METHOD FOR IMPROVING A DRUG OR FEED INTAKE RATE OF SHRIMP

[TECHNICAL FIELD]

The present invention relates to a method for improving drug or feed intake of shrimp. The present invention also relates to drug or feed of which shrimp intake rate is improved.

[BACKGROUND ART]

World shrimp industry has grown until the early 1990' s without any threatening of severe diseases. Vibriosis was a major disease threatening the farming yield of shrimp in the late 1980s and the early 1990s. However, new deadly viruses like yellow head virus, white spot syndrome virus, Taura syndrome virus, etc. were introduced in the 1990s, which exerted a severe bad effect on shrimp farming yield. Particularly, white spot syndrome virus of shrimp is broadly happening from Asia, including Korea and Japan, to Central and South America, and then its economic damage is very severe.

Many studies have being progressed to decrease damage of several shrimp diseases like white spot syndrome virus. Especially, plant extracts which are thought to be relatively safe have been handed down as folk remedy from a long time ago, and recently their effects are scientifically examined. However, conventional plant extracts have unique smell and taste, which

reduces an intake rate of shrimp.

[DISCLOSURE] [TECHNICAL PROBLEM] Accordingly, the object of the present invention is to provide a method for improving a drug or feed intake rate of shrimp, and drug or feed of which shrimp intake is improved.

[TECHNICAL SOLUTION] To achieve the object, the present invention provides a method for improving a drug or feed intake rate of shrimp, wherein the drug or feed is coated with a polymer.

More preferably, the present invention provides the method for improving a drug or feed intake rate of shrimp, wherein the drug or feed is firstly coated with a hydrophilic polymer and secondly coated with a non-hydrophilic polymer.

More preferably, the present invention provides the method for improving a drug or feed intake rate of shrimp, wherein the coating amount of the hydrophilic polymer is 1-30 wt% based on the total weight of drug or feed and the coating amount of the non-hydrophilic polymer is 1-10 wt% based on the total weight of the firstly coated drug or feed.

More preferably, the present invention provides the method for improving a drug or feed intake rate of shrimp, wherein the

drug or feed is made into granules before being coated.

More preferably, the present invention provides the method

for improving a drug or feed intake rate of shrimp, wherein the drug is plant extract. The present invention also provides a drug or feed of which shrimp intake rate is improved, wherein the drug or feed is coated with polymer. More preferably, the present invention provides the drug or feed of which shrimp intake rate is improved, wherein the drug or feed is firstly coated with hydrophilic polymer and then secondly coated with non- hydrophilic polymer.

Hereinafter, the method for improving a drug or feed intake rate of shrimp and the drug or feed having improved intake rate of shrimp will be described in more detail. The present invention is based on the surprising fact that a drug or feed having bad intake rate of shrimp because of its smell, taste, etc. can be efficiently ingested by shrimp if the drug or feed is coated with polymer. This improvement is thought to be achieved by blocking smell, taste, etc. with a coating polymer, but the scope of the present invention is not limited to this presumption.

When considering several aspects, it is preferable to firstly coat the drug or feed with hydrophilic polymer and then secondly coat the firstly coated drug or feed with non-

hydrophilic polymer. In case that the hydrophilic polymer only is used for coating, it is difficult to block smell of the plant extract because the hydrophilic polymer is easily dissolved in water. In case that the non-hydrophilic polymer only is used, a lot of non-hydrophilic polymer must be used to block the smell or taste of drug or feed, and this lots of amount is difficult to be dissolved in water, which may cause the reduction of drug efficacy. In addition, the first coating itself is expected to show the effect blocking bad smell, but the second coating using the non-hydrophilic polymer is expected to improve the stability of the extract as well as the blocking effect of bad smell.

