KR101898508B1 - Food composition for improvement of asthma and pharmaceutical composition for treatment or prevention of asthma with eletric stimulation yeast extract - Google Patents
Food composition for improvement of asthma and pharmaceutical composition for treatment or prevention of asthma with eletric stimulation yeast extract Download PDFInfo
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- KR101898508B1 KR101898508B1 KR1020170056246A KR20170056246A KR101898508B1 KR 101898508 B1 KR101898508 B1 KR 101898508B1 KR 1020170056246 A KR1020170056246 A KR 1020170056246A KR 20170056246 A KR20170056246 A KR 20170056246A KR 101898508 B1 KR101898508 B1 KR 101898508B1
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- yeast
- asthma
- extract
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- electric stimulation
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Abstract
Description
본 발명은 효모 전기 자극 추출물을 포함하는 천식 개선용 식품 조성물 또는 천식 치료 또는 예방용 약학 조성물에 관한 것으로써, 더욱 상세하게는 효모에 전기 자극을 가한 후, 효모로부터 추출한 효모 전기 자극 추출물을 포함하는 것을 특징으로 하는 천식 개선용 식품 조성물 또는 천식 치료 또는 예방용 약학 조성물에 관한 것이다. The present invention relates to a food composition for improving asthma or a composition for preventing or treating asthma, which comprises a yeast electric stimulation extract, and more particularly to a composition for preventing or treating asthma, which comprises an electric stimulation on yeast, And a pharmaceutical composition for treating or preventing asthma.
기관지 천식은 한번 이환되면 일평생 치료를 지속해야 하는 만성질환으로 국민 보건상 부담이 큰 질환이다. 특히 고혈압, 당뇨 등의 다른 만성질환은 중년 이상의 성인에서 발생하는 것과 달리 기관지 천식은 소아부터 노년층에 이르기까지 전 연령층에서 일어나는 질환이다. Bronchial asthma is a chronic disease that should be treated for a lifetime of life once it is taken, which is a burden on public health. Unlike other chronic diseases such as hypertension and diabetes that occur in middle-aged adults, bronchial asthma is a disease that occurs in all age groups, from children to the elderly.
급속한 산업화로 인한 생활환경의 변화 등으로 인해 알레르기성 천식의 유병률이 지속적으로 증가하고, 2조원에 육박하는 막대한 사회적 비용이 소요되는 것으로 추정되고 있다. 또한, 온난화로 인한 지구의 온도, 이산화탄소 수치가 상승함에 따라 인간에게 알레르기를 일으킬 수 있는 식물이 다양해져 꽃가루 알레르기와 천식에 걸릴 위험도 높아지는 것으로 보고되고 있다. It is estimated that the prevalence of allergic asthma continues to increase due to the rapid changes in the living environment caused by industrialization, and the enormous social cost of close to 2 trillion won is required. In addition, as the global temperature and carbon dioxide levels rise due to global warming, it has been reported that the risk of pollen allergy and asthma is increasing with the diversity of plants that can cause allergies to humans.
알레르기성 천식은 병의 원인이 확실치 않아 환경요법, 기관지 확장제와 알레르기 염증을 치료하는 항염증제제를 이용한 약물요법, 알레르기 체질을 개선하는 면역요법 등이 시도되고 있으나, 만성적이고 재발성이어서 치료가 쉽지 않은 실정이다. Allergic asthma is not known to cause the disease, but it is tried to use environmental therapy, bronchodilator and anti-inflammatory drug to treat allergic inflammation, and immunotherapy to improve allergic constitution, but it is chronic, It is true.
알레르기성 천식은 알레르겐에 의하여 기관지가 과민하게 반응함으로써 기관지 수축, 점액 과분비 등에 의해 호흡곤란이 생기는 질환이다. 이 중 점액의 과분비 증상은 기도에 집먼지 진드기나 꽃가루와 같은 항원에 의해 발생한다. 항원이 기도에 침투하면 채내 면역계가 이를 감지하여 항체 IgE를 생성하게 된다. Allergic asthma is a disease in which respiratory distress occurs due to bronchoconstriction and mucus hypersecretion due to sensitization of bronchial tubes by allergens. Mucus hypersecretion is caused by antigens such as house dust mites and pollen in the airway. When the antigen enters the airway, the immune system senses it and produces antibody IgE.
IgE는 기관지 상 비만 세포의 수용체로 작용하며, 추후 항원의 재침투시, 비만 세포가 항원에 반응하여 히스타민과 같은 기도의 염증 반응을 일으키는 여러 물질을 분비하게 한다. 이러한 염증성 물질들로 인해 점막에 염증이 생겨 과다한 점액 분비로, 기도 저항이 증가하며, 폐의 과 팽창 등 호흡곤란을 일으키게 된다.IgE acts as a receptor for bronchial mast cells, and upon subsequent re-infiltration of the antigen, mast cells respond to the antigen and secrete many substances that cause airway inflammation like histamine. These inflammatory substances cause inflammation in the mucous membranes, leading to excessive mucus secretion, increased airway resistance, and breathing difficulties, such as swelling of the lungs.
알레르기성 천식이 유발되면, 정상적인 기도 세포들인 섬유모세포(fibroblast), 근섬유모세포(myofibroblast), 상피세포, 평활근세포 등은 만성 기도 염증과 연관된 사이토카인과 성장인자들을 분비한다. When allergic asthma is induced, normal airways cells fibroblast, myofibroblast, epithelial cells and smooth muscle cells secrete cytokines and growth factors associated with chronic airway inflammation.
