KR20150104482A - A composition comprising the red ginseng extract of postmanufacture for treating or preventing oral squamous cell carcinoma - Google Patents
A composition comprising the red ginseng extract of postmanufacture for treating or preventing oral squamous cell carcinoma Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/25—Araliaceae (Ginseng family), e.g. ivy, aralia, schefflera or tetrapanax
- A61K36/258—Panax (ginseng)
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
- A61P1/02—Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/308—Foods, ingredients or supplements having a functional effect on health having an effect on cancer prevention
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/312—Foods, ingredients or supplements having a functional effect on health having an effect on dental health
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
Abstract
Description
본 발명은 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물에 관한 것으로, 구체적으로는 메탄올을 이용하여 추출된 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물에 관한 것이다. The present invention relates to a composition for preventing, ameliorating or treating oral squamous cell cancer, which comprises an extract of red ginseng as an active ingredient. More specifically, the present invention relates to a composition for oral squamous cell To a composition for preventing, ameliorating or treating cancer.
본 발명에 따른 조성물의 농도는 구강편평세포암의 세포주인 HSC-3 세포 기준으로 0.06~4 mg/ml인 것을 기술적 특징으로 하고 있다.The concentration of the composition according to the present invention is 0.06-4 mg / ml on the basis of HSC-3 cell line of oral squamous cell carcinoma.
본 발명에 따른 조성물은 HSC-3 세포를 사멸시키는 효능, HSC-3 세포의 캐스페이즈(caspase) -3, -8 및 -9의 발현을 촉진하는 효능, HSC-3 세포의 분절 PARP의 발현을 촉진하는 효능, HSC-3 세포의 주기가 세포자멸(apoptosis)기로 진행되는 것을 유도하는 효능, HSC-3 세포의 Bcl-2의 발현을 저해하는 효능, HSC-3 세포의 Bad, Bim 및 Bax를 포함하는 프로-아포토시스(pro-apoptosis) 단백질의 발현을 촉진하는 효능과 ERK, pERK, JNK, pJNK, p38 및 pp38을 포함하는 MAPK 통로 단백질의 발현을 촉진하는 효능을 보유하고 있다.
The composition according to the present invention is effective for killing HSC-3 cells, for promoting the expression of caspase-3, -8 and -9 of HSC-3 cells, for expressing segmental PARP of HSC-3 cells HSC-3 cells, HSC-3 cells, HSC-3 cells, HSC-3 cells, HSC-3 cells, HSC-3 cells, HSC-3 cells, Apoptosis protein and the expression of MAPK pathway proteins including ERK, pERK, JNK, pJNK, p38, and pp38.
구강편평세포암(Oral squamous cell carcinoma)은 구강에 발생하는 악성종양의 95% 이상을 차지하는 질환으로써, 환경학적 요인(흡연, 음주, 동물성 지방 및 포화지방의 과다 섭취 등), 영양학적 요인(비타민, 아연, 구리 또는 철 결핍증 등) 및 병리학적 요인(바이러스 감염 등) 등 다양한 요인에 의하여 발병하는 것으로 알려져 있다.Oral squamous cell carcinoma is a disease that accounts for more than 95% of the malignant tumors that develop in the oral cavity. It is an environmental factor (smoking, drinking, excessive intake of animal fat and saturated fat, etc.) , Zinc, copper, or iron deficiency), and pathological factors (such as viral infection).
구강편평세포암이 발병하면 궤양, 소괴, 균열, 적색반, 과립양상 등 매우 다양한 임상 증상이 발견된다. 병소의 30%는 입술, 25%는 혀이며, 혀에서는 혀 측면,저부, 후면부 또는 상악골에 빈번히 발생한다. When oral squamous cell carcinoma develops, a wide variety of clinical symptoms such as ulcers, fractures, cracks, redness, and granularity are found. 30% of the lesions occur in the lips, 25% in the tongue, and frequently in the tongue in the side, bottom, back, or maxilla.
구강암은 조기발견이 가능함에도 불구하고 5년 생존율이 30%에 불과하며, 혀에 발생할 경우 더욱 예후가 좋지 않다.
Despite early detection of oral cancer, the 5-year survival rate is only 30%, and the prognosis is worse when it occurs in the tongue.
구강암의 치료에는 병소를 냉동 요법 등의 수술적 치료, 방사선 요법 및 화학 요법 등의 비수술적 치료가 수행되고 있으나, 모든 치료 요법에 부작용이 따르는 실정이다.In the treatment of oral cancer, non-surgical treatment such as cryotherapy such as cryotherapy, radiotherapy and chemotherapy is performed, but all therapies have side effects.
냉동 요법은 부종, 수포, 진물, 가피, 동통, 감염 및 출혈 등의 부작용이 있고, 심할 경우 복원을 위한 성형이 필요할 정도로 조직이 괴사될 수 있으며, 방사선 요법은 홍반, 색소침착, 건조성 또는 습윤성 피부박리 및 모세혈관 확장증 등의 부작용이 있다.Cryotherapy has side effects such as edema, blisters, sore throat, scarring, pain, infection and hemorrhage. In severe cases, the tissue may be necrotic enough to require reconstitution. Radiotherapy may include erythema, pigmentation, Skin exfoliation and capillary vasodilation.
더불어 화학 요법은 구역, 구토, 구내염, 설사 및 골수기능장애 등의 부작용이 있다.
In addition, chemotherapy has side effects such as nausea, vomiting, stomatitis, diarrhea, and bone marrow dysfunction.
이에 본 출원인은 부작용이 없으며, 구강편평세포암의 세포주인 HSC-3 세포를 표적으로 함으로써 구강편평세포암을 근본적으로 치료할 수 있는 조성물을 제공할 수 있다고 판단하여, 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암 저해 조성물을 안출하였다.
Thus, the applicant of the present invention has found that it is possible to provide a composition capable of fundamentally treating oral squamous cell cancer by targeting HSC-3 cells, which is a cell line of oral squamous cell carcinoma, The present invention also provides a composition for inhibiting oral squamous cell cancer.
이하에서, 본 발명에 사용된 소재인 홍삼박에 관하여 간략히 설명하고자 한다.
Hereinafter, the red ginseng foil used in the present invention will be briefly described.
인삼(Panax ginseng C. A. Meyer)은 오가피과 인삼속 인삼종에 속하는 다년생식물을 일컬으며, 건조여부와 가공방법에 따라 수삼, 건삼, 홍삼으로 나눌 수 있다. 수삼은 가공되거나 말리지 않는 인삼, 건삼은 햇볕. 열풍 등으로 익히지 않고 말린 인삼, 홍삼은 수삼을 수증기(증삼) 또는 기타 방법으로 쪄서 건조한 것이다. Panax ginseng CA Meyer) is a perennial plant belonging to ginseng belonging to ginseng and ginseng. It can be divided into ginseng, dried ginseng and red ginseng according to the drying condition and processing method. Ginseng is processed or not dried, and sunshine is sunburn. Dried ginseng and red ginseng without being cooked by hot wind are steamed steamed (steamed) or other methods and dried.
