KR20130112128A - Composition for immune enhancement of honey bee - Google Patents
Composition for immune enhancement of honey bee Download PDFInfo
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Abstract
Description
본 발명은 꿀벌의 면역증강용 조성물에 관한 것으로서, 특히 낭충봉아부패병 발생을 억제하는 꿀벌의 면역증강용 조성물에 관한 것이다.
The present invention relates to a composition for enhancing immunity of bees, and more particularly, to a composition for immunizing a bee that inhibits the incidence of insect bite infestation.
우리나라에서는 2009년에 토종벌(동양종꿀벌;Apis Cerana)에서 중국낭충봉아부패병 바이러스(Chinese Sacbrood Virus;CSBV)가 보고된 바 있다.In Korea, the Chinese Sacbrood Virus (CSBV) has been reported in Tochigi bean (Apis Cerana) in 2009.
상기 낭충봉아부패병(Sacbrood)은 꿀벌의 유충에서 발생하는 바이러스성 질병으로 병에 걸린 유충은 번데기가 되지 못하고 말라 죽게 하는 병이다.The Sacbrood is a viral disease caused by a larva of a bee, and the diseased larva is a pupa that does not become a pupa and is killed.
이러한 바이러스는 오염된 화분(花粉; 꽃가루)과 화밀(花蜜; 꽃꿀) 등을 통해 감염된다.These viruses are infected through contaminated pollen (pollen) and flower honey.
일벌들은 병든 유충들이 들어 있는 벌방을 청소하고 말라 죽은 유충들을 제거하게 되는데 이 과정에서 일벌들도 감염되며, 양봉기구와 벌의 교환 등을 통해 벌통에서 벌통으로도 전염이 확산될 수 있다.The worker sweeps the insect larvae and removes the dead larvae. In this process, worker bees are also infected and spread from the beehive to the beehive through the exchange of beekeepers and bees.
유밀(流蜜; 꽃에서 꿀이 분비되는 현상)이 왕성해지고, 벌의 한무리를 형성하는 봉군(蜂群)의 세력이 강하여 양질의 화분과 화밀 등을 공급할 수 있게 되면, 또 산란력이 좋은 여왕벌을 유입하면 대개 병세가 누그러지기도 하나, 질병의 확산이 심해지면 감염된 봉군을 소각하여 감염원을 제거하고, 꿀벌에 영양을 충분히 공급하는 식으로 예방적 조치를 취해야 한다. When the oyster (蜜 蜜; the phenomenon that the honey is secreted from the flower) becomes strong and the power of the bee group forming the bee's beehive is strong and it is possible to supply the high quality flower pots and flower seeds, The infestation of the queen bee usually relieves the disease, but if the spread of the disease becomes worse, preventive measures must be taken, such as incineration of infected populations, removal of infectious agents, and adequate feeding of bees.
그러나, 우리나라에서는 2009년 봄부터 발생하기 시작하여 전국적으로 확산되어 2010년 9월 현재 약 90%의 토종벌이 폐사한 것으로 추정되며, 특히 상기 전염병이 도착한 벌터에서는 약 한 달 안에 모든 봉군이 감염되며 약 두 달이면 모두 폐사되는 실정이다.However, in Korea, it started to occur in spring of 2009 and spread nationwide. It is estimated that about 90% of the native birds died as of September 2010. In particular, in the bull where the infectious disease has arrived, In two months, all are dead.
이와 같이 생산량 급감으로 토종꿀 가격이 크게 상승하는 것은 물론 벌이 사라진 지역은 예전처럼 자연 수정이 어려워져 과수 및 화훼 농가에 2차 피해가 우려된다.
As a result, the price of native honey has risen sharply due to a sharp drop in production, and the area where bees have disappeared has become difficult to naturalize as before and there is a fear of secondary damage to fruit trees and flower farms.
본 발명은 상기와 같은 문제점을 해소하기 위한 목적으로 바이러스에 의한 질병 방제로 인한 꿀벌의 폐사를 방지하는 꿀벌의 면역증강용 조성물을 제공하는데 있다.It is another object of the present invention to provide a composition for immunization of bees, which prevents the death of bees due to disease control by viruses.
특히, 항바이러스 활성뿐만 아니라 항균활성도 갖게 됨으로써 꿀벌의 생리적 환경을 개선하여 꿀벌의 수명 연장 및 면역을 증강시켜 꿀벌 스스로가 이겨낼 수 있도록 하는 꿀벌의 면역증강용 조성물을 제공하는데 있다.In particular, the present invention is to provide a composition for immunity enhancement of a bee, which is capable of enhancing the physiological environment of a bee by enhancing antimicrobial activity as well as antiviral activity, thereby enhancing the life span of the bee and enhancing immunity, thereby allowing the bee to overcome itself.