상피세포는 호산구성 염증을 유지시키고, 비만세포 성장 인자를 분비시키는 사이토카인과 케모카인(GM-CSF, eotaxin-1, RANTES 등)을 분비한다. 이에 대한 기전은 아직 명확하진 않으나, 알레르기성 천식 환자에게서 많이 발생하고 있다. 또한, 천식 환자들 호흡기 점액에서 주로 발견되는 MUC5AC의 분비형 점소의 과다 생성은 천식, 만성 기관지염, 폐기종, 기관지 확장증, 낭포성 섬유증 등의 기도 질환을 악화시키는 주된 요인으로 알려져 있다.Epithelial cells maintain eosinophilic inflammation and secrete cytokines and chemokines (GM-CSF, eotaxin-1, RANTES, etc.) that secrete mast cell growth factors. The mechanism for this is not clear yet, but it is common in patients with allergic asthma. In addition, excessive production of secretory form of MUC5AC, which is mainly found in respiratory mucus in asthmatic patients, is known to be a major factor to exacerbate airway diseases such as asthma, chronic bronchitis, emphysema, bronchiectasis and cystic fibrosis.
본 발명은 알레르기성 천식으로 발생된 염증성 케모카인인 Eotaxin-1과 호흡기 점액에서 주로 발견되는 MUC5AC의 분비형 점소의 발현을 억제하는 성분을 개발함으로써, 알레르기성 천식의 개선 및 예방 또는 치료를 위한 조성물을 개발하여 제공하고자 한다.The present invention relates to a composition for improving or preventing allergic asthma or for preventing or treating allergic asthma by developing a component that inhibits the expression of Eotaxin-1, an inflammatory chemokine generated by allergic asthma, and MUC5AC secretory form, which is mainly found in respiratory mucus And to provide them.
본 발명은 효모에 전기 자극을 가한 후, 효모로부터 추출한 효모 전기 자극 추출물을 포함하는 것을 특징으로 천식 개선용 식품 조성물을 제공한다. The present invention provides a food composition for improving asthma, which comprises an electric stimulation extract of yeast extracted from yeast after electric stimulation is applied to the yeast.
본 발명의 천식 개선용 식품 조성물에 있어서, 상기 효모 전기 자극 추출물은, 바람직하게 효모로부터 추출하고 난 다음, 효모를 제거하고, MWCO 5~10 kDa 한외 여과막을 이용하여 여과하여 여과액을 제거한 것일 수 있다. In the food composition for improving asthma according to the present invention, the yeast electric stimulation extract is preferably extracted from yeast, and then the yeast is removed and filtered using an MWCO 5-10 kDa ultrafiltration membrane to remove the filtrate have.
본 발명의 천식 개선용 식품 조성물에 있어서, 상기 효모는, 바람직하게 효모 배양액일 수 있다. In the food composition for improving asthma of the present invention, the yeast may preferably be a yeast culture broth.
본 발명의 천식 개선용 식품 조성물에 있어서, 상기 전기 자극은, 바람직하게 0.001~20V의 전기로 0.001~10시간 동안 자극을 가하는 것이 좋다. In the asthma-improving food composition of the present invention, it is preferable that the electric stimulation is applied to the stimulation for 0.001 to 10 hours in an electric furnace preferably at 0.001 to 20 V.
본 발명의 천식 개선용 식품 조성물에 있어서, 상기 효모로부터 추출시, 바람직하게 효모의 농도가 1~10%(w/v) 되도록 물을 가하고 교반 추출하는 것이 좋다. In the food composition for improving asthma of the present invention, water is preferably added to the yeast so that the yeast concentration is 1 to 10% (w / v), and the mixture is stirred and extracted.
한편, 본 발명은 효모에 전기 자극을 가한 후, 효모로부터 추출한 효모 전기 자극 추출물을 포함하는 것을 특징으로 천식 치료 또는 예방용 약품 조성물을 제공한다. Meanwhile, the present invention provides a pharmaceutical composition for treating or preventing asthma, which comprises an electric stimulation extract of yeast extracted from yeast after electric stimulation is applied to the yeast.
본 발명의 천식 치료 또는 예방용 약품 조성물에 있어서, 상기 효모 전기 자극 추출물은, 바람직하게 효모로부터 추출하고 난 다음, 효모를 제거하고, MWCO 5~10 kDa 한외 여과막을 이용하여 여과하여 여과액을 제거한 것일 수 있다. In the pharmaceutical composition for treating or preventing asthma according to the present invention, the yeast electric stimulation extract is preferably extracted from yeast, and then the yeast is removed, filtered using an MWCO 5-10 kDa ultrafiltration membrane to remove the filtrate Lt; / RTI >
본 발명의 천식 치료 또는 예방용 약품 조성물에 있어서, 상기 효모는, 바람직하게 효모 배양액일 수 있다. In the pharmaceutical composition for treating or preventing asthma according to the present invention, the yeast may preferably be a yeast culture broth.
본 발명의 천식 치료 또는 예방용 약품 조성물에 있어서, 상기 전기 자극은, 바람직하게 0.001~20V의 전기로 0.001~10시간 동안 자극을 가하는 것이 좋다. In the pharmaceutical composition for treating or preventing asthma according to the present invention, it is preferable that the electric stimulation is applied to the electric stimulation for 0.001 to 10 hours in an electric furnace preferably at 0.001 to 20V.
본 발명의 천식 치료 또는 예방용 약품 조성물에 있어서, 상기 효모로부터 추출시, 바람직하게 효모의 농도가 1~10%(w/v) 되도록 물을 가하고 교반 추출하는 것이 좋다. In the pharmaceutical composition for treating or preventing asthma according to the present invention, water is preferably added to the yeast so that the yeast concentration is 1 to 10% (w / v), and the mixture is stirred and extracted.
