KR20230052574A - A composition for improving, preventing and treating of obesity comprising peanut shell extract - Google Patents
A composition for improving, preventing and treating of obesity comprising peanut shell extract Download PDFInfo
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- KR20230052574A KR20230052574A KR1020210135788A KR20210135788A KR20230052574A KR 20230052574 A KR20230052574 A KR 20230052574A KR 1020210135788 A KR1020210135788 A KR 1020210135788A KR 20210135788 A KR20210135788 A KR 20210135788A KR 20230052574 A KR20230052574 A KR 20230052574A
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- extract
- obesity
- peanut
- extraction
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Abstract
Description
본 발명은 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 개선, 예방 또는 치료용 조성물에 관한 것이다.The present invention relates to a composition for improving, preventing or treating obesity comprising peanut hull extract as an active ingredient.
우리나라는 물론 세계적으로 경제발전 및 의료기술의 발달로 인하여 인간의 평균수명이 길어지고 삶의 질이 빠른 속도로 향상되고 있어 이에 부합하는 적극적인 웰빙시대의 대응전략이 시급히 요청되고 있다. 특히, 가속화되고 있는 노령화, 성인병의 발생에 따른 국민건강의 보호 및 정부와 개인의 의료비 절감의 차원에서도 효과적이고 안전한 의약품 및 건강기능식품의 개발이 필요하다.Due to the development of economic development and medical technology in Korea as well as the world, the average life expectancy of human beings is getting longer and the quality of life is improving at a rapid pace. In particular, it is necessary to develop effective and safe medicines and health functional foods in terms of protecting national health and reducing government and individual medical expenses in accordance with the accelerated aging and occurrence of adult diseases.
심장마비, 고혈압, 동맥경화, 뇌졸중 등의 심혈관질환은 세계적으로 전체 사망률의 30%를 차지하고 있다. 특히 비만, 당뇨병, 고혈압, 동맥경화증, 심장 질환, 뇌졸중 등은 각기 다른 질병으로 보이지만 체지방 증가로 인한 인슐린 저항성이라는 공통 병인을 갖는 하나의 증후군으로 이해되고 있다.Cardiovascular diseases such as heart attack, hypertension, arteriosclerosis, and stroke account for 30% of all mortality worldwide. In particular, obesity, diabetes, hypertension, arteriosclerosis, heart disease, stroke, etc. are understood as a single syndrome having a common etiology, insulin resistance due to increased body fat, although they appear to be different diseases.
비만은 전 세계적으로 가장 흔한 영양장애 중의 하나로서 WHO 통계자료에 의하면 현재 2억 5천여만명이 비만 환자로 분류되며 20년 후에는 약 3억명이 비만으로 고통받을 것으로 예측된다.Obesity is one of the most common nutritional disorders worldwide, and according to WHO statistics, currently about 250 million people are classified as obese patients, and it is predicted that about 300 million people will suffer from obesity in 20 years.
상기 비만은 과량의 에너지 섭취 또는 에너지 소비 저하로 인해 열량 대사가 불균형하여 체지방이 과도하게 축적된 상태를 지칭하며, 최근에는 현대인들의 서구적인 식생활 패턴과 운동 부족으로 인해 비만의 발병율이 급격히 증가하는 추세이다. 또한, 수치상의 개념으로는 BMI(body mass index, 체질량지수) 측정 시 25 이상일 경우를 비만으로 정의하며, BMI 측정법은 체중(kg)을 키의 제곱(㎡)으로 나눈 값을 통해 체지방의 양을 추정하는 비만측정법이다. The obesity refers to a state in which excessive body fat is accumulated due to an imbalance in calorie metabolism due to excessive energy intake or decreased energy consumption. am. In addition, as a numerical concept, obesity is defined as a case of 25 or more when measuring BMI (body mass index). It is a measure of obesity.
비만치료제 중에서 식욕억제제에는 휴터민정, 펜터민 및 로카세린이라는 성분이 있고 부작용으로는 불안감, 현기증과 불면증 등이 있으며 지방분해효소 억제제에는 오르리스타트, 로카세린과 마진돌 등이 있고 부작용으로는 기름변, 복통과 변실금 등이 있는데, 상기 상업화된 약물들의 부작용이 보고됨에 따라 효과가 뛰어나면서 부작용이 적은 천연물 유래의 비만치료제가 요구되는 실정이다.Among the anti-obesity drugs, appetite suppressants include Hutermin, Phentermine, and Lorcaserin. Side effects include anxiety, dizziness, and insomnia. Lipase inhibitors include Orlistat, Lorcaserin, and Marzindol. , Abdominal pain and fecal incontinence, etc. As the side effects of the commercialized drugs are reported, there is a demand for anti-obesity drugs derived from natural products with excellent effects and low side effects.
한편, 땅콩(Peanut, Arachis hypogaea)은 대표적인 유지작물로 콩과에 속하는 일년생의 초본식물이며 한국, 인도와 미국 등 세계 각국에서 재배되어 단백질, 불포화지방산, 비타민과 아미노산 등의 다양한 영양성분을 풍부하게 함유하고 있다. 땅콩 중량의 35~40%를 차지하는 땅콩 겉껍질은 소화율이 낮아 가축 사료로의 사용에 제한이 있어 대부분 폐기되는 실정으로 부산물 처리비용과 환경문제를 발생시키므로 이를 해결하기 위해 땅콩 겉껍질의 활용을 통한 고부가가치 소재 개발이 요구되고 있다.On the other hand, peanut (Peanut, Arachis hypogaea ) is a representative oil crop and is an annual herbaceous plant belonging to the legume family. contains Peanut husk, which accounts for 35-40% of the weight of peanuts, has a low digestibility and is limited in its use as livestock feed, and most of them are discarded, which causes by-product processing costs and environmental problems. The development of high value-added materials is required.
비만은 지방전구세포가 지방세포로 분화되는 정상적인 지방세포의 이상발달이 발생하는데 지방세포로의 과잉분화와 지방세포 내 지방의 축적은 지방세포의 수나 크기를 증가시키고 그에 따라 비만 증상이 더욱 심해지는 것으로 알려져 있다. 지방전구세포가 confluence 상태가 되면 세포 주기가 정지되어 세포의 증식이 중단되고 3-isobutyl-1-methyl-xanthine (IBMX), dexamethasone (DEX) 및 insulin 등의 호르몬에 의해 지방세포로 분화(adipogenisis)가 유도되고 비만은 조직 내 지방세포가 비대(hypertrophy)와 과형성(hyperplasia)으로 초래된다. 지방전구세포의 초기분화 과정은 DEX에 의해 cAMP의 농도가 증가되면서 sterol regulatory element binding proteins-1c (SREBP-1c) 발현이 유도된 후 IBMX에 의해 peroxisome proliferator activated receptor-γ (PPAR-γ)와 CCAAT enhancer binding protein-α (CEBP-α)가 상호 발현되어 분화를 촉진한다. 이들의 발현은 불포화지방산에서 포화지방산으로 합성될 때 포화 지방산을 단일 불포화지방산으로 전환시키는 fatty acid synthase (FAS) 및 stearoyl-CoA desaturase 1 (SCD1) 발현과 함께 지방세포로의 분화를 유도한다. Obesity occurs when abnormal development of normal adipocytes, in which preadipocytes differentiate into adipocytes, occurs. Hyperdifferentiation into adipocytes and accumulation of fat in adipocytes increase the number and size of adipocytes, resulting in more severe obesity symptoms. It is known. When pre-adipocytes reach confluence, the cell cycle is stopped, cell proliferation is stopped, and hormones such as 3-isobutyl-1-methyl-xanthine (IBMX), dexamethasone (DEX), and insulin differentiate into fat cells (adipogenisis). is induced, and obesity is caused by hypertrophy and hyperplasia of fat cells in tissues. In the initial differentiation process of preadipocytes, the expression of sterol regulatory element binding proteins-1c (SREBP-1c) is induced by increasing the concentration of cAMP by DEX, and then peroxisome proliferator activated receptor-γ (PPAR-γ) and CCAAT by IBMX Enhancer binding protein-α (CEBP-α) is co-expressed to promote differentiation. Their expression induces differentiation into adipocytes along with the expression of fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1), which convert saturated fatty acids to monounsaturated fatty acids when synthesized from unsaturated fatty acids to saturated fatty acids.
따라서 지방전구세포 분화에 핵심적인 전사인자의 발현 조절을 통해 지방세포 분화를 억제하는 것은 비만을 예방하는 효과적인 전략이며, 이에 본 연구는 천연물로부터 독성 및 부작용이 없는 천연물질을 이용한 비만치료제가 요구되고 있다.Therefore, suppressing adipocyte differentiation through regulation of the expression of key transcription factors for preadipocyte differentiation is an effective strategy to prevent obesity, and therefore, this study requires an anti-obesity treatment using natural substances without toxicity and side effects from natural products. there is.
본 발명의 목적은 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 예방 또는 치료용 약학 조성물을 제공하는데 있다.An object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of obesity comprising peanut hull extract as an active ingredient.
또한, 본 발명의 다른 목적은 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 예방 또는 개선용 식품 조성물을 제공하는데 있다.In addition, another object of the present invention is to provide a food composition for the prevention or improvement of obesity containing peanut husk extract as an active ingredient.
상기한 목적을 달성하기 위한 본 발명의 비만을 예방 또는 치료할 수 있는 약학 조성물은 땅콩 겉껍질 추출물을 유효성분으로 포함할 수 있다.The pharmaceutical composition capable of preventing or treating obesity of the present invention for achieving the above object may contain peanut hull extract as an active ingredient.
상기 땅콩 겉껍질 추출물은 물, 탄소수 1 내지 4의 저급알코올 또는 이들의 혼합용매로 추출된 것일 수 있다.The peanut husk extract may be extracted with water, lower alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.
상기 혼합용매는 20 내지 99 부피%의 메탄올, 에탄올, 부탄올 또는 프로판올 수용액일 수 있다.The mixed solvent may be an aqueous solution of 20 to 99% by volume of methanol, ethanol, butanol or propanol.
