에 털이 없다. 잎은 타원형으로 길이 5~30cm, 나비 3~9cm 정도이고, 끝은 둔한 것과 뾰족한 것이 있으며 윗면에 털이 없고 짙은 녹색이다. 잎 뒷면은 회백색이고 2~17개로 갈라진 별 모양의 털이 빽빽이 난다. 잎자루는 1~3.6cm이며 잎 가장자리는 물결 모양 또는 굵은 이빨 모양의 톱니로 되어 있다. 꽃은 5월에 피는데 단성화(單性花)이고, 수꽃이삭은 축 늘어지며 5~9개의 화피(花被)와 6~14개의 수술이 있다. 암꽃은 6개의 화피와 2~4개의 암술머리가 있다. 깍정이는 삼각형의 작은돌기로 덮여 있다. 열매는 견과이며 길이 6~23mm, 지름 7~16mm의 타원형이고, 10월에 익으며 식용한다. 어린 잎은 비료로 쓰며, 목재는 기구재, 땔감 등으로 쓰인다. 잎 뒷면에 털이 없는 것을 청갈참(var. pellucida), 톱니가 졸참나무의 톱니같이 생긴 것을 졸갈참(var. acuteserrata), 졸갈참과 같고 잎뒷면에 털이 없는 것을 청졸갈참(var. acuteserrata for. calvescens)이라고 한다. Quercus aliena Blume is also called jade tree, saw blade oak, and big oak tree. Origin is Korea, China, Japan and India. It grows at the foot of a mountain. Its height is 25m and its diameter is about 1m. The bark is shallowly divided like a net. Leaves are oval, 5 ~ 30cm long, 3 ~ 9cm long, with dull and pointed ends, no hairs on the top and dark green. The back side of the leaf is grayish white and the star-shaped hairs are tightly divided into 2 ~ 17 pieces. The petiole is 1 ~ 3.6cm and the edge of leaf has wavy or coarse toothed sawtooth. The flower blooms in May and is a single flower. The ear of a male flower is lanceolate and has 5 ~ 9 flowers and 6 ~ 14 stamens. The female flower has 6 horns and 2 to 4 pistil heads. It is covered with small projections of triangular shape. The fruit is nuts, 6 ~ 23mm in length, 7 ~ 16mm in diameter, elliptical, ripened in October and edible. The young leaves are used as fertilizer, and the wood is used as equipment and firewood. There are no hairs on the back of the leaves (var. Pellucida), no saw teeth (var. Acuteserrata), no saw teeth on the back of the leaves, no acneserrata for calvescens ).
본 발명자들은 이러한 갈참나무가 갖는 다양한 약리활성을 연구하던 중, 갈참나무 추출물이 항비만 효과가 있음을 확인하여 본 발명을 완성하였다. The inventors of the present invention have studied various pharmacological activities possessed by such oak, and confirmed that the oakwood extract has an anti-obesity effect, thus completing the present invention.
본 발명의 목적은 갈참나무 추출물을 함유하는 항비만용 조성물을 제공하는 데에 있다. 보다 자세하게는 본 발명의 목적은 지방세포의 분화억제 및 지방축적 억제 효과와 지방세포 분화 과정에 따른 산화 반응의 억제 효과가 있는 갈참나무의 잎의 알코올 추출물을 함유하는 항비만용 조성물을 제공하는 데에 있다. It is an object of the present invention to provide an anti-obesity composition containing an oak extract. More specifically, it is an object of the present invention to provide an anti-obesity composition containing an alcohol extract of a leaf of oak, which has an inhibitory effect on adipocyte differentiation and inhibition of fat accumulation and an oxidation reaction according to an adipocyte differentiation process .
본 발명은 갈참나무(Quercus aliena Blume) 추출물을 함유하는 항비만용 조성물에 관한 것이다. The present invention relates to an anti-obesity composition containing Quercus aliena Blume extract.
상기 갈참나무 추출물은 갈참나무를 물, C1~C4 알코올 또는 이들의 혼합용액을 용매로 하여 추출할 수 있다. The oakwood extract can be extracted with water, C1-C4 alcohol, or a mixed solution thereof as a solvent.
상기 갈참나무 추출물은 잎을 추출한 것일 수 있다. The above-mentioned oak extract can be obtained by extracting leaves.
본 발명은 또한 갈참나무 추출물을 함유하는 항비만용 건강기능식품을 제공한다. The present invention also provides a health functional food for an anti-obesity containing an oak extract.
이하, 본 발명을 상세하게 설명한다.Hereinafter, the present invention will be described in detail.
