KR20190042835A - Agent for controlling Varroa destructor having Quercetin - Google Patents

Abstract

The present invention relates to a formulation for controlling Varroa jacobsoni comprising quercetin as an active ingredient, a production method thereof, and a control method of Varroa jacobsoni. The formulation of the present invention has excellent water solubility and stability by improving the poor solubility of quercetin and self-emulsifying when mixed with water to form nano-emulsion, can be mass-produced relatively economically, is safe due to less toxic to bees, and has an excellent control effect because Varroa jacobsoni is not tolerant, thereby being able to be useful for controlling the Varroa jacobsoni.

Description

{Agent for controlling Varroa destructor having quercetin}

The present invention relates to a preparation for bee venomous marsal remedy containing quercetin as an active ingredient, a method for producing the same, and a method for relieving honeybee mites.

In general, the beekeeping industry utilizes the ecology and habits of bees to develop high-nutrient, high-quality beef by managing beekeeping such as specification management, breeding and production, distribution of products, remediation of diseases and pests, improvement of beekeeping apparatus, It is a business field that earns income by producing health food.

A typical health food obtained by beekeeping is honey. Honey is a monosaccharide composed mainly of glucose and fructose. It is absorbed directly into the barrier of the body and burned in muscle cells of each part of the body, State. It is a natural comprehensive food containing the necessary minerals in the body, which facilitates physiological action in the body and completes the development of bone tissue.

The beekeeping industry mentioned above is not only able to increase the income by a wide variety of products and rapid increase in demand, but also improves the yield by means of pollen of various crops, and is treated as a major agricultural policy in the world.

In order to continuously develop the beekeeping industry as described above, it is important to protect bees from bacteria, viruses, fungi, parasites and the like, which are diseases of bees, and it is very important to protect bees from bees such as bee mites (ticks).

Bee japonica was originally an external parasite of oriental bees in the Southeast Asia region, but was transferred to western bees and spread to other parts of the world due to the movement of the bee by human activity. It has been empirically proven that it is almost impossible to completely eliminate it if honey bee mite invades an area and settles in years. Therefore, it is necessary to understand the infection route and life history of bee mite and to control its density to minimize the economic loss.

Females of bee mite are parasitized on larvae, pupae and adult bees of bees and inhale lymphatic blood to cause so called tick disease. The weight of the infected bee adult is reduced by 7.1 ~ 30.4% compared with normal, and in the case of severe parasitism, it may become degenerative or developmental arrest. Even in the case of bee pupa, 10% of weight loss is observed when the parasitism (or infection) of bee mite is severe. In severe cases, crippling or pupae of the legs may die (De Jong et al., 1982; Schneider et al., 1987). The bigger damage is due to the weakening of the feud, resulting in a decrease of 30 ~ 46%. Beekeeping farms suffered from ticks average an average of 20,000 places a year, and the estimated amount of damage is increasing year by year, and is estimated to be over 60 billion won last year (as of 2013). When about 3% of bee larvae are infected with ticks, the use of anti-proliferation drugs can minimize the damage.

In Korea, there is a nationwide prevalence that is causing great damage. As a result, more than half of the farmers reported that the direct damage caused by the bee mite is over 30%. In the meantime, in Korea, the use of organic synthetic insecticides has been used more than necessary to control the growth of bees and honey bee mites that cause great damage to bee farmers. Therefore, domestic honey bee mites have a strong tolerance to existing medicines Visible problems are also occurring.

At present, the damage caused by the mite occurs continuously irrespective of the region, and the effective control effect is not obtained, so it is urgently required to develop and introduce a new drug which is resistant to the disease.

KR 10-1384515 B (2014.04.11.) KR 10-0340016 B (2002.06.10.)

The first object to be solved by the present invention is to provide a preparation for bee venomous marsal remedy having no resistance and excellent control effect and a method for producing the same.

A second problem to be solved by the present invention is to provide a method for relieving honeybee mite using the above preparation.

In order to achieve the above object, the present invention provides a pharmaceutical composition comprising quercetin as an active ingredient, at least one solubilizer selected from the group consisting of diethylene glycol monoethyl ether, and polyethylene glycol or a derivative thereof; And a polyoxyethylene surfactant, wherein the emulsion is self-emulsified when mixed with water to form an emulsion.