상기 땅콩 겉껍질 추출물은 20 내지 50 kHz의 진동수 및 50 내지 700 W의 파워의 초음파기로 5 내지 60 분 동안 처리된 것일 수 있다.The peanut husk extract may be treated for 5 to 60 minutes with an ultrasonicator having a frequency of 20 to 50 kHz and a power of 50 to 700 W.
상기 초음파 처리 시 추출온도는 40 내지 100 ℃일 수 있다.During the ultrasonic treatment, the extraction temperature may be 40 to 100 °C.
또한, 상기한 다른 목적을 달성하기 위한 본 발명의 비만을 예방 또는 개선할 수 있는 식품 조성물은 땅콩 겉껍질 추출물을 유효성분으로 포함할 수 있다.In addition, the food composition capable of preventing or improving obesity of the present invention for achieving the above other object may include peanut hull extract as an active ingredient.
본 발명의 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 개선, 예방 또는 치료용 조성물은 리파아제 저해 활성이 시판되는 리파아제 저해제의 저해율에 상당하는 높은 저해 효과를 나타내며, 알파 글루코시데이즈 저해 활성 역시 시판되는 알파 글루코시데이즈 저해제의 저해율보다 높은 저해 효과를 나타내므로 비만에 의해 유발될 수 있는 질환을 개선, 예방 또는 치료할 수 있으므로, 식품 조성물, 나아가 건강기능식품 또는 약학 조성물로 활용될 수 있다.The composition for improving, preventing or treating obesity comprising the peanut husk extract of the present invention as an active ingredient exhibits a high inhibitory effect corresponding to the inhibition rate of commercially available lipase inhibitors in lipase inhibitory activity, and alpha glucosidase inhibitory activity is also commercially available Since it exhibits an inhibitory effect higher than the inhibition rate of alpha glucosidase inhibitors, which can improve, prevent, or treat diseases that can be caused by obesity, it can be used as a food composition, and furthermore, a health functional food or pharmaceutical composition.
또한, 본 발명의 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 개선, 예방 또는 치료용 조성물은 고지방 식이 동물에서 체중 감소효과가 있으므로 항비만 효과 있을 뿐만 아니라 항산화 효과도 우수하다.In addition, the composition for improving, preventing or treating obesity containing the peanut husk extract as an active ingredient of the present invention has an anti-obesity effect and an excellent antioxidant effect because it has a weight loss effect in animals on a high fat diet.
도 1a는 에탄올 농도와 추출시간에 따른 추출물의 총 폴리페놀 활성 변화를 나타내는 반응표면 곡선이며, 도 1b는 에탄올 농도와 추출온도에 따른 추출물의 총 폴리페놀 활성 변화를 나타내는 반응표면 곡선이다.
도 2a는 에탄올 농도와 추출시간에 따른 추출물의 총 플라보노이드 활성 변화를 나타내는 반응표면 곡선이며, 도 2b는 에탄올 농도와 추출온도에 따른 추출물의 총 플라보노이드 활성 변화를 나타내는 반응표면 곡선이다.
도 3a는 에탄올 농도와 추출시간에 따른 추출물의 DPPH 라디칼 소거능 변화를 나타내는 반응표면 곡선이며, 도 3b는 에탄올 농도와 추출온도에 따른 추출물의 DPPH 라디칼 소거능 변화를 나타내는 반응표면 곡선이다.
도 4는 본 발명의 실시예에 따라 제조된 땅콩 겉껍질의 초음파 최적 추출조건을 찾기 위하여 상기 도 1, 도 2 및 도 3의 반응표면곡선을 겹치기기법으로 나타낸 그래프이다.
도 5는 본 발명의 실시예 18에 따라 제조된 땅콩 겉껍질 추출물로 처리 시 항비만 기작의 핵심 단백질인 SREBP-1c, PPAR-γ및 FAS의 발현을 나타내는 웨스턴 블롯이다.
도 6a 내지 도 6c는 상기 도 5에서 SREBP-1c, PPAR-γ및 FAS의 발현을 정량화한 그래프이다. Figure 1a is a response surface curve showing the change in the total polyphenol activity of the extract according to the ethanol concentration and extraction time, Figure 1b is a response surface curve showing the change in the total polyphenol activity of the extract according to the ethanol concentration and extraction temperature.
Figure 2a is a response surface curve showing the change in total flavonoid activity of the extract according to the ethanol concentration and extraction time, Figure 2b is a response surface curve showing the change in total flavonoid activity of the extract according to the ethanol concentration and extraction temperature.
Figure 3a is a response surface curve showing the change in DPPH radical scavenging ability of the extract according to the ethanol concentration and extraction time, Figure 3b is a response surface curve showing the change in the DPPH radical scavenging ability of the extract according to the ethanol concentration and extraction temperature.
Figure 4 is a graph showing the response surface curves of Figures 1, 2 and 3 in an overlapping method in order to find the optimal ultrasonic extraction conditions for peanut shells prepared according to an embodiment of the present invention.
Figure 5 is a Western blot showing the expression of SREBP-1c, PPAR-γ and FAS, which are core proteins of the anti-obesity mechanism, when treated with the peanut hull extract prepared according to Example 18 of the present invention.
6a to 6c are graphs quantifying the expression of SREBP-1c, PPAR-γ, and FAS in FIG. 5 .
본 발명은 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 개선, 예방 또는 치료용 조성물에 관한 것이다.The present invention relates to a composition for improving, preventing or treating obesity comprising peanut hull extract as an active ingredient.
이하, 본 발명을 상세하게 설명한다. Hereinafter, the present invention will be described in detail.
본 발명의 비만을 개선, 예방 또는 치료할 수 있는 조성물은 땅콩 겉껍질 추출물을 유효성분으로 포함한다.The composition capable of improving, preventing or treating obesity of the present invention contains peanut hull extract as an active ingredient.
상기 땅콩(Penut, Arachis hypogaea)에서 발생되는 부산물인 땅콩 겉껍질은 종실을 보호하는 것으로서, 땅콩 겉껍질에는 염증질환이나 암과 같은 여러 질병에 효과가 있는 루테올린과 함께 플라보노이드 계열 화합물로서 강력한 항산화 효능이 있는 에리오딕티올을 함유되어 있다. The peanut shell, a by-product generated from the peanut (Penut, Arachis hypogaea ), protects the seeds, and the peanut shell is a flavonoid-based compound with luteolin, which is effective for various diseases such as inflammatory diseases and cancer, and has a strong antioxidant effect. Contains eriodictyol.
본 발명은 폐기되는 땅콩 겉껍질을 이용하는 것으로서, 종실을 제거한 땅콩 겉껍질만을 이용한다.The present invention uses discarded peanut hulls, and uses only peanut hulls from which seeds have been removed.
상기 땅콩 겉껍질은 추출용매 하에서 초음파처리를 통해 추출되는 것으로서, 구체적으로 땅콩 겉껍질과 추출용매가 1 : 5 내지 25의 중량비, 바람직하게는 1 : 8 내지 15의 중량비로 혼합되어 20 내지 50 kHz, 바람직하게는 30 내지 40 kHz의 진동수 및 50 내지 700 W, 바람직하게는 150 내지 400 W 파워의 초음파기로 40 내지 100 ℃, 바람직하게는 50 내지 95 ℃하에서 5 내지 60분, 바람직하게는 15 내지 50분 동안 처리된다. The peanut husk is extracted through sonication under an extraction solvent, specifically, the peanut husk and the extraction solvent are mixed in a weight ratio of 1: 5 to 25, preferably 1: 8 to 15, and 20 to 50 kHz , preferably with an ultrasonicator having a frequency of 30 to 40 kHz and a power of 50 to 700 W, preferably 150 to 400 W at 40 to 100 ° C., preferably 50 to 95 ° C. for 5 to 60 minutes, preferably 15 to 400 W. Processed for 50 minutes.
땅콩 겉껍질을 추출 시 초음파 추출이 아니라 열수 추출인 경우에는 유효성분이 소량 추출될 뿐만 아니라 항비만 및 항산화 효과가 낮을 수 있다.In the case of hot water extraction rather than ultrasonic extraction when extracting peanut shells, not only a small amount of active ingredients are extracted, but also the anti-obesity and antioxidant effects may be low.
상기 땅콩 겉껍질과 추출용매의 중량비가 상기 범위를 벗어나는 경우에는 추출물에 땅콩 겉껍질의 유효성분이 적은 양으로 추출될 수 있다. When the weight ratio of the peanut hull and the extraction solvent is out of the above range, a small amount of the active ingredient of the husk of the peanut may be extracted in the extract.
또한, 초음파기의 진동수 및 파워가 상기 하한치 미만인 경우에는 땅콩 겉껍질의 유효성분이 적은 양으로 추출될 수 있으며, 상기 상한치 초과인 경우에는 유효물질의 열분해, 중합 또는 유효성분 외에 다른 물질도 다량으로 추출되어 효과가 저하될 수 있다. In addition, when the frequency and power of the sonicator are less than the lower limit, the active ingredient of the peanut shell can be extracted in a small amount, and when the upper limit is exceeded, a large amount of other substances besides the thermal decomposition, polymerization or active ingredient is extracted. effect may be diminished.
또한, 추출온도 및 추출시간이 상기 하한치 미만인 경우에는 땅콩 겉껍질의 유효성분이 적은 양으로 추출될 수 있으며, 상기 상한치 초과인 경우에는 독성물질이 발생할 수 있다.In addition, if the extraction temperature and extraction time are less than the lower limit, the active ingredient of the peanut shell may be extracted in a small amount, and if it exceeds the upper limit, toxic substances may be generated.
상기 추출물을 추출하는 추출용매는 물, 탄소수 1 내지 4의 저급알코올, 에틸렌글리콜, 에틸에테르 또는 이들의 혼합용매이다. 상기 저급알코올로는 20 내지 99 부피%의 메탄올, 에탄올, 부탄올 또는 프로판올 수용액을 들 수 있으며, 바람직하게는 우수한 항비만 효과를 위하여 40 내지 99 부피%의 에탄올 수용액을 들 수 있다.The extraction solvent for extracting the extract is water, lower alcohol having 1 to 4 carbon atoms, ethylene glycol, ethyl ether, or a mixture thereof. Examples of the lower alcohol include 20 to 99 vol% methanol, ethanol, butanol or propanol aqueous solution, preferably 40 to 99 vol% ethanol aqueous solution for excellent anti-obesity effect.