본 발명은 갈참나무 추출물을 함유하는 항비만용 조성물에 관한 것이다. 본 발명에서 갈참나무 추출물을 제조할 때, 갈참나무는 잎, 가지, 잔가지, 줄기, 수피, 목질부, 뿌리, 열매 등의 어느 부위를 사용해도 가능하나, 바람직하게는 잎을 사용하는 것이 가장 좋으며, 갈참나무의 잎 추출물이 다른 부위의 추출물보다 항비만 효과가 가장 우수하다. The present invention relates to an anti-obesity composition containing an oak extract. In the present invention, it is possible to use any part such as a leaf, a branch, a twig, a stem, a bark, a woody part, a root, a fruit, etc., Leaf oak leaf extract has the most excellent anti-obesity effect than other extracts.
상기 갈참나무 추출물은 갈참나무를 물, C1~C4 알코올 또는 이들의 혼합용액을 용매로 하여 추출할 수 있으며, 상기 C1~C4 알코올은 메탄올, 에탄올, 프로판올, 이소프로판올, 부탄올 및 이소부탄올로 이루어진 군에서 선택될 수 있다. 상기 용매는 바람직하게는 C1~C4 알코올의 50~90%(v/v) 수용액, 더 바람직하게는 에탄올의 50~90%(v/v) 수용액일 수 있다. The almond oak extract may be extracted with water, a C1-C4 alcohol or a mixed solution thereof as a solvent. The C1-C4 alcohol may be extracted from methanol, ethanol, propanol, isopropanol, butanol and isobutanol Can be selected. The solvent may preferably be a 50 to 90% (v / v) aqueous solution of a C1 to C4 alcohol, more preferably a 50 to 90% (v / v) aqueous solution of ethanol.
상기 갈참나무 추출물의 제조시 사용되는 물, C1~C4 알코올 또는 이들의 혼합용액은 갈참나무 사용 중량 기준 1~40배 부피(1kg 기준 1~40ℓ)를 사용할 수 있으며, 바람직하게는 5~40배 부피를 사용할 수 있다. 상기 갈참나무 추출물의 추출조건은 20~100℃에서 1~48시간일 수 있다. 상기 과정은 1~4번까지 반복할 수 있다. 그러나 각 과정이 이에 제한되지는 않는다. 추출용 기기로는 통상의 추출기기, 초음파분쇄추출기 또는 분획기를 이용할 수 있다. 이렇게 제조된 갈참나무 추출물은 열풍건조, 감압건조 또는 동결건조하여 용매를 제거할 수 있다. 또한, 상기 갈참나무 추출물은 칼럼크로마토그래피를 이용하여 정제하여 사용할 수 있다. The water, the C1-C4 alcohol, or the mixed solution thereof used in the preparation of the oak extract may be used in an amount of 1 to 40 times by volume (1 to 40 liters per kg), preferably 5 to 40 times The volume can be used. The extracting conditions of the oak extract may be 1 to 48 hours at 20 to 100 ° C. The above process can be repeated 1 to 4 times. However, each process is not limited thereto. As the extraction apparatus, a conventional extraction apparatus, an ultrasonic pulverization extractor, or a fractionator may be used. The thus prepared oak extract can be removed by hot air drying, vacuum drying or freeze drying. The oakwood extract can be purified by using column chromatography.
상기 갈참나무 추출물은 상법에 따라, 유기용매(알코올, 에테르, 아세톤 등)에 의한 추출, 헥산과 물의 분배, 칼럼크로마토그래피에 의한 방법 등, 식물체 성분의 분리 추출에 이용되는 공지의 방법을 단독 또는 적합하게 조합한 방법을 이용하여 분획 또는 정제하여 사용할 수 있다. The alder oak extract can be isolated or isolated by known methods used for the separation and extraction of plant components, such as extraction with an organic solvent (alcohol, ether, acetone, etc.), partition with hexane and water, And may be fractionated or purified by a suitable combination method.
상기 크로마토그래피는 실리카겔 컬럼 크로마토그래피(silica gel column chromatography), 엘에이취-20 컬럼 크로마토그래피(LH-20 column chromatography), 이온교환수지 크로마토그래피(ion exchange resin chromatography), 중압 액체 크로마토그래피(medium pressure liquid chromatography), 박층 크로마토그래피(TLC; thin layer chromatography), 실리카겔 진공 액체 크로마토그래피(silica gel vacuum liquid chromatography) 및 고성능 액체 크로마토그래피(high performance liquid chromatography) 중에서 선택될 수 있다. The chromatography can be carried out using silica gel column chromatography, LH-20 column chromatography, ion exchange resin chromatography, medium pressure liquid chromatography chromatography, thin layer chromatography (TLC), silica gel vacuum liquid chromatography, and high performance liquid chromatography.