인삼 및 홍삼에는 사포닌(Saponin)을 비롯하여 인삼향성분(Panacen), 폴리아세틸렌(Polyacetylen)계 화합물, 함질소성분, 플라보노이드(Flavonoid), 비타민 B군, 효소, 항산화물질, 유기산, 진세노사이드, 아미노산 및 기타 미량원소 등이 함유되어 있다.Ginseng and red ginseng contain saponin as well as Panacen, Polyacetylen, Nitrogen, Flavonoid, Vitamin B, Enzyme, Antioxidant, Organic acid, Ginsenoside, Amino acid And other trace elements.
현재 인삼에는 20~24종의 진세노사이드가 분리 보고된 것에 비하여, 홍삼에는 34~36종의 진세노사이드가 분리 보고되었는데, 홍삼에는 특유의 진세노사이드 성분인 -Rh2, -Rh1, Rg 등의 14종 가량이 더 함유되어 있다고 알려져 있다.At present, 20 ~ 24 species of ginsenosides have been reportedly reported in red ginseng, whereas 34 ~ 36 species of ginsenosides have been reported in red ginseng. In the case of red ginseng, specific ginsenoside components -Rh2, -Rh1, Rg Of which 14 are more known.
홍삼에는 면역력 증진 및 혈압강하 등의 효능이 있으며, 당뇨 및 골다공증 등에도 효과가 있다.
Red ginseng has effects such as improving immunity and lowering blood pressure, and is also effective for diabetes and osteoporosis.
상기와 같은 특징을 보유한 홍삼은 일 회 가공한 후, 또는 복수 회 가공한 후에도 추출하여 홍삼 가공 추출물로써 사용될 수 있다.Red ginseng having the above characteristics can be extracted and used as a red ginseng processing extract even after processing once or multiple times.
홍삼 가공 추출물을 제조하는 방법에는 다양한 방법이 있으며, 일 예로 홍삼박은 홍삼을 물 또는 알코올 등의 용매로 추출한 잔사를 다시 추출하여 제조된다. There are various methods for preparing ginseng processing extract. For example, red ginseng is prepared by re-extracting red ginseng residue extracted with a solvent such as water or alcohol.
홍삼 추출시 사용한 용매에 따라, 상기 용매에 용해되지 않은 홍삼의 유효 잔류 성분인 사포닌, 폴리아세틸렌, 단백질 등이 홍삼박에 대략 20~40% 정도 잔류하게 된다.
Depending on the solvent used for extracting red ginseng, about 20 to 40% of saponin, polyacetylene, protein, and the like, which are effective residues of red ginseng not dissolved in the solvent, remain in the red ginseng.
본 발명과 관련된 문헌을 살펴보면, 대한민국 공개특허공보 제10-2009-0053091호(선행문헌 1)에는 암세포 성장억제용 지용성 인삼추출조성물과 이를 이용한 의약조성물 및 건강식품에 관하여 기재되어 있으며, 대한민국 공개특허공보 제10-2013-0056701호(선행문헌 2)에는 홍삼 및 천연식물 유래 추출물을 포함하는 구강내 항염, 항균 및 소취를 위한 조성물에 관하여 기재되어 있다.Korean Patent Laid-Open Publication No. 10-2009-0053091 (prior art document 1) discloses a composition for extracting fat-soluble ginseng for inhibiting the growth of cancer cells, a pharmaceutical composition using the same, and a health food, Publication No. 10-2013-0056701 (Prior Art 2) discloses compositions for oral anti-inflammatory, antibacterial and deodorant containing red ginseng and natural plant-derived extracts.
또한 대한민국 공개특허공보 제10-2000-0069452호(선행문헌 3)에는 구강 편평세포암에 대한 류코트리엔 길항제에 관하여 기재되어 있다.
Korean Patent Laid-Open Publication No. 10-2000-0069452 (Prior Art 3) discloses a leukotriene antagonist for oral squamous cell cancer.
상기 선행문헌 1 및 2의 내용을 살펴보면, 암세포의 성장을 억제하거나, 구강 내를 항염, 항균 및 소취하는 기술로써, 본 발명과는 소재의 선정 면에서만 미미한 유사점이 있으며 홍삼 가공 추출물을 이용하여 구강편평세포암을 저해하는 기술인 본 발명과는 상이한 기술이다.As a result, the growth of cancer cells is suppressed or anti-inflammation, antibacterial, and deodorization are performed in the oral cavity. Thus, there is a slight similarity with the present invention only in the selection of materials, Is a technique different from the present invention which is a technique for inhibiting squamous cell cancer.
특히 본 발명은 구강편평세포암의 세포주인 HSC-3을 대상으로 하고 있으므로, 적응증이 명확하다 할 수 있다.
In particular, since the present invention targets HSC-3, a cell line of oral squamous cell carcinoma, the indications are clear.
또한 선행문헌 3의 내용을 살펴보면, 구강편평세포암을 치료하는 조성물로써 합성물인 류코트리엔 길항제를 제시하고 있어, 천연물인 홍삼 가공 추출물을 유효성분으로 함유한 조성물을 제시한 본 발명과 상이하다.In addition, in the
결론적으로, 본 발명은 국내에서 시도되지 않은 신규한 발명이라 할 수 있다.
In conclusion, the present invention is a novel invention not attempted in the domestic market.
본 발명의 목적은 구강편평세포암의 세포주인 HSC-3 세포의 세포자멸사를 다각적으로 유도함으로써, 구강편평세포암을 근본적으로 예방, 개선 또는 치료할 수 있는 조성물을 제공하는 것이다.It is an object of the present invention to provide a composition capable of fundamentally preventing, ameliorating, or treating oral squamous cell cancer by variously inducing apoptosis of HSC-3 cell line of oral squamous cell carcinoma.
본 발명의 목적은 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물을 제공하는 것이다.
It is an object of the present invention to provide a composition for preventing, ameliorating or treating oral squamous cell cancer containing red ginseng extract as an active ingredient.
본 발명은 HSC-3 세포를 사멸시키는 효능, HSC-3 세포의 캐스페이즈 -3, -8 및 -9의 발현을 촉진하는 효능, HSC-3 세포의 분절 PARP의 발현을 촉진하는 효능 및 HSC-3 세포의 주기가 세포자멸기로 진행되는 것을 유도하는 효능을 보유한, 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물을 제공함으로써 기술적 과제를 해결하고자 한다.The present invention relates to a method for inhibiting HSC-3 cell death, an effect of promoting expression of caspase-3, -8 and -9 of HSC-3 cells, an effect of promoting expression of segmental PARP of HSC-3 cells, 3 cells having an activity of inducing progression of a cell cycle to a cell apoptosis, and to provide a composition for prevention, improvement or treatment of oral squamous cell cancer which contains red ginseng extract as an active ingredient.