본 발명이 이루고자 하는 기술적 과제들은 이상에서 언급한 기술적 과제들로 제한되지 않으며, 언급되지 않은 다른 기술적 과제들은 아래의 기재로부터 본 발명이 속하는 기술분야에서 통상의 지식을 가진 자에게 명확하게 이해될 수 있을 것이다.
It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory and are not intended to limit the invention to the particular embodiments that are described. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory and are not restrictive of the invention, There will be.
상기 목적을 달성하기 위한 본 발명의 꿀벌의 면역증강용 조성물은 낭충봉아부패병을 일으키는 바이러스에 대해 항바이러스 활성을 갖는 알칼로이드를 유효성분으로 포함하는 것이 특징이다.In order to accomplish the above object, the present invention provides an immunomodulating composition for bees, which comprises an alkaloid having an antiviral activity as an active ingredient against a virus causing insect bite infestation.
상기 알칼로이드는 식물의 화분 유래인 것이 특징이며, 상기 식물의 화분은 유채꽃, 도토리꽃, 밤꽃 중 어느 하나의 화분 또는 이들의 혼합 화분인 것이 특징이다.The alkaloid is characterized in that it is derived from a flower pot of a plant, and the flower pot of the plant is characterized by being a flower pot of any of rape flower, acorn flower, and chestnut flower, or a mixture thereof.
상기 알칼로이드는 조성물 전체 중량을 기준으로 0.001~0.01 중량%로 함유되는 것이 특징이다.The alkaloid is contained in an amount of 0.001 to 0.01% by weight based on the total weight of the composition.
상기 조성물은 항바이러스 활성 및 항균활성을 갖는 플라보노이드를 더 포함하는 것이 특징이다.The composition is characterized by further comprising a flavonoid having an antiviral activity and an antibacterial activity.
상기 플라보노이드는 상기 조성물 전체 중량을 기준으로 0.001~0.05 중량%로 함유되는 것이 특징이다.The flavonoid is contained in an amount of 0.001 to 0.05% by weight based on the total weight of the composition.
상기 조성물은 설탕, 당액, 시트르산, 화분유래 단백질, 비타민, 화분여과 고형물 중 선택된 어느 하나 이상을 더 포함하는 것이 특징이다.Wherein the composition further comprises at least one selected from sugar, sugar solution, citric acid, protein derived from potted plants, vitamin, and filter cake solid.
본 발명의 꿀벌의 낭충봉아부패병 방제방법은 상기의 꿀벌의 면역증강용 조성물을 꿀벌에 급이하여 꿀벌의 낭충봉아부패병을 방제하는 것이 특징이다.
The present invention is characterized in that the composition for controlling immunity of bees is administered to bees to prevent the infestation of insect bites of bees.
본 발명에 의해, 낭충봉아부패병 발생을 억제하는 꿀벌의 면역증강용 조성물이 제공되며 특히, 꿀벌 사육 농가에서 이용할 때 꿀벌산업의 가장 심각한 생산성 저해요인인 질병 발생의 조절이 용이해지고 꿀벌의 생리 개선을 통해 꿀벌을 관리하는데 수월하여 생산물의 생산을 통한 수익창출에 실질적으로 도움이 되어 양봉 농가에도 신소득원 창출의 효과를 얻게 된다.
According to the present invention, there is provided a composition for immunizing a honeybee that inhibits the incidence of insect bite infestation. In particular, when used in a honeybee farm, it is possible to control the occurrence of disease, which is the most serious obstacle to productivity of bees industry, It is easy to manage the bees through the production, and it is beneficial to the profit generation through the production of the product, and the effect of the creation of the new income source is obtained in the beekeeping farmhouse.
도 1은 화분에서 분리한 알칼로이드 성분을 나타낸 TLC 분석을 나타낸 도면이다.
도 2는 본 발명의 꿀벌의 면역증강용 조성물의 항바이러스 시험 결과를 나타낸 도면이다.
도 3은 본 발명의 꿀벌의 면역증강용 조성물에 대한 항균 활성을 나타낸 사진도면이다.
(A) 슈도모나스 에루지노사(Pseudomonas aeruginosa)
(B) 엔테로코커스 패칼리스(Enterococcus faecalis)
(C) 바실러스 서브필러스(Bacillus subtilis)
(D) 스타필로코커스 아우레우스(Staphylococcus aureus , 황색포도상 구균)
(E) 칸디다 알비칸스(Candida albicans)Brief Description of the Drawings Figure 1 shows TLC analysis showing alkaloid components separated from pollen.
2 is a diagram showing the results of an antiviral test of a composition for immunization of bees of the present invention.
3 is a photographic view showing the antibacterial activity of the bee's composition for immunity enhancement of the present invention.
