KR101798425B1 - Insecticidal compositions comprising extract of Dichapetalum moralesii Prance - Google Patents

Abstract

The present invention relates to an insecticidal composition comprising a Dichapetalum extract or a fraction thereof as an active ingredient, and a method of insecticidal use using the same. More specifically, the composition of the present invention comprising Dichapetalum moralesii Prance extract or a fraction thereof as an active ingredient has an excellent insecticidal effect against pests, and is environmentally friendly as a plant natural insecticide and harmless to human , Agricultural pests, hygiene pests, forest pests or plant pathogens.

Description

[0001] The present invention relates to an insecticidal composition comprising extracts of Dicapetalum moralesii Prince,

The present invention relates to an insecticidal composition comprising an extract of Dicapetalum moralesii Prance or a fraction thereof as an active ingredient and a method of insecticidal use using the same.

The mosquito females require the supply of animal protein for the maturation of the egg and access to humans and animals to vomit. The blood-sucking activity of mosquitoes is the mediator of major diseases such as malaria, encephalitis, yellow fever and dengue fever. In addition, due to bloodshed, the bite area causes redness and stunning, and severe itching causes sleep disturbance. In addition, serious scratches can lead to direct damage such as inflammation caused by secondary infection. Domestic mosquito-borne diseases tend to increase the number of patients with encephalitis caused by malaria and small red mosquito ( Culex tritaeniorhynchus ) caused by Chinese Anopheles sinensis .

With the recent warming, mosquitoes have a tendency to increase the number of mosquito species because of the faster cycle of vomiting - resting - spawning. In addition, mosquitoes are living in large buildings such as apartments and department stores due to the expansion of heating facilities.

Methods for controlling mosquitoes that are mediators of disease threatening human health include chemical control and adult control by in vitro fertilization and sterilization by biological and chemical control methods. To date, an important means of controlling mosquitoes has been to rely on pesticides. A liquid containing an insecticide is dissolved in a matte electromagnetism or a wick plug with a linear mosquito repellent which burns the medium containing the insecticide, or the insecticidal component is impregnated in the mat and placed on the electric hot plate. A liquid electron mosquito coil which diffuses an insecticidal component by heating the wick is used.

Domestic insecticides are dominated by pyrethroid insecticides such as permethrin. The pyrethroid system is a representative chemical synthesizer that has been used as a repellent and insecticide effective for mosquitoes, flies, ticks, etc. at a relatively low concentration. It is known that permethrin, allethrin, deltamethrin, Cypermethrin and the like are known. Permethrin is a contact deterrent, rather than a post-emergent repellent, which is toxic to the nervous system of the mosquito, but not toxic to mammals. It is safe as good spawning inhibitors (deterrents) and provides good defense when used in conjunction with DEET.

However, insecticides are mainly used in the form of mats, coils and aerosols rather than applied directly to the skin, thus increasing the permeability of the blood-brain barrier during inhalation, resulting in oxidative damage in the brain, liver and kidney, There is a possibility of health risk in infants and young children, and the damage caused by insecticides such as showing toxicity to the human body due to use of mosquito repellent in closed room is increasing. In addition, the use of pyrethroid insecticides significantly increases the resistance of mosquitoes, and in fact, fungi using the mist method does not affect the mosquitoes at all and rather harms the useful insects, thus adversely affecting the ecosystem.

Although the use of synthetic insecticides for controlling mosquitoes causes side effects such as drug resistance, killing of useful creatures, toxicity to humans and livestock, residuals and various environmental pollution, the use of synthetic insecticides is increasing day by day to be. In this respect, developed countries have long been focusing on the use and application of biochemical intermediates (biocides) in ecosystems from chemical synthetic insecticides, and pest control attempts have been made using them. Examples include microbial means using bacteria ( Bacillus thuringiensis ) and nuclear polyhedrosis virus (granulosis virus, cytoplasmic polyhedrosis virus, etc.), pheromones of insects, toxins of species-specific secondary metabolites present in plants, These are methods of using attractants, feeding or scattering deterrents, repellents and insect growth regulators. Recently, however, the proportion of these is increasing.

As described above, studies for replacing pesticides against mosquito larvae and adults have been carried out all over the world, but no satisfactory materials have been found yet.