본 발명은 HSC-3 세포의 Bcl-2의 발현을 저해하는 효능, HSC-3 세포의 프로-아포토시스 단백질의 발현을 촉진하는 효능 및 HSC-3 세포의 MAPK 통로 단백질의 발현을 촉진하는 효능을 보유한 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물을 제공함으로써 기술적 과제를 해결하고자 한다.
The present invention relates to a method for inhibiting the expression of Bcl-2 in HSC-3 cells, an effect of promoting the expression of pro-apoptotic proteins in HSC-3 cells and an effect of promoting the expression of MAPK channel proteins in HSC-3 cells The present invention aims at solving the technical problem by providing a composition for prevention, improvement or treatment of oral squamous cell cancer which contains red ginseng extract as an active ingredient.
본 발명에 따른 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물은 HSC-3 세포를 사멸시키는 효능, HSC-3 세포의 캐스페이즈 -3, -8 및 -9의 발현을 촉진하는 효능, HSC-3 세포의 분절 PARP의 발현을 촉진하는 효능 및 HSC-3 세포의 주기가 세포자멸기로 진행되는 것을 유도하는 효능을 보유하고 있다.The composition for preventing, ameliorating or treating oral squamous cell carcinoma containing the extract of red ginseng according to the present invention as an active ingredient is effective for killing HSC-3 cells, inhibiting caspase-3, -8, and - 9, the effect of promoting the expression of segmental PARP in HSC-3 cells, and the effect of inducing the cycle of HSC-3 cells to progress to apoptosis.
본 발명에 따른 홍삼 가공 추출물을 유효성분으로 함유하는 구강편평세포암의 예방, 개선 또는 치료를 위한 조성물은 HSC-3 세포의 Bcl-2의 발현을 저해하는 효능, HSC-3 세포의 프로-아포토시스 단백질의 발현을 촉진하는 효능 및 HSC-3 세포의 MAPK 통로 단백질의 발현을 촉진하는 효능을 보유하고 있다.
The composition for preventing, ameliorating or treating oral squamous cell cancer, which comprises the extract of Red Ginseng according to the present invention as an active ingredient, is effective for inhibiting the expression of Bcl-2 in HSC-3 cells, pro-apoptosis of HSC- Promoting the expression of the protein and promoting the expression of the MAPK channel protein of HSC-3 cells.
도 1은 홍삼박 추출물을 처리하였을시 HSC-3 세포의 세포사멸률을 나타낸 그래프이다.
도 2는 홍삼박 추출물을 처리하였을시 HSC-3 세포의 세포사멸률을 나타낸 사진이다.
도 3의 상단은 홍삼박 추출물의 처리 농도에 따른 HSC-3 세포의 캐스페이즈 효소 발현의 변화 양상을 나타낸 그래프이다.
도 3의 하단은 홍삼박 추출물의 처리 시간에 따른 HSC-3 세포의 캐스페이즈 효소 발현의 변화 양상을 나타낸 그래프이다.
도 4는 홍삼박 추출물을 처리하였을시 HSC-3 세포의 분절 PARP 효소 및 캐스페이즈-3 효소의 발현 촉진에 미치는 영향을 웨스턴 블롯을 수행하여 나타낸 사진이다.
도 5는 홍삼박 추출물을 처리하였을시 HSC-3 세포의 세포자멸 유도에 미치는 영향을 플로우 사이토메트리를 이용하여 나타낸 그래프이다.
도 6은 홍삼박 추출물을 처리하였을시 HSC-3 세포 주기의 변화 양상을 나타낸 표이다.
도 7은 홍삼박 추출물을 처리하였을시 HSC-3 세포의 자멸률을 TUNEL assay로 나타낸 그래프이다.
도 8은 홍삼박 추출물을 처리하였을시 HSC-3 세포의 밀도를 cells/ml, 자멸률을 %로 나타낸 표이다.
도 9는 홍삼박 추출물을 HSC-3 세포에 처리하였을시 시간의 경과에 따른 Bcl-2 및 프로-아포토시스 단백질의 발현의 변화 양상을 웨스턴 블롯을 수행하여 나타낸 사진이다.
도 10은 홍삼박 추출물을 농도별로 HSC-3 세포에 처리하였을시 HSC-3 세포의 Bcl-2 및 프로-아포토시스 단백질의 발현에 미치는 영향을 웨스턴 블롯을 수행하여 나타낸 사진이다.
도 11은 홍삼박 추출물을 HSC-3 세포에 처리하였을시 시간의 경과에 따른 MAPK 통로 단백질 발현의 변화 양상을 웨스턴 블롯을 수행하여 나타낸 사진이다.
도 12는 홍삼박 추출물을 농도별로 HSC-3 세포에 처리하였을시 농도에 따른 MAPK 통로 단백질 발현의 변화 양상을 웨스턴 블롯을 수행하여 나타낸 사진이다.FIG. 1 is a graph showing the cell death rate of HSC-3 cells treated with red ginseng root extract. FIG.
2 is a photograph showing the cell death rate of HSC-3 cells when treated with the extract of red ginseng roots.
3 is a graph showing changes in caspase enzyme expression of HSC-3 cells according to the treatment concentration of red ginseng root extract.
3 is a graph showing changes in the expression of caspase enzyme in HSC-3 cells according to the treatment time of red ginseng root extract.
4 is a photograph showing the effect of HSC-3 cells on the promotion of the expression of segmental PARP enzyme and caspase-3 enzyme when treated with red ginseng root extract by Western blotting.
FIG. 5 is a graph showing the effect of HSC-3 cells on induction of apoptosis induced by red ginseng root extract using flow cytometry. FIG.
FIG. 6 is a table showing changes in HSC-3 cell cycle upon treatment with red ginseng root extract.
FIG. 7 is a graph showing the suicide rate of HSC-3 cells treated with red ginseng root extract by TUNEL assay.
8 is a table showing the density of HSC-3 cells in cells / ml and the rate of self-destruction in% when red ginseng roots extract was treated.
FIG. 9 is a photograph showing Western blotting of changes in expression of Bcl-2 and pro-apoptotic proteins over time when HSC-3 cells were treated with red ginseng root extract.
FIG. 10 is a photograph showing Western blotting effect of HSC-3 cells on the expression of Bcl-2 and pro-apoptotic proteins in HSC-3 cells treated with various concentrations of red ginseng root extract.
11 is a photograph showing Western blotting of changes in MAPK channel protein expression over time when HSC-3 cells were treated with red ginseng root extract.
FIG. 12 is a photograph showing Western blotting of changes in MAPK pathway protein expression according to the concentration of HSC-3 cells treated with the extract of red ginseng root at various concentrations.
