KR102551027B1 - Antiviral composition comprising extracts of Phellinus baumii - Google Patents
Antiviral composition comprising extracts of Phellinus baumii Download PDFInfo
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- A—HUMAN NECESSITIES
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Abstract
본 발명은 장수진흙버섯 추출물을 유효성분으로 포함하는 항바이러스용 조성물에 관한 것으로, 본 발명의 장수진흙버섯 추출물은 세포독성 없이 콕사키바이러스(CVB3), 인간 라이노바이러스(HRV1B), 인플루엔자바이러스(Inf/PR8) 및 인간 코로나바이러스(coronavirus 229E)에 대한 항바이러스 효과를 나타내므로, 장수진흙버섯 추출물을 포함하는 조성물을 항바이러스제 및 바이러스성 질환 예방, 치료 또는 개선 용도로 이용할 수 있다.The present invention relates to an antiviral composition comprising a longevity mud mushroom extract as an active ingredient, and the longevity mud mushroom extract of the present invention is free from coxsackievirus (CVB3), human rhinovirus (HRV1B), and influenza virus (Inf /PR8) and human coronavirus (coronavirus 229E), the composition containing the longevity mud mushroom extract can be used as an antiviral agent and for preventing, treating or improving viral diseases.
Description
본 발명은 장수진흙버섯 추출물을 유효성분으로 포함하는 항바이러스용 조성물 및 바이러스성 질환 예방, 치료 또는 개선용 조성물에 관한 것이다.The present invention relates to an antiviral composition comprising a longevity mud mushroom extract as an active ingredient and a composition for preventing, treating, or improving viral diseases.
콕사키바이러스(coxsackievirus) 및 라이노바이러스(Rhinovirus)는 피코르나비리대과(피코르나바이러스과; picornavirus) 엔테로바이러스속(Enterovirus)에 속하는 바이러스로 지름이 27~30 nm 크기의 아주 작은 바이러스이다.Coxsackievirus and Rhinovirus are viruses belonging to the genus Enterovirus of the Picornaviridae family (Picornaviridae) and are very small viruses with a diameter of 27 to 30 nm.
엔테로바이러스는 주로 하절기에 유ㆍ소아 층에 침범하여 감염자의 면역학적 특성에 따라 중증감염을 초래하기도 한다. 특히 위생이 나쁜 환경에서 흔하게 전파되는 전염성 병원체로써 감염경로는 분변-구강이며, 오염된 물과 토양을 통한 경구적인 전파로 이어진다. 감염자에서 배출된 분변속의 바이러스가 오수나 폐수를 통해 지하수, 하천, 해수로 흘러들어가 다시 사람에게로 전염되는데, 드물게는 호흡기 분비물을 통해서도 감염 된다. 엔테로바이러스는 소화기를 통해 감염된 후 인후두 부위나 소장의 림프절에서 일차적으로 증식한 후 신체의 각 장기로 이동 한다. 임상증상은 경증질환으로는 발진, 감기, 근육통, 불명열등을 유발하고 심각한 질환으로는 뇌수막염, 뇌염, 간염, 췌장염, 늑막염, 심근염, 심외막염, 심근증을 유발한다.Enterovirus mainly invades infants and children during the summer season and may cause severe infection depending on the immunological characteristics of the infected person. In particular, it is a contagious pathogen that is commonly transmitted in poor sanitary environments, and the route of infection is fecal-oral, leading to oral transmission through contaminated water and soil. The virus in the feces discharged from the infected person flows into groundwater, rivers, and seawater through sewage or wastewater and is transmitted to humans again. In rare cases, it is also infected through respiratory secretions. After being infected through the digestive tract, enterovirus primarily proliferates in the pharynx or in the lymph nodes of the small intestine, then moves to each organ of the body. Clinical symptoms cause rash, cold, muscle pain, fever of unknown origin as mild diseases, and meningitis, encephalitis, hepatitis, pancreatitis, pleurisy, myocarditis, pericarditis, and cardiomyopathy as serious diseases.
콕사키바이러스는 뉴욕주의 콕사키에서 감염된 어린이의 대변에서 분리되었으며, 엔테로바이러스속에 속하는 바이러스로, 입자는 지름 27~28nm의 정20면체로 유전체는 7,401염기의 (＋)사슬 RNA이며, 경구적으로 감염된다. 콕사키바이러스는 콕사키바이러스 A와 콕사이바이러스 B의 두가지 그룹으로 존재하며, 콕사키바이러스 A는 이완마비, B는 경련성마비를 일으킨다. 콕사키바이러스 B3(coxsackievirus B3; CVB3)는 인간뿐 아니라 마우스에서도 감염이 되는 것으로 알려져 있으며, 간, 심장, 뇌, 척수 및 췌장에 감염되는 것으로 알려져 있다. 유발 질환으로는, 수막염, 근육염, 심근염 마비 등이 있으며, 특히, 콕사키바이러스 B는 심각할 경우 심장이식까지 필요한 특발성 확장성 심근증(idiopathic dilated cardiomyopathy)인 심근염(myocarditis)의 원인으로 알려져 있다. 현재, 효과적인 치료제는 아직 개발되지 않았다.Coxsackievirus was isolated from feces of infected children in Coxsackie, New York. It is a virus belonging to the genus Enterovirus. Infected. Coxsackieviruses exist in two groups, coxsackievirus A and coxsackievirus B. Coxsackievirus A causes flaccid paralysis and B spastic paralysis. Coxsackievirus B3 (CVB3) is known to infect humans as well as mice, and is known to infect the liver, heart, brain, spinal cord and pancreas. Inducing diseases include meningitis, myositis, myocarditis, paralysis, and the like. In particular, coxsackievirus B is known to cause myocarditis, an idiopathic dilated cardiomyopathy that requires heart transplantation in severe cases. Currently, no effective treatment has yet been developed.
인간 라이노바이러스(human rhinovirus)는 사람 대 사람 접촉을 통하여 전염되는 것이 가장 흔하고 때로는 손에 묻은 오염된 분비물에 의하여 자가접종됨으로써 전염될 수 있다. 연중 발생하지만 섭씨 33~35℃에서 가장 잘 증식하는 병원체의 특성상 가을부터 봄에 이르는 기간에 계절성 역학을 보이고, 성인과 소아에서 피로감, 두통, 근육통, 미열 등의 비교적 경한 증상을 보이는 상기도 감염증의 주된 원인 바이러스로 알려져 있었다. 최근에는 기관지 과민성을 가진 환자에서 천명 및 천식 병증의 악화요인 중 하나일 뿐만 아니라 만성 폐쇄성 폐질환의 악화 원인으로도 알려져 왔다.Human rhinovirus is most commonly transmitted through person-to-person contact and can sometimes be transmitted by self-inoculation by contaminated secretions on the hands. Although it occurs year-round, due to the nature of the pathogen, which proliferates best at 33-35 ° C, it shows seasonal dynamics from autumn to spring and shows relatively mild symptoms such as fatigue, headache, muscle pain, and low-grade fever in adults and children. known to be the main causative virus. Recently, it has been known to be one of the exacerbating factors of wheezing and asthma in patients with bronchial hypersensitivity as well as an exacerbating cause of chronic obstructive pulmonary disease.
인플루엔자바이러스(influenza virus)는 오르토믹소비리대과(오르토믹소바이러스과; Orthomyxovirus)의 RNA 바이러스로서 전염성 호흡기 질병을 유발하며, 이러한 바이러스로 인한 호흡기 질병의 종류로는 폐기종, 만성기관지염, 기관지선종, 고립성폐결절, 폐결핵, 농흉, 폐농양, 감기, 독감 및 폐의 조직구 증식증 등이 있다. 상기 인플루엔자바이러스는 A, B 및 C형으로 분류되며, A형 바이러스의 경우 사람에게 주로 감염이 확인되는 바이러스이며, 매년 전세계적으로 독감 유행을 일으켜 높은 비율로 감염 환자를 발생시키며 특히 노인과 영유아, 각종 호흡기 또는 내과 질환이 있는 환자의 사망률을 높이고 있다. Influenza virus is an RNA virus of the Orthomyxoviridae family (Orthomyxovirus) and causes infectious respiratory diseases. , pulmonary tuberculosis, empyema, lung abscess, cold, flu, and histiocytosis of the lung. The influenza virus is classified into types A, B, and C. Type A virus is a virus that is mainly infected with humans, and causes influenza epidemics worldwide every year to cause a high rate of infected patients, especially the elderly and infants, The mortality rate of patients with various respiratory or internal diseases is increasing.