(A) Pseudomonas erosinosa aeruginosa )
(B) Enterococcus faecalis )
(C) Bacillus subtilis subtilis )
(D) Staphylococcus ( Staphylococcus aureus) aureus , Staphylococcus aureus)
(E) Candida albicans ( Candida albicans )
본 명세서에 기재된 용어, 기술 등은 특별한 한정이 없는 한, 본 발명이 속하는 기술 분야에서 일반적으로 사용되는 의미로 사용된다.The terms, techniques, and the like described in this specification are used in the meaning commonly used in the technical field to which the present invention belongs, unless otherwise specified.
본 발명의 꿀벌의 면역증강용 조성물은 낭충봉아부패병을 일으키는 바이러스에 대해 항바이러스 활성을 갖는 알칼로이드를 유효성분으로 포함하는 것이 특징이다.The composition for immunity enhancement of bees of the present invention is characterized by comprising an alkaloid having an antiviral activity as an active ingredient against a virus that causes insect bite infestation.
설명하면, 상기 낭충봉아부패병은 꿀벌의 유충에서 발생하는 바이러스성 질병으로써, 꿀벌의 유충이 바이러스(Sacbrood virus, SBV)에 의해 감염되어 번데기가 되지 못하고 말라 죽게 하는 병이다.As described above, the infectious disease is a viral disease that occurs in a larva of a bee, and the bee larva is infected by a virus (Sacbrood virus, SBV), and is unable to become a pupa.
이에, 양봉농가에서는 현재 상기 바이러스의 감염을 방지하는 방법으로써 그 바이러스가 벌터에 도착하지 않도록 하는 것이며, 감염이 되었을 경우에는 확산방지를 위해서 반경 6킬로미터 안의 모든 토종벌을 살처분하는 방법만을 시행하고 있는 실정이다.Therefore, the beekeeping farmer is currently preventing the virus from infecting the virus, so that the virus does not reach the bulb. In case of infection, only the method of disposing all the nursery beets within a radius of 6 km is carried out It is true.
본 발명은 상기와 같은 손실을 방지하기 위해 낭충봉아부패병을 일으키는 바이러스에 대해 항바이러스 활성을 갖는 물질을 추출해 낸바, 알칼로이드 성분이 낭충봉아부패병을 일으키는 바이러스에 대해 항바이러스 활성을 나타냄을 확인하게 되었다.In order to prevent the above-mentioned loss, the present invention extracts a substance having an antiviral activity against the virus causing the infectious disease, and confirmed that the alkaloid component exhibits antiviral activity against the virus causing the infectious disease of the insect bastard.
이러한 상기 알칼로이드는 질소를 함유하는 염기성 유기화합물로써 일반적으로는 질소의 노폐물, 식물체의 보호, 해독 작용 등에 사용되어 왔다. Such alkaloids are basic organic compounds containing nitrogen and have been generally used for the waste of nitrogen, the protection and detoxification of plants.
본 발명은 상기와 같은 알칼로이드 성분을 화분(꽃가루)으로부터 추출해 낸 것을 사용하며, 특히 식물의 화분을 유기용매 또는 물과 혼합한 혼합물로부터 추출한 추출액에서 분리된 알칼로이드에서 낭충봉아부패병을 일으키는 바이러스에 대해 항바이러스 활성을 나타냄으로써 본 발명에서는 꿀벌의 면역 증강용 조성물의 유효성분으로 상기와 같은 알칼로이드를 포함하게 된다.The present invention uses the above-mentioned alkaloid component extracted from a pollen (pollen), and particularly relates to a virus that causes insect bite corruption in an alkaloid separated from an extract obtained by mixing a pollen of a plant with an organic solvent or water, The present invention includes the alkaloid as an active ingredient of the composition for enhancing immunity of bees.
상기 식물의 화분은 다량의 알칼로이드 성분을 갖고 있는 유채꽃, 도토리꽃, 밤꽃 중 선택된 어느 하나의 화분 또는 이들이 혼합 화분이 섞여 있는 것이며, 상기 알칼로이드 성분은 본 발명의 조성물의 전체 중량을 기준으로 0.001~0.01 중량%로 함유하는 것을 특징으로, 0.001 중량% 미만으로 함유할 경우 항바이러스 활성이 낮아 낭충봉아부패병을 방지하는 효과가 적게 나타나 꿀벌 면역증강용 조성물로 사용하기에 적합하지 않으며, 0.01 중량%를 초과하여 함유할 경우에는 오히려 알칼로이드 성분에 의해 꿀벌유충의 성장에 영향을 미치게 된다.The plant pot may be any one selected from rapeseed flowers, acorn flowers, and chestnut flowers having a large amount of alkaloid components, or a mixture thereof, and the alkaloid component may be present in an amount of 0.001 - 0.01% by weight. When the content is less than 0.001% by weight, the antiviral activity is low and the effect of preventing scabbing of insect pea is less. Therefore, it is not suitable for use as a composition for bee immune enhancement, and 0.01% If it is contained in excess, the alkaloid component rather affects the growth of bee larvae.