또한, 본 발명은 갈참나무 추출물을 함유하는 항비만용 약학 조성물을 제공한다. 상기 갈참나무 추출물은 본 발명의 약학 조성물에 0.001~100 중량%로 하여 첨가될 수 있다.In addition, the present invention provides a pharmaceutical composition for anti-obesity containing an oak extract. The oakwood extract may be added to the pharmaceutical composition of the present invention in an amount of 0.001 to 100% by weight.
상기 약학 조성물은, 각각 통상의 방법에 따라 산제, 과립제, 정제, 캡슐제, 현탁액, 에멀젼, 시럽, 에어로졸 등의 경구형 제형, 외용제, 좌제 및 멸균 주사용액의 형태로 제형화하여 사용될 수 있다. 상기 약학 조성물에 포함될 수 있는 담체, 부형제 및 희석제로는 락토즈, 덱스트로즈, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아 고무, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로즈, 메틸 셀룰로즈, 미정질 셀룰로스, 폴리비닐 피롤리돈, 물, 메틸히드록시벤조에이트, 프로필히드록시벤조에이트, 탈크, 마그네슘 스테아레이트 및 광물유를 들 수 있다. 제제화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 조제된다. 경구투여를 위한 고형제제에는 정제, 환제, 산제, 과립제, 캡슐제 등이 포함되며, 이러한 고형제제는 본 발명의 추출물 또는 화합물에 적어도 하나 이상의 부형제, 예를 들면, 전분, 탄산칼슘, 수크로스 또는 락토오스, 젤라틴 등을 섞어 조제된다. 또한 단순한 부형제 이외에 마그네슘 스테아레이트, 탈크 같은 윤활제들도 사용된다. 경구를 위한 액상 제제로는 현탁제, 내용액제, 유제, 시럽제 등이 해당되는데 흔히 사용되는 단순희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다. 비경구 투여를 위한 제제에는 멸균된 수용액, 비수성용제, 현탁제, 유제, 동결건조 제제, 좌제가 포함된다. 비수성용제, 현탁제로는 프로필렌글리콜, 폴리에틸렌글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르 등이 사용될 수 있다. 좌제의 기제로는 위텝솔(witepsol), 마크로골, 트윈(tween) 61, 카카오지, 라우린지, 글리세로제라틴 등이 사용될 수 있다. The pharmaceutical compositions may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories and sterilized injection solutions according to conventional methods. Examples of carriers, excipients and diluents that can be contained in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose , Methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient, such as starch, calcium carbonate, sucrose, Lactose, gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. Examples of the suppository base include witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like.
본 발명의 약학 조성물의 투여량은 치료받을 대상의 연령, 성별, 체중과, 치료할 특정 질환 또는 병리 상태, 질환 또는 병리 상태의 심각도, 투여경로 및 처방자의 판단에 따라 달라질 것이다. 이러한 인자에 기초한 투여량 결정은 당업자의 수준 내에 있으며, 일반적으로 투여량은 0.01㎎/㎏/일 내지 대략 2000㎎/㎏/일의 범위이다. 더 바람직한 투여량은 1㎎/㎏/일 내지 500㎎/㎏/일이다. 투여는 하루에 한번 투여할 수도 있고, 수회 나누어 투여할 수도 있다. 상기 투여량은 어떠한 면으로든 본 발명의 범위를 한정하는 것은 아니다. The dosage of the pharmaceutical composition of the present invention will depend on the age, sex, body weight of the subject to be treated, the particular disease or condition to be treated, the severity of the disease or condition, the route of administration and the judgment of the prescriber. Dosage determinations based on these factors are within the level of ordinary skill in the art and generally the dosage ranges from 0.01 mg / kg / day to approximately 2000 mg / kg / day. A more preferable dosage is 1 mg / kg / day to 500 mg / kg / day. The administration may be carried out once a day or divided into several times. The dose is not intended to limit the scope of the invention in any way.