The hydrophilic polymer of the present invention means a polymer that can be easily dissolved in water even if lots of polymer is used for coating. This hydrophilic polymer includes,

but is not limited to, hydroxymethylcellulose,

hydroxyethylcellulose, hydroxyethylmethylcellulose, hydroxypropylmethylcellulose, hydroxypropylethylcellulose, carboxymethylcellulose, hydroxypropylcellulose, cellulose derivatives, sodium alginate, alginic acid, soybean protein, wheat protein, carrageenan, tragacanth gum, agar, arabia gum, guar gum, xanthan gum, gums, pectin, polyvinylalcohol, polymethylmethacrylate derivatives, polyvinylpyrrolidone, polyalkeneoxide, polyalkeneoxide derivatives, polyalkeneglycol, polyethylene-polypropylene polymer, polyoxyethylene-

polyoxypropylene polymer, zein, diethylaminoacetate, aminoalkylmethacrylate copolymer, cyclodextrin, chitin, chitosan and gelatin. Hydroxymethylcellulose, hydroxyethylcellulose, hydroxyethylmethylcellulose, hydroxypropylmethylcellulose, hydroxypropylethylcellulose, carboxymethylcellulose, hydroxypropylcellulose, cellulose derivatives, sodium alginate, alginic acid, polyvinylalcohol and polyvinylpyrrolidone are more preferable .

The non-hydrophilic polymer of the present invention means a polymer that is difficult to be dissolved or is very slowly dissolved in water so that the polymer can prevent the drug or feed from being dissolved into water in a short time. This non- hydrophilic polymer includes, but is not limited to, methylcellulose, ethylcellulose, shellac, polyvinylacetate, poly-L-lysine, hydroxypropylmethylcellulose phthalate, high- viscous cellulose derivatives, hydroxypropylmethylcellulose acetate succinate, cellulose acetate phthalate, corn protein extract, corn protein extract derivatives, soybean protein, wheat protein, chitin, chitic acid, agar, pectin, gums and polymethylmethacrylate derivatives. Ethylcellulose,

methylcellulose and shellac are more preferable.

Preferably, the amount of the first coating using the hydrophilic polymer is 1-30 wt% based on the total weight of drug or feed, and more preferably, 4-15 wt%. Preferably, the

amount of the second coating using the non-hydrophilic polymer is 1-10 wt% based on the total weight of the first coated granules, and more preferably 2-8 wt%.

In case of the first coating, water can be used for making a coating solution. In case of the second coating, ethanol, methanol, isopropanol, acetonitrile, acetone, ether, hexane, chloroform, 1,4-dioxane, dimethylsulfoxide, tetrahydrofuran, ethylacetate, methylacetate or their mixture can be used for making a coating solution.

The coating can be performed with conventional machines well known in the art, like a fluidized bed granulator, a spray drier, C/F granulator, etc. The scope of the present invention is not limited to these machines.

Preferably, the drug or feed according to the present invention is prepared into granules for easy coating process before polymer coating. Methods for preparing granules are well known in the art to which the present invention pertains, and

for example, dry granulation method, wet granulation method, etc. can be used for the present invention. If the shrimp intake rate of a drug or feed is bad because of its smell, taste, etc., the method for improving shrimp intake rate according to the present invention can be applied to the drug or feed. For example, Artemisia japonica extract, Artemisia japonica var. hallaisanensis extract, Artemisia

gmelini extract, Artemisia capillaris extract, Artemisia annua extract, Saururus chinensis extract, Clinacanthus nutans extract, Zanthoxylum piperitum extract, Houttuynia cordata extract,

Glycyrrhiza uralensis extract, Cnidium officinale extract, peanut extract, Fagopyrum esculentum extract, Codium fragile

extract, Porphyra tenera extract, Dictammus dasycarpus extract, glucan, lipopolysaccharide, fucoidan, saponin, lactoferrin, inulin, dextran, egg yolk antibody, antibody protein, 5-amino levulinic acid, etc. can be used in the present invention. Particularly, there are many studies using plant extracts, being expected to be safe, to treat shrimp's diseases. However, most plant extracts have unique smells, and therefore the present invention can be usefully used in the treatment for shrimp. Plant extracts can be made according to extracting methods well known in the art. For example, plant extracts are prepared as follows: about 1-15 times volume of solvent such as water, ethanol, methanol, propanol, butanol, acetone, ethylacetate or their mixture are added to a plant, and then plant extracts are prepared by cold water immersion method, heat extraction method, reflux-cooling extraction method, etc. Then, the plant extracts are filtered by filter paper having an appropriate size (for example, filter paper having 10 um of pore size) , and the filtrate are concentrated in vacuo, distillation-concentrated

under reduced pressure or concentrated by spray-drying at a

proper temperature (for example, about 30-70 ^° C ) to get a final plant extract.