본 발명의 땅콩 겉껍질물 추출물은 약학 조성물 외에 건강기능식품에도 사용될 수 있다.Peanut husk extract of the present invention can be used in health functional foods in addition to pharmaceutical compositions.
또한, 본 발명은 반응표면분석법을 이용하여 항산화용 땅콩 겉껍질 초음파 추출물을 제조하는 방법을 제공한다.In addition, the present invention provides a method for preparing an ultrasonic extract of peanut hull for antioxidant using a response surface analysis method.
본 발명의 반응표면분석법을 이용한 땅콩 겉껍질 초음파 추출물의 제조방법은 (A) 물, 탄소수 1 내지 4의 저급알코올 또는 이들의 혼합용매인 추출용매, 40 내지 100 ℃의 추출온도, 5 내지 60분의 추출시간의 추출조건 하에서 20 내지 50 kHz의 진동수 및 50 내지 700 W의 파워의 초음파기로 추출한 후 원심분리하여 수득한 상등액의 DPPH 라디칼 소거능에 대한 실험값을 획득하는 단계; (B) 상기 (A)단계의 실험값을 이용하여 2차 회귀식 모델을 도출하는 단계; (C) 상기 (B)단계에서 도출된 2차 회귀식 모델을 변량분석(ANOVA)하여 신뢰도를 입증하고 상기 추출조건에 대한 반응표면곡선을 수득하는 단계; (D) 상기 (A)단계에서 수득한 상등액의 총 폴리페놀 함량에 대한 실험값을 획득하는 단계; (E) 상기 (D)단계의 실험값을 이용하여 하기 2차 회귀식 모델을 도출하는 단계; (F) 상기 (E)단계에서 도출된 2차 회귀식 모델을 변량분석(ANOVA)하여 신뢰도를 입증하고 상기 추출조건에 대한 반응표면곡선을 수득하는 단계; (G) 상기 (A)단계에서 수득한 상등액의 총 플라보노이드 함량에 대한 실험값을 획득하는 단계; (H) 상기 (G)단계의 실험값을 이용하여 2차 회귀식 모델을 도출하는 단계; (I) 상기 (H)단계에서 도출된 2차 회귀식 모델을 변량분석(ANOVA)하여 신뢰도를 입증하고 상기 추출조건에 대한 반응표면곡선을 수득하는 단계; 및 (J) 상기 (C), (F) 및 (I)단계에서 수득된 반응표면곡선을 겹치기기법(superimposing)으로 DPPH 라디칼 소거능, 총 폴리페놀 함량 및 총 플라보노이드 함량에 대한 공통의 최적화 조건을 예측하는 단계;를 포함한다. 이렇게 예측된 조건을 이용하여 땅콩 겉껍질을 초음파 추출하면 DPPH 라디칼 소거능, 총 폴리페놀 함량 및 총 플라보노이드 함량에 대하여 우수한 효과를 보인다. Method for producing ultrasonic extract of peanut husk using response surface analysis of the present invention is (A) water, lower alcohol having 1 to 4 carbon atoms or an extraction solvent that is a mixed solvent thereof, extraction temperature of 40 to 100 ℃, 5 to 60 minutes Obtaining an experimental value for the DPPH radical scavenging ability of the supernatant obtained by centrifugation after extraction with an ultrasonicator having a frequency of 20 to 50 kHz and a power of 50 to 700 W under extraction conditions of an extraction time of; (B) deriving a quadratic regression model using the experimental values of step (A); (C) verifying reliability by performing analysis of variance (ANOVA) on the second-order regression model derived in step (B) and obtaining a response surface curve for the extraction conditions; (D) obtaining an experimental value for the total polyphenol content of the supernatant obtained in step (A); (E) deriving the following quadratic regression model using the experimental values of step (D); (F) verifying reliability by performing analysis of variance (ANOVA) on the second-order regression model derived in step (E) and obtaining a response surface curve for the extraction conditions; (G) obtaining an experimental value for the total flavonoid content of the supernatant obtained in step (A); (H) deriving a quadratic regression model using the experimental values of step (G); (I) verifying reliability by performing analysis of variance (ANOVA) on the second-order regression model derived in step (H) and obtaining a response surface curve for the extraction conditions; and (J) predict common optimization conditions for DPPH radical scavenging activity, total polyphenol content and total flavonoid content by superimposing the response surface curves obtained in steps (C), (F) and (I). including; Ultrasonic extraction of peanut shells using these predicted conditions shows excellent effects on DPPH radical scavenging ability, total polyphenol content, and total flavonoid content.
상기 DPPH 라디칼 소거능, 총 폴리페놀 함량 및 총 플라보노이드 함량에 대한 공통의 최적화 조건에 따른 추출조건은 추출 시간 35.8분, 추출 온도 82.7 ℃ 및 에탄올 농도 96.0 부피%이다.The extraction conditions according to the common optimization conditions for the DPPH radical scavenging activity, total polyphenol content, and total flavonoid content were extraction time of 35.8 minutes, extraction temperature of 82.7 °C, and ethanol concentration of 96.0% by volume.
본 발명에서 사용되는 용어 ‘추출물’은 상기 용매를 이용하여 땅콩 겉껍질에 포함된 성분을 추출한 추출물, 이들로부터 분획한 분획물, 이들 추출물 또는 분획물을 추가적으로 농축한 농축물, 이를 정제 또는 분리한 정제물도 포함하고, 상기 추출물, 분획물, 농축물 또는 정제물을 건조한 건조물 또는 그를 분쇄한 분말을 포함하는 의미로 사용된다. The term 'extract' as used in the present invention refers to extracts obtained by extracting components contained in peanut shells using the solvent, fractions fractionated therefrom, concentrates obtained by additionally concentrating these extracts or fractions, and purified or separated purified products. Including, it is used in the sense of including the extract, fraction, concentrate or purified product dried or pulverized powder thereof.
상기 정제물의 제조를 위해 분자량 컷-오프 값을 갖는 한외 여과막을 통과시키거나, 또는 다양한 크로마토그래피(크기, 전하, 소수성 또는 친화성에 따른 분리를 위해 제작된 것)에 의한 분리 등, 추가적으로 실시된 다양한 정제 방법을 부가할 수 있다.For the preparation of the purified product, a variety of additionally carried out, such as passing through an ultrafiltration membrane having a molecular weight cut-off value, or separation by various chromatography (made for separation according to size, charge, hydrophobicity or affinity). A purification method may be added.
본 발명은 땅콩 겉껍질 추출물을 유효성분으로 포함하는 비만의 예방 또는 개선용 식품 조성물에 관한 것이다.The present invention relates to a food composition for preventing or improving obesity comprising peanut husk extract as an active ingredient.
상기 ‘식품 조성물’은 유효성분으로 땅콩 겉껍질 추출물 이외에, 식품 제조에 통상적으로 사용되는 식품의 기준 및 규격(‘식품공전’)에 기재된 식품으로 사용가능한 식품 원료, 식품첨가물 공전에 기재된 식품첨가물을 포함한다.The 'food composition' includes, in addition to the peanut husk extract as an active ingredient, food raw materials usable as food described in the standards and specifications of food commonly used in food manufacturing ('Food Code'), and food additives described in the Food Additive Code include
특별히 한정할 필요는 없으나 예를 들어 단백질, 탄수화물, 지방, 영양소, 조미제 및 향미제를 포함한다. 상기 탄수화물은 단당류, 예를 들어, 포도당, 과당 등; 이당류, 예를 들어 말토스, 설탕, 유당 등; 올리고당 또는 폴리사카라이드, 예를 들어 덱스트린, 물엿, 사이클로덱스트린 등; 당알코올, 예를 들어 자일리톨, 소르비톨, 에리트리톨 등을 사용할 수 있다. 상기 향미제는 천연 향미제[타우마틴, 스테비아 추출물(예를 들어 레바우디오시드 A, 글리시르히진 등]) 및 합성 향미제(사카린, 아스파르탐 등)를 사용할 수 있다.It does not need to be particularly limited, but includes, for example, proteins, carbohydrates, fats, nutrients, seasonings and flavoring agents. The carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sugar, lactose and the like; oligosaccharides or polysaccharides such as dextrin, starch syrup, cyclodextrin and the like; Sugar alcohols such as xylitol, sorbitol, erythritol and the like can be used. As the flavoring agent, natural flavoring agents (thaumatin, stevia extract (eg, rebaudioside A, glycyrrhizin, etc.)) and synthetic flavoring agents (saccharin, aspartame, etc.) may be used.
상기 땅콩 겉껍질 추출물을 유효성분으로 식품 조성물을 제조하는 경우에 땅콩 겉껍질 추출물은 비만 질환을 예방 또는 치료할 수 있는 함량이면 특별히 한정할 필요는 없으나, 예를 들어 0.1 내지 99 중량%, 0.5 내지 95 중량%, 1 내지 90 중량%, 2 내지 80 중량%, 3 내지 70 중량%, 4 내지 60 중량%, 5 내지 50 중량%로 포함될 수 있다.In the case of preparing a food composition using the peanut hull extract as an active ingredient, the peanut hull extract does not need to be particularly limited as long as it can prevent or treat obesity diseases, but, for example, 0.1 to 99% by weight, 0.5 to 95 % by weight, 1 to 90% by weight, 2 to 80% by weight, 3 to 70% by weight, 4 to 60% by weight, may be included in 5 to 50% by weight.