본 발명의 약학 조성물은 쥐, 가축, 인간 등의 포유동물에 다양한 경로로 투여될 수 있다. 투여의 모든 방식은 예상될 수 있는데, 예를 들면, 경구, 직장 또는 정맥, 근육, 피하, 자궁내 경막 또는 뇌혈관내 주사에 의해 투여될 수 있다. 본 발명의 화합물은 독성 및 부작용이 거의 없으므로 예방 목적으로 장기간 복용시에도 안심하고 사용할 수 있는 약제이다. The pharmaceutical composition of the present invention can be administered to mammals such as rats, livestock, humans, and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine dural or intracerebral injection. Since the compound of the present invention has little toxicity and side effects, it can be safely used even for long-term administration for preventive purposes.
또한, 본 발명은 갈참나무 추출물 및 식품학적으로 허용 가능한 식품보조 첨가제를 포함하는 항비만용 건강기능식품을 제공한다. 상기 갈참나무 추출물은 본 발명의 건강기능식품에 0.001~100 중량%로 하여 첨가될 수 있다. 본 발명의 건강기능식품은 정제, 캡슐제, 환제 또는 액제 등의 형태를 포함하며, 본 발명의 추출물을 첨가할 수 있는 식품으로는, 예를 들어, 각종 드링크제, 육류, 소세지, 빵, 캔디류, 스넥류, 면류, 아이스크림, 유제품, 스프, 이온음료, 음료수, 알코올 음료, 껌, 차 및 비타민 복합제 등이 있다. In addition, the present invention provides an anti-obesity health functional food comprising an almond oak extract and a pharmaceutically acceptable food-aid additive. The oakwood extract may be added in an amount of 0.001 to 100% by weight to the health functional food of the present invention. The health functional food of the present invention includes forms such as tablets, capsules, pills, and liquids. Examples of the foods to which the extract of the present invention can be added include various kinds of drinks, meat, sausage, bread, Snacks, noodles, ice cream, dairy products, soups, ionic drinks, beverages, alcoholic beverages, gums, tea and vitamin complexes.
본 발명은 갈참나무 추출물을 함유하는 항비만용 조성물에 관한 것으로서, 상기 갈참나무 추출물이 포함된 약학 조성물 또는 건강기능식품은 비만과 같은 대사질환의 치료용 조성물 또는 이들 질환의 예방과 개선을 위한 건강기능식품으로서 용이하게 이용될 수 있다. 참나무류를 활용한 선행기술로 일본공개특허 제1999-299451호, 한국공개특허 제10-2011-0111568호, 한국등록특허 제10-1500836호 등이 확인되기는 하지만, 본 발명에서처럼 갈참나무의 잎 추출물을 제조하고, 세포독성이 없는 추출물의 농도를 확인하고, 이 농도에서 항비만 효과가 있는 추출물의 함량을 제시한 결과는 확인할 수 없다. The present invention relates to an anti-obesity composition containing an alopecum oak extract, wherein the pharmaceutical composition or health functional food containing the alopecum oak extract comprises a composition for the treatment of metabolic diseases such as obesity, And can be easily used as a functional food. Japanese Unexamined Patent Publication No. 1999-299451, Korean Patent Publication No. 10-2011-0111568, Korean Patent No. 10-1500836 and the like have been confirmed as prior art utilizing oak, but in the present invention, leaf oak leaf extract , The concentration of extract without cytotoxicity was confirmed, and the result of presenting the content of extract having anti-obesity effect at this concentration was not confirmed.
도 1은 갈참나무 추출물의 세포독성 확인 결과를 나타낸다.
도 2는 갈참나무 추출물의 지방 세포 분화 억제 확인결과를 나타낸다.
도 3은 지방세포 분화과정에서의 갈참나무 추출물의 ROS 소거활성 측정 결과를 나타낸다. Fig. 1 shows the result of cytotoxicity of the oakwood extract.
Fig. 2 shows the result of inhibition of adipocyte differentiation by the oak extract of Goa.
FIG. 3 shows the results of measurement of ROS scavenging activity of Ganoderma lucidum extract in adipocyte differentiation.
이하 본 발명의 바람직한 실시예를 상세히 설명하기로 한다. 그러나, 본 발명은 여기서 설명되는 실시예에 한정되지 않고 다른 형태로 구체화될 수도 있다. 오히려, 여기서 소개되는 내용이 철저하고 완전해지고, 당업자에게 본 발명의 사상을 충분히 전달하기 위해 제공하는 것이다. Hereinafter, preferred embodiments of the present invention will be described in detail. However, the present invention is not limited to the embodiments described herein but may be embodied in other forms. Rather, the intention is to provide an exhaustive, complete, and complete disclosure of the principles of the invention to those skilled in the art.