본 명세서 및 청구 범위에 사용된 용어나 단어는 통상적이거나 사전적인 의미로 한정해서 해석되어서는 안 되며, 발명자는 그 자신의 발명을 가장 최선의 방법으로 설명하기 위해 용어의 개념을 적절하게 정의할 수 있다는 원칙에 입각하여 본 발명의 기술적 사상에 부합하는 의미와 개념으로 해석되어야만 한다.The terms and words used in the present specification and claims should not be construed as limited to ordinary or dictionary terms and the inventor may properly define the concept of the term in order to best describe its invention It should be construed as meaning and concept consistent with the technical idea of the present invention.
따라서 본 명세서에 기재된 실시예, 참조예 및 도면에 기술된 사항은 본 발명의 가장 바람직한 일 예에 불과할 뿐이고 본 발명의 기술적 사상을 모두 대변하는 것은 아니므로, 본 출원시점에 있어서 이들을 대체할 수 있는 다양한 균등물과 변형예들이 있을 수 있음을 이해하여야 한다.
Therefore, the embodiments described in the present specification, the reference examples, and the drawings are merely the most preferred examples of the present invention, and not all of the technical ideas of the present invention are described. Therefore, It should be understood that various equivalents and modifications may be present.
실시예Example
1. 홍삼 가공 추출물의 제조 1. Preparation of red ginseng processing extract
홍삼을 가공한 후 다시 추출하여 홍삼 가공 추출물을 제조하며, 홍삼 가공 추출물에는 홍삼박이 포함된다.After processing red ginseng, it is re-extracted to produce red ginseng processing extract. Red ginseng processing extract contains red ginseng.
또한 홍삼 가공 추출물에는 복수 회 가공한 후의 추출물이 포함한다.
Also, the red ginseng processing extract contains the extract after processing multiple times.
홍삼 가공 추출물을 추출할시 추출 방법에는 용매 추출법, 수증기 증류법, 이산화탄소 초임계 추출법, 수증기 증류법, 마이크로 웨이브 공정 추출법, 퍼콜레이션 추출법 등의 다양한 추출 방법이 사용될 수 있으며, 바람직하게는 용매 추출법을 이용할 수 있다. 추출시 시간 및 온도가 달라질 수 있다.Various extraction methods such as a solvent extraction method, a steam distillation method, a carbon dioxide supercritical extraction method, a steam distillation method, a microwave process extraction method, and a percolation extraction method may be used as the extraction method when extracting the red ginseng processing extract. have. The extraction time and temperature may vary.
용매 추출시, 추출 용매의 종류에 따라 추출 온도, 추출 시간, 용매의 양 및 잔류 성분 처리 방식 등을 달리하여 처리한다. 추출 용매 또한 다양한 용매가 사용될 수 있다. 추출 가능한 용매에는 물, 에탄올, 메탄올, 지방유, 글리세린, 마유, 플로필렌글리콜, 에테르, 클로르포름, 석유에테르, 헥산, 벤젠, 메틸렌클로라이드, 에틸아세테이트, 아세톤, 부탄올 및 이소프로판올 등이 있으며, 바람직하게는 메탄올을 사용할 수 있으나 이에 한정되는 것은 아니다. When the solvent is extracted, the extraction temperature, the extraction time, the amount of the solvent and the treatment method of the residual component are treated differently depending on the type of the extraction solvent. Extraction solvents. Various solvents may also be used. Examples of the extractable solvent include water, ethanol, methanol, fatty oil, glycerin, hemp, flouroprene glycol, ether, chloroform, petroleum ether, hexane, benzene, methylene chloride, ethyl acetate, acetone, butanol and isopropanol, Methanol may be used, but is not limited thereto.
또한 용매의 농도 역시 달라질 수 있으며, 바람직하게는 70 ~ 90 중량% 농도의 메탄올을 사용할 수 있으나 이에 한정되는 것은 아니다.
The concentration of the solvent may also be varied, and preferably 70 to 90% by weight of methanol may be used, but is not limited thereto.
또한 설계 조건에 따라 추출 후 농축, 여과 및 동결건조 등의 과정을 1회 이상 수행할 수 있으며, 상기 공정을 수행한 후 용매에 용해할 수도 있다.
In addition, the extraction, filtration, freeze-drying, and the like may be performed one or more times according to the design conditions, and may be dissolved in the solvent after performing the above-mentioned process.
실험예Experimental Example
1. One.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포의 사멸 -3 cell death
(1) 실험 준비(1) Preparing the experiment
1) 홍삼박의 준비1) Preparation of red ginseng
홍삼 가공 추출물의 효과를 극대화하기 위하여, 홍삼 가공 추출물의 종류 중 가장 하급에 속하며 주로 폐기처분되는 홍삼박을 택하여 홍삼박의 HSC-3 사멸 효능을 알아보고자 하였다.
In order to maximize the effect of red ginseng processing extracts, we examined the HSC-3 killing efficacy of red ginseng roots by selecting the red ginseng roots which are the lowest among the red ginseng processing extracts.
2) HSC-3 세포의 준비2) Preparation of HSC-3 cells
혀에 발생한 구강편평세포암에 위치한 HSC-3 세포를 준비하였다. HSC-3 세포는 JCRB(Japanese Cancer Research Bank)에서 제공받았다.Located in the oral squamous cell carcinoma of the tongue HSC-3 cells were prepared. HSC-3 cells were obtained from JCRB (Japanese Cancer Research Bank).
이하 모든 실험예에서, 실험예 1과 동일한 과정으로 준비된 HSC-3 세포를 사용하였음을 밝혀둔다.
It is to be noted that HSC-3 cells prepared in the same manner as in Experimental Example 1 were used in all of the following Experimental Examples.
(2) 실험 과정(2) Experimental process
MTT assay를 수행하여 홍삼박 추출물의 HSC-3 세포사멸 효능을 알아보고자 하였다.MTT assay was performed to investigate the HSC-3 cell killing effect of red ginseng roots extract.
홍삼박 메탄올 추출물의 농도를 각각 조정한 후, 농도를 조정한 각 홍삼박 추출물을 HSC-3 세포에 처리하여 MTT assay를 수행하였다. After adjusting the concentrations of the methanol extracts of red ginseng roots, each of the red ginseng roots extracts adjusted to the concentration was treated with HSC-3 cells and MTT assay was performed.
MTT assay는 통상적인 방법으로 수행되었다.
The MTT assay was performed in a conventional manner.
또한 실험예 1에 사용된 홍삼박 추출물은 실시예 1에 기재된 과정과 동일하게 제조하였으며, 90 중량% 메탄올을 사용하여 제조되었다. The red ginseng root extract used in Experimental Example 1 was prepared in the same manner as described in Example 1, and was prepared using 90% by weight methanol.