본 발명의 방법에 의하여 제조한 전기 자극 효모추출물은 인체에 사용시 부작용이 없고, 알레르기성 천식의 예방, 치료 및 완화에 효과를 발휘함이 확인되었다. 더욱 구체적으로, 본 발명의 전기 자극 효모추출물은 알레르기성 천식으로 발생하는 염증성 케모카인 Eotaxin-1의 발현과 점액 분비형 점소인 MUC5AC의 발현을 억제하는 효과가 있는 것으로 확인되었다. 따라서, 본 발명의 전기 자극 효모추출물은 LPS로 유발된 알레르기성 천식의 염증반응의 예방과 치료용 조성물, 화장품 소재 및 기능성 식품의 신소재로 활용될 수 있다.It has been confirmed that the electric stimulated yeast extract prepared by the method of the present invention has no side effects when used in the human body, and is effective in preventing, treating and alleviating allergic asthma. More specifically, it was confirmed that the electric stimulating yeast extract of the present invention has an effect of inhibiting the expression of the inflammatory chemokine Eotaxin-1, which is caused by allergic asthma, and the expression of MUC5AC, which is a mucus-secreting type point. Therefore, the electric stimulating yeast extract of the present invention can be utilized as a composition, a cosmetic material and a functional food new material for prevention and treatment of an inflammatory reaction of allergic asthma induced by LPS.
도 1은 본 발명 전기 자극 효모추출물의 추출 방식에 따른 세포독성효과에 대한 결과 (도 1의 a는 전기 자극 효모추출물 단독 처리, 도 1의 b는 LPS와 함께 처리)와 Eotaxin-1, ICAM-1, MUC5AC의 발현 양상(도 1의 c)을 보여주고 있다.
도 2는 본 발명 전기 자극 효모추출물의 추출온도에 따른 세포독성효과에 대한 결과를 보여주고 있다.
도 3은 본 발명 전기 자극 효모추출물의 추출온도에 따른 티오레독신의 함유량을 보여주고 있다.
도 4는 본 발명 전기 자극 효모추출물의 추출온도에 따른 Eotaxin-1의 발현량을 보여주고 있다.Fig. 1 shows the results of cytotoxic effect according to the extraction method of the electric stimulation yeast extract according to the present invention (Fig. 1 (a) shows electric stimulation yeast extract alone treatment, Fig. 1 (b) shows treatment with LPS) and Eotaxin- 1, and MUC5AC (Fig. 1 (c)).
FIG. 2 shows the results of the cytotoxic effect of the electric stimulating yeast extract according to the present invention on the extraction temperature.
3 shows the content of thioredoxin according to the extraction temperature of the electric stimulation yeast extract of the present invention.
FIG. 4 shows the expression level of Eotaxin-1 according to the extraction temperature of the electric stimulating yeast extract of the present invention.
본 발명에서는 효모에 외부 자극을 가하여 효모가 이 자극에 대한 반응으로써 티오레독신을 합성 및 분비하게 하는 과정에 의하여 티오레독신의 함량을 현저하게 증가시킨 효모 전기 자극 추출물을 제공한다. The present invention provides a yeast electric stimulation extract in which the content of thioredoxin is remarkably increased by applying an external stimulus to the yeast and causing yeast to synthesize and secrete thioredoxin as a reaction to the stimulation.
효모에 가해지는 외부 자극으로는 전기, 압력, rpm, 초음파, 온도 등 다양한 요인이 있는데, 본 발명에서는 전기를 주요 인자로 적용하여 티오레독신의 함량을 증가시킨 효모추출물을 완성하였다. 효모에 가하여 주는 전기는 전압을 기준으로 효모슬러리에 0.001~20V 이내로, 시간은 0.001~10시간 동안 적용하였는데, 이와 같은 조건에서 천식에 우수한 효과를 나타내는 효모추출물을 수득할 수 있었다. 다만, 추출 조건 중 온도는 일반적인 범위에서 거의 영향을 주지 않는 것으로 확인되었다. As external stimuli applied to yeast, there are various factors such as electricity, pressure, rpm, ultrasonic wave and temperature. In the present invention, yeast extract having increased content of thioredoxin by applying electricity as a main factor was completed. The electricity applied to the yeast was applied to the yeast slurry within the range of 0.001 to 20 V and the time was 0.001 to 10 hours based on the voltage. Under these conditions, a yeast extract showing excellent effect on asthma could be obtained. However, it was confirmed that the temperature of extracting conditions had little effect on the general range.
본 발명의 전기 자극 효모 추출물은, 전기 자극을 가한 후 물을 추가로 가하여 효모의 농도가 1~10%(w/v)가 되도록 한 다음, 교반기로 교반하면서 추출하고, 이를 원심분리하여 효모를 제거하여 1차 추출액을 얻고, 분자량 크기(MWCO) 5 kDa 또는 10 kDa을 걸러내는 한외여과막을 이용하여 사용하여 부피를 1/10으로 줄였다. 한외여과막을 통과한 액은 제거하고 한외여과 후 남은 농축액을 회수하여 본 발명의 효모추출물로 제조하기 위하여 동결건조하고, 최종적으로 옅은 황색의 분말을 얻었다. 최종적으로 얻어지는 수율은 원료인 건조효모분말을 기준으로 1~5%(w/w)의 범위였다. The electric stimulation yeast extract of the present invention is prepared by adding electric water after addition of electric stimulus so that the yeast concentration is 1 to 10% (w / v), and then the mixture is stirred with a stirrer and centrifuged to remove yeast To obtain a primary extract, and the volume was reduced to 1/10 by using an ultrafiltration membrane having a molecular weight (MWCO) of 5 kDa or 10 kDa. The solution passed through the ultrafiltration membrane was removed, and the concentrated solution remaining after the ultrafiltration was recovered and lyophilized to prepare the yeast extract of the present invention, and finally a pale yellow powder was obtained. The final yield was in the range of 1 to 5% (w / w) based on dry yeast powder.