On the other hand, Dichapetalum It has been disclosed that plant extracts belonging to the genus can be used as a therapeutic agent for AIDS (Application No. 1020097016128 (Sep. 11, 2003)) mixed with other plant extracts. In addition, it has been found that toxic components are present in the seeds of Dichapetalum toxicarium and the leaves of D. cymosum (RA PETERS and RW WAKELIN, 1959, Biochem. J., February; 71 (2): 245- 248), the insecticidal effect of Dichapetalum moralesii Prance extract and its fractions has not been elucidated yet.

Accordingly, the present inventors have completed the present invention in order to develop an environmentally friendly and highly insecticidal insecticide capable of replacing a chemical insecticide which damages the environment and damages human life.

It is an object of the present invention to provide an insecticidal composition comprising as an active ingredient an extract of Dicapetalum moralesii Prance or a fraction thereof.

Another object of the present invention is to provide a method for insecticidal insemination using the insecticidal composition.

The present invention relates to a process for the preparation of Dichapetalum Which comprises a plant extract belonging to the genus Escherichia or a fraction thereof as an active ingredient.

The plant is Dicapateralum moralesii Prance.

In the present invention, the insecticidal composition and the insecticide have the same meaning unless otherwise stated.

The Dicapetalum moralesii Prance extract contained in the composition of the present invention can be extracted by a known extraction method, and the method is not limited thereto. For example, the extract may be prepared by washing, drying and pulverizing Dicapetalum moralesii Prance; And extracting the pulverized dicapateralum moralecia france with a solvent selected from water, an organic solvent or a mixed solvent thereof.

The extract can be extracted using water or an organic solvent, and the kind of the organic solvent is not limited. For example, those extracted with a solvent selected from the group consisting of water, lower alcohols having 1 to 4 carbon atoms, and mixed solvents thereof, preferably methanol, ethanol or butanol. More preferably, the extract is extracted with methanol. In addition, in the present invention, the extract includes any one of the extract obtained by the extraction treatment, the diluted solution of the extract, the concentrate, the dried product obtained by drying the extract, and the adjusted product or the purified product.

The extraction method is not particularly limited, and extraction can be carried out at room temperature or with heating under conditions where the active ingredient is not destroyed or minimized. More specifically, in the present invention, a method for obtaining a dicapateralum morlareifrance extract is as follows. The dried dicapateralum moraleciferum is dissolved in a polar solvent of a C1 to C4 lower alcohol such as water, methanol, ethanol, and butanol, or the like, having a volume of about 2 to 20 times, preferably about 3 to 5 times the dry weight, A mixed solvent having a mixing ratio of about 1: 0.1 to 1:10 is used as an elution solvent, and the extraction temperature is 20 to 100 캜, preferably 25 캜, the extraction period is about 3 to 10 days, preferably 4 Extraction is carried out using extraction methods such as shaking, hot water extraction, cold extraction, reflux cooling, or ultrasonic extraction for 10 hours. According to a specific embodiment of the present invention, the naturally dried powder of Dicapateralum Moraleshifranus was extracted with methanol at room temperature, filtered using a filter paper, and evaporated at 35 DEG C under vacuum.

The Dichapetalum moralesii Prance extract can be extracted from various organs of natural, hybrid, and variant plants and can be extracted from various organs such as roots, ground parts, stems, leaves, flowers, fruit bodies, But it can be extracted from plant tissue culture, and it is preferable to extract from the aerial part of the plant.

In the present invention, the solvent fraction of Dichapetalum moralesii Prance extract is suspended in distilled water and then fractionated using a polar solvent such as hexane, ethyl acetate or a nonpolar solvent such as chloroform or weakly polar butanol, A solvent fraction and a non-polar solvent fraction, respectively, can be obtained. In one embodiment of the present invention, the solvent fraction is an ethyl acetate fraction, a hexane fraction, a chloroform fraction or a butanol fraction. The fraction may be obtained by a method known in the art, for example, by suspending a methanol extract of Dichapetalum moralesii Prance in distilled water, and then adding about 1 to 100 times, preferably A non-polar solvent such as hexane, ethyl acetate or chloroform at a volume of about 1 to 5 times the volume of the non-polar solvent-soluble layer, and extracting and separating the non-polar solvent-soluble layer from 1 to 10 times, preferably 2 to 5 times. In addition, conventional fractionation processes can also be performed (Harborne JB Phytochemical methods: A guide to modern techniques of plant analysis, 3rd Ed. P6-7, 1998).