코로나바이러스(coronavirus)는 외피가 있는 단일가닥의 양성 RNA 바이러스이며 게놈 크기는 25-32kb로 지금까지 알려진 RNA 바이러스 중에서는 비교적 큰 바이러스에 속한다. 외피에 곤봉모양의 돌기인 스파이크(spike) 단백질이 박혀 있어 화염 또는 왕관 모양의 특이적 구조를 가지고 있고, 라틴어 Corona로부터 바이러스 이름이 유래되었다고 한다. 사람에게 감염되는 인간 코로나바이러스는 1960대 발견된 HCoV-229E와 HCoV-OC43, 사스 대유행 이후 발견된 HCoV-NL63(2004년)과 HCoV-HKU1(2005년) 등이 알려져있다. 최근의 코로나바이러스감염증-19(coronavirus disease 2019, COVID-19) 바이러스는 2019년 중국 후베이성의 우한시에서 처음 발견된 코로나바이러스(Coroviridae) 군에 속하는 바이러스이다. 이들로 인한 증상은 일반적으로 상기도감염증과 관계가 있으며, 발열을 동반한 기침, 호흡곤란, 숨가쁨, 가래 등 호흡기 증상을 주로 보이고, 이외에도 두통, 오한, 콧물, 근육통뿐만 아니라 식욕부진, 메스꺼움, 구토, 복통, 설사 등 소화기 증상도 나타날 수 있다.Coronavirus is an enveloped, single-stranded positive RNA virus with a genome size of 25-32 kb, which is one of the relatively large RNA viruses known so far. It is said that the spike protein, a club-shaped protrusion, is embedded in the outer coat, so it has a specific flame or crown-shaped structure, and the virus name is derived from the Latin word Corona. Human coronaviruses that infect humans are known, such as HCoV-229E and HCoV-OC43 discovered in the 1960s, and HCoV-NL63 (2004) and HCoV-HKU1 (2005) discovered after the SARS pandemic. The recent coronavirus disease 2019 (COVID-19) virus is a virus belonging to the family of coronaviruses (Coroviridae) first discovered in 2019 in Wuhan City, Hubei Province, China. Symptoms caused by these are generally related to upper respiratory infections, and mainly show respiratory symptoms such as fever accompanied by cough, shortness of breath, shortness of breath, and phlegm. Gastrointestinal symptoms such as abdominal pain and diarrhea may also occur.
이러한 각종 바이러스는 인체에서 각종 질환을 유발시키는 임상적으로 매우 중요한 바이러스임에도 불구하고 치료법에 관한 연구는 거의 없고 치료법 역시 아직 확정되어 있지 않다. 현재까지 바이러스 질환의 치료방법은 백신에 의한 예방과 극소수의 약제만 존재하나, 바이러스 내성의 증가로 인하여 사용에 제한이 있다. 그러므로 최근에는 안전성과 보다 높은 효과, 그리고 저렴한 생산비용을 갖는 항바이러스제제들이 개발 중이지만 아직도 많은 바이러스들에서 치료 효과를 갖는 제제는 없다.Although these various viruses are clinically very important viruses that cause various diseases in the human body, there is little research on treatment methods, and treatment methods have not yet been confirmed. Until now, only vaccine prevention and very few drugs exist as treatment methods for viral diseases, but their use is limited due to increased viral resistance. Therefore, in recent years, antiviral agents having safety, higher efficacy, and low production cost are being developed, but there is still no agent having a therapeutic effect on many viruses.
이에, 본 발명자들은 부작용과 내성의 잠재성이 적고 생산비용이 낮은 약용식물을 기반으로 한 항바이러스 제제를 개발하기 위하여 연구한 결과, 장수진흙버섯 추출물이 항바이러스 활성을 나타냄을 확인하여 본 발명을 완성하였다.Accordingly, the present inventors studied to develop an antiviral preparation based on medicinal plants with low potential for side effects and resistance and low production cost, and as a result, it was confirmed that the longevity mushroom extract exhibited antiviral activity, and the present invention was achieved. completed.
본 발명의 목적은 항바이러스용 조성물을 제공하는 것이다.An object of the present invention is to provide an antiviral composition.
본 발명의 다른 목적은 바이러스성 질환 예방 또는 치료용 약학 조성물을 제공하는 것이다.Another object of the present invention is to provide a pharmaceutical composition for preventing or treating viral diseases.
본 발명의 또 다른 목적은 바이러스성 질환 예방 또는 개선용 식품 조성물을 제공하는 것이다.Another object of the present invention is to provide a food composition for preventing or improving viral diseases.
상기 목적을 달성하기 위하여,In order to achieve the above purpose,
본 발명은 장수진흙버섯(Phellinus baumii) 추출물을 유효성분으로 포함하는 항바이러스용 조성물을 제공한다.The present invention provides an antiviral composition comprising an extract of longevity mud mushroom ( Phellinus baumii ) as an active ingredient.
일 구현예에서, 상기 항바이러스용 조성물은 유효성분으로서 장수진흙버섯 추출물을 포함하므로, 피코르나바이러스(피코르나비리대과, picornaviridae), 오르토믹소바이러스(오르토믹소비리대과, Orthomyxoviridae) 및 코로나바이러스(코로나비리대과, Coronaviridae)로 이루어진 군으로부터 선택된 하나 이상의 바이러스에 대한 항바이러스 활성을 가지는 것일 수 있다.In one embodiment, since the antiviral composition contains the longevity mud mushroom extract as an active ingredient, picornavirus (picornaviridae, picornaviridae), orthomyxovirus (orthomyxoviridae, Orthomyxoviridae) and corona It may have antiviral activity against one or more viruses selected from the group consisting of viruses (Coronaviridae).
일 구현예에서, 상기 바이러스는 콕사키바이러스(coxsackievirus), 라이노바이러스(rhinovirus), 인플루엔자바이러스(influenza virus) 및 알파 코로나바이러스(alpha coronavirus)로 이루어진 군으로부터 선택된 하나 이상의 바이러스일 수 있다.In one embodiment, the virus may be one or more viruses selected from the group consisting of coxsackievirus, rhinovirus, influenza virus and alpha coronavirus.
또한, 본 발명은 장수진흙버섯 추출물을 포함하는 바이러스성 질환 예방 또는 치료용 약학 조성물을 제공한다.In addition, the present invention provides a pharmaceutical composition for preventing or treating viral diseases comprising the longevity mud mushroom extract.
나아가, 본 발명은 장수진흙버섯 추출물을 포함하는 바이러스성 질환 예방 또는 개선용 식품 조성물을 제공한다.Furthermore, the present invention provides a food composition for preventing or improving viral diseases comprising the longevity mud mushroom extract.
일 구현예에서, 상기 바이러스성 질환은 피코르나바이러스(피코르나비리대과, picornaviridae), 오르토믹소바이러스(오르토믹소비리대과, Orthomyxoviridae) 및 코로나바이러스(코로나비리대과, Coronaviridae)로 이루어진 군으로부터 선택된 하나 이상의 바이러스로 인하여 유발되는 질환일 수 있고, 바람직하게는, 콕사키바이러스(coxsackievirus), 라이노바이러스(rhinovirus), 인플루엔자바이러스(influenza virus) 및 알파 코로나바이러스(alpha coronavirus)로 이루어진 군으로부터 선택된 하나 이상의 바이러스로 인하여 유발되는 질환일 수 있다.In one embodiment, the viral disease is from the group consisting of picornavirus (picornaviridae), orthomyxovirus (Orthomyxoviridae) and coronavirus (Coronaviridae). It may be a disease caused by one or more selected viruses, preferably one selected from the group consisting of coxsackievirus, rhinovirus, influenza virus and alpha coronavirus It may be a disease caused by more than one virus.
일 구현예에서, 상기 바이러스성 질환은 바이러스성 수막염, 간염, 근육염, 심근염, 심근증, 췌장염, 당뇨, 유행성 근육통, 뇌염, 감기, 독감, 천식, 만성 폐쇄성 폐질환, 폐렴, 상기도감염증, 폐기종, 만성기관지염, 기관지선종, 고립성폐결절, 폐결핵, 농흉, 폐농양, 폐의 조직구 증식증, 축농증, 중이염, 소아마비, 포진성 구협염, 구제역, 급성출혈성 결막염, 수족구병 및 수포병으로 이루어진 군에서 선택된 하나 이상일 수 있다.In one embodiment, the viral disease is viral meningitis, hepatitis, myositis, myocarditis, cardiomyopathy, pancreatitis, diabetes, mumps, encephalitis, cold, flu, asthma, chronic obstructive pulmonary disease, pneumonia, upper respiratory tract infection, emphysema, chronic At least one day selected from the group consisting of bronchitis, bronchial adenoma, solitary pulmonary nodule, pulmonary tuberculosis, empyema, lung abscess, lung histiocytosis, sinusitis, otitis media, poliomyelitis, stomatitis herpes, foot-and-mouth disease, acute hemorrhagic conjunctivitis, hand-foot-and-mouth disease, and bullous disease. can
본 발명의 장수진흙버섯(Phellinus baumii) 추출물은 세포독성 없이 콕사키바이러스 B3형(coxsackievirus B3; CVB3), 인간 라이노바이러스 1B형(human rhinovirus 1B; HRV1B), 인플루엔자바이러스 A형 H1N1(influenza virus A/H1N1/PR8; Inf/PR8) 및 인간 코로나바이러스 229E(human coronavirus 229E)에 대하여 항바이러스 효과를 나타내므로, 상기 장수진흙버섯 추출물을 포함하는 조성물을 항바이러스제 및 바이러스성 질환 예방, 치료 또는 개선 용도로 이용할 수 있다.The longevity mud mushroom ( Phellinus baumii ) extract of the present invention is coxsackievirus B3 type (CVB3), human rhinovirus type 1B (HRV1B), influenza virus type A H1N1 (influenza virus A/ H1N1 / PR8; Available.