이상의 과정을 정리한, 본 발명의 효모 전기 자극 추출물을 수득하기 위한 바람직한 일 실시예는, 효모에 0.001~20V의 전기로 0.001~10시간 동안 전기 자극을 가하는 단계 (a); 상기 단계 (a) 후 물을 추가로 가하여 효모의 농도가 1~10 (w/v)이 되도록 조정한 다음, 교반하면서 추출하는 단계 (b); 상기 단계 (b) 후, 효모를 제거하고, MWCO 5~10 kDa 한외 여과막을 이용하여 여과액을 제거함으로써 추출물을 농축하는 단계 (c); 를 포함하는 과정이다. A preferred embodiment for obtaining the yeast electric stimulation extract according to the present invention comprises: (a) applying electric stimulation to the yeast for 0.001 to 10 hours in an electric furnace at 0.001 to 20 V; (B) adding water after the step (a) to adjust the yeast concentration to 1 to 10 (w / v) and then extracting with stirring; (C) removing the yeast after the step (b) and concentrating the extract by removing the filtrate using an ultrafiltration membrane of 5 to 10 kDa MWCO; .
이와 같은 과정을 통하여 농축된 본 발명의 효모추출물에 함유된 티오레독신은 분자량을 기준으로 SDS-PAGE를 이용하여 정성적인 분리를 통하여 확인하였고, 이를 다시 ELISA를 통하여 정량하였다. 최종적으로 함유된 티오레독신의 함량은 0.01~3%(w/w)의 범위였고, 이와 같은 수치는 일반적인 효모추출물에서 불검출인 것과 비교하면 현저하게 높은 함량임을 알 수 있었다. Thioredoxin contained in the concentrated yeast extract of the present invention was identified through qualitative separation using SDS-PAGE on the basis of molecular weight, and quantified by ELISA. The content of thioredoxin finally contained was in the range of 0.01 to 3% (w / w), and this value was remarkably higher than that of the non-detected yeast extract.
한편, 본 발명의 조성물에 있어서, 기관지 알러지에 유효한 효과를 얻기 위해서는 상기 전기 자극 효모 추출물의 농도가 10~50 μg/ml인 것이 좋으며, 더욱 바람직하게는 25~50 μg/ml인 것이 좋다. 상기 범위를 만족할 시, 기도 상피세포에서 Eotaxin-1, MUC5AC의 발현을 억제할 수 있어, 알레르기성 천식의 개선에 현저히 우수한 효과를 발휘할 수 있기 때문이다.Meanwhile, in the composition of the present invention, the concentration of the electric stimulating yeast extract is preferably 10 to 50 μg / ml, more preferably 25 to 50 μg / ml in order to obtain an effective effect on bronchial allergy. When the above range is satisfied, the expression of Eotaxin-1 and MUC5AC in airway epithelial cells can be inhibited and a remarkably excellent effect can be exerted in the improvement of allergic asthma.
본 발명의 천식 개선용 식품 조성물은 일 예로, 드링크류, 카페인 음료, 일반음료, 초콜렛, 빵류, 스낵류, 과자류, 사탕, 피자, 젤리, 면류, 껌류, 유제품류, 아이스크림류, 알코올성 음료, 술, 비타민 복합체 및 그 밖의 건강 보조식품류 중 선택되는 어느 하나일 수 있으나, 이에 한정되는 것은 아니다.The food composition for improving asthma according to the present invention may be in the form of a drink, a caffeinated beverage, a general beverage, a chocolate, a bread, a snack, a confectionery, a candy, a pizza, a jelly, a noodle, a gum, a milk product, an ice cream, Vitamin complexes, and other health supplement foods. However, the present invention is not limited thereto.
한편, 본 발명의 천식 치료 또는 예방용 약학 조성물은 상기 본 발명의 전기 자극 효모추출물 외에 약제학적으로 허용 가능한 담체, 희석제 또는 부형제를 더욱 포함할 수 있다. 사용가능한 담체, 부형제 또는 희석제로는, 락토즈, 텍스트로즈, 수크로즈, 솔비톨, 만니톨, 자이리톨, 에리스리톨, 말티톨, 전분, 아카시아 고무, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀로로즈, 메틸 셀로로즈, 미정질 셀롤로즈, 폴리비닐피롤리돈, 물, 메틸하이드록시벤조에이트, 프로필하이드록시벤조에이트, 탈크, 마그네슘 스테아레이트 및 광물유가 있으며, 이 중 선택되는 하나 이상을 사용할 수 있다.In addition, the pharmaceutical composition for treating or preventing asthma of the present invention may further comprise a pharmaceutically acceptable carrier, diluent or excipient in addition to the electric stimulating yeast extract of the present invention. Examples of usable carriers, excipients or diluents include lactose, textol, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, , Microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and one or more selected from these may be used.