The insecticidal object of the composition is not limited and may include agricultural pests, hygiene pests and forest pests. In addition, the insecticidal object includes both adults and larvae. The hygiene pests are mosquitoes, wheels, flies, mites are preferred, more preferably, Culex Phi Enschede arm lances (Culex pipiens pallens), Culex tree other enioh hinge kusu (Culex tritaeniorhynchus), Culex biswinuyi (Culex from the group consisting of vishnui , Culex gelidus , Culex fuscocephala , Aedes aegypti , Togo forest mosquito ( Aedes togo ) and Anopheles stephensi It is one or more mosquito selected.

When used as an active ingredient of the insecticide of the present invention, Dicapetalum moralesii Prance extract can be used as such, without the addition of any other ingredients. The insecticide of the present invention may also be prepared by mixing Dichapetalum moralesii Prance extract with a conventional solid carrier, liquid carrier, gas carrier or bait, or absorbing it into a basic substance such as a porous ceramic plate or nonwoven fabric A surfactant and if necessary other adjuvants are added to the composition to formulate it in various forms such as an oil spray, a mat, a coil, an emulsifiable concentrate, a powder, a wettable powder, a milk animal, a microcapsule, , Aerosols, fumigants (e.g., fogging), vaporizable formulations, smoking formulations, toxic handouts, anti-mite sheets or resin formulations.

The insecticide of the present invention usually contains 0.01 to 95% by weight of Dicapetalum moralesii Prance extract as an active ingredient. More preferably, the extract of Dicapetalum moralesii Prance is contained in the composition at a concentration of 1 to 100 ppm.

Solid carriers that can be used in the insecticides of the present invention include fine powders or granules of clay materials such as kaolin clay, diatomaceous earth, synthetic hydrated silicon oxide, bentonite, clay and clay; Various talc, ceramics and other inorganic materials such as sericite, quartz, sulfur, activated carbon, calcium carbonate and hydrated silica; And chemical fertilizers such as ammonium sulphate, ammonium phosphate, ammonium nitrate, urea and ammonium chloride.

Liquid carriers include water; Alcohols such as methanol and ethanol; Ketones such as acetone and methyl ethyl ketone; Aromatic hydrocarbons such as benzene, toluene, xylene, ethylbenzene and methylnaphthalene; Aliphatic hydrocarbons such as hexane, cyclohexane, kerosene and light oil; esters such as ethyl acetate and butyl acetate; Nitriles such as acetonitrile and isobutyrnitrile; Ethers such as diisopropyl ether and dioxane; Acid amides such as N, N-dimethylformamide and N, N-dimethylacetamide; Halogenated hydrocarbons such as dichloromethane, trichloroethane and carbon tetrachloride; Dimethyl sulfoxide; And vegetable oils, such as soybean oil and cottonseed oil.

The gas carrier or propellant may include Freon gas, butane gas, LPG (liquefied petroleum gas), dimethyl ether, and carbon dioxide.

Base materials for use in toxic handouts include handouts, such as grain powders, vegetable oils, sugars and crystalline cellulose; Antioxidants such as dibutylhydroxytoluene and nordihydroguaiaretic acid; Preservatives, such as dehydroacetic acid; Non-foraging materials, such as red pepper powder; And an attractiveness flavor, such as cheese flavor and onion flavor.

Surfactants may include alkyl sulfates, alkyl sulfonates, alkyl aryl sulfonates, alkyl aryl ethers and polyoxyethylene derivatives thereof, polyethylene glycol ethers, polyhydric alcohol esters and sugar alcohol derivatives.

Adjuvants such as adhesives or dispersing agents include casein, gelatin; Polysaccharides such as starch, gum arabic, cellulose derivatives and alginic acid; Lignin derivatives, bentonites, sugars and synthetic water-soluble polymers such as polyvinyl alcohol, polyvinyl pyrrolidone and polyacrylic acid.