도 1은 본 발명의 장수진흙버섯 추출물의 콕사키바이러스 B3형(CVB3)에 대한 항바이러스 활성을 확인하기 위하여, 세포 대조구(Ctrl)와 바이러스 대조구(Veh) 및 장수진흙버섯 추출물 처리 농도에 따른 vero 세포의 감염후 세포 생존율(cell viability, %)을 측정한 결과를 나타낸 그래프이다.
도 2는 본 발명의 장수진흙버섯 추출물의 인간 라이노바이러스 1B형(HRV1B)에 대한 항바이러스 활성을 확인하기 위하여, 장수진흙버섯 추출물 처리 농도에 따른 hela 세포의 감염후 세포 생존율(cell viability, %)을 측정한 결과를 나타낸 그래프이다.
도 3은 본 발명의 장수진흙버섯 추출물의 인플루엔자바이러스 A형(Inf/PR8)에 대한 항바이러스 활성을 확인하기 위하여, 세포 대조구(Ctrl)와 바이러스 대조구(Veh) 및 장수진흙버섯 추출물 처리 농도에 따른 A549 세포의 감염후 세포 생존율(cell viability, %)을 측정한 결과를 나타낸 그래프이다.
도 4는 본 발명의 장수진흙버섯 추출물의 인간 알파 코로나바이러스(coronavirus 229E)에 대한 항바이러스 활성을 확인하기 위하여, 세포 대조구(Ctrl)와 바이러스 대조구(Veh) 및 장수진흙버섯 추출물 처리 농도에 따른 Huh7 세포의 감염후 세포 생존율(cell viability, %)을 측정한 결과를 나타낸 그래프이다.
도 5는 본 발명의 장수진흙버섯 추출물의 세포독성을 확인하기 위하여, 장수진흙버섯 추출물 처리 농도에 따른 A549 세포, Huh7 세포, hela 세포 및 vero 세포의 세포 생존율(cell viability, %)을 측정한 결과를 나타낸 그래프이다.1 is a cell control (Ctrl), a virus control (Veh) and vero according to the treatment concentration of the longevity mud mushroom extract in order to confirm the antiviral activity of the longevity mud mushroom extract of the present invention against coxsackie virus type B3 (CVB3) It is a graph showing the results of measuring cell viability (%) after infection of cells.
Figure 2 is a cell viability (cell viability, %) after infection of hela cells according to the treatment concentration of the longevity mud mushroom extract to confirm the antiviral activity against human rhinovirus type 1B (HRV1B) of the longevity mud mushroom extract of the present invention. It is a graph showing the result of measuring .
3 is a cell control (Ctrl), a virus control (Veh), and a treatment concentration of the longevity mud mushroom extract in order to confirm the antiviral activity of the longevity mushroom extract of the present invention against influenza virus type A (Inf/PR8). It is a graph showing the results of measuring the cell viability (%) of A549 cells after infection.
Figure 4 is a cell control (Ctrl), a virus control (Veh) and Huh7 according to the treatment concentration of the longevity mushroom extract to confirm the antiviral activity of the longevity mushroom extract of the present invention against human alpha coronavirus (coronavirus 229E). It is a graph showing the results of measuring cell viability (%) after infection of cells.
Figure 5 is a result of measuring the cell viability (%) of A549 cells, Huh7 cells, hela cells and vero cells according to the treatment concentration of the longevity mud mushroom extract in order to confirm the cytotoxicity of the longevity mud mushroom extract of the present invention. is a graph showing
이하, 본 발명을 상세히 설명한다.Hereinafter, the present invention will be described in detail.
본 발명의 일 측면에서, 장수진흙버섯(Phellinus baumii) 추출물을 유효성분으로 포함하는 항바이러스용 조성물을 제공한다.In one aspect of the present invention, there is provided an antiviral composition comprising a longevity mud mushroom ( Phellinus baumii ) extract as an active ingredient.
본 발명의 '장수진흙버섯(Phellinus baumii)'은 뽕나무, 느릅나무, 사시나무, 오리나무 등의 활엽수의 심재부에 자생하는 버섯으로 소나무비늘버섯과(Hymenochaetaceae)의 진흙버섯속(Phellinus Ouel. Em.lmaz)에 속하는 백색부후 균이다. 장수진흙버섯은 옛날부터 자궁출혈, 월경불순 등의 부인병치료제로 사용되어 왔으며 최근에는 높은 항암 효과 때문에 각광을 받고 있다.The 'longevity mud mushroom ( Phellinus baumii )' of the present invention is a mushroom that grows wild in the heartwood of broad-leaved trees such as mulberry, elm, aspen, alder, etc. It is a white rot fungus belonging to .lmaz). Longevity mud mushroom has been used as a treatment for gynecological diseases such as uterine bleeding and menstrual irregularity since ancient times, and has recently been in the limelight due to its high anticancer effect.
본 발명에 있어서, 상기 장수진흙버섯은 재배한 것 또는 시판되는 것 등 제한 없이 사용할 수 있으며, 장수진흙버섯의 자실체 및 균사체 모두를 이용하는 것일 수 있고, 바람직하게는 자실체를 이용하는 것일 수 있으나, 이에 제한되지는 않는다.In the present invention, the longevity mud mushroom can be used without limitation, such as cultivated or commercially available, and may use both fruiting bodies and mycelia of longevity mud mushrooms, preferably fruiting bodies, but limited thereto It doesn't work.
본 발명에서 사용되는 용어 "추출물(extract)"은 생약을 적절한 침출액으로 짜내고 침출액을 증발시켜 농축한 제제를 의미하는 것으로, 이에 제한되지는 않으나, 추출 처리에 의해 얻어지는 추출액, 추출액의 희석액 또는 농축액, 추출액을 건조하여 얻어지는 건조물, 이들의 조정제물 또는 정제물일 수 있다. 상기 추출물은 통상의 기술분야에 공지된 일반적인 추출방법, 분리 및 정제방법을 이용하여 제조할 수 있다. 상기 추출방법으로는, 이에 제한되지는 않으나, 바람직하게 열탕 추출, 열수 추출, 냉침 추출, 환류 냉각 추출 또는 초음파 추출 등의 방법을 사용할 수 있다. The term "extract" used in the present invention refers to a preparation obtained by squeezing a crude drug into an appropriate leachate and evaporating the leachate to concentrate, but is not limited thereto, such as, but not limited to, an extract obtained by extraction treatment, a dilution or concentrate of the extract, It may be a dried product obtained by drying the extract, a product of a crude product or a purified product thereof. The extract can be prepared using a general extraction method, separation and purification method known in the art. The extraction method is not limited thereto, but preferably, methods such as hot water extraction, hot water extraction, cold needle extraction, reflux cooling extraction, or ultrasonic extraction may be used.
본 발명에 있어서, 상기 장수진흙버섯 추출물은 추출용매로 추출하거나 추출용매로 추출하여 제조한 추출물에 분획용매를 가하여 분획함으로써 제조할 수 있다. 상기 추출용매는 이에 제한되지 않으나, 물, 유기용매 또는 이들의 혼합용매 등을 사용할 수 있으며, 상기 유기용매는 탄소수 1 내지 4의 알코올이나, 에틸아세테이트 또는 아세톤 등의 극성용매, 헥산 또는 디클로로메탄의 비극성용매 또는 이들의 혼합용매를 사용할 수 있다. 또한, 바람직하게는 물, 탄소수 1 내지 탄소수 4의 알코올 또는 이들의 혼합용매를 사용할 수 있으며, 보다 바람직하게는 에탄올을 사용할 수 있다. In the present invention, the longevity mud mushroom extract can be prepared by extraction with an extraction solvent or fractionation by adding a fractionation solvent to an extract prepared by extraction with an extraction solvent. The extraction solvent is not limited thereto, but water, an organic solvent, or a mixed solvent thereof may be used, and the organic solvent may be an alcohol having 1 to 4 carbon atoms, a polar solvent such as ethyl acetate or acetone, hexane or dichloromethane. Non-polar solvents or mixed solvents thereof may be used. Also, preferably, water, alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof may be used, and more preferably ethanol may be used.