한편, 본 발명의 천식 치료 또는 예방용 약학 조성물의 제형은 사용방법에 따라 바람직한 형태일 수 있으며, 특히 포유동물에 투여된 후 활성 성분의 신속, 지속 또는 지연된 방출을 제공할 수 있도록 당업계에 공지된 방법을 채택하여 제형화 하는 것이 좋다. 구체적인 제형의 예로는 경고제(PLASTERS), 과립제(GRANULES), 로션제(LOTIONS), 리니멘트제(LINIMENTS), 리모나데제(LEMONADES), 산제(POWDERS), 시럽제(SYRUPS), 액제(LIQUIDS AND SOLUTIONS), 에어로솔제(AEROSOLS), 엑스제(EXTRACTS), 엘릭실제(ELIXIRS), 연고제(OINTMENTS), 유동엑스제(FLUIDEXTRACTS), 유제(EMULSIONS), 현탁제(SUSPENSIONS), 전제(DECOCTIONS), 정제(TABLETS), 좌제(SUPPOSITORIES), 주사제(INJECTIONS), 주정제(SPIRITS), 카타플라스마제(CATAPLSMA), 캅셀제(CAPSULES), 크림제(CREAMS), 트로키제(TROCHES), 틴크제(TINCTURES), 파스타제(PASTES), 환제(PILLS), 연질 또는 경질 젤라틴 캅셀 중 선택되는 어느 하나일 수 있다.Meanwhile, the pharmaceutical composition for the treatment or prevention of asthma of the present invention may be in a form suitable for the method of use, and may be in a form suitable for oral, parenteral, It is recommended to adopt the method of formulation. Examples of specific formulations include, but are not limited to, PLASTERS, GRANULES, LOTIONS, LINIMENTS, LEMONADES, POWDERS, SYRUPS, LIQUIDS AND SOLUTIONS, AEROSOLS, EXTRACTS, ELIXIRS, OINTMENTS, FLUIDEXTRACTS, EMULSIONS, SUSPENSIONS, DECOCTIONS, (TABLETS), SUPPOSITORIES, INJECTIONS, SPIRITS, CATAPLSMA, CAPSULES, CREAMS, TROCHES, TINCTURES, Pastes (PASTES), pills (PILLS), soft or hard gelatin capsules.
한편, 본 발명의 천식 치료 또는 예방용 약학 조성물의 투여량은 투여방법, 복용자의 연령, 성별, 체중, 및 질환의 중증도 등을 고려하여 결정하는 것이 좋다. 일 예로, 본 발명의 천식 치료 또는 예방용 약학 조성물은 유효성분을 기준으로 하였을 때, 1일 0.00001 내지 100 ㎎/㎏(체중)으로 1회 이상 투여 가능하다. 그러나 상기의 투여량은 예시하기 위한 일 예에 불과하며, 복용자의 상태에 따라 의사의 처방에 의해 변화될 수 있다.On the other hand, the dosage of the pharmaceutical composition for treating or preventing asthma of the present invention is determined in consideration of the administration method, the age, sex, weight, and severity of the disease. For example, the pharmaceutical composition for treating or preventing asthma according to the present invention may be administered at least once per day at a dose of 0.00001 to 100 mg / kg (body weight) based on the active ingredient. However, the dosage is only an example and may be changed by a doctor's prescription depending on the condition of the recipient.
본 발명은 전기 자극 효모추출물을 함유하는 조성물은 알레르기성 천식을 예방, 치료 및 개선시킬 수 있다. 또한, 본 발명의 전기 자극 효모추출물은 알레르기성 천식으로 발생하는 염증을 억제하여 알레르기성 천식의 예방과 치료에 기능성 신소재로 활용될 수 있다. The present invention can prevent, treat and ameliorate allergic asthma in a composition containing an electric stimulating yeast extract. In addition, the electric stimulating yeast extract of the present invention can be used as a functional new material for prevention and treatment of allergic asthma by inhibiting inflammation caused by allergic asthma.
본 발명의 전기 자극 효모추출물은 알레르기 천식에 의해 발현되는 염증성 케모카인인 Eotaxin-1을 억제하며, 점액 점소 분비형 단백질인 MUC5AC 발현 억제 효과를 보인다. 더욱 구체적으로 본 발명의 전기 자극 효모추출물을 UF 10K을 사용하여 저분자 물질들을 제거하고 60℃로 추출한 결과, 세포 독성을 보이지 않을뿐더러, Eotaxin-1의 발현을 억제함이 확인되었다.The electric stimulating yeast extract of the present invention inhibits Eotaxin-1, an inflammatory chemokine expressed by allergic asthma, and exhibits an effect of inhibiting MUC5AC expression, a mucin-releasing protein. More specifically, it was confirmed that the electric stimulation yeast extract of the present invention was not cytotoxic and inhibited the expression of Eotaxin-1 when
[[ 실시예Example 1: 효모에 외부 자극으로 전기 충격을 가하여 효모 내 1: Electric shock is applied to the yeast by external stimulation, 티오레독신Thioredoxin 함량 증진] Increased content]
본 실험에서는 효모 (Saccharomyces cerevisiae)에 외부 자극을 가하여 효모가 이 자극에 반응함으로써 티오레독신을 합성하여 분비하는 과정을 탐구함으로써, 티오레독신의 함량을 현저하게 증가시킨 효모 추출물을 개발하고자 하였다. In this experiment, yeast ( Saccharomyces To investigate the process of synthesizing and secretion of thioredoxin by applying an external stimulus to the cerevisiae and reacting the yeast with the stimulus, the inventors sought to develop a yeast extract that significantly increases thioredoxin content.
효모에 다양한 외부 자극을 가할 수 있는데, 본 실시예에서는 외부 자극으로 전기를 가하여 티오레독신의 함량이 증가하는지 여부를 확인하고자 하였다. Various external stimuli can be applied to the yeast. In this embodiment, it was tried to determine whether the content of thioredoxin is increased by applying electricity to an external stimulus.
효모에 가하여 주는 전기는 전압을 기준으로 효모슬러리(yeat slurry, 효모를 배양한 배양액)에 10V로 시간은 5시간 동안 적용하였고, 온도는 40℃ 또는 60℃에서 수행하였다. The electricity applied to the yeast was applied to yeast slurry (yeast slurry, yeast culture) at 10 V for 5 hours, and the temperature was 40 ° C or 60 ° C.