<실시예 1. 갈참나무 추출물의 제조>≪ Example 1: Preparation of goose oak extract >
갈참나무 추출물은 100 g의 잎을 70%(v/v) 에탄올 수용액 1000 ㎖에 침지시킨 후 실온(25℃)에서 24시간 동안 추출하였다. 상기 추출 과정을 3회 반복하고, 얻어진 추출액은 모두 합하였다. 추출물은 필터 페이퍼(Whatman, No.3, Maidstone, Kent, UK)로 여과한 후, 회전진공농축기(EYELA, N-3000, Tokyo, Japan)를 사용하여 감압농축한 후, 동결건조(Ilshinbiobase Co., Ltd, Yangju, Korea) 하였다. The oak extract was prepared by dipping 100 g of leaf in 1000 ml of 70% (v / v) ethanol solution and then extracting at room temperature (25 ℃) for 24 hours. The extraction procedure was repeated three times, and the obtained extracts were all combined. The extract was filtered with a filter paper (Whatman, No.3, Maidstone, Kent, UK), concentrated under reduced pressure using a rotary vacuum concentrator (EYELA, N-3000, Tokyo, Japan) and then lyophilized (Ilshinbiobase Co., Ltd.). , Ltd, Yangju, Korea).
<실시예 2. 갈참나무 추출물의 항비만 활성 확인>Example 2. Identification of anti-obesity activity of gooseberry extract
실시예 2-1. 시료의 제조Example 2-1. Preparation of sample
실험에 사용한 시료인 실시예 1에서 얻은 갈참나무 추출물을 100% DMSO(dimethyl sulfoxide)를 이용하여 세포 배양 상태에서의 최종농도가 25 ㎍/㎖, 50 ㎍/㎖, 100 ㎍/㎖가 되도록 희석하여 사용하였다.The oak extract obtained in Example 1 used in the experiment was diluted with 100% DMSO (dimethyl sulfoxide) to a final concentration of 25 μg / ml, 50 μg / ml and 100 μg / ml in a cell culture condition Respectively.
실시예 2-2. 3T3-L1 세포 배양 Example 2-2. 3T3-L1 cell culture
3T3-L1 세포는 ATCC에서 구입하여 고농도 포도당 DMEM(Dulbecco's modified Eagle's medium)에 10% BS(bovine serum), 1% Penicillin/streptomycin(P/S) 존재 하에 37℃, 5% CO2 배양기에서 배양하였다. 세포배양접시에서 전-지방세포(Preadipocyte) 상태의 3T3-L1 세포가 100% 가득 차게 되면 지방세포(adipocyte)로의 분화를 위하여 10% FBS(fetal bovine serum), 1% P/S가 함유된 고농도 포도당 DMEM을 사용하였으며, 0.5 mM Isobutylmethylxanthine(IBMX), 1 μM Dexamethasone(DEX), 1 ㎍/㎖ Insulin을 처리하여 48시간 배양하여 분화시켰다. 세포는 실험목적에 따라 96웰플레이트(96-well plate)와 24웰플레이트(24-well plate)에 세포를 분주하여 배양 및 실험하였다.3T3-L1 cells were purchased from ATCC and cultured in high-glucose DMEM (Dulbecco's modified Eagle's medium) in the presence of 10% BS (bovine serum) and 1% penicillin / streptomycin (P / S) at 37 ° C in a 5% CO 2 incubator . When the 3T3-L1 cells in the preadipocyte state are filled to 100% in the cell culture dish, a high concentration of 10% FBS (fetal bovine serum) and 1% P / S are added for differentiation into adipocytes Glucose DMEM was used and 0.5 mM Isobutylmethylxanthine (IBMX), 1 μM Dexamethasone (DEX) and 1 μg / ㎖ insulin were added for 48 hours. Cells were cultured and tested in a 96-well plate and a 24-well plate by dividing cells according to the purpose of the experiment.
실시예 2-3. XTT 어세이를 이용한 세포 독성평가 Examples 2-3. Evaluation of cytotoxicity using XTT assays
3T3-L1 세포에 대한 시료의 세포 독성평가는 XTT 어세이 키트(2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide innersalt assay kit)를 이용하여 측정하였다. The cytotoxicity of 3T3-L1 cells was assessed using an XTT assay kit (2-methoxy-4-nitro-5-sulfophenyl) -2H-tetrazolium-5-carboxanilide innersalt assay kit Respectively.