더불어, 이하 모든 실험예에 사용된 홍삼박 추출물 역시 실험예 1에 사용된 홍삼박 추출물과 동일한 것임을 밝혀둔다.
In addition, the red ginseng root extract used in all the following Experimental Examples was also evaluated in Experimental Example 1 It is the same as the red ginseng roots extract used.
(3) 실험 결과(3) Experimental results
도 1은 홍삼박 추출물을 처리하였을시 HSC-3 세포의 세포사멸률을 나타낸 그래프이다.
FIG. 1 is a graph showing the cell death rate of HSC-3 cells treated with red ginseng root extract. FIG.
실험 결과, 홍삼박 추출물의 처리 농도가 증가할 수록 HSC-3 세포의 생존율 역시 감소하는 것을 발견할 수 있었다. As a result, the survival rate of HSC-3 cells was found to decrease with increasing concentration of red ginseng root extract.
특히 홍삼박 추출물 0.125 mg/ml 처리군에서는 HSC-3 세포의 생존율이 50% 이하로 감소하기 시작하였으며, 0.25, 0.5 mg/ml로 처리 농도를 높임에 따라 점차 감소하였음을 발견할 수 있었다. In particular, the survival rate of HSC-3 cells decreased to less than 50% in the group treated with 0.125 mg / ml of red ginseng extract, and gradually decreased with increasing treatment concentration to 0.25 and 0.5 mg / ml.
또한 홍삼박 추출물 1 mg/ml 처리군에서는 0%에 가깝게 세포 생존율이 감소한 것을 발견할 수 있었다.
In addition, it was found that the cell survival rate in the group treated with 1 mg / ml of red ginseng extract decreased to 0%.
도 2는 홍삼박 추출물을 처리하였을시 HSC-3 세포의 세포사멸률을 나타낸 사진이다.
2 is a photograph showing the cell death rate of HSC-3 cells when treated with the extract of red ginseng roots.
실험 결과, 도 1에서와 동일하게, 홍삼박 추출물의 처리 농도가 증가할 수록 HSC-3 세포의 생존률이 감소하는 것을 발견할 수 있었다.As shown in FIG. 1, the survival rate of HSC-3 cells was found to decrease as the treatment concentration of red ginseng roots extract increased.
홍삼박 추출물을 처리하지 않았을시의 HSC-3 세포를 관찰해 보면 세포의 밀집도가 높은 것을 알 수 있으나, 홍삼박 추출물을 투여한 후부터 밀집도가 감소하는 것을 발견할 수 있었다.When HSC-3 cells were not treated with red ginseng roots extract, it was found that the cell density was high, but it was found that the density decreased after administration of red ginseng roots extract.
특히 홍삼박 추출물 1 mg/ml 투여군에서는 세포의 밀집도가 약 1/2로 감소하였으며, 2 mg/ml 투여군에서는 약 1/4로 감소하였음을 발견할 수 있었다.
Especially, the density of cells in the group treated with 1 mg / ml of red ginseng extract decreased to about ½ and decreased to about ¼ in the group treated with 2 mg / ml.
실험예Experimental Example
2. 2.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포에서의 -3 cells
캐스페이즈CAS PHASE
효소 발현 촉진 Promoting enzyme expression
(1) 실험 결과(1) Experimental results
캐스페이즈는 세포자멸을 유도하는 대표적인 효소이므로, 홍삼박 추출물을 HSC-3 세포에 처리하였을시 캐스페이즈의 발현 양상을 확인하였다.
Since caspase is a typical enzyme inducing apoptosis, When HSC-3 cells were treated, caspase expression was observed.
도 3의 상단은 홍삼박 추출물의 처리 농도에 따른 HSC-3 세포의 캐스페이즈 효소 발현의 변화 양상을 나타낸 그래프이다.
3 is a graph showing changes in caspase enzyme expression of HSC-3 cells according to the treatment concentration of red ginseng root extract.
실험 결과, 홍삼박 추출물의 처리 시간이 증가할 수록 HSC-3 세포의 캐스페이즈 -3, -8 및 -9의 발현율은 모두 증가하는 것을 관찰할 수 있었다.As a result, the expression rates of caspase-3, -8 and -9 in HSC-3 cells were increased with increasing treatment time of red ginseng root extract.
홍삼박 추출물 처리 전 캐스페이즈 -3, -8 및 -9의 발현율은 모두 약 1에 불과하였으나, 홍삼박 추출물 처리 24시간 후 캐스페이즈 -3은 약 15, 캐스페이즈 -8은 약 11, 캐스페이즈 -9는 약 13으로 현저히 증가하였다.The expression rates of caspase-3, -8 and -9 before treatment of red ginseng roots extract were all about 1, but after 24 hours of treatment with red ginseng roots extract, caspase-3 was about 15, caspase-8 was about 11, -9 was increased to about 13.
특히 캐스페이즈 효소 중에서도 세포자멸 유도에 가장 깊은 연관이 있는 캐스페이즈 -3의 발현율이 가장 높은 것을 관찰할 수 있었다.
Especially, the highest expression of caspase-3 was found to be most related to apoptosis induction among caspase enzymes.
도 3의 하단은 홍삼박 추출물의 처리 시간에 따른 HSC-3 세포의 캐스페이즈 효소 발현의 변화 양상을 나타낸 그래프이다.
3 is a graph showing changes in the expression of caspase enzyme in HSC-3 cells according to the treatment time of red ginseng root extract.
실험 결과, 홍삼박 추출물의 처리 농도가 증가할 수록 HSC-3 세포의 캐스페이즈 -3, -8 및 -9의 발현율은 모두 증가하는 것을 관찰할 수 있었다.As a result, it was observed that the expression levels of caspase-3, -8 and -9 in HSC-3 cells were increased with increasing concentration of red ginseng root extract.
홍삼박 추출물 처리 전 캐스페이즈-3, -8 및 -9의 발현율은 모두 약 1에 불과하였으나, 홍삼박 추출물 4mg/ml 처리 후 캐스페이즈-3은 약 11, 캐스페이즈 -8은 약 8.5, 캐스페이즈-9는 약 8로 현저히 증가하였다.The expression rates of caspase-3, -8 and -9 before treatment with red ginseng root extract were only about 1. However, after treatment with 4 mg / ml of red ginseng root extract, caspase-3 was about 11, caspase-8 was about 8.5, Phase-9 was significantly increased to about 8.
특히 홍삼박 추출물의 처리 농도에 따른 캐스페이즈 효소의 발현율을 알아보는 상기 실험에서도 캐스페이즈 -3의 발현율이 가장 높은 것을 관찰할 수 있었다.
In particular, it was observed that the expression rate of caspase-3 was the highest in the above-described experiment in which the expression level of caspase enzyme was determined by the treatment concentration of red ginseng root extract.