본 발명의 일실시예에서는 추출용매로서 주정(에탄올)을 이용하여 분쇄된 장수진흙버섯을 추출한 뒤 여과 및 농축하여 장수진흙버섯 추출물을 제조하였다. 이때, 장수진흙버섯 분쇄물 중량 기준 1 내지 10배 부피에 해당하는 양의 주정을 혼합하여 2시간 이상, 바람직하게는 20 내지 30시간 침지하여 상온에서 추출하는 것일 수 있으나, 이에 제한되지는 않는다.In one embodiment of the present invention, the longevity mud mushroom extract was prepared by extracting the pulverized longevity mud mushroom using alcohol (ethanol) as an extraction solvent, followed by filtration and concentration. At this time, it may be extracted at room temperature by mixing alcohol in an amount corresponding to 1 to 10 times the volume of the pulverized longevity mushroom and immersing for 2 hours or more, preferably 20 to 30 hours, but is not limited thereto.
일 구현예에서, 상기 항바이러스용 조성물은 유효성분으로서 장수진흙버섯 추출물을 포함하므로, 피코르나바이러스(피코르나비리대과, picornaviridae), 오르토믹소바이러스(오르토믹소비리대과, Orthomyxoviridae) 및 코로나바이러스(코로나비리대과, Coronaviridae)로 이루어진 군으로부터 선택된 하나 이상의 바이러스에 대한 항바이러스 활성을 가지는 것일 수 있다. In one embodiment, since the antiviral composition contains the longevity mud mushroom extract as an active ingredient, picornavirus (picornaviridae, picornaviridae), orthomyxovirus (orthomyxoviridae, Orthomyxoviridae) and corona It may have antiviral activity against one or more viruses selected from the group consisting of viruses (Coronaviridae).
일 구현예에서, 상기 피코르나바이러스는 바람직하게는 엔테로바이러스(enterovirus)일 수 있고, 보다 바람직하게는, 콕사키바이러스(coxsackievirus) 및 라이노바이러스(rhinovirus)로 이루어진 군으로부터 선택된 하나 이상의 바이러스일 수 있다.In one embodiment, the picornavirus may be preferably an enterovirus, more preferably, one or more viruses selected from the group consisting of coxsackievirus and rhinovirus there is.
일 구현예에서, 상기 오르토믹소바이러스는 인플루엔자바이러스(influenza virus)일 수 있고, 상기 코로나바이러스는 알파 코로나바이러스(alpha coronavirus)일 수 있다.In one embodiment, the orthomyxovirus may be an influenza virus and the coronavirus may be an alpha coronavirus.
특히 바람직하게는, 상기 항바이러스용 조성물은 콕사키바이러스 B3형(coxsackievirus B3; CVB3), 인간 라이노바이러스 1B형(human rhinovirus 1B; HRV1B), 인플루엔자바이러스 A형(influenza virus A/H1N1/PR8; Inf/PR8) 및 인간 코로나바이러스 229E(human coronavirus 229E)로 이루어진 군으로부터 선택된 하나 이상의 바이러스에 대한 항바이러스 활성을 가지는 것일 수 있다.Especially preferably, the antiviral composition is coxsackievirus B3 (CVB3), human rhinovirus 1B (HRV1B), influenza virus A (influenza virus A / H1N1 / PR8; Inf /PR8) and human coronavirus 229E (human coronavirus 229E) may have antiviral activity against one or more viruses selected.
본 발명의 일 실시예에서는 콕사키바이러스 B3형(CVB3), 인간 라이노바이러스 1B형(HRV1B), 인플루엔자바이러스 A형(Inf/PR8) 및 인간 코로나바이러스 229E(coronavirus 229E)으로 각각 감염된 vero 세포, hela 세포, A549 세포 및 Huh7 세포에서, 장수진흙버섯 추출물이 바이러스 증식에 의한 세포의 생존율 감소를 억제하여 세포 생존율을 효과적으로 증가시키는 것을 확인함으로써, 장수진흙버섯 추출물이 우수한 항바이러스 활성을 가짐을 입증하였다(실시예 2 및 도 1 내지 4 참조).In one embodiment of the present invention, vero cells infected with coxsackievirus type B3 (CVB3), human rhinovirus type 1B (HRV1B), influenza virus type A (Inf / PR8) and human coronavirus 229E (coronavirus 229E), hela In cells, A549 cells and Huh7 cells, it was confirmed that the longevity mushroom extract effectively increased cell viability by inhibiting the decrease in cell viability caused by virus proliferation, demonstrating that the longevity mushroom extract had excellent antiviral activity ( See Example 2 and Figures 1 to 4).
일 구현예에서, 상기 장수진흙버섯 추출물은 바이러스의 증식을 억제시킴으로써 콕사키바이러스, 라이노바이러스, 인플루엔자바이러스 및 알파 코로나바이러스로 이루어진 군으로부터 선택된 어느 하나 이상의 바이러스에 대한 항바이러스 효과를 발휘하는 것일 수 있다.In one embodiment, the longevity mud mushroom extract may exert an antiviral effect against any one or more viruses selected from the group consisting of coxsackie virus, rhinovirus, influenza virus, and alpha coronavirus by inhibiting the proliferation of viruses. .
본 발명의 일실시예에서는 vero 세포, hela 세포, A549 세포 및 Huh7 세포에 장수진흙버섯 추출물을 처리하여도 세포독성이 나타나지 않음을 확인함으로써, 부작용과 내성의 잠재성이 낮은 항바이러스제로로 유용하게 사용될 수 있음을 확인하였다(실시예 2 및 도 5 참조).In one embodiment of the present invention, by confirming that cytotoxicity does not occur even when vero cells, hela cells, A549 cells, and Huh7 cells are treated with the longevity mud mushroom extract, it is useful as an antiviral agent with low potential for side effects and resistance. It was confirmed that it can be used (see Example 2 and FIG. 5).
또한, 본 발명의 다른 측면에서, 장수진흙버섯 추출물을 포함하는 바이러스성 질환 예방 또는 치료용 약학 조성물을 제공한다.In addition, in another aspect of the present invention, a pharmaceutical composition for preventing or treating viral diseases containing the longevity mud mushroom extract is provided.
일 구현예에서, 상기 바이러스성 질환은 피코르나바이러스(피코르나비리대과, picornaviridae), 오르토믹소바이러스(오르토믹소비리대과, Orthomyxoviridae) 및 코로나바이러스(코로나비리대과, Coronaviridae)로 이루어진 군으로부터 선택된 하나 이상의 바이러스로 인하여 유발되는 질환일 수 있고, 바람직하게는, 콕사키바이러스(coxsackievirus), 라이노바이러스(rhinovirus), 인플루엔자바이러스(influenza virus) 및 알파 코로나바이러스(coronavirus)로 이루어진 군으로부터 선택된 하나 이상의 바이러스로 인하여 유발되는 질환일 수 있다.In one embodiment, the viral disease is from the group consisting of picornavirus (picornaviridae), orthomyxovirus (Orthomyxoviridae) and coronavirus (Coronaviridae). It may be a disease caused by one or more selected viruses, preferably, one or more selected from the group consisting of coxsackievirus, rhinovirus, influenza virus and alpha coronavirus It may be a disease caused by a virus.
특히 바람직하게는, 상기 바이러스성 질환은 콕사키바이러스 B3형(coxsackievirus B3; CVB3), 인간 라이노바이러스 1B형(human rhinovirus 1B; HRV1B), 인플루엔자바이러스 A형(influenza virus A/H1N1/PR8; Inf/PR8) 및 인간 코로나바이러스 229E(human coronavirus 229E)로 이루어진 군으로부터 선택된 하나 이상의 바이러스로 인하여 유발되는 질환일 수 있다.Particularly preferably, the viral disease is coxsackievirus B3 (CVB3), human rhinovirus 1B (HRV1B), influenza virus A / H1N1 / PR8; Inf/ It may be a disease caused by one or more viruses selected from the group consisting of PR8) and human coronavirus 229E.