Stabilizers include PAT (isopropyl acid phosphate), BHT (2,6-di-tert-butyl-4-methylphenol), BHA (2-tert- 4-methoxyphenol), vegetable oils, mineral oils, surfactants, fatty acids and esters thereof.

When the insecticides of the present invention are used as agricultural insecticides, fungicides or nematocides, their application amount is usually 0.1 to 100 g per 10 acres. For emulsifiable concentrates, wettable powders, animal and other similar formulations used after dilution with water, the application concentration is usually in the range of 1 to 100,000 ppm. Application of granules, dust or other similar formulations is carried out as an undiluted formulation. When the insecticides of the present invention are used as insecticides, fungicides or nematocides for the prevention of epidemics, they may be present in concentrations of from 0.1 to 500 ppm in water for emulsifiable concentrates, wettable powders, animal or other similar formulations Diluted or applied in the case of oil sprays, aerosols, fumigants, toxic handouts, antidicking sheets or other similar formulations. Such application amount and concentration may vary depending on the form of the formulation, application time, place and method, type of pest, degree of damage, and other factors, so that it is not limited to the above range and can be increased or decreased.

When the insecticide of the present invention is used as a live insecticidal or acaricidal agent for the control of parasites of domestic animals such as cattle and pigs, or pets such as cats and dogs, the composition or a salt thereof may be administered by known veterinary methods, Tablets, capsules, solutions for submersion, boli, food incorporation, suppositories or injections for control; Or by spraying, pouring (dripping or dripping) the treatment of oily or aqueous solutions, or by using molded articles of suitable shape such as collar and ear tags (tags) for non-systematic control. In this case, the present compound is usually applied in an amount of 1 to 100 mg per kg body weight of the host.

The insecticides of the present invention may be used in combination with other insecticides, nematicides, fungicides, fungicides, fungicides, herbicides, plant growth regulators, synergists, fertilizers, soil conditioners and / or animal feeds, But can be used at the same time.

In one embodiment of the present invention, the methanol extract of Dichapetalum moralesii Prance was treated with a third larva of Aedes aegypti and Culex pipiens pallens , and the concentration of the extract Dependent mosquito insecticidal effect. First, the Dichapetalum moralesii Prance extract was fractionated using ethyl acetate (EtOAc), hexane (Hexane), chloroform (CHCl ₃ ), butanol (BuOH) and water, And the insecticidal effect was confirmed (Examples 3 and 4). More specifically, in the case of butanol, ethyl acetate and hexane fractions, the insecticidal effect was found only in the Aedes aegypti . In the case of the methanol extract and the chloroform fraction, the Aedes aegypti and the red house mosquito Culex pipiens pallens ) showed insecticidal activity. On the other hand, the water fraction of Dichapetalum moralesii Prance did not show the insecticidal effect in the Aedes aegypti and Culex pipiens pallens .

Based on the above results, it was confirmed that the extract and fraction of Dicapetalum moralesii Prance of the present invention had excellent insecticidal effect against insects, especially mosquitoes.

In another aspect, the present invention relates to a method for insect pest insect using an insecticidal composition comprising the extract of Dicapetalum moralesii Prance or a fraction thereof as an active ingredient. In a specific embodiment, the insecticidal method of the present invention includes application of the insecticide of the present invention in consideration of various factors such as formulation of insecticides, kinds of insect pests to be insecticides, objects to be insecticides, places and methods, Treating the pesticide to an insect pest or habitat in an effective amount.

The type of insect pests to be insect pests is not limited, and may include agricultural pests, hygiene pests and forest pests. In addition, the insecticidal object includes both adults and larvae. The hygiene insect is preferably a mosquito, a wheel, a fly, and a mite, and in particular, a mosquito is preferable. More preferably, the mosquito is selected from the group consisting of Culex pipiens pallens, Culex tritaeniorhynchus , Culex vishnui , Culex gelidus , One or more mosquitoes selected from the group consisting of Culex fuscocephala , Aedes aegypti , Aedes togo , and Anopheles stephensi .

The method of use is preferably a spraying treatment, a water surface treatment, a matt treatment or the like, and the applied dose may be used in a capacity to which a conventional insecticide is applied. In addition, when the insecticidal composition of the present invention is sprayed in a place naturally inhabited by a mosquito larva, a mosquito larva can be killed due to respiratory insufficiency by forming a coating on a contact poison or water surface.