XTT에 Election coupling agent 역할을 하는 PMS (phenazine methosulfate)를 첨가하여 Bioreduction을 증가시키게 한 후, 세포에 XTT와 PMS를 첨가하여 살아있는 세포의 Mitochondrial dehydrogenase에 의한 XTT의 Tetrazolium ring이 분해시켜 Formazan crystal을 형성하게 되면, Formazan crystal은 수용액에 녹아 노란색을 나타나게 된다. 이 노란색을 ELISA(VersaMax ELISA Microplate Reader, Molecular Devices, Sunnyvale, USA)로 측정하여 세포 독성 평가에 이용하였다. XTT and PMS were added to the cells to increase the bioreduction by adding PMS (phenazine methosulfate), which serves as an Election coupling agent to XTT, and then the Tetrazolium ring of XTT by mitochondrial dehydrogenase of living cells was decomposed to form Formazan crystal Formazan crystals are dissolved in aqueous solution and become yellow. This yellow was measured by ELISA (VersaMax ELISA Microplate Reader, Molecular Devices, Sunnyvale, USA) and used for cytotoxicity evaluation.
이를 위해, 3T3-L1 preadipocyte cell을 실험 전날 1×106 농도로 96웰플레이트에 분주하여 24 시간 동안 배양하였다. 갈참나무 추출물의 농도는 각각 25 ㎍/㎖, 50 ㎍/㎖, 100 ㎍/㎖이 되도록 처리한 후 세포배양기에서 24 시간 동안 배양하였다. 그 후 XTT 1 ㎖, PMS reagent 20 ㎕ 비율로 혼합하여 배양액(media) 부피의 20%를 첨가하여 세포배양기에서 4시간 반응시킨 후, 분광광도계(spectrophotometer)를 이용하여 450 nm와 690 nm의 흡광도를 측정하고, 결과값을 계산하여 도 1에 나타내었다.For this purpose, 3T3-L1 preadipocyte cells were plated in 96-well plates at a concentration of 1 × 10 6 on the day before the experiment and cultured for 24 hours. The concentration of oak extract was 25 ㎍ / ㎖, 50 ㎍ / ㎖ and 100 ㎍ / ㎖, respectively, and cultured in a cell incubator for 24 hours. Then, 1 ㎖ of XTT and 20 ㎕ of PMS reagent were mixed, and 20% of the media volume was added. After reacting for 4 hours in a cell incubator, absorbance at 450 nm and 690 nm was measured using a spectrophotometer The results are shown in Fig.
도 1을 참고하면, 갈참나무 추출물을 각각 25 ㎍/㎖, 50 ㎍/㎖, 100 ㎍/㎖ 농도를 처리하여 XTT 어세이 방법으로 3T3-L1 세포의 생존율에 미치는 영향을 측정한 결과, 갈참나무 추출물이 100 ㎍/㎖까지 세포독성을 보이지 않는 것으로 나타난다. As shown in FIG. 1, the effect of the oak extract on the survival rate of 3T3-L1 cells was examined by XTT assay at a concentration of 25 μg / ml, 50 μg / ml and 100 μg / The extract shows no cytotoxicity up to 100 [mu] g / ml.
실시예 2-4. 오일 레드 O 염색을 이용한 지방세포의 분화 억제 및 지방축적 억제 효능평가 Examples 2-4. Inhibition of adipocyte differentiation and evaluation of fat accumulation inhibition by oil red O staining
오일 레드 O 염색(Oil red O staining)을 이용하여 분화과정에 따른 3T3-L1 세포내 지방축적량을 측정하고자 갈참나무 추출물 시료를 처리하여 24웰플레이트에서 8일 동안 분화된 3T3-L1 세포의 배양액을 제거한 후, 10% 포르말린(formalin) 용액 0.5 ㎖를 첨가하여 5분간 실온에서 방치하였다. 동량의 포르말린 용액으로 분화된 3T3-L1 세포 1시간 이상 실온에서 방치한 후, 포르말린을 제거하고 60% 이소프로필 알코올 용액 0.5 ㎖로 세척하여 세포를 완전히 건조시켰다. 완전히 건조된 세포들은 미리 제조해 둔 오일 레드 O 용액(working solution - Oil red O : DDW = 6 : 4)으로 세포 내 축적된 지방성분들을 충분히 염색한 후, 증류수를 이용하여 세포를 4회 세척하고 완전히 건조시켰다. 세포내 축적된 지방성분과 결합한 오일 레드 O는 100% 이소프로필 알코올 용액을 이용하여 모두 용출시킨 후, 분광광도계를 이용하여 490 nm에서 흡광도를 측정하였다. 이때 양성 대조군으로 5mM NAC(N-acetyl-L-cysteine)를 사용하여 3T3-L1 세포의 분화 정도를 비교 분석하였고 이에 대한 결과는 도 2에 나타내었다. In order to measure the amount of fat accumulation in 3T3-L1 cells by oil red o staining, 3T3-L1 cells were cultured in 24 well plates for 8 days. After removing, 0.5 ml of a 10% formalin solution was added and left at room temperature for 5 minutes. 3T3-L1 cells differentiated with the same amount of formalin solution were left at room temperature for more than 1 hour, formalin was removed, and cells were washed with 0.5 ml of 60% isopropyl alcohol solution to completely dry the cells. The fully dried cells were stained with a pre-prepared working solution (Oil Red O: DDW = 6: 4), and the cells were washed 4 times with distilled water It was completely dried. The oily red O bound to the lipid components accumulated in the cells was eluted with a 100% isopropyl alcohol solution and absorbance was measured at 490 nm using a spectrophotometer. At this time, the degree of differentiation of 3T3-L1 cells was compared and analyzed using 5 mM NAC (N-acetyl-L-cysteine) as a positive control. The results are shown in FIG.