일 구현예에서, 상기 바이러스성 질환은 바이러스성 수막염, 간염, 근육염, 심근염, 심근증, 췌장염, 당뇨, 유행성 근육통, 뇌염, 감기, 독감, 천식, 만성 폐쇄성 폐질환, 폐렴, 상기도감염증, 폐기종, 만성기관지염, 기관지선종, 고립성폐결절, 폐결핵, 농흉, 폐농양, 폐의 조직구 증식증, 축농증, 중이염, 소아마비, 포진성 구협염, 구제역, 급성출혈성 결막염, 수족구병 및 수포병으로 이루어진 군에서 선택된 하나 이상일 수 있다.In one embodiment, the viral disease is viral meningitis, hepatitis, myositis, myocarditis, cardiomyopathy, pancreatitis, diabetes, mumps, encephalitis, cold, flu, asthma, chronic obstructive pulmonary disease, pneumonia, upper respiratory tract infection, emphysema, chronic At least one day selected from the group consisting of bronchitis, bronchial adenoma, solitary pulmonary nodule, pulmonary tuberculosis, empyema, lung abscess, lung histiocytosis, sinusitis, otitis media, poliomyelitis, stomatitis herpes, foot-and-mouth disease, acute hemorrhagic conjunctivitis, hand-foot-and-mouth disease, and bullous disease. can
상기 “예방”은 병리학적 현상의 발생 빈도 또는 정도를 감소시키는 모든 행위를 의미한다. 예방은 완전할 수 있으며 또는 부분적일 수도 있다. 이 경우에는 개체 내의 콕사키바이러스 및/또는 라이노바이러스에 의해 발생되는 증상이 상기 조성물을 사용하지 않은 경우와 비교하여 감소하는 현상을 의미할 수 있다.The "prevention" means any action that reduces the frequency or severity of pathological phenomena. Prevention can be complete or partial. In this case, it may mean a phenomenon in which symptoms caused by coxsackievirus and / or rhinovirus in the subject are reduced compared to the case where the composition is not used.
상기 “치료”는 치료하고자 하는 대상(또는 개체) 또는 세포의 천연 과정을 변경시키기 위하여 임상적으로 개입하는 모든 행위를 의미하며, 임상 병리 상태가 진행되는 동안 또는 이를 예방하기 위하여 수행할 수 있다. 목적하는 치료 효과는 질병의 발생 또는 재발을 예방하거나, 증상을 완화시키거나, 질병에 따른 모든 직접 또는 간접적인 병리학적 결과를 저하시키거나, 전이를 예방하거나, 질병 진행 속도를 감소시키거나, 질병 상태를 경감 또는 일시적 완화시키거나, 예후를 개선시키는 것을 포함할 수 있다.The "treatment" means any action that clinically intervenes to change the natural process of a target (or individual) or cell to be treated, and can be performed while a clinical pathological state is progressing or to prevent it. The desired therapeutic effect is to prevent the occurrence or recurrence of the disease, alleviate the symptoms, reduce any direct or indirect pathological consequences of the disease, prevent metastasis, reduce the rate of disease progression, or reduce the rate of disease progression. alleviating or palliating the condition, or improving the prognosis.
상기, "개체"란, 상기 질환이 발병하였거나 발병할 수 있는 인간을 포함한 원숭이, 소, 말, 양, 돼지, 닭, 칠면조, 메추라기, 고양이, 개, 마우스, 쥐, 토끼 또는 기니아 피그를 포함한 모든 동물을 의미하는 것일 수 있고, 본 발명의 약학적 조성물을 개체에게 투여함으로써 상기 질환들을 효과적으로 예방 또는 치료할 수 있다.The term "individual" refers to all monkeys, cows, horses, sheep, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits, or guinea pigs, including humans who have or may have the disease. It may mean an animal, and the above diseases can be effectively prevented or treated by administering the pharmaceutical composition of the present invention to a subject.
상기 조성물은 경구적 전달, 비경구적 전달의 형태로 투여될 수 있다. 상기 조성물은 전신 또는 국소 투여될 수 있으며, 상기 투여는 경구 투여 및 비경구 투여를 포함할 수 있다. 상기 조성물은 적절한 투여 형태를 제공하도록 적합한 양의 약학적으로 허용되는 비히클 또는 담체와 함께 제형화될 수 있다.The composition may be administered in the form of oral delivery or parenteral delivery. The composition may be administered systemically or locally, and the administration may include oral administration and parenteral administration. The composition may be formulated with a pharmaceutically acceptable vehicle or carrier in suitable amounts to provide an appropriate dosage form.
상기 조성물은 약학 조성물의 제조에 사용되는 담체, 부형제 및 희석제를 더 포함할 수 있다. 상기 담체, 부형제 및 희석제로는 락토즈, 덱스트로즈, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아 고무, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로즈, 메틸 셀룰로즈, 미정질 셀룰로스, 폴리비닐 피롤리돈, 물, 메틸히드록시벤조에이트, 프로필히드록시벤조에이트, 탈크, 마그네슘 스테아레이트 또는 광물유를 들 수 있으나, 이에 제한되는 것은 아니다.The composition may further include carriers, excipients and diluents used in the preparation of pharmaceutical compositions. The carriers, excipients and diluents include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, or mineral oil.
또한, 상기 조성물은 산제, 과립제, 정제, 캡슐제, 현탁액, 에멀젼, 시럽, 에어로졸 등의 경구형 제형, 외용제, 좌제 및 멸균 주사용액의 형태로 제제화 하여 사용할 수 있다.In addition, the composition may be formulated and used in the form of oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories and sterile injection solutions.
경구 투여를 위한 고형제제는 정제, 환제, 산제, 과립제, 캡슐제 등이 사용될 수 있고, 상기 고형제제는 상기 추출물에 적어도 하나 이상의 부형제, 예컨대, 전분, 칼슘카보네이트, 수크로스, 락토오스, 또는 젤라틴 등을 혼합하여 조제할 수 있다. 또한, 상기 부형제 이외에 마그네슘 스티레이트, 탈크 같은 윤활제가 사용될 수 있다.Solid preparations for oral administration may include tablets, pills, powders, granules, capsules, etc., and the solid preparations may include at least one excipient added to the extract, such as starch, calcium carbonate, sucrose, lactose, or gelatin. It can be prepared by mixing. In addition to the above excipients, lubricants such as magnesium styrate and talc may be used.
경구 투여를 위한 액상 제제는 현탁제, 내용액제, 유제, 시럽제 등이 사용될 수 있고, 단순희석제인 물, 리퀴드 파라핀 외에 여러 가지 부형제, 예컨대 습윤제, 감미제, 방향제, 보존제 등이 사용될 수 있다.Liquid preparations for oral administration may include suspensions, internal solutions, emulsions, syrups, and the like, and various excipients such as wetting agents, sweeteners, fragrances, and preservatives in addition to simple diluents such as water and liquid paraffin.
비경구 투여를 위한 제제는 멸균된 수용액, 비수성용제, 현탁제, 유제, 동결건조 제제, 좌제가 사용될 수 있다.Preparations for parenteral administration may include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried preparations, and suppositories.
상기 비수성용제, 현탁제는 프로필렌글리콜(propylene glycol), 폴리에틸렌 글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르가 사용될 수 있다. 상기 좌제의 기제로는 위텝솔(witepsol), 마크로골, 트윈(tween) 61, 카카오지, 라우린지, 글리세로제라틴이 사용될 수 있다.Propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used as the non-aqueous solvent or suspending agent. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin fat, and glycerogeratin may be used.
상기 약학 조성물은 약제학적으로 유효한 양이 대상체에 투여될 수 있다. 상기 “약제학적으로 유효한 양”은 의학적 치료에 적용 가능한 합리적인 수혜/위험 비율로 질환을 치료하기에 충분한 양을 의미하며, 유효 용량 수준은 환자의 질환의 종류, 중증도, 약물의 활성, 약물에 대한 민감도, 투여 시간, 투여 경로 및 배출 비율, 치료 기간, 동시 사용되는 약물을 포함한 요소 및 기타 의학 분야에 잘 알려진 요소에 따라 결정될 수 있다.The pharmaceutical composition may be administered to a subject in a pharmaceutically effective amount. The "pharmaceutically effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level is based on the type and severity of the patient's disease, the activity of the drug, and the drug It may be determined according to factors including sensitivity, time of administration, route of administration and excretion rate, duration of treatment, drugs used concurrently, and other factors well known in the medical arts.
상기 약학 조성물은 개별 치료제로 투여하거나 다른 치료제와 병용하여 투여될 수 있고, 종래의 치료제와 순차적으로 또는 동시에 투여될 수 있으며, 단일 또는 다중 투여될 수 있다. 상기 요소들을 모두 고려하여 부작용 없이 최소한의 양으로 최대 효과를 얻을 수 있는 양을 투여하는 것이 바람직하며, 이는 당업자에 의해 용이하게 결정될 수 있다.The pharmaceutical composition may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple times. Considering all of the above factors, it is preferable to administer an amount that can obtain the maximum effect with the minimum amount without side effects, which can be easily determined by those skilled in the art.
일 구현예에서, 본 발명은 장수진흙버섯 추출물을 필요로 하는 개체에게 투여하는 단계를 포함하는, 바이러스성 질환을 예방 또는 치료하는 방법을 제공한다.In one embodiment, the present invention provides a method for preventing or treating a viral disease, comprising administering the longevity mud mushroom extract to a subject in need thereof.