The insecticidal composition of the present invention contains Dicapetalum moralesii Prance extract as an active ingredient and is environmentally friendly, harmless to human body and can solve various problems caused by abuse of chemical insecticide. The insecticidal composition of the present invention can be effectively used for controlling pests resistant to chemical insecticides and for controlling agricultural pests, hygiene pests, forest pests, and plant pathogens.

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

Example 1: Insect insects

Experiments were carried out in 2005 on the third larva of Aedes aegypti and Culex pipiens pallens, which had been sold from the National Institutes of Health in 2005. The mosquito adults were fed 5% sugar water at a temperature of 28 ± 2 ° C and a relative humidity of 80 ± 5% in a mosquito net (50 cm × 50 cm) at 16: 8 (L: D) The blood of the rats was vigorously bred. For the larvae hatching from laying eggs, chickens were fed with a mixture of artificial diets (1: 1 dry weight) mixed with powdered liver and yeast powder.

Example 2: Dicapetalum Moraleshifran ( Dichapetalum moralesii Prance) extracts and fractions

Plants were provided by Instituto Nacional de Biodiversida in Costa Rica and samples from the Biotech Research Institute Overseas Plant Extract Bank (control number PB008002) were used. The extracts were prepared by adding 200 ml of methanol to 20 g of the ground sample and ultrasonically extracting the extract with an ultrasonic extractor for 4 hours. This was repeated three times, and the resulting extract was filtered and concentrated under reduced pressure using a vacuum concentrator to obtain 2.0 g of a methanol extract. To obtain the fraction of the extract, 1.78 g of the methanol extract of the above plant was suspended in 75 ml of distilled water, and the fraction was subjected to solvent fractionation with 75 ml of hexane to obtain a hexane fraction. This was repeated twice to obtain a hexane fraction (573.8 mg). Then, chloroform, ethyl acetate and n-butanol were added to the aqueous layer which was successively left in the same manner to obtain a chloroform fraction (386.2 mg), ethyl acetate fraction (63 mg), butanol fraction (171 mg) and a water fraction ).

Example 3: Dicapetalum Moraleshifran ( Dichapetalum moralesii Prance) Investigation of insecticidal activity of methanol extract

The experiment was conducted by modifying the mosquito larvae test method (WHO, 1970), which is the standard method of the World Health Organization. Each methanol extract of Dicapetalum moralesii Prance was diluted to 20,000 ppm with dimethyl sulfoxide (DMSO) to prepare a stock solution. In each cell of a 6-well cell culture plate, 10 mosquito larvae of the third or fourth instar group were placed. The stock solution was then treated at different concentrations and exposed for 24 hours before the mortality rate was recorded. The concentrations were tested at 100 ppm, 50 ppm, 25 ppm, 12.5 ppm, 6.25 ppm, 3.12 ppm, 1.56 ppm, 0.78 ppm. A control group was used for each experiment. When the control group had a mortality rate of 20% or more, the control group was nullified and retested. The results are shown in Table 1 below.

Methanol extracts of Dicapetalum moralesii Prance showed excellent insecticidal activity in both Aedes aegypti and Culex pipiens pallens in a concentration dependent manner.

Experimental results of methanol extract MeOH density
(ppm) Insecticide rate (mean ± standard deviation,%) Egyptian forest mosquito
( Aedes aegypty ) Red house mosquito
( Culex pipiens pallens) 100 100 100 50 70 ± 1.5 100 25 40 100 12.5 20 80 ± 0.6 6.25 10 50 ± 0.6 3.12 0 20 ± 1.0 1.56 0 10 0.78 0 0

Example 4: Dicapetalum Moralescifer ( Dichapetalum moralesii Prance) Investigation of insecticidal activity of fractions

The experiment was conducted by modifying the mosquito larvae test method (WHO, 1970), which is the standard method of the World Health Organization. First, the Dichapetalum moralesii Prance extract was fractionated using ethyl acetate (EtOAc), hexane (Hexane), chloroform (CHCl ₃ ), butanol (BuOH) and water. Each fraction was diluted to 20,000 ppm with dimethyl sulfoxide (DMSO) to prepare a stock solution. Then, 10 mosquito larvae of 3 ~ 4th instar were placed in each cell of a 6-well cell culture plate, and the stock solution was treated at different concentrations for 24 hours, and the mortality rate was recorded. The concentrations were tested at 50 ppm, 25 ppm, 12.5 ppm, 6.25 ppm, 3.12 ppm and 1.56 ppm.