According to one embodiment of the present invention, the quercetin may be contained in an amount of 0.5 to 3% by weight based on the total weight of the preparation.

According to one embodiment of the present invention, the solubilizer may be contained in an amount of 10 to 30 parts by weight based on 1 part by weight of the quercetin.

According to one embodiment of the present invention, the surfactant may be included in an amount of 20 to 40 parts by weight based on 1 part by weight of the quercetin.

According to one embodiment of the present invention, the surfactant is selected from the group consisting of polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), poly Polyoxyethylene sorbitan monopalmitate (tween 40), and polyoxyethylene sorbitan monolaurate (tween 20). The polyoxyethylene sorbitan monolaurate (tween 20) may be selected from the group consisting of polyoxyethylene sorbitan monopalmitate (tween 40) and polyoxyethylene sorbitan monolaurate (tween 20).

According to one embodiment of the present invention, the formulation comprises 0.5-3% by weight of quercetin, 10-30 parts by weight of the quercetin relative to 1 part by weight of diethylene glycol monoethyl ether and 20-40 parts by weight of polyoxyethylene sorbitan monol (Polyoxyethylene sorbitan monooleate (Tween 80)).

According to one embodiment of the present invention, when the preparation is diluted with water, the average particle size may be 50 nm or less.

(1) mixing quercetin with one or more solubilizers selected from the group consisting of diethylene glycol monoethyl ether, and polyethylene glycol or a derivative thereof; And (2) mixing the mixture of step (1) with polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate Mixing at least one surfactant selected from the group consisting of polyoxyethylene sorbitan monopalmitate (tween 40) and polyoxyethylene sorbitan monolaurate (tween 20) To provide a method for producing a preparation for bee mite relief.

In addition, the present invention relates to a method for producing a bee mite relieving agent, which comprises mixing a formulation for bee mating remover with 10 to 50 times by weight water to form a nano emulsion; And spraying the nanoemulsion onto a bee infected with bee mite. The present invention also provides a method of relieving bee mite.

The preparation of the present invention improves the poor solubility of quercetin and self-emulsifies upon mixing with water to form a nano emulsion, thereby being excellent in water solubility and stability, being economically mass-producible, less toxic to bees, Bee mites are not tolerant and have excellent control effect, so they can be useful for relieving bee mites.

Fig. 1 is a photograph showing the appearance of bee mites that are parasitized with adult bees.
Fig. 2 is a photograph showing the appearance of bee mites parasitized on the larvae of bees.
FIG. 3 is a graph showing the size distribution of nanoparticles formed after mixing a preparation for bee mite remover with water according to an embodiment of the present invention.
FIG. 4 is a photograph showing a state in which the preparation for bee venomous remedy is mixed with water according to an embodiment of the present invention.

Hereinafter, the present invention will be described in detail.

According to one aspect of the present invention, there is provided a preparation for bee venomous moth salvage comprising quercetin as an active ingredient. The present inventors have found that quercetin, which is mainly used as a food additive, has a strong insecticidal effect against bee mite, and devised the present invention. However, quercetin has a problem in that it is formulated into liquid form for use as a spraying method due to the property of being insoluble in water. In order to overcome this problem, the present invention has been completed by applying a self-microemulsifying drug delivery system (SMEDDS).

The basic principle of the " self-emulsifying drug delivery system " in the present invention is that when the system is in contact with water, it automatically forms an oil-in-water microemulsion under mild mechanical agitation.

The present inventors have sought to develop a self emulsifying drug delivery system to improve the poor availability of quercetin and to increase penetration into bee mite. As a result, when a preparation is prepared by mixing quercetin with a solubilizer and a surfactant, quercetin is present in a nano-emulsion state in an aqueous solution due to self-emulsification upon mixing with water and dilution, and solubilization is greatly improved since the drug particles are minimized. It is confirmed that the penetration rate and insecticidal power against mite greatly increase and the present invention has been completed.

The quercetin may be contained in an amount of 0.5 to 3% by weight, preferably 1 to 2.5% by weight, more preferably 2% by weight based on the total weight of the preparation.