이러한, 상기 본 발명의 꿀벌의 면역 증강용 조성물에는 항바이러스활성뿐만 아니라 항균활성도 갖기 위해 플라보노이드를 더 포함하기도 한다. The composition for immunizing a bee of the present invention may further contain a flavonoid to have antibacterial activity as well as antiviral activity.
상기 플라보노이드는 식품에 널리 분포하는 노란색 계통의 색소로서 항바이러스, 항암, 항바이러스, 항알레르기 및 항염증 활성을 지니며, 독성이 거의 없는 것으로 보고되어 있다.The flavonoids are yellow-based pigments widely distributed in foods and have antiviral, anticancer, antiviral, antiallergic and anti-inflammatory activities, and are reported to have little toxicity.
이러한 플라보노이드는 본 발명의 꿀벌의 면역 증강용 조성물 전체 중량을 기준으로 0.001~0.05 중량%로 함유하는 것을 특징으로, 상기 조성물 전체 중량을 기준으로 0.001 중량% 미만으로 함유될 경우에는 항바이러스 및 항균활성이 낮아 상기 알칼로이드만을 사용한 경우보다 더 높은 상승 효과를 기대하기 어려우며, 항균활성도 낮게 나타나 꿀벌의 면역증강용 조성물로 사용하기에 적합하지 않으며, 0.05 중량%를 초과하여 함유할 경우에는 전체적으로 꿀벌의 급이를 용이하게 하기 위해 포함될 타 성분들이 적게 함유되게 되어 꿀벌유충 성장을 오히려 방해하게 된다. Such a flavonoid is contained in an amount of 0.001 to 0.05% by weight based on the total weight of the composition for immunizing a bee of the present invention. When the flavonoid is contained in an amount of less than 0.001% by weight based on the total weight of the composition, antiviral activity and antimicrobial activity It is difficult to expect a higher synergistic effect than the use of the alkaloid alone, and the antimicrobial activity is also low. Therefore, it is not suitable for use as a composition for enhancing the immunity of bees. When the content exceeds 0.05% by weight, Lt; RTI ID = 0.0 > bee larvae < / RTI >
이와 같이 본 발명의 꿀벌의 낭충봉아부패병 방제방법은 상기 꿀벌의 면역증강용 조성물을 꿀벌에 급이를 통해 항바이러스 활성을 나타내어 꿀벌의 낭충봉아부패병을 방제하는 것을 특징으로 꿀벌들이 상기 본 발명의 꿀벌의 면역 증강용 조성물을 용이하게 급이하도록 하는 보조 성분으로 설탕, 당액, 시트르산, 화분유래 단백질, 비타민, 화분여과 고형물 중 어느 하나 이상을 더 포함하기도 하며, 특히 화분유래 단백질과 화분 여과 고형물은 화분자체가 껍질구조의 특이성으로 인해 난소화성을 갖고 있으므로 이를 보완해주는 역할을 함과 동시에, 단백질을 공급해줌으로써 추가적으로 유충의 성장에 도움을 주게 된다.
As described above, the method for controlling insect annoying disease of bees according to the present invention is characterized in that the composition for immunity enhancement of the bees exhibits antiviral activity through feeding to a bee, thereby controlling insect bite annoyance of the bee. The protein content of the pollen-derived protein and the pollen-filtered solid matter may be more than the pollen-derived protein, the pollen-derived protein, the vitamin- Because it has ovariability due to the specificity of the shell structure, it plays a role of complementing it and at the same time it helps the growth of the larva by supplying protein.
이하, 본 발명에 대하여 실시예 및 실험예를 통하여 상세히 설명하나, 이들이 본 발명의 범위를 제한하는 것은 아니다.Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples, but the scope of the present invention is not limited thereto.
<실시예 1> 본 발명의 꿀벌의 면역 증강용 조성물 제조1Example 1 Preparation of composition for immunity enhancement of bees of the present invention 1
알칼로이드 성분을 하기와 같은 방법으로 분리해 내었다. The alkaloid components were separated in the following manner.
먼저 화분 20kg(잡화)을 준비한 후 여기에 에탄올(EtOH) 72ℓ와 1차 증류수 72ℓ를 넣고 침전시킨 후 2틀이 경과된 다음 거즈(Miracloth 1R)로 1회 걸러 내었다. First, 20 kg of pots (miscellaneous) was prepared, 72 ℓ of ethanol (EtOH) and 72 ℓ of primary distilled water were added and precipitated.