나아가, 본 발명의 또 다른 측면에서, 장수진흙버섯 추출물을 포함하는 바이러스성 질환 예방 또는 개선용 식품 조성물을 제공한다.Furthermore, in another aspect of the present invention, a food composition for preventing or improving viral diseases containing the longevity mud mushroom extract is provided.
일 구현예에서, 상기 바이러스성 질환은 엔테로바이러스(enterovirus)로 인하여 유발되는 질환일 수 있고, 바람직하게는 콕사키바이러스(coxsackievirus) 및 라이노바이러스(rhinovirus)로 이루어진 군으로부터 선택된 하나 이상의 바이러스로 인하여 유발되는 질환일 수 있다.In one embodiment, the viral disease may be a disease caused by an enterovirus, preferably caused by one or more viruses selected from the group consisting of coxsackievirus and rhinovirus it could be a disease.
일 구현예에서, 상기 바이러스성 질환은 바이러스성 수막염, 간염, 근육염, 심근염, 심근증, 췌장염, 당뇨, 유행성 근육통, 뇌염, 감기, 천식, 만성 폐쇄성 폐질환, 폐렴, 축농증, 중이염, 소아마비, 포진성 구협염, 구제역, 급성출혈성 결막염, 수족구병 및 수포병으로 이루어진 군에서 선택된 하나 이상일 수 있다.In one embodiment, the viral disease is viral meningitis, hepatitis, myositis, myocarditis, cardiomyopathy, pancreatitis, diabetes, mumps, encephalitis, cold, asthma, chronic obstructive pulmonary disease, pneumonia, sinusitis, otitis media, poliomyelitis, herpes zoster It may be one or more selected from the group consisting of stomatitis, foot-and-mouth disease, acute hemorrhagic conjunctivitis, hand-foot-and-mouth disease, and blisters.
본 발명의 조성물을 식품 조성물로 사용하는 경우, 상기 장수진흙버섯 추출물을 그대로 첨가하거나 다른 식품 또는 식품 성분과 함께 사용할 수 있고, 통상의 방법에 따라 적절하게 사용할 수 있다.When the composition of the present invention is used as a food composition, the longevity mud mushroom extract may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method.
상기 조성물은 유효성분 이외에 식품학적으로 허용 가능한 식품보조첨가제를 포함할 수 있으며, 유효성분의 혼합량은 사용목적(예방, 건강 또는 치료적 처치)에 따라 적합하게 결정될 수 있다.The composition may include food additives acceptable in food science in addition to the active ingredient, and the mixing amount of the active ingredient may be appropriately determined depending on the purpose of use (prevention, health or therapeutic treatment).
본 발명에서 사용되는 용어 "식품보조첨가제"란 식품에 보조적으로 첨가될 수 있는 구성요소를 의미하며, 각 제형의 건강기능식품을 제조하는데 첨가되는 것으로서 당업자가 적절히 선택하여 사용할 수 있다. 식품보조첨가제의 예로는 여러 가지 영양제, 비타민, 광물(전해질), 합성 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 충진제, 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알콜, 탄산음료에 사용되는 탄산화제 등이 포함되지만, 상기 예들에 의해 본 발명의 식품보조첨가제의 종류가 제한되는 것은 아니다.The term "food supplement additive" used in the present invention refers to a component that can be added to food supplementally, and can be appropriately selected and used by those skilled in the art as being added to prepare a health functional food of each formulation. Examples of food additives include various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and fillers, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners , pH regulators, stabilizers, preservatives, glycerin, alcohol, carbonation agents used in carbonated beverages, etc. are included, but the types of food additives of the present invention are not limited by the above examples.
본 발명의 식품 조성물은 건강기능식품을 포함하는 의미인 것일 수 있다. 본 발명에서 사용되는 용어 "건강기능식품"이란 인체에 유용한 기능성을 가진 원료나 성분을 사용하여 정제, 캅셀, 분말, 과립, 액상 및 환 등의 형태로 제조 및 가공한 식품을 말한다. 여기서 '기능성'이라 함은 인체의 구조 및 기능에 대하여 영양소를 조절하거나 생리학적작용 등과 같은 보건용도에 유용한 효과를 얻는 것을 의미한다. The food composition of the present invention may be meant to include health functional food. The term "health functional food" used in the present invention refers to food prepared and processed in the form of tablets, capsules, powders, granules, liquids and pills using raw materials or ingredients having useful functionalities for the human body. Here, 'functionality' means obtaining useful effects for health purposes, such as adjusting nutrients for the structure and function of the human body or physiological actions.
본 발명의 건강기능식품은 통상의 기술분야에서 통상적으로 사용되는 방법에 의하여 제조 가능하며, 상기 제조 시에는 통상의 기술분야에서 통상적으로 첨가하는 원료 및 성분을 첨가하여 제조할 수 있다. 또한 상기 건강기능식품의 제형 또한 건강기능식품으로 인정되는 제형이면 제한 없이 제조될 수 있다. The health functional food of the present invention can be manufactured by a method commonly used in the conventional technical field, and can be prepared by adding raw materials and components commonly added in the conventional technical field during the manufacture. In addition, the formulation of the health functional food may also be manufactured without limitation as long as the formulation is recognized as a health functional food.
본 발명의 식품용 조성물은 다양한 형태의 제형으로 제조될 수 있으며, 일반약품과는 달리 식품을 원료로 하여 약품의 장기 복용 시 발생할 수 있는 부작용 등이 없는 장점이 있고, 휴대성이 뛰어나, 본 발명의 건강기능식품은 항바이러스제 또는 바이러스성 질환 치료제의 효과를 증진 시키기 위한 보조제로 섭취가 가능하다.The composition for food of the present invention can be prepared in various types of formulations, and unlike general drugs, it has the advantage of not having side effects that may occur when taking drugs for a long time by using food as a raw material, and has excellent portability. Health functional foods of the above can be consumed as supplements to enhance the effectiveness of antiviral agents or viral disease treatments.
또한, 본 발명의 조성물이 사용될 수 있는 건강식품의 종류에는 제한이 없다. 아울러 본 발명의 장수진흙버섯 추출물을 유효성분으로 포함하는 조성물은 당업자의 선택에 따라 건강기능식품에 함유될 수 있는 적절한 기타 보조 성분과 공지의 첨가제를 혼합하여 제조할 수 있다. In addition, there is no limitation on the type of health food in which the composition of the present invention can be used. In addition, the composition containing the longevity mud mushroom extract of the present invention as an active ingredient can be prepared by mixing suitable other auxiliary ingredients that can be contained in health functional foods and known additives according to the selection of those skilled in the art.
첨가할 수 있는 식품의 예로는 육류, 소세지, 빵, 쵸코렛, 캔디류, 스낵류, 과자류, 피자, 라면, 기타 면류, 껌류, 아이스크림 류를 포함한 낙농제품, 각종 스프, 음료수, 차, 드링크제, 알콜 음료 및 비타민 복합제 등이 있으며, 본 발명에 따른 추출물을 주성분으로 하여 제조한 즙, 차, 젤리 및 주스 등에 첨가하여 제조할 수 있다.Examples of foods that can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, chewing gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and There are vitamin complexes, etc., and it can be prepared by adding the extract according to the present invention to juice, tea, jelly, juice, etc. prepared as a main component.
본 발명의 장수진흙버섯 추출물은 천연 약용 식물을 원료로 하므로 약학 조성물 또는 식품 조성물로 사용할 경우에도 일반적인 합성 화합물에 비하여 부작용 및 내성이 덜할 수 있으므로, 안전하게 약학적 조성물 및 건강기능식품에 포함되어 유용하게 사용될 수 있다.Since the longevity mud mushroom extract of the present invention is made from natural medicinal plants as raw materials, even when used as a pharmaceutical composition or food composition, side effects and tolerance may be less than those of general synthetic compounds, so it is safely included in pharmaceutical compositions and health functional foods and is useful. can be used
하기의 실시예를 통하여 본 발명을 보다 상세하게 설명한다. 그러나 하기 실시예는 본 발명의 내용을 구체화하기 위한 것일 뿐 이에 의해 본 발명이 한정되는 것은 아니다.The present invention will be described in more detail through the following examples. However, the following examples are only for specifying the content of the present invention, and the present invention is not limited thereto.
<실시예 1> 장수진흙버섯 추출물의 제조<Example 1> Preparation of longevity mud mushroom extract
장수진흙버섯(장수상황버섯, Phellinus baumii)을 잘게 분쇄한 다음, 상기 장수진흙버섯 분쇄물에 주정을 추출용매로 부가하여 24시간 동안 침지하여 추출하였다. 추출된 추출물을 여과하고 여과물을 농축한 후 동결건조하여 장수진흙버섯 추출물을 제조하였다.Longevity mud mushroom ( Phellinus baumii ) was finely pulverized, and then alcohol was added as an extraction solvent to the longevity mud mushroom pulverization, followed by soaking for 24 hours for extraction. The extracted extract was filtered, the filtrate was concentrated, and lyophilized to prepare an extract of longevity mud mushroom.