When the content of quercetin is less than the lower limit, there is a possibility that the effect of remedying honey bees mite may be insignificant. When the content of quercetin exceeds the upper limit, there may be quercetin suspended or precipitated due to not being enclosed in the nanoparticle formulation, It becomes impossible to spray.

The preparation for bee mite relief of the present invention comprises diethyleneglycol monoethylether as a solubilizer of quercetin. The diethylene glycol monoethyl ether may be contained in an amount of 10 to 30 parts by weight, preferably 15 to 25 parts by weight, relative to 1 part by weight of the quercetin.

When the content of the diethylene glycol monoethyl ether is less than the lower limit, there is a fear that the formulation stability is lowered due to the undissolved quercetin. When the upper limit is exceeded, there is no problem in dissolving quercetin as the main ingredient. However, , The production cost is increased and the concentration of the main component is lowered, resulting in an increase in logistics cost and inconvenience of storage due to the enlargement of the packaging container.

The preparation may further comprise 20 to 40 parts by weight, preferably 25 to 35 parts by weight, of a surfactant containing a polyoxyethylene compound, based on 1 part by weight of the quercetin. When the content of the surfactant is less than the lower limit, nanoparticles can not be formed and the product becomes an emulsified product. If it exceeds the above upper limit value, unnecessary use of the excipient excessively lowers the concentration of the main component, which is not preferable because the volume of the product becomes large, which is unsuitable for storage and transportation.

The surfactant may be selected from the group consisting of polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate sorbitan monopalmitate (tween 40), and polyoxyethylene sorbitan monolaurate (tween 20).

The bee mite remedy of the present invention is characterized in that it contains 0.5 to 3 wt% of quercetin, 10 to 30 parts of diethylene glycol monoethyl ether per 1 part of quercetin and 20 to 40 parts of polyoxyethylene sorbitan monooleate sorbitan monooleate (Tween 80)).

When the preparation is diluted by mixing with water, a nano emulsion having an average particle size of 50 nm or less can be formed. Thus, when quercetin, an active ingredient of the present invention, is present in a nanoemulsion state in an aqueous solution, solubilization is greatly improved, and penetration rate and insecticidal power against bee sprouting can be greatly increased.

In addition, the bee mite remedy formulations containing quercetin as an active ingredient and using the self-emulsifying drug delivery system of the present invention are easy to mix with water, have excellent adhesion and adhesion, and can be sprayed or sprayed on bees infected with bee mite It is not easy to volatilize and has excellent efficacy.

(1) mixing at least one solubilizing agent selected from the group consisting of a main component quercetin, diethylene glycol monoethyl ether, and polyethylene glycol or a derivative thereof at a temperature of 40 to 70 ° C; And (2) mixing the mixture of step (1) with polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate Mixing at least one surfactant selected from the group consisting of polyoxyethylene sorbitan monopalmitate (tween 40) and polyoxyethylene sorbitan monolaurate (tween 20) To provide a method for producing a preparation for bee mite relief.

The present invention also relates to a process for preparing a pharmaceutical composition, comprising mixing said formulation with 10 to 500 times by weight water to form a nanoemulsion; And spraying the nanoemulsion onto a bee infected with bee mite. The present invention also provides a method of relieving bee mite. As described above, when the preparation for bee mite remedy according to the present invention is diluted by mixing with water, the insecticidal effect against honeybee mite is effectively exhibited even in a dilute solution having a concentration of quercetin of 0.01 to 1% by weight.

The bee mite relieving method of the present invention is excellent in the relieving effect on bee mite even when spraying the preparation for bee mite relief only once.

It is preferable that the spraying is performed once a day, and it is more preferable to spray continuously for three days.

Hereinafter, the present invention will be described in more detail with reference to preferred embodiments. It will be apparent, however, to those skilled in the art that these embodiments are for further explanation of the present invention and that the scope of the present invention is not limited thereby.

Example

Examples and Comparative Examples 1 to 5

Among the excipients which are harmless to the human body, quercetin is used as a solubilizing agent such as diethyleneglycolmonoethylether (Carbitol), polyethylene glycol (PEG200) or propylene glycol (PPG) To prepare a preparation for bee mite relief as shown in Table 1 below.