도 2를 참고하면, 갈참나무 추출물에 대해 농도 의존적으로 지방 축적량이 유의적으로 감소하는 것으로 나타나, 상기 갈참나무 추출물이 지방세포의 분화 억제 및 지방 축적 억제에 효능이 있는 것으로 확인된다(도 2의 con - 추출물을 처리하지 않은 군).2, the amount of fat accumulation was significantly decreased in a concentration-dependent manner in the oak extract of Goa, and the oak extract was effective in inhibiting the differentiation of adipocytes and suppressing the accumulation of fat (see Fig. 2 con - extract without treatment).
한편, 기존에 약용으로 사용되는 참나무 유래 조성물은 통상적으로 참나무과 유래 열매인 도토리가 주로 사용되기에, 갈참나무로부터 잎 외에 도토리 과육 이용하여 추출물을 제조한 후 100㎍/㎖ 조건에서 오일 레드 O 염색 결과를 비교한 바, 잎 추출물이 도 2와 같은 약 75% 내외의 지방축적량을 보일 때, 도토리 과육 추출물은 95% 이상의 지방축적량을 보여, 도토리 추출물보다 잎 추출물이 지방축적 억제 효과가 더 우수함을 알 수 있다. On the other hand, since the oak-derived composition conventionally used for medicinal purposes is generally used as an acorn which is a fruit derived from oak, the extract is prepared from acorn oak, When the leaf extract showed a fat accumulation amount of about 75% as shown in FIG. 2, the acorn extract showed 95% or more fat accumulation and the leaf extract had better fat accumulation inhibitory effect than the acorn extract .
실시예 2-5. NBT 어세이를 이용한 ROS 저감 효능평가 Examples 2-5. Evaluation of ROS abatement efficacy using NBT assay
NBT 어세이(Nitroblue tetrazolium assay)를 이용하여 지방세포의 분화과정에 따른 ROS(reactive oxygen species) 생성량을 측정하기 위하여 먼저 24웰플레이트에 8일 동안 갈참나무 추출물을 농도별(25, 50, 100 μg/mL)로 처리하면서 분화를 유도하였다. 이 후 3T3-L1 세포의 배양액을 제거한 후 멸균된 PBS(phosphate-buffered saline, pH 7.4)를 이용하여 2회 세척, 0.2%(w/v) NBT(nitroblue tetrazolium) 용액 0.2 ㎖를 첨가하여 CO2 배양기 안에서 90분간 반응시켰다. 염색 후 증류수로 2회 세척 후 건조시키고, DMSO(세포내 축적된 ROS들은 NBT 용액과 반응하여 dark blue formazan을 생성하게 되며, 이를 세포로부터 용출하여 흡광도를 측정하고자 함)와 1 N 수산화칼륨을 7:3(v:v) 비율로 혼합한 용액을 0.3 ㎖ 첨가하여 용출시킨 후 동량의 증류수를 첨가하여 분광광도계를 이용하여 570 nm에서 흡광도를 측정하였고 이에 대한 결과를 도 3에 나타내었다.To determine the amount of reactive oxygen species (ROS) produced by the differentiation process of adipocytes using the Nitroblue tetrazolium assay, 24-well plates were seeded with 25, 50, 100 μg / mL) to induce differentiation. By using this after 3T3-L1 After removing the culture medium of the cells with sterile PBS (phosphate-buffered saline, pH 7.4) 2 times wash, 0.2% (w / v) NBT was added to 0.2 ㎖ (nitroblue tetrazolium) solution, CO 2 And incubated for 90 minutes in the incubator. After the staining, the plate was rinsed twice with distilled water and dried. DMSO (the ROS accumulated in the cells reacted with the NBT solution to produce dark blue formazan, which was eluted from the cells to measure the absorbance) and 1 N potassium hydroxide : 3 (v: v). The same amount of distilled water was added thereto, and the absorbance at 570 nm was measured using a spectrophotometer. The results are shown in FIG.