상기 식품 조성물에서 유효성분인 땅콩 겉껍질 추출물은 섭취자의 상태, 체중, 질병의 유무나 정도 및 기간에 따라 다르지만, 통상의 기술자에 의해 적절하게 선택될 수 있다. 예들 들어 1일 투여량을 기준으로 1 내지 5,000 mg, 바람직하게는 5 내지 2,000 mg, 더욱 바람직하게는 10 내지 1,000 mg, 더더욱 바람직하게는 20 내지 800 mg, 가장 바람직하게는 50 내지 500 mg일 수 있고, 투여 횟수는 특별히 한정할 필요는 없으나 1일 3회 내지 1주일에 1회의 범위 내에서 통상의 기술자가 조절할 수 있다. 건강 및 위생을 목적으로 하거나 또는 건강 조절을 목적으로 하는 장기간의 섭취의 경우에는 상기 범위 이하일 수 있다.Peanut husk extract, which is an active ingredient in the food composition, varies depending on the condition, body weight, presence or absence of disease, degree and duration of the eater, but may be appropriately selected by a person skilled in the art. For example, it may be 1 to 5,000 mg, preferably 5 to 2,000 mg, more preferably 10 to 1,000 mg, still more preferably 20 to 800 mg, and most preferably 50 to 500 mg based on the daily dose. There is, and the number of administrations need not be particularly limited, but can be adjusted by a person skilled in the art within the range of three times a day to once a week. In the case of long-term intake for the purpose of health and hygiene or health control, it may be less than the above range.
상기 식품 조성물은 특별히 한정할 필요는 없으나 예를 들어 산제, 과립제, 정제, 캡슐제, 환제, 엑스제, 젤리 제형, 티백 제형 또는 음료 제형일 수 있다.The food composition does not need to be particularly limited, but may be, for example, a powder, granule, tablet, capsule, pill, extract, jelly formulation, tea bag formulation or beverage formulation.
또한 일반 식품에 비만의 예방 또는 개선의 기능성을 부여하기 위하여 상기 땅콩 겉껍질 추출물을 첨가할 수 있으며, 첨가가 가능한 식품은, 특별히 한정할 필요는 없으나 예를 들어 식품위생법 제7조에 따른 식품의 기준 및 규격(‘식품공전’)에 예시된 과자류, 빵 또는 떡류, 코코아가공품류 또는 초콜릿류, 식육 또는 알가공품, 어육가공품, 두부류 또는 묵류, 면류, 다류, 커피, 음료류, 특수용도식품, 장류, 조미식품, 드레싱류, 김치류, 젓갈류, 절임식품, 조림식품, 주류, 건포류, 기타 식품류 등에 첨가될 수 있다. 또한 축산물위생관리법 제4조에 따른 축산물의 가공기준 및 성분규격(‘축산물공전’)에 예시된 유가공품, 식육가공품 및 포장육, 알가공품에 첨가될 수 있다.In addition, the peanut husk extract can be added to general foods to provide functionality for preventing or improving obesity. and confectionery, bread or rice cakes, processed cocoa products or chocolates, processed meat or egg products, processed fish meat products, tofu or jelly, noodles, teas, coffee, beverages, special purpose foods, pastes, It can be added to seasoning foods, dressings, kimchi, salted fish, pickled foods, stewed foods, alcoholic beverages, raisins, and other foods. In addition, it can be added to dairy products, processed meat products, packaged meat, and egg products exemplified in the processing standards and ingredient specifications of livestock products ('livestock product code') according to Article 4 of the Livestock Products Sanitation Control Act.
한편, 상기 땅콩 겉껍질 추출물을 유효성분으로 하는 식품 조성물은 단독으로 “비만의 예방 또는 개선용 건강기능식품”으로 이용될 수 있다. On the other hand, the food composition containing the peanut husk extract as an active ingredient can be used alone as "health functional food for preventing or improving obesity".
상기 ‘건강기능식품’은 인체에 유용한 기능성을 가진 원료나 성분을 사용하여 법적 기준에 따라 제조(가공을 포함)한 식품(건강기능식품에 관한 법률 제3조 제1호)을 말한다. 상기 ‘건강기능식품’은 국가마다 용어나 범위에 차이가 있을 수 있으나, 미국의 ‘식이 보충제(Dietary Supplement)’, 유럽의 ‘식품 보충제(Food Supplemnet)’, 일본의 ‘보건기능식품’ 또는 ‘특정보건용식품(Food for Special Health Use, FoSHU)’, 중국의 ‘보건식품’ 등에 해당할 수 있다.The above ‘functional health food’ refers to food manufactured (including processing) in accordance with legal standards using raw materials or ingredients that have functional properties useful for the human body (
상기 식품 조성물 또는 건강기능식품은 식품첨가물을 추가로 포함할 수 있으며, 식품첨가물로서의 적합여부는 다른 규정이 없는 한 ‘식품첨가물공전’의 총칙 및 일반시험법 등에 따라 해당 품목에 관한 규격 및 기준에 따른다.The food composition or health functional food may additionally contain food additives, and the suitability as a food additive is determined according to the standards and standards for the item in accordance with the general rules and general test methods of the 'Food Additive Code' unless otherwise specified. follow
또한 상기 건강기능식품에는 상기 땅콩 겉껍질 추출물과 함께 “비만의 예방 또는 개선용 건강기능식품”에 사용되는 ‘기능성 원료’로 고시된 원료 또는 개별인정된 원료로서, Lactobacillus gasseri BNR17, L-카르니틴타르트 레이트, 가르시니아캄보지아껍질추출물, 공액리놀렌산(유리지방산), 공액리놀렌산(트리글리세라이드), 그린마떼추출물, 그린커피빈추출물, 깻잎추출물, 녹차추출물, 대두배아추출물 등 복합물, 돌외잎주정추출분말, 락토페린(우유정제단백질), 레몬 밤 추출물 혼합분말, 마테열수추출물, 미역 등 복합추출물(잔티젠), 발효식초석류복합물, 보이차추출물, 서목태(쥐눈이콩) 펩타이드 복합물, 식물성유지 디글리세라이드, 와일드망고 종자추출물, 중쇄지방산(MCFA)함유 유지, 콜레우스포스콜리추출물, 키토산, 키토올리고당, 풋사과추출폴리페놀, 핑거루트추출분말, 히비스커스 복합추출물 등의 체지방감소와 관련된 건강기능식품 소재를 복합하여 비만의 예방 또는 개선용 건강기능식품을 제조할 수 있다.In addition, the health functional food is a raw material notified as a 'functional raw material' or individually recognized raw material used in "health functional food for preventing or improving obesity" together with the peanut shell extract, Lactobacillus gasseri BNR17, L-carnitine tart Latex, Garcinia cambogia peel extract, conjugated linolenic acid (free fatty acid), conjugated linolenic acid (triglyceride), green mate extract, green coffee bean extract, sesame leaf extract, green tea extract, soybean embryo extract, etc. Milk purified protein), lemon balm extract mixed powder, mate hot water extract, seaweed, etc. complex extract (Xanthigen), fermented vinegar pomegranate complex, puerh tea extract, seomoktae (swine eye bean) peptide complex, vegetable oil diglyceride, wild mango seed Prevention of obesity by combining health functional food ingredients related to body fat reduction, such as extract, medium-chain fatty acid (MCFA)-containing oil, Coleus forskoli extract, chitosan, chitooligosaccharide, green apple-extracted polyphenol, finger root extract powder, and hibiscus complex extract Or health functional food for improvement can be manufactured.
또한 본 발명은 인간, 또는 인간을 제외한 동물에게 상기 조성물을 투여하는 비만의 치료방법을 제공한다.In addition, the present invention provides a method for treating obesity by administering the composition to humans or non-human animals.
또한 본 발명은 비만의 예방 또는 치료용 의약, 또는 동물용 의약 제조를 위한 땅콩 겉껍질 추출물의 신규 용도를 제공한다.In addition, the present invention provides a new use of the peanut hull extract for the preparation of drugs for preventing or treating obesity, or veterinary drugs.
상기 ‘약학 조성물’또는 ‘의약’은 유효성분으로 땅콩 겉껍질 추출물 이외에, 약학 조성물 등의 제조에 통상적으로 사용하는 적절한 담체, 부형제 및 희석제를 더 포함할 수 있다.The 'pharmaceutical composition' or 'medicine' may further include appropriate carriers, excipients, and diluents commonly used in the manufacture of pharmaceutical compositions, etc., in addition to peanut husk extract as an active ingredient.
상기 ‘담체’는 세포 또는 조직 내로의 화합물의 부가를 용이하게 하는 화합물이다. 상기 ‘희석제’는 대상 화합물의 생물학적 활성 형태를 안정화시킬 뿐만 아니라, 화합물을 용해시키게 되는 물에서 희석되는 화합물이다. The 'carrier' is a compound that facilitates the addition of the compound into cells or tissues. The 'diluent' is a compound diluted in water that not only stabilizes the biologically active form of the target compound, but also dissolves the compound.
상기 담체, 부형제 및 희석제로는 특별히 한정할 필요는 없으나 예를 들어, 유당, 포도당, 설탕, 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 광물유 등을 들 수 있다.The carrier, excipient, and diluent are not particularly limited, but, for example, lactose, glucose, sugar, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose , methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.
상기 약학 조성물, 의약, 동물용 약학 조성물 또는 동물용 의약의 사용량은 환자 또는 치료대상 동물의 나이, 성별, 체중에 따라 달라질 수 있으며, 무엇보다도, 치료대상 개체의 상태, 치료 대상 질환의 특정한 카테고리 또는 종류, 투여 경로, 사용되는 치료제의 속성에 의존적일 것이다.The amount of the pharmaceutical composition, drug, veterinary pharmaceutical composition, or veterinary drug used may vary depending on the age, sex, and weight of the patient or animal to be treated, and above all, the condition of the subject to be treated, a specific category of disease to be treated, or It will depend on the type, route of administration, and nature of the therapeutic used.