여기서 걸러진 액만을 보아 엠버리스트(Amberlist 15)를 통해 알칼로이드 성분만을 흡착해 낸 후 0.5N NH4OH로 내린 다음 감압농축한 후 농축액을 Dowexⓡ1×2,50-100mesh 로 흡착하여 3차증류수로 내려은 다음 3차 멸균수로 10배 희석한 화분추출액(알칼로이드)을 포함하는 본 발명의 꿀벌의 면역 증강용 조성물을 제조하였다.
After only the alkaloid component was adsorbed through Amberlist 15, it was reduced with 0.5N NH 4 OH and concentrated under reduced pressure. The concentrate was adsorbed on Dowex 占 1 × 2, 50-100 mesh, The composition for immunity enhancement of bees of the present invention containing the pollen extract (alkaloid) diluted 10 times with the third sterilized water down was prepared.
<실시예 2> 본 발명의 꿀벌의 면역 증강용 조성물 제조2Example 2 Preparation of composition for immunity enhancement of bees of the present invention 2
물 10ℓ, 설탕 15Kg, 플라보노이드액(플라보노이드 0.1%함유)1000㎖, 당액(fructose/glucose=1.5) 21ℓ, 시트르산(Citric acid) 200g, 비타민 조성물 400g, 상기 실시예1에서 얻은 화분추출액(알칼로이드) 100㎖, 화분단백질 10%, 화분여과고형물 1%로 이루어져 총 40kg인 본 발명의 꿀벌의 면역 증강용 조성물을 제조하였다.(Alkaloid) 100 (100 g) obtained in Example 1, 10 liters of water, 15 kg of sugar, 1000 ml of flavonoid solution (containing 0.1% of flavonoid), 21 liters of fructose / glucose = 1.5, 200 g of citric acid, Ml, 10% of pollen protein, 1% of pollen filtration solid, and a total weight of 40 kg, to prepare the bee immune enhancing composition of the present invention.
이때, 상기 비타민 조성물(본제 1kg중)은 하기와 같은 조성으로 이루어져 있다.At this time, the vitamin composition (in 1 kg of the present invention) has the following composition.
Vitamin A 5,000,000 I.UVitamin A 5,000,000 I.U
Vitamin D 1,000,000 I.U Vitamin D 1,000,000 I.U
Vitamin E 1,000 I.UVitamin E 1,000 I.U
Vitamin C 1,000 mgVitamin C 1,000 mg
Vitamin K 400 mgVitamin K 400 mg
Vitamin B 1,000 mgVitamin B 1,000 mg
Vitamin B6 400 mgVitamin B6 400 mg
Vitamin B12 400 mcgVitamin B12 400 mcg
Niacin 400 mgNiacin 400 mg
Ca-pantothenate 2,000 mgCa-pantothenate 2,000 mg
Folic acid 20,000 mcgFolic acid 20,000 mcg
Sodium chloride (Na로써) 3,740 mgSodium chloride (as Na) 3,740 mg
Potassium chloride (K로써) 870 mgPotassium chloride (with K) 870 mg
Ferrous Sulfate (Fe로써) 2,000 mgFerrous Sulfate (as Fe) 2,000 mg
Manganese Sulfate (Mn으로써) 360,000 mcgManganese Sulfate (as Mn) 360,000 mcg
Zinc Sulfate (Zn으로써) 160,000 mcgZinc Sulfate (as Zn) 160,000 mcg
Potassium Iodide (I로써) 80,000 mcgPotassium Iodide (as I) 80,000 mcg
Cobalt Sulfate (Co로써) 40,000 mcgCobalt Sulfate (as Co) 40,000 mcg
Selenium (Se로써) 20,000 mcg
Selenium (as Se) 20,000 mcg
또한, 상기 화분단백질은 하기와 같은 추출방법을 통해 얻은 것을 사용하였다. In addition, the pollen protein obtained by the following extraction method was used.
설명하면, 먼저 화분 10g 준비한 후, 90㎖의 0.1M sodium phospate buffer 넣고 30분간 저어준 다음, 0.5N NaOH를 이용하여 pH 7로 맞춘 후, 60℃에서 Alcalase 0.2unit을 넣고 2시간 동안 교반한 다음, 90℃에서 20분간 끓여서 효소반응을 없앤 후, 1200g에서 원심분리를 15분간 한 다음 paper filter로 걸러서 여과액을 최종 조성물의 10% 첨가하여 사용하였다.
Prepare 10g of pollen first, add 90ml of 0.1M sodium phospate buffer, stir for 30 minutes, adjust to pH 7 with 0.5N NaOH, add 0.2unit of Alcalase at 60 ℃ and stir for 2 hours , Boiled at 90 ° C for 20 minutes to remove the enzyme reaction, centrifuged at 1200 g for 15 minutes, filtered with a paper filter, and 10% of the final composition was added to the filtrate.