As said above, it is preferable to make the plant extract into granules for easiness of coating process before coating.

The extract is mixed with conventionally used diluents in the

art (for example, maltodextrin, lactose, microcrystalline cellulose, sucrose, mannitol and starch) or general shrimp feed, and then prepared into granules according to well-known methods in the art like dry-granulation method, wet-granulation method, simple mixing method, etc. This method is preferable in aspects of mixing efficiency with general feed, easiness in handling and the following coating process. Furthermore, it is preferable for the following processes to add about 0.1-5% (based on the total weight of granules) of lubricant (for example, silicone dioxide and magnesium stearate) into prepared granules.

[BRIEF DESCRIPTION OF THE DRAWINGS]

Figure 1 shows efficacy evaluation results of Artemisia japonica extract and Saururus chinensis extract on white spot syndrome virus of shrimp.

Figures 2 and 3 show efficacy evaluation results of the coated Artemisia japonica extract and the coated Saururus chinensis extract on white spot syndrome virus of shrimp.

[MODES FOR CARRYING OUT THE INVENTION]

Hereinafter, the present invention is described in considerable detail to help those skilled in the art understand the present invention. However, the following examples are

offered by way of illustration and are not intended to limit the

scope of the invention.

<Example 1> Preparation of extracts 5 kg of dried Artemisia japonica and Saururus chinensis, respectively, were immersion-extracted at room temperature with 50 liter of water, ethanol and their 1:1 mixture. The immersion-

extraction was once performed for 3 days, and then re-performed for 2 days with no delay. Extracts were filtered by 10 um of

filter paper and the filtrate was concentrated in vacuo at 60 ^° C until the weight decreased to 5% of the initial weight. The concentrate was made into granules with about 2.5 times of lactose based of the weight of the concentrate and about 2% of silicone dioxide based on the total weight. When considering the yield of the concentrate, ethanol extraction was most efficient. Thus, extracts made with ethanol were used in the following all examples.

<Example 2> Efficacy evaluation on white spot syndrome virus of shrimp

Efficacy of a mixture of Artemisia japonica extract and

Saururus chinensis extract on white spot syndrome virus was evaluated, and a proper mixing ratio of the mixture was investigated as follows: Shrimp: Palaemon paucidens was used and their average body weight was 0.3g. Shrimps were accustomed to experimental environments for one week before tests. During that

accommodation period, general feed was supplied the shrimps.

Test samples: Artemisia japonica extract and Saururus chinensis extract prepared in the example 1 were used.

Test group: Control group was provided with general feed having no Artemisia japonica extract and Saururus chinensis extract. Five test groups were provided with mixtures of Artemisia japonica extract and Saururus chinensis extract having mixing ratios of 10:0, 6:4, 5:5, 4:6 and 0:10, respectively. 30 shrimps were included in each group.

Feeding of test samples: Test samples were supplied everyday for 7 days before infection of the virus to evaluate preventing and treating effect on white spot syndrome virus. In addition, test samples were supplied everyday for 7 days after

the infection of the virus. Feeding was performed at a.m. 10 o'clock everyday.

Raising conditions of shrimp: Shrimps were raised in 10

liter of water bath. The temperature of the bath was kept at 20+

re .

Infection of white spot syndrome virus: Shrimps were

artificially infected with white spot syndrome virus and died finally. Three died shrimps were finely ground in 10 ml of distilled water. The ground solution was added to the water bath as 3500:1 of ratio to infect shrimps in water bath with white spot syndrome virus. After the infection, survival rates of shrimps were evaluated.