상기 약학 조성물, 의약, 동물용 약학 조성물 또는 동물용 의약은 체내에서 활성성분의 흡수도, 배설속도, 환자 또는 치료대상 동물의 연령 및 체중, 성별 및 상태, 치료할 질병의 중증정도 등에 따라 적절히 선택되나, 일반적으로 1일 0.1 내지 1,000 mg/kg, 바람직하게는 1 내지 500 mg/kg, 더욱 바람직하게는 5 내지 250 mg/kg, 가장 바람직하게는 10 내지 100 mg/kg으로 투여하는 것이 바람직하다. 이렇게 제형화 된 단위 투여형 제제는 필요에 따라 일정시간 간격으로 수회 투여할 수 있다.The pharmaceutical composition, drug, veterinary pharmaceutical composition or veterinary drug is appropriately selected according to the absorption rate of the active ingredient in the body, the rate of excretion, the age and weight, sex and condition of the patient or animal to be treated, the severity of the disease to be treated, etc. , It is generally preferred to administer 0.1 to 1,000 mg/kg, preferably 1 to 500 mg/kg, more preferably 5 to 250 mg/kg, and most preferably 10 to 100 mg/kg per day. The unit dosage formulation thus formulated can be administered several times at regular time intervals as needed.
상기 약학 조성물, 의약, 동물용 약학 조성물 또는 동물용 의약은 개별적으로 예방제 또는 치료제로서 투여하거나 다른 치료제와 병용하여 투여될 수 있고, 종래의 치료제와는 순차적 또는 동시에 투여될 수 있다.The pharmaceutical composition, medicament, veterinary pharmaceutical composition, or veterinary medicament may be administered individually as a prophylactic or therapeutic agent, or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents.
상기 약학조성물, 의약, 동물용 약학 조성물 또는 동물용 의약은 각각 통상의 방법에 따라 산제, 과립제, 정제, 캡슐제, 트로키제, 현탁액, 에멀젼, 시럽, 에어로졸 등의 경구 제형으로 제형화하여 사용될 수 있다. 제형화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 조제될 수 있다.The pharmaceutical composition, drug, veterinary pharmaceutical composition, or veterinary drug may be formulated into oral dosage forms such as powders, granules, tablets, capsules, troches, suspensions, emulsions, syrups, aerosols, etc. according to conventional methods and used. there is. When formulated, it may be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants.
경구 투여를 위한 고형 제제에는 정제, 환제, 산제, 과립제, 캡슐제, 트로키제 등이 포함되며, 이러한 고형 제제는 상기 화합물에 적어도 하나 이상의 부형제 예를 들면, 전분, 칼슘 카보네이트, 설탕 또는 유당, 젤라틴 등을 섞어 조제될 수 있다. 또한 단순한 부형제 이외에 마그네슘 스테아레이트, 탈크 같은 윤활제들도 사용될 수 있다. 경구를 위한 액상 제제로는 현탁제, 내용액제, 유제, 시럽제 등이 해당되는데 흔히 사용되는 단순 희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다.Solid preparations for oral administration include tablets, pills, powders, granules, capsules, troches, etc., and these solid preparations contain at least one excipient such as starch, calcium carbonate, sugar or lactose, and gelatin in the compound. It can be prepared by mixing etc. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Liquid preparations for oral use include suspensions, solutions for oral use, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, aromatics, and preservatives may be included. .
상기 비만의 치료방법은 인간, 또는 인간을 제외한 동물, 특히 포유동물에게 상기 조성물을 투여 하는 것으로, 예를 들어 비만인 치료대상 개체에게 상기 조성물을 투여하는 것이다.The method for treating obesity is to administer the composition to humans or non-human animals, particularly mammals, for example, to an obese subject to be treated.
상기 치료를 위한 투여량, 투여 방법 및 투여 횟수는 상기 약학 조성물, 의약, 동물용 약학 조성물 또는 동물용 의약의 투여량, 투여 방법 및 투여 횟수를 참고할 수 있다.The dosage, administration method and number of administrations for the treatment may refer to the dosage, administration method and number of administrations of the pharmaceutical composition, medicine, veterinary pharmaceutical composition or veterinary medicine.
이하, 본 발명의 이해를 돕기 위하여 바람직한 실시예를 제시하나, 하기 실시예는 본 발명을 예시하는 것일 뿐 본 발명의 범주 및 기술사상 범위 내에서 다양한 변경 및 수정이 가능함은 당업자에게 있어서 명백한 것이며, 이러한 변형 및 수정이 첨부된 특허청구범위에 속하는 것도 당연한 것이다.Hereinafter, preferred embodiments are presented to aid understanding of the present invention, but the following examples are merely illustrative of the present invention, and various changes and modifications are possible within the scope and spirit of the present invention. It is obvious to those skilled in the art, It goes without saying that these variations and modifications fall within the scope of the appended claims.
실시예 1 내지 17. Examples 1 to 17.
국내산 피땅콩을 수세하여 종실을 제거한 후 얻은 땅콩 겉껍질을 60 ℃ 건조오븐에서 24 시간 건조하여 더 이상의 중량이 낮아지지 않는 것을 확인하고 건조한 시료를 분쇄기로 분쇄한 후 25 mesh로 여과하였다. 상기 분쇄된 땅콩 겉껍질과 농도가 상이한 에탄올 수용액을 1 : 10의 중량비로 혼합하여 초음파기에 투입 후 초음파(40 kHz, 200 W)를 이용하여 추출하였다. After washing domestic peanuts with water to remove seeds, the peanut shells obtained were dried in a drying oven at 60 ° C for 24 hours to confirm that the weight did not decrease any more, and the dried samples were pulverized with a grinder and filtered through a 25 mesh. The pulverized peanut shells and aqueous ethanol solutions having different concentrations were mixed at a weight ratio of 1: 10, put into a sonicator, and then extracted using ultrasonic waves (40 kHz, 200 W).
이때 추출 공정의 추출시간, 추출온도 및 에탄올 농도를 다르게 하여 각각의 추출물을 수득하였다.At this time, each extract was obtained by varying the extraction time, extraction temperature, and ethanol concentration of the extraction process.
[항산화][antioxidant]
<시험예 I><Test Example I>
시험예 1. DPPH 라디칼 소거능, 총 폴리페놀(TPC) 및 총 플라보노이드(TFC) 측정Test Example 1. Measurement of DPPH radical scavenging activity, total polyphenols (TPC) and total flavonoids (TFC)
1-1. DPPH 라디칼 소거능(%): DPPH(2,2-Diphenyl-1-picrylhydrazyl) free radical 소거활성을 측정하는 Lee등의 방법을 변형하여 사용하였다. 라디칼이 항산화 물질로 인하여 환원되면 고유의 보라색을 잃고 노란색으로 변색되는데 이를 흡광도로 계산하여 라디칼 소거능을 측정하였다. DPPH 시약을 MtOH로 희석하여 분광광도계를 이용해 517 nm에서 O.D 값을 1.0로 맞춰 DPPH 희석액을 준비한다. 준비한 시료 0.25 ml에 희석한 DPPH 시약 1.25 ml를 가하여 암실에서 20분간 정치반응 후 원심분리하여 517 nm에서 흡광도를 측정하고 대조군과 비교하여 아래와 같이 free radical 소거활성을 백분율로 표현하였다. 대조군으로는 ascorbic acid를 이용하였다.1-1. DPPH radical scavenging activity (%): The method of measuring DPPH (2,2-Diphenyl-1-picrylhydrazyl) free radical scavenging activity was modified and used. When radicals are reduced due to antioxidants, they lose their original purple color and turn yellow, which was calculated by absorbance to measure radical scavenging ability. Dilute the DPPH reagent with MtOH and adjust the O.D value to 1.0 at 517 nm using a spectrophotometer to prepare a DPPH diluted solution. 1.25 ml of diluted DPPH reagent was added to 0.25 ml of the prepared sample, left for 20 minutes in a dark room, centrifuged, and the absorbance was measured at 517 nm. The free radical scavenging activity was expressed as a percentage as shown below compared to the control group. As a control, ascorbic acid was used.
[수학식 1][Equation 1]
라디칼 소거활성(%)={1-(Abs(test)-Abs(color))/Abs(control)}×100Radical scavenging activity (%) = {1-(Abs (test) -Abs (color) ) / Abs (control) } × 100
1-2. 총 폴리페놀: 총 폴리페놀(Total Polyphenol Content) 함량은 변형된 Folin-Ciocalteu’s 방법을 이용하여 측정하였다. Folin-Ciocalteu’s 페놀 용액을 시료에 첨가하여 폴리페놀 화합물에 의해 환원되어 청색으로 발색되는 원리에 근거하여 흡광도 변화로 시료의 총 폴리페놀 농도를 측정하였다. 각 시료 0.14 mL를 취하고 0.2 N F.C 페놀용액을 0.7 mL 가하여 8분간 실온에 반응시켰다. 0.56 mL의 7.5% Sodium carbonate 용액을 첨가하여 실온에서 1시간 반응시킨 후 10초간 vortexing하고 분광광도계로 756 nm에서 흡광도를 측정하였다. 표준물질로 Gallic acid를 사용하였고 총 폴리페놀 함량은 gallic acid를 농도별로 희석하여 검량선을 작성한 후 mg Gallic Acid Equivalent (GAE) / g Dried Material (DM)으로 표시하였다. 1-2. Total Polyphenol: Total Polyphenol Content was measured using a modified Folin-Ciocalteu's method. Folin-Ciocalteu's phenol solution was added to the sample, and the total polyphenol concentration of the sample was measured based on the absorbance change based on the principle that it is reduced by the polyphenol compound and develops blue color. Take 0.14 mL of each sample, add 0.7 mL of 0.2 N F.C. phenol solution, and react at room temperature for 8 minutes. After adding 0.56 mL of 7.5% sodium carbonate solution, reacting at room temperature for 1 hour, vortexing for 10 seconds, and measuring absorbance at 756 nm with a spectrophotometer. Gallic acid was used as a standard material, and the total polyphenol content was expressed as mg Gallic Acid Equivalent (GAE) / g Dried Material (DM) after preparing a calibration curve by diluting gallic acid by concentration.