또한, 상기 화분여과고형물은 먼저 여과지에 걸러진 고형물을 회수한 후, 이 고형물을 동결건조 하여 최종 조성물의 1%로 첨가하여 사용하였다.In addition, the pollen filtration solids were first recovered from the filter paper, and the solids were lyophilized and added to 1% of the final composition.
<실험예 1> 화분내 함유하는 5종의 알칼로이드 확인 및 급이를 통한 꿀벌 바이러스에 대한 항바이러스 활성 확인
<Experimental Example 1> Identification of 5 kinds of alkaloids contained in the pollen and confirmation of antiviral activity against bee virus through feeding
1. 실험방법1. Experimental Method
1) 알칼로이드 확인1) Identification of alkaloids
TLC 검정을 시행하여 도 1과 같이 나타났다.TLC assay was performed as shown in Fig.
설명하면, TLC 플레이트(plate) 전개를 위하여 실리카겔(silica gel) 윗부분 1.5cm정도의 silica gel은 칼로 긁어서 제거하고 아래에서 1.5cm 부분에 연필로 선을 긋고 용액의 시작점을 표시하였다.For explanation, silica gel of 1.5 cm on the upper part of silica gel was scratched off with a knife, and a line with a pencil was drawn at 1.5 cm from the bottom to mark the starting point of the solution for the development of the TLC plate.
시작점에 용액을 떨어뜨리고 전개용매에 0.5cm정도 잠기게 silica gel을 넣고 1시간정도 전개시켰으며, 전개용매로는 이소프로판올(iso prophanol) : 아세트산(acetic acid) : H2O = 4 : 1 : 1의 비율로 조성한 액을 사용하여 전개 후 TLC판을 말린 다음 발색용액를 분무하고 말린 다음 발색용액을 분무하고 말려서 확인하였다.The solution was dropped at the starting point, and silica gel was immersed in the developing solvent to a depth of about 0.5 cm and developed for about 1 hour. Isopropanol: acetic acid: H 2 O = 4: 1: 1 , The TLC plate was dried and then the coloring solution was sprayed and dried, followed by spraying and drying the color developing solution.
발색용액은 O-톨리딘(O-Tolidine) 32mg을 acetic acid 3ml에 녹여 H2O로 총 볼륨(total volume) 50ml로 만들고 요오드화칼륨(Potassium iodide) 0.2g H2O로 total volume 50ml 만든 것을 섞은 것이며, 발색용액은 50% 치아염소산나트륨을 사용하였다.
The coloring solution was prepared by dissolving 32 mg of O-Tolidine in 3 ml of acetic acid, making a total volume of 50 ml with H 2 O, and making a total volume of 50 ml with potassium iodide (0.2 g H 2 O) And 50% sodium hypochlorite was used as a coloring solution.
2) 항바이러스 활성실험2) Antiviral activity experiments
항바이러스 효과 검정을 위하여 꿀벌을 채집하여 PCR법을 이용하여 바이러스에 감염이 된 꿀벌을 모아서 30마리씩 유발을 이용하여 5ml의 10mM KP buffer로 마쇄하고 미라클로스(Miracloth)로 여과한 후 투과된 액을 10,000g으로 1시간 동안 원심분리하여 상층액을 다시 180,000g의 속도로 2시간 30분간 원심분리한 후 10mM의 KP buffer 1ml에 침전물을 녹여서 당농도구배 초원심분리하여 바이러스만을 따로 모은 후 바이러스 농도를 계산하여 꿀벌간이 사육용 캐이지에 꿀벌 100마리씩 넣고 설탕물에 타서 바이러스를 꿀벌에 먹인 후 면역증강제를 처리하여 꿀벌 바이러스 감소 정도를 조사하였다.
In order to test the antiviral effect, bees were collected, and the virus-infected bees were collected using PCR method. Then, 30 bees were ground with 5 ml of 10 mM KP buffer using induction, filtered with Miracloth, After centrifuging at 10,000 g for 1 hour, the supernatant was centrifuged at 180,000 g for 2 hours and 30 minutes. The precipitate was dissolved in 1 ml of 10 mM KP buffer, In order to investigate the reduction of honey bee virus, 100 bees were added to the bee cat breeding cage, and the virus was fed to bees and then treated with the immune enhancer.