도 3을 참고하면, 갈참나무 추출물을 처리한 지방세포에서 농도의존적으로 유의하게 ROS 함량이 감소하는 것으로 나타난다(도 3의 con - 추출물을 처리하지 않은 군).Referring to FIG. 3, the ROS content of the oak extract-treated adipocytes was significantly decreased in a concentration-dependent manner (the con-extract-treated group of FIG. 3).
<제제예 1. 약학적 제제>≪ Formulation Example 1 >
본 발명의 갈참나무 추출물 200g을 락토오스 175.9g, 감자전분 180g 및 콜로이드성 규산 32g과 혼합하였다. 이 혼합물에 10% 젤라틴 용액을 첨가시킨 후, 분쇄해서 14 메쉬체를 통과시켰다. 이것을 건조시키고 여기에 감자전분 160g, 활석 50g 및 스테아린산 마그네슘 5g을 첨가해서 얻은 혼합물을 정제로 만들었다. 200 g of the oak extract of the present invention was mixed with 175.9 g of lactose, 180 g of potato starch and 32 g of colloidal silicic acid. To this mixture was added a 10% gelatin solution, which was pulverized and passed through a 14-mesh sieve. This was dried, and a mixture obtained by adding 160 g of potato starch, 50 g of talc and 5 g of magnesium stearate was made into tablets.
<제제예 2. 식품 제조><Formulation Example 2: Food Preparation>
제제예 2-1. 조리용 양념의 제조Formulation Example 2-1. Manufacture of cooking seasonings
본 발명의 갈참나무 추출물을 조리용 양념에 1 중량%로 첨가하여 건강 증진용 조리용 양념을 제조하였다.The oak extract of the present invention was added to the cooking seasoning at 1 wt% to prepare a cooking sauce for health promotion.
제제예 2-2. 밀가루 식품의 제조Formulation Example 2-2. Manufacture of flour food products
본 발명의 갈참나무 추출물을 밀가루에 0.1 중량%로 첨가하고, 이 혼합물을 이용하여 빵, 케이크, 쿠키, 크래커 및 면류를 제조하여 건강 증진용 식품을 제조하였다.The oak extract of the present invention was added to wheat flour in an amount of 0.1% by weight, and bread, cake, cookies, crackers and noodles were prepared using the mixture to prepare a food for health promotion.
제제예 2-3. 스프 및 육즙(gravies)의 제조Preparation Example 2-3. Manufacture of soups and gravies
본 발명의 갈참나무 추출물을 스프 및 육즙에 0.1 중량%로 첨가하여 건강 증진용 수프 및 육즙을 제조하였다.The health promotion soup and the juice were prepared by adding the oak extract of the present invention to the soup and juice at 0.1 wt%.
제제예 2-4. 유제품(dairy products)의 제조Formulation Example 2-4. Manufacture of dairy products
본 발명의 갈참나무 추출물을 우유에 0.1 중량%로 첨가하고, 상기 우유를 이용하여 버터 및 아이스크림과 같은 다양한 유제품을 제조하였다.The oakwood extract of the present invention was added to milk in an amount of 0.1 wt%, and various dairy products such as butter and ice cream were prepared using the milk.
제제예 2-5. 야채주스 제조Formulation Example 2-5. Vegetable juice manufacturing
본 발명의 갈참나무 추출물 0.5g을 토마토주스 또는 당근주스 1,000㎖에 가하여 건강 증진용 야채주스를 제조하였다.0.5 g of the oak extract of the present invention was added to 1,000 ml of tomato juice or carrot juice to prepare health promotion vegetable juice.
제제예 2-6. 과일주스 제조Formulation Example 2-6. Manufacture of fruit juice
본 발명의 갈참나무 추출물 0.1g을 사과주스 또는 포도주스 1,000㎖에 가하여 건강 증진용 과일주스를 제조하였다.Healthy fruit juice was prepared by adding 0.1 g of the oak extract of the present invention to apple juice or 1,000 ml of grape juice.
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