1-3. 총 플라보노이드: 총 플라보노이드(Total Flavonoid Content) 함량은 Zhishen 등의 방법을 변형하여 사용하였다. 플라보노이드에 알칼리를 작용시키면 황색으로 발색되는 원리에 근거하여 흡광도를 측정해 총 플라보노이드 농도를 측정하였다. 각 시료 0.5 mL에 DW 2.5 ml와 99% 에탄올 1.5 ml 가한 후 1M potassium acetate 0.1 ml와 10% aluminum chloride 0.1 ml를 가하여 vortexing 후 실온에서 30분간 방치하였다. 415 nm에서 흡광도를 측정하였으며 quercetin을 200, 100, 50, 25 ul/ml로 농도 별로 희석하여 검량선을 구하여 플라보노이드 함량을 mg Quercetin Equivalent (QE) / g Dried Material (DM)로 나타내었다. 1-3. Total Flavonoids: The total flavonoid content was used by modifying the method of Zhishen et al. The total flavonoid concentration was measured by measuring the absorbance based on the principle that flavonoids are colored yellow when alkali is applied. After adding 2.5 ml of DW and 1.5 ml of 99% ethanol to 0.5 mL of each sample, 0.1 ml of 1M potassium acetate and 0.1 ml of 10% aluminum chloride were added, vortexed, and allowed to stand at room temperature for 30 minutes. Absorbance was measured at 415 nm, and quercetin was diluted by concentration to 200, 100, 50, and 25 ul/ml to obtain a calibration curve, and the flavonoid content was expressed as mg Quercetin Equivalent (QE) / g Dried Material (DM).
(min)extraction time
(min)
(℃)extraction temperature
(℃)
(mg GAE/mg DW)total polyphenols
(mg GAE/mg DW)
(mg QE/mg DW)total flavonoids
(mg QE/mg DW)
총 폴리페놀 함량에 미치는 조건 탐색Exploration of conditions affecting total polyphenol content
각각의 추출 조건에 따른 추출물의 활성 변화를 반응표면 곡선으로 나타낸 결과는 도 1과 같다. 도 1a는 에탄올 농도와 추출시간에 따른 추출물의 총 폴리페놀 활성 변화를 나타내는 반응표면 곡선이며, 도 1b는 에탄올 농도와 추출온도에 따른 추출물의 총 폴리페놀 활성 변화를 나타내는 반응표면 곡선이다.Results showing the change in activity of the extract according to each extraction condition as a response surface curve are shown in FIG. 1. Figure 1a is a response surface curve showing the change in the total polyphenol activity of the extract according to the ethanol concentration and extraction time, Figure 1b is a response surface curve showing the change in the total polyphenol activity of the extract according to the ethanol concentration and extraction temperature.
위 표 1 및 도 1에 도시된 바와 같이, 실험값을 기반하여 최적화 기법으로 예측된 땅콩 겉껍질 추출물의 총 폴리페놀의 최대 수율은 추출 시간 55.0 분, 추출 온도 94.0 ℃ 및 에탄올 농도 74.3%의 조건에서 10.2 mg GAE/g DM으로 확인되었다. As shown in Table 1 and FIG. 1 above, the maximum yield of total polyphenols in the peanut hull extract predicted by the optimization technique based on the experimental values was under the conditions of an extraction time of 55.0 minutes, an extraction temperature of 94.0 ° C, and an ethanol concentration of 74.3%. It was confirmed as 10.2 mg GAE/g DM.
총 플라보노이드 함량에 미치는 조건 탐색Exploration of conditions affecting total flavonoid content
각각의 추출 조건에 따른 추출물의 활성 변화를 반응표면 곡선으로 나타낸 결과는 도 2와 같다. 도 2a는 에탄올 농도와 추출시간에 따른 추출물의 총 플라보노이드 활성 변화를 나타내는 반응표면 곡선이며, 도 2b는 에탄올 농도와 추출온도에 따른 추출물의 총 플라보노이드 활성 변화를 나타내는 반응표면 곡선이다.The results showing the change in activity of the extract according to each extraction condition as a response surface curve are shown in FIG. 2. Figure 2a is a response surface curve showing the change in total flavonoid activity of the extract according to the ethanol concentration and extraction time, Figure 2b is a response surface curve showing the change in total flavonoid activity of the extract according to the ethanol concentration and extraction temperature.
플라보노이드는 다양한 생리적 기능을 갖는 페놀계 화합물의 총칭이며 유리 상태로 존재하거나 당류와 결합한 배당체의 형태로 존재하며 열수와 유기용매를 이용한 추출을 통해 생산되어 식품, 화장품 및 의약품 산업에서 널리 이용된다.Flavonoids are a generic term for phenolic compounds with various physiological functions and exist in a free state or in the form of glycosides combined with sugars. They are produced through extraction using hot water and organic solvents and are widely used in the food, cosmetic and pharmaceutical industries.
위 표 1 및 도 2에 도시된 바와 같이, 땅콩 겉껍질 추출물의 총 플라보노이드 활성에 최적 추출 조건을 최적화 기법으로 예측한 결과, 총 플라보노이드의 최대 수율은 추출 시간 49.0 분, 추출 온도 94.0 ℃ 및 에탄올 농도 77.1%의 조건에서 1.18 mg QE/g DM으로 확인되었다. As shown in Table 1 and FIG. 2 above, as a result of predicting the optimal extraction conditions for the total flavonoid activity of the peanut hull extract with the optimization technique, the maximum yield of total flavonoids was 49.0 minutes for extraction time, 94.0 ° C. for extraction temperature, and ethanol concentration. It was confirmed as 1.18 mg QE/g DM in 77.1% of conditions.
추출 온도가 총 플라보노이드 함량에 유의한 영향을 미치나 추출 시간이 미치는 영향이 크지 않다는 것을 재차 확인하였으며 추출 온도에 비해 에탄올 농도에 의존성이 높다는 것을 확인하였다. Although the extraction temperature had a significant effect on the total flavonoid content, it was confirmed again that the effect of the extraction time was not significant, and it was confirmed that the dependence on the ethanol concentration was higher than the extraction temperature.
DPPH 라디칼 소거능에 미치는 조건 탐색Exploration of conditions affecting DPPH radical scavenging activity
각각의 추출 조건에 따른 추출물의 활성 변화를 반응표면 곡선으로 나타낸 결과는 도 3과 같다. 도 3a는 에탄올 농도와 추출시간에 따른 추출물의 DPPH 라디칼 소거능 변화를 나타내는 반응표면 곡선이며, 도 3b는 에탄올 농도와 추출온도에 따른 추출물의 DPPH 라디칼 소거능 변화를 나타내는 반응표면 곡선이다.The results showing the change in activity of the extract according to each extraction condition as a response surface curve are shown in FIG. 3. Figure 3a is a response surface curve showing the change in DPPH radical scavenging ability of the extract according to the ethanol concentration and extraction time, Figure 3b is a response surface curve showing the change in the DPPH radical scavenging ability of the extract according to the ethanol concentration and extraction temperature.
위 표 1 및 도 3에 도시된 바와 같이, 땅콩 겉껍질 추출물의 총 플라보노이드 활성에 최적 추출 조건을 최적화 기법으로 예측한 결과, DPPH 라디칼 소거능의 최대값은 추출 시간 55.0 분, 추출 온도 81.2 ℃ 및 에탄올 농도 73.2%의 조건에서 89.4%로 확인되었다.As shown in Table 1 and FIG. 3 above, as a result of predicting the optimal extraction conditions for the total flavonoid activity of the peanut husk extract with the optimization technique, the maximum value of DPPH radical scavenging ability was 55.0 minutes for extraction time, 81.2 ° C. for extraction temperature, and ethanol. It was confirmed as 89.4% under the condition of 73.2% concentration.
최적조건 탐색Optimal condition search
땅콩 겉껍질 추출물에 있어 주요 3 조건인 DPPH 라디칼 소거능(RSA), 총 폴리페놀(TPC) 및 총 플라보노이드(TFC)의 개별 최적화 결과를 바탕으로 Design Expert를 이용하여 반응표면곡선을 겹치기기법(superimposing)으로 3개 변수의 공통 최적화 조건을 예측하여 도 6(X1: 추출시간, X2: 추출온도)에 나타내었다. Based on the individual optimization results of DPPH radical scavenging ability (RSA), total polyphenols (TPC), and total flavonoids (TFC), which are the three main conditions in peanut hull extract, design expert was used to superimpose response surface curves 6 (X 1 : extraction time, X 2 : extraction temperature) by predicting the common optimization conditions of the three variables.
도 4에 도시된 바와 같이, 모든 종속 변수(TPC, TFC, RSA)의 최대화를 동시에 만족하는 최적 조건 중 경제적인 운전 조건을 위한 추출 온도가 최소화된 지점을 선택했을 때, 추출 시간은 35.8 분, 추출 온도는 82.7 ℃ 및 에탄올 농도는 96.0 부피%로 예측되었다. 이때, TPC, TFC 및 RSA는 각각 8.67 mg GAE/g DM, 0.84 mg QE/g DM 및 85.3%로 확인되었다. 통계학적 최적화를 통해 생리활성물질 추출의 최적 조건인 실시예 18은 추출 시간 49.0 분, 추출 온도 94.0 ℃ 및 에탄올 농도 77.1 부피%이며 이를 이용하여 향후 실험을 진행하였다 As shown in FIG. 4, when the point at which the extraction temperature is minimized for economical operating conditions is selected among the optimal conditions that simultaneously satisfy the maximization of all dependent variables (TPC, TFC, and RSA), the extraction time is 35.8 minutes, The extraction temperature was predicted to be 82.7 °C and the ethanol concentration to be 96.0% by volume. At this time, TPC, TFC and RSA were identified as 8.67 mg GAE/g DM, 0.84 mg QE/g DM and 85.3%, respectively. Example 18, which is the optimal condition for extracting bioactive substances through statistical optimization, has an extraction time of 49.0 minutes, an extraction temperature of 94.0 ° C, and an ethanol concentration of 77.1% by volume, and future experiments were conducted using this.
[항비만][Anti-obesity]
<시험예 Ⅱ><Test Example Ⅱ>
시험예 2. 리파아제 활성 억제능(LAI) 및 α-글루코시다아제 활성 억제능(GAI)Test Example 2. Lipase activity inhibitory activity (LAI) and α-glucosidase activity inhibitory activity (GAI) 측정measurement
실시예 1 내지 18에 따라 제조된 추출물을 이용하여 비만의 지표인 리파아제 활성 억제능(Lipase activity inhibition , LAI)와 α-글루코시다아제 활성 억제능(α-Glucosidase activity inhibition, GAI)를 측정하였다.Lipase activity inhibition (LAI) and α-glucosidase activity inhibition (GAI), which are indicators of obesity, were measured using the extracts prepared according to Examples 1 to 18.