한편, 효모 추출물은 하기의 과정을 통해 수득하였다. 우선, 상기의 전기 자극 후, 물을 추가로 가하여 효모의 농도가 5%(w/v)가 되도록 한 다음, 교반기로 교반하면서 추출하였다. 그 후, 원심분리하여 효모를 제거하여 1차 추출액을 얻고, 이를 한외여과막을 이용하여 분자량 크기(MWCO) 5 kDa (이하, 'UF-5K'라 함) 또는 10 kDa (이하, 'UF-10K'라 함)을 사용하여, 부피를 1/10으로 줄였다. 이후, 한외여과막을 통과한 액은 제거하고, 한외여과 후 남은 농축액을 회수하여, 동결건조하고 최종적으로 옅은 황색의 분말을 얻었다. 최종적으로 얻어지는 수율은 원료인 건조효모분말을 기준으로 1~5%(w/w)의 범위였다. On the other hand, the yeast extract was obtained through the following procedure. After the electric stimulation, water was further added to make the yeast concentration 5% (w / v), and the mixture was extracted with a stirrer while stirring. Thereafter, the yeast was removed by centrifugation to obtain a primary extract, which was then subjected to ultrafiltration using a molecular weight MWCO of 5 kDa (hereinafter referred to as' UF-5K ') or 10 kDa (hereinafter referred to as' UF-10K Quot;), the volume was reduced to 1/10. Thereafter, the liquid passing through the ultrafiltration membrane was removed, the ultrafiltration liquid remaining after the ultrafiltration was recovered, and lyophilized to finally obtain a pale yellow powder. The final yield was in the range of 1 to 5% (w / w) based on dry yeast powder.
[[ 실험예Experimental Example 1: 본 발명 전기 자극 효모추출물에 함유된 1: Electrically stimulated yeast extract of the present invention 티오레독신Thioredoxin 함량 측정] Content measurement]
상기 실시예 1에서 수득한 본 발명의 전기 자극 효모추출물에 함유된 티오레독신은 분자량을 기준으로 SDS-PAGE를 이용한 정성적인 분리를 통하여 확인하였고, 이를 다시 ELISA를 통하여 정량하였다. Thioredoxin contained in the electric stimulating yeast extract of the present invention obtained in Example 1 was confirmed by qualitative separation using SDS-PAGE on the basis of molecular weight, and quantified by ELISA again.
전기 자극 효모추출물에 최종적으로 함유된 티오레독신의 함량은 0.01~3%(w/w)의 범위였고, 이는 일반적인 효모추출물에서 불검출인 것과 비교하면, 현저하게 높은 함량임을 확인할 수 있었다. The content of thioredoxin finally contained in the electric stimulating yeast extract was in the range of 0.01 to 3% (w / w), which was significantly higher than that of the non-detected yeast extract.
[[ 실험예Experimental Example 2: 본 발명 전기 자극 효모추출물의 천식 효과 확인] 2: Identification of asthmatic effect of the electric stimulation yeast extract of the present invention]
1. 실험 방법 1. Experimental Method
(1) (One) 기도상피세포Airway epithelial cells 배양실험 Culture experiment
기도상피세포는, 10% FBS(fetal bovin serum), L-글루타민 2 mmol/L, 페니실린 100 KU/L, 인슐린 2.5 mg/L가 보충된 스트렙토마이신 100 mg/L, 하이드로코르티손 0.361 mg/L, 아포트랜스페린 2.5 mg/L와 상피성장인자 20 μg/L가 포함된 25 mmol/L의 HEPES-buffered M199 배지에서 배양되었다. BEAS-2B 세포는 37℃ 5% CO2 인큐베이터에서 배양하였으며, 48시간 주기로 배양액을 교체하였다.Airway epithelial cells were treated with 100 mg / L streptomycin supplemented with 10% FBS (fetal bovine serum), 2 mmol / L L-glutamine, 100 KU / L penicillin, 2.5 mg / L insulin, 0.361 mg / L hydrocortisone, Were cultured in 25 mmol / L HEPES-buffered M199 medium containing 2.5 mg / L apotransferrin and 20 μg / L epithelial growth factor. BEAS-2B cells were cultured in a 37 ° C 5% CO 2 incubator and the culture medium was changed at 48-hour intervals.
(2) (2) 세포생존률Cell survival rate 조사 Research
기도상피세포를 LPS와의 세포배양실험을 거친 후, 세포 생존률을 측정하기 위하여 자동 마이크로플레이트 리더 스펙트로포토미터 (microplate reader spectrophotometer)를 이용한 MTT 분석이 이루어졌다. 이 분석법은 테트라즐리움(tetrazlium) MTT 염 (3-(4,5-dimethylthiazol-2-y1)-2,5- di-phenyltetrazolium bromide)이 미토콘드리아 효소인 숙시네이트 데하이드로지나아제 (succinate dehydrogenase)에 의하여 불용성의 포르마잔(formazan)으로 변하는 과정을 이용함으로써 측정하였다. After airway epithelial cells were subjected to cell culture with LPS, MTT analysis was performed using an automatic microplate reader spectrophotometer to measure cell viability. This assay is based on the observation that the tetrazlium MTT salt (3- (4,5-dimethylthiazol-2-y1) -2,5-di-phenyltetrazolium bromide) is a mitochondrial enzyme, succinate dehydrogenase By using a process that changes to insoluble formazan.
기도상피세포를 전기 자극 효모추출물(CPI) 및 LPS로 배양실험을 거친 후 세척한 다음, MTT(5mg/ml stock solution)가 첨가된 페놀 레드 (phenol red)가 들어있지 않는 배지에서 3시간 배양시켰다. 그 후에, 이소프로필 알코올 (isopropyl alcohol)을 가하여 조심스럽게 흔들어 주면서 형성된 포르마잔(formazan)을 용해시키고 λ=570nm에서 흡광도를 측정하였다.The airway epithelial cells were cultured with electrical stimulation yeast extract (CPI) and LPS, washed and then cultured in a medium containing no phenol red added with MTT (5 mg / ml stock solution) for 3 hours . Thereafter, the formed formazan was dissolved while isopropyl alcohol was added and carefully shaken, and the absorbance was measured at? = 570 nm.