2. 실험결과2. Experimental results
상기 실시예1의 본 발명의 꿀벌의 면역 증강용 조성물에 대하여 항바이러스 활성실험을 확인 한 결과, 도 2에 나타나 있듯이, 1달 후 꿀벌바이러스가 발생하지 않은 것을 확인하였다. 이러한 결과를 토대로 농가 실증시험을 실시하여 면역증강제의 항바이러스활성을 확인하였다.As a result of checking the antiviral activity of the composition for immunity enhancement of bees of the present invention of Example 1, it was confirmed that bee virus did not occur after one month as shown in FIG. Based on these results, we conducted a farm test to confirm the antiviral activity of the immune enhancer.
또한, 하기 표 1과 같이 낭충봉아부패병을 일으키는 바이러스에 대해 항바이러스 활성을 나타냄을 알 수 있었다.In addition, as shown in Table 1, antiviral activity against viruses causing cystic fibrosis was shown.
Treated group
Survival group (+ proliferative group)
음성(15군)Positive (group 15)
Voice (group 15)
<실험예 2> 본 발명의 꿀벌의 면역력 증강용 조성물의 항균력 확인<Experimental Example 2> Determination of the antibacterial activity of the composition for enhancing immunity of bees of the present invention
1. 실험방법1. Experimental Method
실시예 2의 꿀벌의 면역 증강용 조성물의 항균력을 검사하기 위해 (A) 슈도모나스 에루지노사(Pseudomonas aeruginosa), (B) 엔테로코커스 패칼리스(Enterococcus faecalis), (C) 바실러스 서브필러스(Bacillus subtilis), (D) 스타필로코커스 아우레우스(Staphylococcus aureus , 황색포도상 구균), (E) 칸디다 알비칸스(Candida albicans)를 검사대상으로 하였다.To examine the antibacterial activity of the composition for immunity enhancement of bee of Example 2, (A) Pseudomonas aeruginosa aeruginosa , (B) Enterococcus faecalis , (C) Bacillus subtilis subtilis ), (D) Staphylococcus ( Staphylococcus aureus) aureus , Staphylococcus aureus), and (E) Candida albicans .
면역증강제의 항균력 검사는 각각의 균주가 2.5×104의 농도가 되도록 하여 영양 배지에 도말하고 종이 디스크를 놓은 후 대조구(항균물질)과 멸균수, 액상배지, 면역증강제를 각각 100ul씩 처리 후 1일 후 디스크 주변의 투명환을 확인하여 항균능을 검정하였다.
The antimicrobial activity of the immune enhancer was determined by treating the control (antimicrobial substance), sterilized water, liquid medium and immune enhancer in a ratio of 2.5 × 10 4 per 100 μl each The antimicrobial activity was examined by confirming the transparent ring around the disc.
2. 실험결과2. Experimental results
상기 실험결과 도 3에 나타나 있듯이, 본 발명의 꿀벌의 면역 증강용 조성물은 슈도모나스 에루지노사(Pseudomonas aeruginosa), 엔테로코커스 패칼리스(Enterococcus faecalis), 바실러스 서브필러스(Bacillus subtilis), 스타필로코커스 아우레우스(Staphylococcus aureus , 황색포도상 구균), 칸디다 알비칸스(Candida albicans) 균에 대해 항균활성을 나타냄을 알 수 있었다.
As a result of the above experiment, as shown in FIG. 3, the composition for immunization of bees of the present invention is Pseudomonas aeruginosa aeruginosa , enterococcus faecalis faecalis , Bacillus subtilis, subtilis ), Staphylococcus ( Staphylococcus aureus) aureus , Staphylococcus aureus), Candida albicans ( Candida albicans ) against the bacteria.
상기의 결과를 토대로 본 발명에 의해, 바이러스에 의한 질병 방제로 인한 꿀벌의 폐사를 방지하며, 특히 항바이러스 활성뿐만 아니라 항균활성도 갖게 됨으로써 꿀벌의 생리적 환경을 개선하여 꿀벌의 수명 연장 및 면역을 증강시켜 꿀벌 스스로가 이겨낼 수 있도록 하는 꿀벌의 면역증강용 조성물이 제공된다.
Based on the above results, it is possible to prevent the death of bees due to the disease control by viruses, and in particular to have antiviral activity as well as antiviral activity, thereby improving the physiological environment of bees, thereby enhancing the life span and immunity of bees There is provided a composition for enhancing immunity of bees, which enables the bees to overcome themselves.
상기의 본 발명은 바람직한 실시예 및 시험예를 중심으로 살펴보았으며, 본 발명이 속하는 기술분야에서 통상의 지식을 가진 자는 본 발명의 본질적 기술 범위 내에서 상기 본 발명의 상세한 설명과 다른 형태의 실시예들을 구현할 수 있을 것이다. 여기서 본 발명의 본질적 기술범위는 특허청구범위에 나타나 있으며, 그와 동등한 범위 내에 있는 모든 차이점은 본 발명에 포함된 것으로 해석되어야 할 것이다.While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments. You can implement the examples. The scope of the present invention is defined by the appended claims, and all differences within the scope of the claims are to be construed as being included in the present invention.