A control group was used for each experiment. When the control group had a mortality rate of 20% or more, the control group was nullified and retested. The results are shown in Tables 2 to 6 below. Methanol extracts of butanol, ethyl acetate and hexane fractions showed only insecticidal effects against Aedes aegypti . In the case of chloroform fractions, both Aedes aegypti and Culex pipiens pallens And showed insecticidal activity. On the other hand, the water fraction of Dichapetalum moralesii Prance did not show the insecticidal effect in the Aedes aegypti and Culex pipiens pallens.

Experimental results of ethyl acetate (EtOAc) fraction of methanol extract EtOAc density
(ppm) Insecticide rate (mean ± standard deviation,%) Egyptian forest mosquito
( Aedes aegypty ) Red house mosquito
( Culex pipiens pallens) 50 100 0 25 93 ± 0.6 0 12.5 83 ± 0.6 0 6.25 77 ± 0.6 0 3.12 37 ± 2.9 0 1.56 0 0

Experimental results of hexane fraction of methanol extract Hexane density
(ppm) Insecticide rate (mean ± standard deviation,%) Egyptian forest mosquito
( Aedes aegypty ) Red house mosquito
( Culex pipiens pallens) 50 87 ± 0.6 0 25 83 ± 0.6 0 12.5 83 ± 0.6 0 6.25 70 ± 1.7 0 3.12 53 ± 0.6 0 1.56 43 ± 1.2 0

Chloroform (CHCl ₃ ) fractions of methanol extracts CHCl ₃ density
(ppm) Insecticide rate (mean ± standard deviation,%) Egyptian forest mosquito
( Aedes aegypty ) Red house mosquito
( Culex pipiens pallens) 50 100 100 25 100 100 12.5 97 ± 0.6 80 ± 1.0 6.25 77 ± 0.6 53 ± 0.6 3.12 37 ± 1.2 23 ± 1.2 1.56 17 ± 0.6 0

Experimental results of butanol (BuOH) fraction of methanol extract BuOH density
(ppm) Insecticide rate (mean ± standard deviation,%) Egyptian forest mosquito
( Aedes aegypty ) Red house mosquito
( Culex pipiens pallens) 50 17 ± 0.6 0 25 3 ± 0.6 0 12.5 0 0 6.25 0 0 3.12 0 0 1.56 0 0

Experimental results of water fraction of methanol extract Water density
(ppm) Insecticide rate (mean ± standard deviation,%) Egyptian forest mosquito
( Aedes aegypty ) Red house mosquito
( Culex pipiens pallens) 50 0 0 25 0 0 12.5 0 0 6.25 0 0 3.12 0 0 1.56 0 0

Claims (10)

An insecticidal composition comprising an over-the-ground extract of Dicapetalum moralesii Prance or a fraction thereof as an active ingredient. delete The composition according to claim 1, wherein the extract is an extract extracted with a solvent selected from the group consisting of lower alcohols having 1 to 4 carbon atoms and a mixed solvent thereof. 4. The composition of claim 3, wherein the lower alcohol solvent is methanol. The composition of claim 1 wherein said fraction is an ethyl acetate fraction, a hexane fraction, a chloroform fraction or a butanol fraction. The composition of claim 1, wherein the composition has an insecticidal effect on mosquito or mosquito larvae. 7. The method of claim 6, wherein the mosquito is selected from the group consisting of Culex pipiens pallens, Culex tritaeniorhynchus , Culex vishnui , Culex gelidus , Wherein the composition is at least one mosquito selected from the group consisting of Culex fuscocephala , Aedes aegypti , Aedes togo and Anopheles stephensi . The composition of claim 1, wherein said extract is comprised at a concentration of 1 to 100 ppm. 9. A method for insect pest control using the composition of any one of claims 1 to 8. 10. The method of claim 9, wherein the pest is a mosquito.