(3) (3) 웨스턴블랏팅Western Blotting
기도상피세포로부터 10% SDS, 1% 글리세로포스페이트(glycerophosphate), 0.1 mol/L Na3VO4, 0.5 mol/L NaF와 프로테아제 인히비터 칵테일 (protease inhibitor cocktail)이 포함된 1 mol/L, Tris-HCl (pH 6.8) 라이시스 버퍼 (lysis buffer)를 이용하여 세포용해물을 얻은 후, 로우리 어세이 (Lowry assay) 방법을 이용하여 단백질 함량을 정량하였다. L of Tris (1 mol / L) containing 10% SDS, 1% glycerophosphate, 0.1 mol / L Na 3 VO 4 , 0.5 mol / L NaF and protease inhibitor cocktail The cell lysate was obtained using lysis buffer of HCl (pH 6.8), and the protein content was quantitated using the Lowry assay method.
세포용해액은 5X dye와 혼합하여 95℃에서 10분간 끓인 후 8~15% SDS-폴리아크릴아미드 겔에 전기영동하여 단백질을 분리하였다. 전기영동이 끝난 후 니트로셀룰로오스 멤브레인 (nitrocellulose membrane)에 전이시켰다.The cell lysate was mixed with 5X dye, boiled at 95 ° C for 10 minutes, and then separated by electrophoresis on 8-15% SDS-polyacrylamide gel. After electrophoresis, the cells were transferred to a nitrocellulose membrane.
멤브레인은 5% 탈지 건조 우유 (nonfat dry milk)와 보바인 시럼 알부민 (bovine serum albumin)에서 3시간 블록킹(blocking)하고, 특정 1차 항체를 붙인 후 4℃에서 오버나이트하였다. TBS-T로 세척한 후 이차항체 anti-goat, anti-mouse IgG conjugated horse-radish peroxidase와 1시간 반응시킨 후, ECL 용액을 이용하여 X-ray 필름으로 밴드 사진을 촬영하였다.Membranes were blocked for 3 h in 5% nonfat dry milk and bovine serum albumin, and then incubated at 4 ° C with a specific primary antibody attached. After washing with TBS-T, secondary antibody anti-goat and anti-mouse IgG conjugated horse-radish peroxidase were reacted for 1 hour and band images were taken with X-ray film using ECL solution.
(4) (4) 쿠마시블루Kumasi Blue 염색 dyeing
추출온도를 다르게 한 효모추출물을 15% SDS-폴리아크릴아미드 겔에 전기영동하여 단백질을 분리하였다. 전기영동이 끝난 후 쿠마시블루 용액을 이용하여 1시간 염색 후 데스타인(destain) 버퍼로 1시간 RT로 세척한 후 크기별로 분리된 단백질을 확인하였다.The yeast extract with different extraction temperature was electrophoresed on 15% SDS-polyacrylamide gel to separate proteins. After electrophoresis, the cells were stained with Coomassie blue for 1 hour, washed with destain buffer for 1 hour, and then isolated proteins were identified by size.
2. 실험 결과 2. Experimental results
실험 결과는 도 1 내지 4와 같이 나타났다. Experimental results are shown in Figs. 1 to 4.
도 1은 전기 자극 효모추출물을 단순추출, UF-5K, UF-10K로 추출하여 기도 상피세포에 단독 (도 1의 a) LPS와 함께 처리 (도 1의 b)한 결과로써, UF-10K 저분자로 추출한 물질의 경우, 세포 독성이 없는 것으로 확인되었다. 또한, Eotaxin-1, ICAM-1, MUC5AC의 발현을 UF-10K 추출물에서 억제하는 것을 확인할 수 있었다 (도 1의 c).Fig. 1 shows the results of the UF-10K low-molecular-weight assay (Fig. 1) as a result of treatment with LPS alone (Fig. 1 a) and airway epithelial cells Were not found to be cytotoxic. In addition, it was confirmed that the expression of Eotaxin-1, ICAM-1, and MUC5AC was inhibited by the UF-10K extract (FIG.
도 2는 추출온도에 따른 전기 자극 효모 추출물의 세포 독성을 확인한 결과로써, 40~60℃에서 세포 독성이 없는 것을 확인할 수 있었다.FIG. 2 shows the cytotoxicity of the electric stimulated yeast extract according to the extraction temperature. As a result, it was confirmed that there was no cytotoxicity at 40 to 60 ° C.
도 3은 추출온도에 따른 전기 자극 효모 추출물 내 티오레독신의 함유랑을 측정한 결과로써, 티오레독신의 함유량이 60℃에서 높은 것을 확인할 수 있었다.FIG. 3 shows the content of thioredoxin in the electric stimulated yeast extract according to the extraction temperature. As a result, it was confirmed that the content of thioredoxin was high at 60 ° C.
도 4는 추출온도 40℃, 60℃에서, 본 발명 전기 자극 효모추출물이 천식 마커인 Eotaxin-1의 발현을 억제하는 것을 보여준다.4 shows that the electric stimulation yeast extract of the present invention suppresses the expression of Eotaxin-1, which is an asthma marker, at extraction temperatures of 40 캜 and 60 캜.
Claims (10)
상기 효모 전기 자극 추출물은 티오레독신의 함량이 증대된 것을 특징으로 하는 천식 개선용 식품 조성물.
After the yeast was subjected to electric stimulation and extracted, the yeast was removed, and the yeast electric stimulation extract was obtained by filtration using an MWCO 10 kDa ultrafiltration membrane to remove the filtrate.
Wherein the yeast electric stimulation extract has an increased thioredoxin content.