Polyoxyethylene sorbitan monooleate (Tween 80) and Cremophor EL were used as the surfactant of the preparation for the bee mite remedy.

As a concrete manufacturing method, quercetin powder is put into a container in a solvent according to the composition shown in the following Table 1, a solubilizing agent is added, and quercetin is completely dissolved while heating to 60 캜. After the solution was cooled to room temperature, a surfactant was added and mixed thoroughly.

Category (g) Example Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Comparative Example 5 Active ingredient Quercetin 0.4 0.4 0.4 0.4 0.4 0.4 Solvent Carbitol 7.8 7.8 - - - - PEG200 - - 7.8 7.8 - - PPG - - - - 7.8 7.8 Interface
Activator Tween80 11.8 - 11.8 - 11.8 - Cremophor EL - 11.8 - 11.8 - 11.8 Gross weight 20 20 20 20 20 20

Comparative Example 6

It is a product launched in the existing market, and citric acid is the active ingredient.

Experimental Example

Experimental Example 1. Stability test of formulation

Examples and comparative examples were diluted 100 times (v / v) in water and mixed by intermittent shaking. After mixing for 6 hours or longer, the particle size was measured using a particle analyzer. The 'stability' was evaluated as '○' when the shape and particle size were compared immediately after dilution with water and after 6 hours or more, and the shape and particle size did not change over time. The measurement and evaluation results are shown in Table 2 below.

division Example Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Comparative Example 5 shape
(Appearance) 나노 입자 emulsion 나노 입자 emulsion 나노 입자 emulsion Average particle size
(nm) 46 480 80 55 320 2100 stability
(Stability) ○ ○ × × × ×

As shown in Table 2, it was confirmed that the nano-particle formulations were produced only in the case of Examples, Comparative Examples 2 and 4. The nano-particle preparation is advantageous in that it can be uniformly sprayed because it does not precipitate when diluted with water. On the other hand, in the case of the emulsion preparation, uniformity is lowered in the liquid state over time, and uniform spraying becomes impossible, which is not preferable.

In addition, it was confirmed that 'nano-particle preparation' was evaluated as 'stability' only in the case of 'Example' because the shape and particle size did not change over time.

Experimental Example 2. Safety Test

A safety test is a test that confirms and verifies the risk of bee injury caused by drugs or excipients before confirming the effectiveness of honey bee mite relief.

An Italian species ( Apis melliffera ) bee, which is commonly cultivated in domestic beekeeping farms, was used for the experiment. In order to stabilize the bees to be injected into the experiment, the placement of the beads was completed 7 days before the experiment. The number of experimental beads was 10, and the beehive number was assigned to each beehive, and the boundary between beehives and beehives was clearly distinguished. In addition, the floor was laid with fibers so that the number of bows could be grasped.

Experimental Example 2-1. Safety evaluation according to single administration

First, ten bees from three prepared beads were captured, put into three bees collecting tanks for experiments, and diluted with 10 volumes of tap water, respectively. In the samples of the examples, excipients except for quercetin were used The excipient sample, the sample of Comparative Example 1, and the control sample consisting only of tap water were each transferred to an atomizer, and then sprayed into each of the above-described bee trapping baskets. An atomizer with a spray amount of about 0.8 mL was used, and the spray was set to a set of 5 times. Five sets were repeatedly sprayed to wet the bees completely. All of the bees wet on the specimen could not fly and were in a state of crawling, but after the water had evaporated, they could fly and returned to their beehives. As a result, the safety of the sample of the example can be confirmed.

Experimental Example 2-2. Safety evaluation according to repeated administration

Each of the prepared samples was repeatedly sprayed at intervals of 4 days in a manner of spraying four times (twice before and after the consumption) per consumption of each hive (巢 spleen). The results are shown in Table 3 below. The number of beads shown in Table 3 is the cumulative value of dead bees around the beehive after 48 hours from spraying each sample.

division Number of beads (cumulative value) Number of bees remaining in beehive One-time spray Two-time spray 3 times spray Control group (Tap water) 35 55 80 8,241 Excipient experimental group 43 62 91 10,052 Example 34 60 88 9.018 Comparative Example 1 39 63 90 9,517

As can be seen from Table 3, the number of dead bees according to the formulation of the Example was not much different from that of the control water spraying experiment, and it was confirmed that the bees were safe.