<준비예 1> 바이러스와 세포주 및 시약 준비<Preparation Example 1> Preparation of viruses, cell lines and reagents
항바이러스 활성 분석에 이용한 콕사키바이러스 B3형(CVB3), 인간 라이노바이러스 1B형(HRV1B), 인플루엔자바이러스 A형 H1N1/PR8(Inf/PR8) 및 인간 코로나바이러스 229E(coronavirus 229E)는 ATCC (American Type Culture Collection, Manassas, VA, USA)에서 구매하였다. 콕사키바이러스 B3형은 vero 세포에 감염시켜 37℃에서 증식시켰고, 라이노바이러스 1B형은 hela 세포에 감염시켜 32℃에서 증식시켰으며, 인플루엔자바이러스 PR8은 A549 세포에, 인간 코로나바이러스 229E는 Huh7 세포에 각각 감염시켜 37℃에서 증식시켰다. Coxsackievirus type B3 (CVB3), human rhinovirus type 1B (HRV1B), influenza virus type A H1N1/PR8 (Inf/PR8) and human coronavirus 229E (coronavirus 229E) used for antiviral activity analysis were ATCC (American Type Culture Collection, Manassas, VA, USA). Coxsackievirus type B3 was infected with vero cells and grown at 37°C, rhinovirus type 1B was infected with hela cells and grown at 32°C, influenza virus PR8 was infected with A549 cells, and human coronavirus 229E was grown with Huh7 cells. Each was infected and grown at 37°C.
Vero 세포, hela세포, A549 세포 및 Huh7 세포는 각각 1% 소태아혈청(FBS) 및 0.01% 항생제가 첨가된 최소 필수 배지(MEM media)에서 유지하였다. 항생제와 trypsin-EDTA, FBS 및 MEM media는 Gibco사 제품을 사용 하였다. 조직 배양 플레이트(Tissue culture plates)는 Falcon(BD Biosciences, San Jose, CA, 미국)에서 구입하였다. 술포로다민 B(sulforhodamine B; SRB)는 Sigma-Aldrich(St. Louis, MO, 미국)에서 구매 하였다. 트립신-EDTA를 사용하여 세포를 탈착시켰다.Vero cells, hela cells, A549 cells and Huh7 cells were maintained in minimum essential medium (MEM media) supplemented with 1% fetal bovine serum (FBS) and 0.01% antibiotics, respectively. Antibiotics, trypsin-EDTA, FBS, and MEM media were supplied by Gibco. Tissue culture plates were purchased from Falcon (BD Biosciences, San Jose, CA, USA). Sulforhodamine B (SRB) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Cells were detached using trypsin-EDTA.
추출물은 DMSO를 용해용액으로 사용하여 MEM 배지(MEM media)에 희석하였다. DMSO의 최종 농도는 세포배양에 전혀 영향이 없는 최대 농도인 0.1%로 사용하였다. 또한 0.1% DMSO는 세포대조군으로 사용하였다.The extract was diluted in MEM media using DMSO as a dissolving solution. The final concentration of DMSO was used as 0.1%, which is the maximum concentration that does not affect cell culture at all. In addition, 0.1% DMSO was used as a cell control.
<실시예 2> 장수진흙버섯 추출물의 항바이러스 활성 및 세포독성 측정<Example 2> Measurement of antiviral activity and cytotoxicity of longevity mud mushroom extract
바이러스에 의해 일어나는 세포 병변을 억제시키는 능력을 분석하는 방법인 SRB assay를 이용하여, 장수진흙버섯 추출물의 콕사키바이러스 B3형, 라이노바이러스 1B형, 인플루엔자바이러스 A형(PR8) 및 인간 코로나바이러스(229E)에 대한 항바이러스 활성 및 세포독성을 평가하였다.Coxsackie virus type B3, rhinovirus type 1B, influenza virus type A (PR8) and human coronavirus (229E ) was evaluated for antiviral activity and cytotoxicity.
96-웰 배양 플레이트(Nunc, Denmark) 각각의 웰(well)에 콕사키바이러스 B3형(CVB3) 및 라이노바이러스 1B형(HRV1B) 각 바이러스에 감수성을 갖는 세포인 vero 세포와 hela 세포를 3×104의 수로 준비하였다. 24시간 후 96-웰 배양 플레이트에 각각의 세포가 90%정도 자랐을 때 실험에 이용하였다. 각 웰의 배지(media)를 제거한 후, 각 세포에 대하여 PBS로 1회 세척하였다. 그 후, 1% FBS 및 각각의 바이러스가 포함된 90μL의 MEM을 각 웰에 넣고, 장수진흙버섯 추출물을 0.4, 2, 10 및 50 μg/ml의 농도가 되도록 각 웰에 10μL 첨가하였다. 한편, 4개의 웰은 추출물을 처리하지 않고 바이러스만 처리한 바이러스대조구(Vehicle; Veh)로 사용하였고 또 다른 4개의 웰은 바이러스와 추출물을 모두 처리하지 않고 추출물 대신 1% DMSO를 처리한 세포대조구(Control; Ctrl)로 사용하였다. 각 웰은 CO2 배양기 (VS-9108MS, Vision, Korea)에서 37℃, 5% CO2의 조건으로 2일 동안 배양하였다.In each well of a 96-well culture plate (Nunc, Denmark), 3 × 10 vero cells and hela cells, which are cells sensitive to coxsackie virus type B3 (CVB3) and rhinovirus type 1B (HRV1B) viruses, were placed in each well. prepared in the number of 4 . After 24 hours, when each cell grew to about 90% in a 96-well culture plate, it was used for the experiment. After removing the media of each well, each cell was washed once with PBS. Thereafter, 90 μL of MEM containing 1% FBS and each virus was put into each well, and 10 μL of the longevity mushroom extract was added to each well to have concentrations of 0.4, 2, 10 and 50 μg/ml. On the other hand, 4 wells were used as a virus control (Vehicle; Veh) in which only virus was treated without extract treatment, and another 4 wells were used as cell control cells treated with 1% DMSO instead of extract without both virus and extract treatment ( Control; Ctrl) was used. Each well was cultured for 2 days in a CO 2 incubator (VS-9108MS, Vision, Korea) under conditions of 37° C. and 5% CO 2 .
또한, 추출물의 세포독성을 알아보기 위하여, 바이러스가 첨가되지 않은 배양액을 이용하여 상기와 동일한 방법으로 hela 세포 및 vero 세포에 각각 0.4~50 μg/ml 농도의 추출물 또는 1% DMSO를 첨가하고 37℃, 5% CO2의 조건으로 CO2 배양기에서 2일 동안 배양하였다.In addition, in order to examine the cytotoxicity of the extract, the extract or 1% DMSO at a concentration of 0.4 to 50 μg/ml was added to hela cells and vero cells in the same manner as above using the culture medium to which no virus was added, respectively, and 37 ° C. , and cultured for 2 days in a CO 2 incubator under conditions of 5% CO 2 .
각 배양종료 후, 각 웰의 배양 상등액을 제거하고 웰에 남아있는 세포를 70% 아세톤 용액 100 μL로 고정하고 SRB 용액 100 μL을 각 웰에 가하여 세포단백질을 염색시켰다. 세포단백질과 결합하지 않은 SRB 염색제를 세척하여 제거한 후 10 mM 트리스 용액(pH 10.5) 100 μL를 각 웰에 가하여 세포와 결합되어 있는 염색제를 용해시켜서 565 nm에서 ELISA reader로 각 웰의 흡광도를 측정하여 항바이러스 활성 및 세포독성을 계산하였다. 이때, 항바이러스 활성의 측정은 바이러스를 감염시키지 않은 웰의 흡광도 (B), 바이러스를 감염시킨 웰의 흡광도 (C), 추출물과 바이러스를 같이 투여한 웰의 흡광도 (D)로 표시하고 하기 수학식 1에 따라 감염후 세포 생존율(cell viability, %)을 계산하여 평가하였다. 여기서 생존율이 100%인 경우 항바이러스 효과 100%이고, 생존율이 0%인 경우 항바이러스 효과가 없는 것을 의미한다.After completion of each culture, the culture supernatant of each well was removed, the cells remaining in the well were fixed with 100 μL of a 70% acetone solution, and 100 μL of SRB solution was added to each well to stain cell proteins. After washing and removing the SRB dye that is not bound to cell protein, 100 μL of 10 mM Tris solution (pH 10.5) is added to each well to dissolve the dye that is bound to the cell, and the absorbance of each well is measured with an ELISA reader at 565 nm. Antiviral activity and cytotoxicity were calculated. At this time, the measurement of antiviral activity is expressed as the absorbance of wells not infected with the virus (B), the absorbance of the wells infected with the virus (C), and the absorbance of the wells administered with the extract and the virus (D), and the following formula After infection according to 1, cell viability (%) was calculated and evaluated. Here, when the survival rate is 100%, the antiviral effect is 100%, and when the survival rate is 0%, it means that there is no antiviral effect.