Results were shown in figure 1. As shown in the figure 1,

Artemisia japonica extract, Saururus chinensis extract and their mixture increased the survival rate of shrimps in comparison with control. Mixture of Artemisia japonica extract and Saururus chinensis extract having almost equal mixing ratio, particularly

6:4 weight ratio mixture of Artemisia japonica extract and Saururus chinensis extract, showed superior survival rate than the other test samples.

However, when Artemisia japonica extract and Saururus chinensis extract were administered by simple mixing with general feed, lots of test samples were not eaten by shrimps and remained, which was thought to be caused by the unique smell of the extracts. Because of this problem, difficulty was expected in letting shrimps eat enough extracts and there was a need to improve a drug or feed intake rate of shrimp.

<Example 3> Coating of extract

Extracts were coated with polymer like the below examples to block the unique smell of the extracts, that is, to increase

shrimp's intake rate of Artemisia japonica extract and Saururus

chinensis extract. <Example 3-l>

φ The first coating

100 g of hydroxypropylmethylcellulose (HPMC2910) was dissolved into 1,000 ml of distilled water to make a coating solution. 1,000 g of 6:4 (weight) mixture of Artemisia japonica extract granule and Saururus chinensis extract granule, prepared in the example 1, were fluidized in a fluidized bed granulator (GX40, Freund, Japan) and coated by spraying the coating solution. The preparing conditions of the fluidized bed granulator were as follows:

Inlet air temperature: about 60 ^° C

Flow rate of inlet air: 0.3 1/minute

Flow rate of outlet air: 0.5 1/minute

Rotor rotation number: 220 rpm

Spraying speed: 10 ml/minute

© The second coating

49.5 g of ethylcellulose was dissolved in a mixture of 385 ml of acetone and 110 ml of ethanol to make a coating solution. The above coated granules were fluidized in the fluidized bed granulator and coated by spraying the second coating solution.

The preparing conditions of the fluidized bed granulator were as follows :

Inlet air temperature: about 50 ^° C

Flow rate of inlet air: 0.3 1/minute Flow rate of outlet air: 0.5 1/minute

Rotor rotation number: 230 rpm

Spraying speed: 12 ml/minute

<Example 3-2>

(D The first coating 3 g of sodium alginate was dissolved in 1,000 ml of distilled water to make a coating solution. 1,000 g of 6:4

(weight) mixture of Artemisia japonica extract granule and Saururus chinensis extract granule, prepared in the example 1, and 97 g of lactose were fluidized in the fluidized bed granulator and coated by spraying the coating solution. The preparing conditions of the fluidized bed granulator were similar to those of the first coating of the example 3-1.

(2) The second coating

49.5 g of hydroxypropylmethylcellulose phthalate (HPMCP) was dissolved in a mixture of 247.5 ml of acetone and 247.5 ml of ethanol to make a coating solution. The above coated granules were fluidized in the fluidized bed granulator and coated by spraying the second coating solution. The preparing conditions of the fluidized bed granulator were similar to those of the

second coating of the example 3-1.

<Example 3-3>

φ The first coating

100 g of hydroxypropylmethylcellulose (HPMC2910) was dissolved in 1,000 ml of distilled water to make a coating solution. 1,000 g of 6:4 (weight) mixture of Artemisia japonica extract granule and Saururus chinensis extract granule, prepared in the example 1, was fluidized in the fluidized bed granulator and coated by spraying the coating solution. The preparing conditions of the fluidized bed granulator were similar to those of the first coating of the example 3-1.

(2) The second coating

49.5 g of shellac was dissolved in 495 ml of ethanol to make a coating solution. The above coated granules were fluidized in the fluidized bed granulator and coated by spraying the second coating solution. The preparing conditions of the fluidized bed granulator were similar to those of the second coating of the example 3-1.