중심합성계획법(central composite design, CCD)을 이용하여 추출시간(X1), 추출온도(X2), 용매농도(X3)에 대하여, 5단계의 -1.68, -1, 0, 1 및 1.68로 코드화하여 17개 실험범위를 설계하고, 설계된 하기 조건과 앞선 최적화 조건(실시예 18)를 통해 도출된 조건에 따라 항비만 실험을 통해 실험값을 수득하였다.Extraction time (X 1 ), extraction temperature (X 2 ), and solvent concentration (X 3 ) using central composite design (CCD) were -1.68, -1, 0, 1 and 1.68 in 5 steps. 17 experimental ranges were designed by coding, and experimental values were obtained through anti-obesity experiments according to the conditions derived through the designed following conditions and the preceding optimization conditions (Example 18).
2-1. 리파아제 활성 억제능(%): 리파아제 효소 활성 억제능이 우수하면 장내에서 지방의 흡수를 억제할 수 있어 비만 예방에 효과가 있다. 돼지 췌장 리파아제로 lipase 활성 억제능을 측정하는 실험을 수행하였다. 시료를 0.1 mg/mL의 농도로 희석한 후에 0.167 mM의 p-nitrophenylpalmitate용액, 0.061 M의 Tris-HCl buffer (pH 8.5) 및 0.3 mg/mL의 리파아제와 함께 plate에 넣고 25 ℃에서 10 분간 반응시켰다. 반응 후, 405 nm에서 흡광도를 측정하였으며, 대조군(control)은 시료를 용매로 대체하여 실험을 수행하였다. 이후 리파아제 억제활성은 하기 수학식으로 계산하였다.2-1. Lipase activity inhibitory ability (%): Excellent ability to inhibit lipase enzyme activity can inhibit the absorption of fat in the intestine, which is effective in preventing obesity. An experiment was performed to measure the lipase activity inhibitory ability with porcine pancreatic lipase. After diluting the sample to a concentration of 0.1 mg/mL, it was placed on a plate with 0.167 mM p-nitrophenylpalmitate solution, 0.061 M Tris-HCl buffer (pH 8.5), and 0.3 mg/mL lipase, and reacted at 25 °C for 10 minutes. . After the reaction, absorbance was measured at 405 nm, and as a control, the experiment was performed by replacing the sample with a solvent. After that, the lipase inhibitory activity was calculated by the following formula.
[수학식 2][Equation 2]
리파아제 억제 활성(%) = 1-(시료의 흡광도/control의 흡광도) X 100Lipase inhibitory activity (%) = 1-(absorbance of sample/absorbance of control)
2-2. α-글루코시다아제 활성 억제능(%): 시료 50 ㎕에 0.5 U/ml 알파-글루코시데이즈 효소액 50 ㎕를 첨가하고 여기에 200 mM 인산나트륨 완충액(pH 6.8) 50 ㎕를 혼합하여 37 ℃에서 10 분간 예비 배양한 후, 인산나트륨 완충액(pH 6.8)을 100 ㎕ 가하여 37 ℃에서 10 분간 반응시켰다. 반응액에 100 mM Na2CO3 0.75 ml를 첨가하여 반응을 정지시키고 420 nm에서 흡광도를 측정하였으며 음성 대조구는 시료 대신 증류수를 사용하여 동일한 방법으로 진행하여 흡광도의 차이를 하기 수학식에 의해 백분율(%)로 산출하였다.2-2. α-glucosidase activity inhibitory ability (%): 50 μl of 0.5 U/ml alpha-glucosidase enzyme solution was added to 50 μl of the sample, mixed with 50 μl of 200 mM sodium phosphate buffer (pH 6.8), pre-incubated at 37 ° C. for 10 minutes, and then mixed with sodium phosphate buffer (pH 6.8). 6.8) was added and reacted at 37°C for 10 minutes. The reaction was stopped by adding 0.75 ml of 100 mM Na 2 CO 3 to the reaction solution, and the absorbance was measured at 420 nm. The negative control proceeded in the same manner using distilled water instead of the sample, and the difference in absorbance was calculated as a percentage ( %) was calculated.
[수학식 3][Equation 3]
알파-글루코시다아제 저해활성(%) = [1-(음성대조구 흡광도 ÷ 실험구 흡광도)] X 100Alpha-glucosidase inhibitory activity (%) = [1-(absorbance of negative control group ÷ absorbance of experimental group)]
(min)extraction time
(min)
(℃)extraction temperature
(℃)
위 표 2에 나타낸 바와 같이, 리파아제 활성 억제능에 대한 실험값은 실시예 18에 따라 제조된 추출물이 다른 군에 비하여 우수한 것을 확인하였고, α-글루코시다아제 활성 억제능은 실시예 15에 따라 제조된 추출물이 다른 군에 비하여 우수한 것을 확인하였다. 항비만에 효과를 가지는 두 가지 효소에 대한 억제 활성은 실시예 18이 다른 군에 비하여 우수한 것으로 나타났다. 특히, 리파아제 저해제로 시판되고 있는 orlistat는 85.74±1.35%의 저해율을 나타내므로 실시예 18에 따라 제조된 추출물이 orlistat 대비 유의적으로 높은 저해 활성을 나타내는 것을 확인하였다. 양성대조군에 상당하는 높은 억제효과를 나타내며 리파아제 저해 활성이 높은 땅콩 겉껍질 추출물로부터 항비만 활성을 기대할 수 있다.As shown in Table 2 above, the experimental values for the lipase activity inhibitory ability confirmed that the extract prepared according to Example 18 was superior to the other groups, and the α-glucosidase activity inhibitory ability was confirmed by the extract prepared according to Example 15. It was confirmed that it was superior to other groups. Inhibitory activity for the two enzymes having an anti-obesity effect was found to be superior to that of Example 18 compared to the other groups. In particular, since orlistat, which is commercially available as a lipase inhibitor, exhibits an inhibition rate of 85.74±1.35%, it was confirmed that the extract prepared according to Example 18 exhibits significantly higher inhibitory activity than orlistat. Anti-obesity activity can be expected from the peanut hull extract, which shows a high inhibitory effect equivalent to that of the positive control group and has high lipase inhibitory activity.
또한, 알파 글루코시데이즈 저해제로 알려진 acarbose는 86.65±0.24%의 저해율을 나타내므로 실시예 18에 따라 제조된 추출물이 acarbose 대비 유의적으로 높은 저해 활성을 보여 항비만 및 식후 혈당 조절제로서의 활용 가능성이 뛰어날 것으로 판단된다.In addition, since acarbose, known as an alpha glucosidase inhibitor, exhibits an inhibition rate of 86.65 ± 0.24%, the extract prepared according to Example 18 shows a significantly higher inhibitory activity than acarbose, so it is highly likely to be used as an anti-obesity and postprandial blood glucose regulator. It is judged to be
반면, 정제수를 사용한 실시예 13은 리파아제 활성 억제능 및 α-글루코시다아제 활성 억제능 모두 수치가 낮은 것을 확인하였다. On the other hand, in Example 13 using purified water, it was confirmed that both lipase activity inhibitory ability and α-glucosidase activity inhibitory ability were low.
리파아제 활성 억제능 및 α-글루코시다아제 활성 억제능이 모두 고르게 우수한 군은 실시예 7, 15와 18인 것을 확인되었으며 실시예 18에서 최대값을 가짐을 확인하였다.It was confirmed that examples 7, 15, and 18 were equally excellent in both lipase activity inhibitory ability and α-glucosidase activity inhibitory ability, and it was confirmed that Example 18 had the maximum value.
시험예 3. 항비만 유전자 발현 Test Example 3. Anti-obesity gene expression
분화가 완료된 지방전구세포를 인산완충용액으로 세척한 후 추출 키트를 사용하여 전체 RNA를 추출한 후 상기 추출된 전체 RNA를 diethyl pyrocarbonate (DEPC)로 희석하고 260 ㎚에서 흡광도를 측정하여 정량한 다음 RNA (1 μg)에 cDNA synthesis 완충용액과 cDNA synthsis enzyme mix를 혼합하여 역전사 반응하여 cDNA를 합성하였다. 항비만 핵심유전자인 SREBP-1c, PPAR-γ와 FAS 프라이머를 첨가해 RT-PCR을 수행하였다. 지방전구세포는 지방세포로 분화되면서 중성지방 축적에 관여하는 전사인자를 유도하며, 분화 초기과정에서 cAMP의 농도가 증가하게되면 SREMP-1c 발현이 유도됨에 따라 PPAR-γ와 CEBP-α가 상호 발현되고 중성지방 합성에 관여하는 효소인 FAS와 함께 지방세포의 분화를 유도하게 된다.After washing differentiated preadipocytes with phosphate buffer, total RNA was extracted using an extraction kit, the extracted total RNA was diluted with diethyl pyrocarbonate (DEPC), absorbance was measured at 260 nm, and RNA was quantified ( 1 μg) was mixed with cDNA synthesis buffer and cDNA synthesis enzyme mix, and cDNA was synthesized by reverse transcription reaction. RT-PCR was performed by adding SREBP-1c, PPAR-γ and FAS primers, which are key anti-obesity genes. As preadipocytes differentiate into adipocytes, transcription factors involved in triglyceride accumulation are induced, and when the concentration of cAMP increases during the early stage of differentiation, SREMP-1c expression is induced, resulting in mutual expression of PPAR-γ and CEBP-α. It induces the differentiation of adipocytes together with FAS, an enzyme involved in triglyceride synthesis.
도 5는 본 발명의 실시예 18에 따라 제조된 땅콩 겉껍질 추출물(1 mg/ml)로 처리 시 SREBP-1c, PPAR-γ및 FAS의 발현을 나타내는 웨스턴 블롯이다. 도 6a 내지 도 6c는 상기 도 5에서 SREBP-1c, PPAR-γ및 FAS의 발현을 정량화한 그래프이다. Figure 5 is a Western blot showing the expression of SREBP-1c, PPAR-γ and FAS when treated with peanut hull extract (1 mg/ml) prepared according to Example 18 of the present invention. 6a to 6c are graphs quantifying the expression of SREBP-1c, PPAR-γ, and FAS in FIG. 5 .