실험예Experimental Example
3. 3.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포에서의 분절 -3 Segments in cells
PARPPARP
효소 발현 촉진 Promoting enzyme expression
(1) 실험 과정(1) Experimental process
PARP 효소는 세포자멸을 억제하는 단백질로써, PARP 효소를 분절하여 불활성화하면 세포자멸가 유도된다.
PARP enzyme is a protein that inhibits apoptosis. PARP enzyme is induced to induce apoptosis by fragmenting and inactivating it.
1) 웨스턴 블롯(western blot)1) western blot
홍삼박 추출물을 HSC-3 세포에 처리하였을시 분절 PARP 효소에 미치는 영향을 알아보고자 하였다.To investigate the effect of red ginseng roots extract on segmental PARP enzyme treated with HSC-3 cells.
홍삼박 추출물의 농도를 각각 조정한 후, 농도를 조정한 각 홍삼박 추출물을 HSC-3 세포에 처리하여 웨스턴 블롯을 수행하였다. After adjusting the concentration of the red ginseng roots extract, each concentration of the red ginseng roots extract was treated with HSC-3 cells to perform Western blotting.
웨스턴 블롯은 통상적인 방법으로 수행되었다.
Western blotting was performed in a conventional manner.
(2) 실험 결과(2) Experimental results
도 4는 홍삼박 추출물을 처리하였을시 HSC-3 세포의 분절 PARP 효소 및 캐스페이즈-3 효소의 발현 촉진에 미치는 영향을 웨스턴 블롯을 수행하여 나타낸 사진이다.
4 is a photograph showing the effect of HSC-3 cells on the promotion of the expression of segmental PARP enzyme and caspase-3 enzyme when treated with red ginseng root extract by Western blotting.
실험 결과, HSC-3 세포의 분절 PARP 효소 및 캐스페이즈-3 효소의 밴드가 진하고 두께가 굵었으므로, 분절 PARP 효소 및 캐스페이즈-3 효소의 발현율이 높음을 관찰할 수 있었다.
As a result of the experiment, it was observed that the fraction of PARP and caspase-3 enzymes of HSC-3 cells were highly expressed and thus the PARP enzyme and the caspase-3 enzyme were highly expressed.
실험예Experimental Example
4. 4.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포에서의 세포 주기 확인 Cell Cycle Identification in Cells
(1) 실험 과정(1) Experimental process
1) 세포 주기 분석1) Cell cycle analysis
세포 주기는 subG1기, G0/G1기, S기 및 G2/M기의 4단계로 분류할 수 있다. The cell cycle can be divided into four stages: subG1, G0 / G1, S, and G2 / M.
그 중 subG1기는 자멸이 진행되는 세포에서만 발견되는 시기로써, 특정 세포의 주기를 상기 subG1기가 되도록 유도하면 특정 세포는 자멸된다.Among them, the subG1 group is found only in apoptotic cells. When a certain cell cycle is induced to be the subG1 group, certain cells are self-destructed.
또한 G2/M기는 분열이 활성화되는 시기이다.
Also, the G2 / M group is the period when the cleavage is activated.
홍삼박 추출물을 처리한 HSC-3 세포에서의 세포 주기를 확인하기 위하여, 홍삼박 추출물의 세포 주기 분석(cell cycle analysis)을 수행하였다.Cell cycle analysis of red ginseng roots extract was performed to confirm cell cycle in HSC-3 cells treated with red ginseng roots extract.
세포 주기 분석은 플로우 사이토메트리(flow cytometry) 기법을 사용하여 수행되었다.
Cell cycle analysis was performed using a flow cytometry technique.
① PI 염색① PI staining
홍삼박 추출물의 농도를 조정하여 HSC-3 세포에 처리한 후, 통상적인 방법으로 PI(propidium iodide) 염색을 수행하였다.
The concentration of red ginseng root extract was adjusted to treat HSC-3 cells, and PI (propidium iodide) staining was performed by a conventional method.
② 수치화② Quantification
PI 염색이 수행된 HSC-3 세포의 세포자멸률을 WinMDI 2.8 프로그램을 이용하여 수치화하였다.
Cellular apoptosis rate of HS-3 cells subjected to PI staining was quantified using WinMDI 2.8 program.
(2) 실험 결과(2) Experimental results
도 5는 홍삼박 추출물을 처리하였을시 HSC-3 세포의 세포자멸 유도에 미치는 영향을 플로우 사이토메트리를 이용하여 나타낸 그래프이다.
FIG. 5 is a graph showing the effect of HSC-3 cells on induction of apoptosis induced by red ginseng root extract using flow cytometry. FIG.
실험 결과, 홍삼박 추출물의 처리 농도를 증가시킬 수록 G2/M기의 HSC-3 세포가 감소하고 있음을 관찰할 수 있었다.As a result, it was observed that HSC-3 cells of G2 / M group decreased as the treatment concentration of red ginseng root extract was increased.
특히 홍삼박 추출물 1 mg/ml 처리군에서는 subG1기의 HSC-3 세포 비율이 현저히 증가하였음을 관찰할 수 있었다.
In particular, the ratio of HSC-3 cells in subG1 group was significantly increased in the group treated with 1 mg / ml of red ginseng extract.
도 6은 홍삼박 추출물을 처리하였을시 HSC-3 세포 주기의 변화 양상을 나타낸 표이다.
FIG. 6 is a table showing changes in HSC-3 cell cycle upon treatment with red ginseng root extract.
실험 결과, 홍삼박 추출물의 처리 농도를 증가시킬 수록 G0/G1기 및 G2/M기인 HSC-3 세포 비율이 감소함을 관찰할 수 있었다. As a result of the experiment, it was observed that the ratio of G0 / G1 and G2 / M HSC-3 cells decreased as the concentration of the red ginseng root extract was increased.
특히 홍삼박 추출물 1 mg/ml 처리군에서는 subG1기의 HSC-3 세포 비율이 현저히 증가하였음을 관찰할 수 있었다.
In particular, the ratio of HSC-3 cells in subG1 group was significantly increased in the group treated with 1 mg / ml of red ginseng extract.
실험예Experimental Example
5. 5.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포에서의 -3 cells
세포자멸률Cell apoptosis rate
확인 Confirm
(1) 실험 결과(1) Experimental results
도 7은 홍삼박 추출물을 처리하였을시 HSC-3 세포의 자멸률을 TUNEL assay로 나타낸 그래프이다.
FIG. 7 is a graph showing the suicide rate of HSC-3 cells treated with red ginseng root extract by TUNEL assay.
실험 결과, 홍삼박 추출물의 처리 농도를 증가시킬 수록 그래프 내에서의 HSC-3 세포의 early apop 지수 및 late apop 지수가 점점 상승하며, live 지수는 점점 하강함을 관찰할 수 있었다.