상기 단계 (a) 후, 물을 가하고 교반하면서 효모를 추출하는 단계 (b);
상기 단계 (c) 후, 효모를 제거하고, MWCO 10kDa 한외 여과막을 이용하여 여과액을 제거하여 효모 전기 자극 추출물을 제조하는 단계 (c);를 포함하고,
상기 효모 전기 자극 추출물은 티오레독신의 함량이 증대되는 것을 특징으로 하는 천식 개선용 식품 조성물의 제조방법.
(A) applying electrical stimulation to the yeast;
(B) after the step (a), water is added and the yeast is extracted while stirring;
(C) a step of removing the yeast and removing the filtrate using an MWCO 10 kDa ultrafiltration membrane to prepare a yeast electric stimulation extract; and (c)
Wherein the yeast electric stimulation extract has an increased content of thioredoxin.
상기 효모는,
효모 배양액인 것을 특징으로 하는 천식 개선용 식품 조성물.
The method according to claim 1,
The yeast,
Wherein the composition is a yeast culture.
상기 전기 자극은,
0.001~20V의 전기로 0.001~10시간 동안 자극을 가하는 것을 특징으로 하는 천식 개선용 식품 조성물.
The method according to claim 1,
Wherein the electrical stimulation comprises:
And a stimulus is applied for 0.001 to 10 hours in an electric furnace at 0.001 to 20 V.
상기 효모로부터 추출시,
효모의 농도가 1~10%(w/v)가 되도록 물을 가하고 교반 추출하는 것을 특징으로 하는 천식 개선용 식품 조성물.
The method according to claim 1,
Upon extraction from the yeast,
And water is added thereto so that the yeast concentration is 1 to 10% (w / v), followed by stirring and extraction.
상기 효모 전기 자극 추출물은 티오레독신의 함량이 증대된 것을 특징으로 하는 천식 치료 또는 예방용 약품 조성물.
After the yeast was subjected to electric stimulation and extracted, the yeast was removed, and the yeast electric stimulation extract was obtained by filtration using an MWCO 10 kDa ultrafiltration membrane to remove the filtrate.
Wherein the yeast electric stimulation extract has an increased content of thioredoxin.
상기 단계 (a) 후, 물을 가하고 교반하면서 효모를 추출하는 단계 (b);
상기 단계 (c) 후, 효모를 제거하고, MWCO 10kDa 한외 여과막을 이용하여 여과액을 제거하여 효모 전기 자극 추출물을 제조하는 단계 (c);를 포함하고,
상기 효모 전기 자극 추출물은 티오레독신의 함량이 증대되는 것을 특징으로 하는 천식 치료 또는 예방용 약학 조성물의 제조방법.
(A) applying electrical stimulation to the yeast;
(B) after the step (a), water is added and the yeast is extracted while stirring;
(C) a step of removing the yeast and removing the filtrate using an MWCO 10 kDa ultrafiltration membrane to prepare a yeast electric stimulation extract; and (c)
Wherein the yeast electric stimulation extract has an increased content of thioredoxin.
상기 효모는,
효모 배양액인 것을 특징으로 하는 천식 치료 또는 예방용 약품 조성물.
The method according to claim 6,
The yeast,
Wherein the composition is a yeast culture solution.
상기 전기 자극은,
0.001~20V의 전기로 0.001~10시간 동안 자극을 가하는 것을 특징으로 하는 천식 치료 또는 예방용 약품 조성물.
The method according to claim 6,
Wherein the electrical stimulation comprises:
And a stimulus is applied for 0.001 to 10 hours in an electric furnace at 0.001 to 20 V.
상기 효모로부터 추출시,
효모의 농도가 1~10%(w/v)가 되도록 물을 가하고 교반 추출하는 것을 특징으로 하는 천식 치료 또는 예방용 약품 조성물. The method according to claim 6,
Upon extraction from the yeast,
And water is added to the mixture so that the yeast concentration is 1 to 10% (w / v), followed by stirring and extraction.
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| KR102080498B1 (en) * | 2019-08-23 | 2020-02-24 | 정영옥 | Food composition for improving or preventing osteopenia comprising ferulic acid and lactic acid bacteria as active ingredient |
| KR20200089374A (en) * | 2019-01-16 | 2020-07-27 | 한림대학교 산학협력단 | Composition of improvement, prevention and treatment in Chronic obstructive pulmonary disease with yeast extract |
| WO2021071413A1 (en) * | 2019-10-07 | 2021-04-15 | Milmed Unico Ab | Yeast for the treatment of allergy |
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| CN103951727A (en) * | 2014-04-25 | 2014-07-30 | 华南理工大学 | Method for extracting active components of yeast based on high voltage pulse discharge technology |
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| CN103951727A (en) * | 2014-04-25 | 2014-07-30 | 华南理工大学 | Method for extracting active components of yeast based on high voltage pulse discharge technology |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20200089374A (en) * | 2019-01-16 | 2020-07-27 | 한림대학교 산학협력단 | Composition of improvement, prevention and treatment in Chronic obstructive pulmonary disease with yeast extract |
| KR102318414B1 (en) | 2019-01-16 | 2021-10-29 | 한림대학교 산학협력단 | Composition of improvement, prevention and treatment in Chronic obstructive pulmonary disease with yeast extract |
| KR102080498B1 (en) * | 2019-08-23 | 2020-02-24 | 정영옥 | Food composition for improving or preventing osteopenia comprising ferulic acid and lactic acid bacteria as active ingredient |
| WO2021071413A1 (en) * | 2019-10-07 | 2021-04-15 | Milmed Unico Ab | Yeast for the treatment of allergy |
| CN114514029A (en) * | 2019-10-07 | 2022-05-17 | 密美德尤尼克有限公司 | Yeast for treating allergy |
| CN114514029B (en) * | 2019-10-07 | 2024-03-08 | 密美德尤尼克有限公司 | Yeast for treating allergy |
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