<Example 4> Intake rate evaluation of coated extract granules

Test feeds for shrimp were made with coated extract granules prepared in the above examples 3-1 to 3-3. Intake rates according to coating polymers were evaluated as follows:

Shrimp: Palaemon paucidens was used and their average body

weight was 0.3g. Shrimps were accustomed to experimental environments for one week before tests. During that accommodation period, general feed was supplied the shrimps.

Test group: Control group was provided with feed having 6:4 (weight) mixture of uncoated Artemisia japonica extract granules and uncoated Saururus chinensis extract granules, prepared in example 1. 30 shrimps were included in each group.

Feeding of test samples: Test samples were supplied from the 0 ^th day to the 5 ^th day, wherein the 0 ^th day was the day after one week of the above accommodation period ended. 2 g of feed

was supplied once a day, and feeding was performed at a.m. 10 o'clock everyday.

Raising conditions of shrimp: Shrimps were raised in 10

liter of water bath. The temperature of the bath was kept at 20+ I ⁰ C.

Evaluation items: From the 0 ^th day to the 5 ^th day of feeding test samples, the remaining amount of feed supplied in the previous day was recorded everyday at the same time of feeding as 5 points (from 1 to 5) according to the remaining amount in water bath. After the evaluation of the remaining amount, water bath were cleaned and shrimps were supplied new feeds. After dividing the bottom of the bath into 6 parts (3 partitions in the width and 2 partitions in the length) , points were granted as follows: grade 0 (-) - all 6 parts have no

remaining feed; grade 1 (+ ) - one part among 6 parts has remaining feed; grade 2 (++) - two parts have remaining feed; grade 3 (+++) - three parts have remaining feed; grade 4 (++++) - four parts have remaining feed; and grade 5 (+++++) - five or all parts have remaining feed. Results were shown in table 1 below.

[Table 1]

As shown in the table 1, groups provided with examples 3-1 to 3-3 having coated samples showed higher intake rate than control group provided with uncoated extracts. Particularly, example 3-1 showed the highest intake rate of feed.

<Example 5> Evaluation on the medicinal effect of coated extract

As said in the followings, it was evaluated whether the coated extract showed a good medicinal effect or not.

Shrimp: Kuruma shrimp (Maruspenaeus japonicus) was used and the average body weight of the shrimps was 10.5-18.9 g. 17-

20 shrimps were included in each group. Shrimps were accustomed to experimental environments for two weeks before test. During the accommodation period, general feed was supplied.

Test sample: The coated extract granules prepared in the

example 3-1 were used.

Test feed: Gold Prawn (Higashimaru, Kagoshima, Japan) was used as test feed. 0 g, 125 g, 250 g, 500 g or 1000 g of the test sample was added per one ton of the test feed. The supply amount of test feed was 1% (per one day) of the body weight per one shrimp.

Feeding of test feed: Test feed was supplied everyday from 7 days before infection of the virus to 14 days after infection of the virus. Feeding of test feed was performed p.m. 6 o'clock

everyday.

Raising conditions of shrimp: Shrimps were raised in 60

liter of water bath. The temperature of the bath was kept at 21+

HC.

Infection of white spot syndrome virus: Shrimps were artificially infected with white spot syndrome virus and died finally. Three died shrimps were finely ground in 10 ml of sterilized sea water. The ground solution was diluted with sterilized sea water as 8000:1 of ratio to make a virus infection solution. Infection of virus was performed by soaking shrimps of each group in the virus infection solution for two hours and then again transferring the shrimps into the 60 liter of water bath. After the infection, mortality rate was evaluated for 10 days.

Evaluation item: After the infection of virus, the

mortality rate of each group and conventional symptoms of white spot syndrome virus were evaluated.

Example 5 was twice performed and results were shown in figures 2 and 3, respectively. As shown in figures 2 and 3, groups administered with test samples showed higher survival rate than control group. Especially, groups administered with

500 g and 1,000 g of test samples showed much higher survival rate than control group. These results show that the coated extract according to the present invention has a good medicinal effect.

[INDUSTRIAL APPLICABILITY]

As shown above, the present invention can improve shrimp's intake rate of drug or feed by coating drug or feed with polymer.