As the results of the experiment, it was observed that as the treatment concentration of red ginseng roots extract increased, the early apop index and late apop index of the HSC-3 cells in the graph gradually increased and the live index gradually decreased.
도 8은 홍삼박 추출물을 처리하였을시 HSC-3 세포의 밀도를 cells/ml, 자멸률을 %로 나타낸 표이다.
8 is a table showing the density of HSC-3 cells in cells / ml and the rate of self-destruction in% when red ginseng roots extract was treated.
실험 결과, 홍삼박 추출물의 처리 농도를 증가시킬 수록 HSC-3 세포의 밀도는 감소하고, 자멸률은 현저히 증가하는 것을 관찰할 수 있었다.As a result, the density of HSC-3 cells decreased and the rate of self-destruction increased with increasing concentration of red ginseng roots extract.
특히 자멸률은 홍삼박 추출물 0.0625 mg/ml 처리군에서 10.60%였으나, 홍삼박 추출물의 처리 농도를 증가시킬 수록 꾸준히 증가하여 홍삼박 추출물 1 mg/ml 처리군에서는 82.95%였다.
Especially, the rate of self - destruction was 10.60% in the group treated with 0.0625 mg / ml of red ginseng extract, but increased steadily with increasing concentration of red ginseng extract, which was 82.95% in the group treated with 1 mg / ml of red ginseng extract.
실험예Experimental Example
6. 6.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포에서의 -3 cells
BCLBCL
-2 발현 저해 및 프로--2 expression inhibition and pro-
아포토시스Apoptosis
단백질 발현 촉진 확인 Confirmation of protein expression promotion
(1) 실험 과정(1) Experimental process
Bcl-2는 세포자멸을 저해하는 역할을 하는 단백질이며, Bad, Bax 및 Bim 등의 프로-아포토시스 단백질은 세포자멸을 촉진하는 역할을 한다.
Bcl-2 is a protein that inhibits apoptosis, and pro-apoptotic proteins such as Bad, Bax, and Bim play a role in promoting apoptosis.
홍삼박 추출물을 HSC-3 세포에 처리하였을시 Bcl-2의 발현과, Bad, Bax 및 Bim을 포함하는 프로-아포토시스 단백질의 발현에 미치는 영향을 알아보고자 하였다.
To investigate the effect of red ginseng roots extract on the expression of Bcl-2 and the expression of pro-apoptotic proteins including Bad, Bax and Bim when treated with HSC-3 cells.
1) 홍삼박 추출물 처리 시간 조정 및 웨스턴 블롯1) Treatment time of red ginseng extract and western blotting
홍삼박 추출물 0.25 mg/ml를 HSC-3 세포에 처리하고 각각 설정된 시간이 경과한 후, 웨스턴 블롯을 수행하였다. The HSC-3 cells were treated with 0.25 mg / ml of red ginseng extract and subjected to Western blotting after a predetermined time had elapsed.
웨스턴 블롯은 통상적인 방법으로 수행되었다.
Western blotting was performed in a conventional manner.
2) 홍삼박 추출물 농도 조정 및 웨스턴 블롯2) Concentration adjustment of red ginseng roots extract and western blotting
홍삼박 추출물의 농도를 각각 조정한 후, 각 홍삼박 추출물을 HSC-3 세포에 처리하여 웨스턴 블롯을 수행하였다. After adjusting the concentration of the red ginseng roots extract, each ginseng roots extract was treated with HSC-3 cells to perform Western blotting.
웨스턴 블롯은 통상적인 방법으로 수행되었다.
Western blotting was performed in a conventional manner.
(2) 실험 결과(2) Experimental results
도 9는 홍삼박 추출물을 HSC-3 세포에 처리하였을시 시간의 경과에 따른 Bcl-2 및 프로-아포토시스 단백질의 발현의 변화 양상을 웨스턴 블롯을 수행하여 나타낸 사진이다.
FIG. 9 is a photograph showing Western blotting of changes in expression of Bcl-2 and pro-apoptotic proteins over time when HSC-3 cells were treated with red ginseng root extract.
실험 결과, Bcl-2은 홍삼박 추출물 처리 후 시간이 경과함에 따라 밴드가 점점 옅어지는 것을 관찰할 수 있었다. As a result of the experiment, it was observed that the bands gradually became thinner with time after treatment of the red ginseng roots extract with Bcl-2.
반면 Bad, Bim 및 Bax는 홍삼박 추출물 처리 후 시간이 경과함에 따라 밴드가 진해지는 것을 관찰할 수 있었다.
On the other hand, Bad, Bim and Bax were observed to be thickened with time after treatment with red ginseng extract.
도 10은 홍삼박 추출물을 농도별로 HSC-3 세포에 처리하였을시 HSC-3 세포의 Bcl-2 및 프로-아포토시스 단백질의 발현에 미치는 영향을 웨스턴 블롯을 수행하여 나타낸 사진이다.
FIG. 10 is a photograph showing Western blotting effect of HSC-3 cells on the expression of Bcl-2 and pro-apoptotic proteins in HSC-3 cells treated with various concentrations of red ginseng root extract.
실험 결과, Bcl-2는 홍삼박 추출물의 처리 농도가 높아질 수록 밴드가 점점 옅어지는 것을 관찰할 수 있었다.As a result, the band of Bcl-2 was gradually decreased as the treatment concentration of red ginseng extract increased.
반면 Bad, Bim 및 Bax는 홍삼박 추출물의 처리 농도가 높아질 수록 밴드가 점점 짙어지는 것을 관찰할 수 있었다.
On the other hand, in the case of Bad, Bim and Bax, it was observed that as the treatment concentration of the red ginseng extract increased, the band became thicker.
실험예Experimental Example
7. 7.
홍삼박Red ginseng
추출물을 처리한 Extract-treated
HSCHSC
-3 세포에서의 -3 cells
MAPKMAPK
통로 단백질 발현 촉진 확인 Confirming Promotion of Pathway Protein Expression
(1) 실험 과정(1) Experimental process
ERK, pERK, JNK, pJNK, p38 및 pp38을 포함하는 MAPK 통로 단백질은 세포자멸을 유도하는 역할을 한다.MAPK pathway proteins including ERK, pERK, JNK, pJNK, p38 and pp38 play a role in inducing apoptosis.
홍삼박 추출물을 HSC-3 세포에 처리하였을시 ERK, pERK, JNK, pJNK, p38, pp38을 포함하는 MAPK 통로 단백질의 발현에 미치는 영향을 알아보고자 하였다.
To investigate the effect of red ginseng roots extract on the expression of MAPK pathway proteins including ERK, pERK, JNK, pJNK, p38, and pp38 when treated with HSC-3 cells.