Claims (8)
꿀벌의 면역증강용 조성물.
An alkaloid having antiviral activity against a virus causing cystic beetle decay disease as an active ingredient,
Bee honeycomb composition.
상기 알칼로이드는 식물의 화분유래인 것을 특징으로 하는,
꿀벌의 면역증강용 조성물.
The method of claim 1,
The alkaloid is characterized in that the plant-derived,
Bee honeycomb composition.
상기 식물의 화분은 유채꽃, 도토리꽃, 밤꽃 중 어느 하나의 화분 또는 이들의 혼합 화분인 것을 특징으로 하는,
꿀벌의 면역증강용 조성물.
3. The method of claim 2,
The pollen of the plant is characterized in that any one of the rapeseed, acorn flowers, chestnut flower pots or a mixture of these,
Bee honeycomb composition.
상기 알칼로이드는 조성물 전체 중량을 기준으로 0.001~0.01 중량%로 함유되는 것이 특징인,
꿀벌의 면역증강용 조성물.
The method of claim 1,
The alkaloid is characterized in that it is contained in 0.001 to 0.01 wt% based on the total weight of the composition,
Bee honeycomb composition.
상기 조성물은 항바이러스 활성 및 항균활성을 갖는 플라보노이드를 더 포함하는 것이 특징인,
꿀벌의 면역증강용 조성물.
The method of claim 1,
The composition is characterized in that it further comprises a flavonoid having antiviral activity and antimicrobial activity,
Bee honeycomb composition.
상기 플라보노이드는 상기 조성물 전체 중량을 기준으로 0.001~0.05 중량%로 함유되는 것이 특징인,
꿀벌의 면역증강용 조성물.
The method of claim 5,
The flavonoid is characterized in that it is contained in 0.001 to 0.05% by weight based on the total weight of the composition,
Bee honeycomb composition.
상기 조성물은 설탕, 당액, 시트르산, 화분유래 단백질, 비타민, 화분여과 고형물 중 선택된 어느 하나 이상을 더 포함하는 것이 특징인,
꿀벌의 면역증강용 조성물.
7. The method according to any one of claims 1 to 6,
The composition is characterized in that it further comprises any one or more selected from sugar, sugar, citric acid, pollen-derived protein, vitamins, pollen filtration solids,
Bee honeycomb composition.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101600983B1 (en) * | 2014-10-15 | 2016-03-08 | 이우철 | Therapeutic and preventive agent for sacbrood of bees |
| CN109393254A (en) * | 2017-08-18 | 2019-03-01 | 彦臣生技药品股份有限公司 | Composition and method for improving insect immunity |
| KR20190073127A (en) * | 2017-12-18 | 2019-06-26 | 전남대학교산학협력단 | Honey bee alternative feed composition using onion, and its manufacturing method |
| KR20210036897A (en) * | 2018-12-12 | 2021-04-05 | 대한민국(농촌진흥청장) | Functional saccharide feed composition for bee |
| CN114712370A (en) * | 2022-05-07 | 2022-07-08 | 中国农业科学院麻类研究所 | Application of tomatidine in preparation of drug for resisting Israel acute paralysis virus |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR102070362B1 (en) | 2018-10-19 | 2020-01-28 | (주)이엘티사이언스 | The composition of immune enhancement for honey bee, and method for manufacturing thereof |
| KR102251942B1 (en) * | 2021-02-25 | 2021-05-17 | 하성섭 | The composition of immune enhancement for beekeeping |
| KR102610687B1 (en) | 2023-04-17 | 2023-12-06 | 주식회사 메디풀 | Immunity enhancer composition for beekeeping and use thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2009103276A2 (en) | 2008-02-18 | 2009-08-27 | Sven Buchholz | Method for combined application of two substances for simultaneous prevention and monitoring of differing types of pest |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101600983B1 (en) * | 2014-10-15 | 2016-03-08 | 이우철 | Therapeutic and preventive agent for sacbrood of bees |
| CN109393254A (en) * | 2017-08-18 | 2019-03-01 | 彦臣生技药品股份有限公司 | Composition and method for improving insect immunity |
| KR20190073127A (en) * | 2017-12-18 | 2019-06-26 | 전남대학교산학협력단 | Honey bee alternative feed composition using onion, and its manufacturing method |
| KR20210036897A (en) * | 2018-12-12 | 2021-04-05 | 대한민국(농촌진흥청장) | Functional saccharide feed composition for bee |
| CN114712370A (en) * | 2022-05-07 | 2022-07-08 | 中国农业科学院麻类研究所 | Application of tomatidine in preparation of drug for resisting Israel acute paralysis virus |
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