[수학식 1][Equation 1]
그 결과, 콕사키바이러스 B3형(CVB3)에 대하여 장수진흙버섯 추출물이 50 μg/mL의 농도에서 약 25%의 항바이러스 활성을 나타내었다(도 1). 또한 라이노바이러스 1B형(HRV1B)에 대하여, 장수진흙버섯 추출물 50 μg/mL의 농도에서 약 70%, 10 μg/mL의 농도에서 약 40%, 2 μg/mL 및 0.4 μg/mL의 농도에서 각각 약 20%의 항바이러스 활성을 나타내는 것을 확인하였다(도 2). 또한, 인플루엔자 바이러스 A형(Inf/PR8)에 대하여, 장수진흙버섯 추출물 50 μg/mL의 농도에서 약 80%, 10 μg/mL의 농도에서 약 30%, 2 μg/mL의 농도에서 약 20%의 항바이러스 활성을 나타내는 것을 확인하였다(도 3). 마지막으로, 인간 코로나바이러스 229E(Coronavarius 229E)에 대하여, 장수진흙버섯 추출물 50 μg/mL의 농도에서 약 40%의 항바이러스 활성을 나타내었다(도 4).As a result, the longevity mud mushroom extract exhibited an antiviral activity of about 25% against coxsackievirus type B3 (CVB3) at a concentration of 50 μg/mL (FIG. 1). In addition, for rhinovirus type 1B (HRV1B), about 70% at a concentration of 50 µg/mL of rhinovirus extract, about 40% at a concentration of 10 µg/mL, and at a concentration of 2 µg/mL and 0.4 µg/mL, respectively. It was confirmed that it exhibited about 20% of antiviral activity (FIG. 2). In addition, against influenza virus type A (Inf/PR8), about 80% at a concentration of 50 μg/mL, about 30% at a concentration of 10 μg/mL, and about 20% at a concentration of 2 μg/mL of longevity mushroom extract It was confirmed that the antiviral activity of (FIG. 3). Finally, with respect to human coronavirus 229E (Coronavarius 229E), antiviral activity of about 40% was exhibited at a concentration of 50 μg/mL of longevity mushroom extract (FIG. 4).
한편, 장수진흙버섯 추출물의 세포독성을 각각의 항바이러스 활성 실험에 사용한 vero 세포와 hela 세포 및 A549세포에서 확인한 결과, 장수진흙버섯 추출물은 본 발명의 실험에서 사용한 0.4 내지 50 μg/mL의 농도에서 거의 모두 세포독성을 나타내지 않는 것을 확인하였다(도 5). 따라서, 본 발명의 실시예의 방법으로 제조된 장수진흙버섯 추출물을 CVB3, HRV1B, Inf/PR8, Coronavirus 229E 등에 의해 유발된 바이러스 감염을 예방 또는 치료하기 위한 효과적인 항바이러스제로 이용할 수 있음을 확인하였다.On the other hand, as a result of confirming the cytotoxicity of the longevity mushroom extract in vero cells, hela cells and A549 cells used in each antiviral activity test, the longevity mushroom extract was used in the present experiment at a concentration of 0.4 to 50 μg / mL. It was confirmed that almost all showed no cytotoxicity (FIG. 5). Therefore, it was confirmed that the longevity mud mushroom extract prepared by the method of Example of the present invention can be used as an effective antiviral agent for preventing or treating viral infections caused by CVB3, HRV1B, Inf/PR8, Coronavirus 229E, etc.
이제까지 본 발명에 대하여 그 바람직한 실시예들을 중심으로 살펴보았다. 본 발명이 속하는 기술 분야에서 통상의 지식을 가진 자는 본 발명이 본 발명의 본질적인 특성에서 벗어나지 않는 범위에서 변형된 형태로 구현될 수 있음을 이해할 수 있을 것이다. 그러므로 개시된 실시예들은 한정적인 관점이 아니라 설명적인 관점에서 고려되어야 한다. 본 발명의 범위는 전술한 설명이 아니라 특허 청구범위에 나타나 있으며, 그와 동등한 범위 내에 있는 모든 차이점은 본 발명에 포함된 것으로 해석되어야 할 것이다.So far, the present invention has been looked at with respect to its preferred embodiments. Those skilled in the art to which the present invention pertains will be able to understand that the present invention can be implemented in a modified form without departing from the essential characteristics of the present invention. Therefore, the disclosed embodiments should be considered from an illustrative rather than a limiting point of view. The scope of the present invention is shown in the claims rather than the foregoing description, and all differences within the equivalent scope will be construed as being included in the present invention.
Claims (11)
상기 추출물은 장수진흙버섯 분쇄물 중량 기준 1 내지 10배 부피에 해당하는 양의 에탄올을 혼합하여 20시간 내지 30시간 침지하여 상온에서 추출한 것이고,
상기 바이러스는 콕사키바이러스 B3형(coxsackievirus B3; CVB3), 인간 라이노바이러스 1B형(human rhinovirus 1B; HRV1B) 및 인간 코로나바이러스 229E(human coronavirus 229E)로 이루어진 군으로부터 선택된 하나 이상의 바이러스인 것인, 항바이러스용 조성물.Longevity mud mushroom ( Phellinus baumii ) An antiviral composition comprising an ethanol extract as an active ingredient,
The extract is extracted at room temperature by mixing ethanol in an amount corresponding to 1 to 10 times the volume of the pulverized longevity mushroom and immersing for 20 to 30 hours,
The virus is one or more viruses selected from the group consisting of coxsackievirus B3 (CVB3), human rhinovirus 1B (HRV1B) and human coronavirus 229E, Composition for viruses.
상기 추출물은 상기 바이러스의 증식을 억제하는 것인, 항바이러스용 조성물.According to claim 1,
The extract is to inhibit the proliferation of the virus, antiviral composition.
상기 추출물은 장수진흙버섯 분쇄물 중량 기준 1 내지 10배 부피에 해당하는 양의 에탄올을 혼합하여 20시간 내지 30시간 침지하여 상온에서 추출한 것이고,
상기 바이러스로 인하여 유발되는 바이러스성 질환은 근육염, 심근염, 심근증, 당뇨, 유행성근육통, 감기, 독감, 폐렴, 상기도감염증, 포진성 구협염, 급성출혈성 결막염, 수족구병 및 수포병으로 이루어진 군에서 선택된 하나 이상의 바이러스성 질환인 것인, 약학 조성물.For preventing or treating viral diseases caused by at least one virus selected from the group consisting of coxsackie virus type B3 (CVB3), human rhinovirus type 1B (HRV1B) and human coronavirus 229E containing an ethanol extract of longevity mud mushroom As a pharmaceutical composition,
The extract is extracted at room temperature by mixing ethanol in an amount corresponding to 1 to 10 times the volume of the pulverized longevity mushroom and immersing for 20 to 30 hours,
The viral disease caused by the virus is one selected from the group consisting of myositis, myocarditis, cardiomyopathy, diabetes, epidemic myalgia, cold, flu, pneumonia, upper respiratory infection, stomatitis herpes, acute hemorrhagic conjunctivitis, hand foot and mouth disease, and blistering disease. More than one viral disease, the pharmaceutical composition.
상기 추출물은 장수진흙버섯 분쇄물 중량 기준 1 내지 10배 부피에 해당하는 양의 에탄올을 혼합하여 20시간 내지 30시간 침지하여 상온에서 추출한 것이고,
상기 바이러스로 인하여 유발되는 바이러스성 질환은 근육염, 심근염, 심근증, 당뇨, 유행성근육통, 감기, 독감, 폐렴, 상기도감염증, 포진성 구협염, 급성출혈성 결막염, 수족구병 및 수포병으로 이루어진 군에서 선택된 하나 이상의 바이러스성 질환인 것인, 식품 조성물.For preventing or improving viral diseases caused by one or more viruses selected from the group consisting of coxsackie virus type B3 (CVB3), human rhinovirus type 1B (HRV1B) and human coronavirus 229E containing an ethanol extract of longevity mud mushroom As a food composition,
The extract is extracted at room temperature by mixing ethanol in an amount corresponding to 1 to 10 times the volume of the pulverized longevity mushroom and immersing for 20 to 30 hours,
The viral disease caused by the virus is one selected from the group consisting of myositis, myocarditis, cardiomyopathy, diabetes, epidemic myalgia, cold, flu, pneumonia, upper respiratory infection, stomatitis herpes, acute hemorrhagic conjunctivitis, hand foot and mouth disease, and blistering disease. More than one viral disease, food composition.
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