도 5 및 도 6에 도시된 바와 같이, 지방전구세포에서 지방세포로 분화된 세포를 대조군 1로 이용하였으며, 실시예 18에 따라 제조된 땅콩 겉껍질 추출물 처리군은 대조군 1 대비 SREBP-1c, PPAR-γ와 FAS의 유전자 발현이 유의적으로 감소하는 것을 확인하였다(p < 0.05). As shown in Figures 5 and 6, cells differentiated from preadipocytes into adipocytes were used as control 1, and the peanut hull extract treatment group prepared according to Example 18 was compared to control 1 SREBP-1c, PPAR It was confirmed that the gene expressions of -γ and FAS were significantly decreased ( p < 0.05).
이에 따라, 땅콩 겉껍질 추출물이 보유한 항비만 활성이 지방전구세포 분화과정에 관여하는 핵심 유전자 발현 저해 확인을 통해 지방전구세포와 지방축적 억제에 효능이 있음을 확인하였다. Accordingly, it was confirmed that the anti-obesity activity possessed by the peanut husk extract inhibits the expression of key genes involved in the differentiation process of pre-adipocytes and is effective in inhibiting pre-adipocytes and fat accumulation.
시험예 4. 동물실험Test Example 4. Animal testing
200~250 g의 sprague-dawley계 마우스(5주령, 숫컷)를 실험에 사용하였다. 아크릴 케이지(45 X 60 X 25 cm)에서 사육되었으며, 충분한 사료와 물이 공급되며 적절한 인공조도로 12시간의 낮, 밤을 조절하였다(am 8:00부터 낮). 그리고 일정한 온도(20~24℃) 및 습도(45~65%)를 유지시켜 주었다. 바뀐 환경에 적응하도록 일주일간 살펴보며 수면주기를 유지하고, 이상행동을 확인하였다. 200-250 g sprague-dawley mice (5 weeks old, male) were used in the experiment. Raised in an acrylic cage (45 X 60 X 25 cm), sufficient feed and water were supplied, and 12 hours of day and night were controlled with appropriate artificial lighting (day from 8:00 am). And a constant temperature (20 ~ 24 ℃) and humidity (45 ~ 65%) was maintained. The sleep cycle was maintained for a week to adapt to the changed environment, and abnormal behavior was checked.
동물의 프로토콜은 선문대학교의 기관 동물 관리 및 사용위원회(IACUC)에 의해 승인되었다. Animal protocols were approved by the Institutional Animal Care and Use Committee (IACUC) of Sunmoon University.
실험동물은 체중변화가 일정하고 건강한 동물만을 선별하여 임의 배치법에 의해 일반식이를 급여한 정상군, 고지방식이를 급여한 대조군 2, 고지방식이 및 실시예 4의 추출물을 급여한 군으로 나누었으며 실험군마다 8마리씩 사용하였다. The experimental animals were selected only for healthy animals with constant weight change, and were divided into a normal group fed a normal diet by random arrangement method, a control group 2 fed a high-fat diet, and a group fed a high-fat diet and the extract of Example 4. Eight rats were used in each experimental group.
실시예 4의 추출물은 총 식이를 기준으로 5 중량%로 혼합하여 사용하였다.The extract of Example 4 was mixed and used at 5% by weight based on the total diet.
-3개 군의 마우스--3 groups of mice-
정상군: 일반식이 급여 8마리Normal group: 8 animals fed with normal diet
대조군: 고지방식이 급여 8마리 Control group: 8 animals fed a high-fat diet
실시예 18: 고지방식이 + 실시예 18의 추출물 급여 8마리Example 18: High-fat diet + 8 animals fed with the extract of Example 18
정상군, 대조군 및 실시예 18에 급여된 식이의 조성은 하기와 같다.The composition of the diet fed to the normal group, control group and Example 18 is as follows.
4-1. 마우스의 무게 측정4-1. Weigh the mouse
마우스의 초기 무게와 4주 후의 무게를 측정하였다.The initial weight of the mouse and the weight after 4 weeks were measured.
위 표 4에 나타낸 바와 같이, 정상군, 대조군 및 실시예 18군에 있어서 세 군 모두 초기 체중 조건 및 식이 섭취량이 유사하였으나, 4주 후 최종 체중에 있어 실시예 18군이 대조군에 비하여 체중감소 효과가 있는 것을 확인하였다. As shown in Table 4 above, in the normal group, the control group, and the Example 18 group, all three groups had similar initial body weight conditions and food intake, but in the final body weight after 4 weeks, the Example 18 group had a weight loss effect compared to the control group confirmed that there is
즉, 실시예 18군의 체중 증감을 보면 정상군에 비해서는 증가하였으나 대조군 2에 비해서는 감소된 것을 확인하였다. That is, looking at the weight gain and decrease in the Example 18 group, it was confirmed that it increased compared to the normal group but decreased compared to the control group 2.
아래에 본 발명의 추출물을 포함하는 조성물의 제제예를 설명하나, 본 발명은 이를 한정하고자 함이 아닌 단지 구체적으로 설명하고자 함이다.Examples of formulations of compositions containing the extract of the present invention are described below, but the present invention is not intended to limit them, but is intended to be specifically described.
제제예 1: 산제의 제조Formulation Example 1: Preparation of powder
실시예 4의 추출물 분말 20 mg20 mg of extract powder of Example 4
유당 100 mg
탈크 10 mg
상기의 성분들을 혼합하고 기밀포에 충진하여 산제를 제조한다.A powder is prepared by mixing the above ingredients and filling them in an airtight bag.
제제예 2: 정제의 제조Formulation Example 2: Preparation of tablets
실시예 4의 추출물 분말 10 mg10 mg of extract powder of Example 4
옥수수전분 100 mg
유당 100 mg
스테아린산 마그네슘 2 mgMagnesium stearate 2 mg
상기의 성분들을 혼합한 후 통상의 정제의 제조방법에 따라서 타정하여 정제를 제조한다.After mixing the above ingredients, tablets are prepared by tableting according to a conventional tablet manufacturing method.
제제예 3: 캡슐제의 제조Formulation Example 3: Preparation of capsule formulation
실시예 4의 추출물 분말 10 mg10 mg of extract powder of Example 4
결정성 셀룰로오스 3 mg3 mg of crystalline cellulose
락토오스 14.8 mgLactose 14.8 mg
마그네슘 스테아레이트 0.2 mgMagnesium stearate 0.2 mg
통상의 캡슐제 제조방법에 따라 상기의 성분을 혼합하고 젤라틴 캡슐에 충전하여 캡슐제를 제조한다.Capsules are prepared by mixing the above ingredients and filling them into gelatin capsules according to a conventional capsule preparation method.
제제예 4: 과립제의 제조Formulation Example 4: Preparation of granules
실시예 4의 추출물 분말 1,000 mg1,000 mg of extract powder of Example 4
비타민 혼합물 적량Appropriate amount of vitamin mixture
비타민 A 아세테이트 70 ㎍Vitamin A Acetate 70 μg
비타민 E 1.0 mgVitamin E 1.0 mg
비타민 B1 0.13 mgVitamin B1 0.13 mg
비타민 B2 0.15 mgVitamin B2 0.15 mg
비타민 B6 0.5 mgVitamin B6 0.5 mg
비타민 B12 0.2 ㎍Vitamin B12 0.2 μg
비타민 C 10 mg
비오틴 10 ㎍10 μg of biotin
니코틴산아미드 1.7 mgNicotinamide 1.7 mg
엽산 50 ㎍
판토텐산 칼슘 0.5 mgCalcium Pantothenate 0.5 mg
무기질 혼합물 적량Appropriate amount of mineral mixture
황산제1철 1.75 mgFerrous sulfate 1.75 mg
산화아연 0.82 mgZinc Oxide 0.82 mg
탄산마그네슘 25.3 mgMagnesium Carbonate 25.3 mg
제1인산칼륨 15 mg
제2인산칼슘 55 mg
구연산칼륨 90 mg
탄산칼슘 100 mg
염화마그네슘 24.8 mgMagnesium Chloride 24.8 mg
상기의 비타민 및 미네랄 혼합물의 조성비는 건강기능식품에 적합한 성분을 바람직한 실시예로 혼합 조성하였지만, 그 배합비를 임의로 변형 실시하여도 무방하며, 통상의 건강기능식품 제조방법에 따라 상기의 성분을 혼합한 다음, 과립을 제조하고, 통상의 방법에 따라 건강기능식품 조성물 제조에 사용할 수 있다.The composition ratio of the above vitamin and mineral mixture is a mixture of ingredients suitable for health functional food in a preferred embodiment, but the mixing ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional health functional food manufacturing method. Next, granules can be prepared and used in the preparation of health functional food compositions according to conventional methods.
제제예 5: 음료 제형의 제조Formulation Example 5: Preparation of beverage formulation
실시예 4의 추출물 분말 1,000 mg1,000 mg of extract powder of Example 4
구연산 1,000 mgCitric Acid 1,000 mg
올리고당 100 g100 g of oligosaccharides
매실농축액 2 g2 g plum concentrate
타우린 1 g1 g of taurine
정제수를 가하여 전체 900 mLAdd purified water to total 900 mL
통상의 음료 제조방법에 따라 상기의 성분을 혼합한 다음, 약 1 시간 동안 85 ℃에서 교반 가열한 후, 만들어진 용액을 여과하여 멸균된 2 L 용기에 취득하여 밀봉 멸균한 뒤 냉장 보관한 다음 본 발명의 기능성 음료 조성물 제조에 사용한다.After mixing the above ingredients according to the usual beverage manufacturing method, stirring and heating at 85 ° C. for about 1 hour, the resulting solution is filtered and collected in a sterilized 2 L container, sealed and sterilized, and then refrigerated according to the present invention. It is used for the preparation of functional beverage compositions.
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