1) 홍삼박 추출물 처리 시간 조정 및 웨스턴 블롯1) Treatment time of red ginseng extract and western blotting
홍삼박 추출물 0.25 mg/ml를 HSC-3 세포에 처리하고 각각 설정된 시간이 경과한 후, 웨스턴 블롯을 수행하였다. The HSC-3 cells were treated with 0.25 mg / ml of red ginseng extract and subjected to Western blotting after a predetermined time had elapsed.
웨스턴 블롯은 통상적인 방법으로 수행되었다.
Western blotting was performed in a conventional manner.
2) 홍삼박 추출물 농도 조정 및 웨스턴 블롯2) Concentration adjustment of red ginseng roots extract and western blotting
홍삼박 추출물의 농도를 각각 조정한 후, 각 홍삼박 추출물을 HSC-3 세포에 처리하여 웨스턴 블롯을 수행하였다. After adjusting the concentration of the red ginseng roots extract, each ginseng roots extract was treated with HSC-3 cells to perform Western blotting.
웨스턴 블롯은 통상적인 방법으로 수행되었다.
Western blotting was performed in a conventional manner.
(2) 실험 결과(2) Experimental results
도 11은 홍삼박 추출물을 HSC-3 세포에 처리하였을시 시간의 경과에 따른 MAPK 통로 단백질 발현의 변화 양상을 웨스턴 블롯을 수행하여 나타낸 사진이다.
11 is a photograph showing Western blotting of changes in MAPK channel protein expression over time when HSC-3 cells were treated with red ginseng root extract.
실험 결과, 홍삼박 추출물을 처리한 HSC-3 세포에서의 ERK, pERK, JNK, pJNK, p38 및 pp38은 모두 밴드가 진한 것을 관찰할 수 있었다. As a result, ERK, pERK, JNK, pJNK, p38 and pp38 in the HSC-3 cells treated with the extract of red ginseng roots were observed to be rich in band.
특히 pJNK는 처리 후 시간이 경과함에 따라 밴드가 점점 진해지는 것을 관찰할 수 있었다.
Especially, pJNK was observed to gradually increase in band as time elapsed after treatment.
도 12는 홍삼박 추출물을 농도별로 HSC-3 세포에 처리하였을시 농도에 따른 MAPK 통로 단백질 발현의 변화 양상을 웨스턴 블롯을 수행하여 나타낸 사진이다.
FIG. 12 is a photograph showing Western blotting of changes in MAPK pathway protein expression according to the concentration of HSC-3 cells treated with the extract of red ginseng root at various concentrations.
실험 결과, 농도별로 홍삼박 추출물을 처리한 HSC-3 세포에서의 ERK, pERK, JNK, pJNK, p38 및 pp38은 모두 밴드가 진한 것을 관찰할 수 있었다.As a result, it was observed that ERK, pERK, JNK, pJNK, p38 and pp38 in the HSC-3 cells treated with the extract of red ginseng roots were rich in band.
특히 pJNK는 처리 농도가 증가함에 따라 밴드가 점점 진해지는 것을 관찰할 수 있었다. Especially, pJNK could be observed to increase gradually as the treatment concentration increased.
Claims (5)
A composition for preventing, ameliorating or treating oral squamous cell cancer, which contains red ginseng extract as an active ingredient.
상기 조성물에 함유되는 홍삼 가공 추출물의 농도는 구강편평세포암의 세포주인 HSC-3 세포 기준으로 0.06~4 mg/ml인 것을 특징으로 하는 조성물.
The method according to claim 1,
Wherein the concentration of the ginseng processing extract contained in the composition is 0.06 to 4 mg / ml based on the HSC-3 cell line of the oral squamous cell carcinoma.
상기 조성물은 HSC-3 세포를 사멸시키는 효능, HSC-3 세포의 캐스페이즈(caspase) -3, -8 및 -9의 발현을 촉진하는 효능, HSC-3 세포의 분절 PARP의 발현을 촉진하는 효능, HSC-3 세포의 주기가 세포자멸(apoptosis)기로 진행되는 것을 유도하는 효능, HSC-3 세포의 Bcl-2의 발현을 저해하는 효능, HSC-3 세포의 프로-아포토시스(pro-apoptosis) 단백질의 발현을 촉진하는 효능 및 HSC-3 세포의 MAPK 통로 단백질의 발현을 촉진하는 효능을 보유하는 것을 특징으로 하는 조성물.
The method according to claim 1 or 2,
The composition is effective in killing HSC-3 cells, promoting the expression of caspase-3, -8 and -9 of HSC-3 cells, and enhancing the expression of segmental PARP in HSC-3 cells , An effect of inducing the cycle of HSC-3 cells to progress to apoptosis, an effect of inhibiting the expression of Bcl-2 of HSC-3 cells, a pro-apoptosis protein of HSC-3 cells And promoting the expression of the MAPK channel protein of HSC-3 cells.
A health functional food for preventing or improving oral squamous cell carcinoma, to which a food additive additive is added to the composition according to any one of claims 1 to 3.
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|---|---|---|---|---|
| KR20000069452A (en) | 1996-12-13 | 2000-11-25 | 피터 지. 스트링거 | Leukotriene Antagonists for Oral Squamous Cell Carcinoma |
| KR100485936B1 (en) * | 2002-10-23 | 2005-04-27 | 주식회사 바이오사포젠 | Anticarcinogenic constituents of ginsenoside Rh2 and Rg3 |
| KR20090053091A (en) | 2007-11-22 | 2009-05-27 | 주식회사 비지오코프풍정 | Side mirror having puddle lamp with function of rear view lamp |
| KR20100098208A (en) * | 2009-02-27 | 2010-09-06 | 한국생명공학연구원 | Black ginseng with increased rg3 and rh2 contents and process for preparing the same |
| KR20130056701A (en) | 2011-11-22 | 2013-05-30 | 주식회사 피비에스 | Composition for antiinflammatory, antibacterial and removing stench comprising red ginseng extracts and extracts derived from natural materials |
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| KR20000069452A (en) | 1996-12-13 | 2000-11-25 | 피터 지. 스트링거 | Leukotriene Antagonists for Oral Squamous Cell Carcinoma |
| KR100485936B1 (en) * | 2002-10-23 | 2005-04-27 | 주식회사 바이오사포젠 | Anticarcinogenic constituents of ginsenoside Rh2 and Rg3 |
| KR20090053091A (en) | 2007-11-22 | 2009-05-27 | 주식회사 비지오코프풍정 | Side mirror having puddle lamp with function of rear view lamp |
| KR20100098208A (en) * | 2009-02-27 | 2010-09-06 | 한국생명공학연구원 | Black ginseng with increased rg3 and rh2 contents and process for preparing the same |
| KR20130056701A (en) | 2011-11-22 | 2013-05-30 | 주식회사 피비에스 | Composition for antiinflammatory, antibacterial and removing stench comprising red ginseng extracts and extracts derived from natural materials |
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