Experimental Example 3: Efficacy test

Each sample diluted to 50 times by weight with tap water was immersed in an atomizer, and then sprayed onto the corresponding rod. At this time, each individual was sprayed at a distance of about 30 cm, twice on the front and back sides respectively, and sprayed on all the consumptions in the bag. The number of consumption in each group was the same, and the cluster size of the bee was about 10,000. The bees belonging to each group have no other diseases, and the groups with similar infectivity of bee mite were prepared and used in the experiment.

In order to count the bees and bee mite that fall on the bottom of the beehive, the commercially available sheet is cut to the size corresponding to the area of the beehive and the sticky part with the adhesive of the sheet is placed on the bottom of the beehive so that the bee mite falls . At the time when 24 hours had elapsed from the introduction of the sheet, the sheet was collected and the number of bee mites attached on the sheet was counted using a microscope. Further, after a new sheet was placed on the bottom of each beehive, the sheet was collected after 48 hours from that time, and the number of honeybee mites attached on the sheet was counted to confirm whether the efficacy was sustainable.

Table 4 shows the results of counting the numbers of bee flies attached on the sheet. The results shown in Table 4 are the average values obtained by repeating trials for each sample.

division Bee mite death number Remaining bee caterpillars Insecticidal rate (%) After 1 day 3 days later system Control group (Tap water) 24 35 59 1,197 4.7 Excipient experimental group 83 47 130 1,086 10.7 Example 995 212 1,207 54 95.7 Comparative Example 1 347 96 443 657 40.3 Comparative Example 6 530 85 615 655 48.4

As shown in Table 4, the bee mite remedy according to the present invention showed a mite mite mortality of 95% or more within three days even by one spraying, and thus it was confirmed that there is a very excellent remedy effect.

Although the present invention has been described in terms of the preferred embodiments mentioned above, it is possible to make various modifications and variations without departing from the spirit and scope of the invention. It is also to be understood that the appended claims are intended to cover such modifications and changes as fall within the true spirit of the invention.

Claims (9)

It contains quercetin as an active ingredient,
Diethylene glycol monoethyl ether, and polyethylene glycol or a derivative thereof; And
A surfactant for polyoxyethylene (polyoxyethylene), which is self-emulsified when mixed with water to form a nano emulsion. The method according to claim 1,
Wherein the quercetin is contained in an amount of 0.5 to 3% by weight based on the total weight of the preparation. The method according to claim 1,
Wherein the solubilizer is contained in an amount of 5 to 30 parts by weight based on 1 part by weight of the quercetin. The method according to claim 1,
Wherein the surfactant is present in an amount of 10 to 40 parts by weight based on 1 part by weight of the quercetin. The method according to claim 1,
The surfactant may be selected from the group consisting of polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate sorbitan monopalmitate (tween 40), and polyoxyethylene sorbitan monolaurate (tween 20). The present invention relates to a preparation for bee venomous mildew. The method according to claim 1,
0.5 to 3% by weight of quercetin,
5 to 30 parts by weight of diethylene glycol monoethyl ether and 10 to 40 parts by weight of polyoxyethylene sorbitan monooleate (Tween 80) relative to 1 part by weight of quercetin. Remedies. The preparation according to claim 1, wherein the formulation has an average particle size of 50 nm or less when diluted in water. (1) mixing at least one solubilizer selected from the group consisting of quercetin, diethylene glycol monoethyl ether, and polyethylene glycol or a derivative thereof at a temperature of 40 to 70 ° C; And
(2) To the mixture of step (1), polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate Mixing at least one surfactant selected from the group consisting of polyoxyethylene sorbitan monopalmitate (tween 40) and polyoxyethylene sorbitan monolaurate (tween 20) (JP) METHOD FOR MANUFACTURING A MEDICAMENT FOR MORTAR AID 10. A method for producing a honey bee deodorant according to any one of claims 1 to 7, which comprises mixing 10 to 50 times by weight of water to form a nano emulsion; And
Spraying the nanoemulsion onto a bee infected with bee mite. ≪ RTI ID = 0.0 > 21. < / RTI >

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