JPS63196514A - Preventive and remedy for protozoological disease - Google Patents

Abstract

PURPOSE:To obtain a preventive and remedy having safe and economic effects on protozoological diseases, containing tannin of condensation type. CONSTITUTION:The titled preventive and remedy containing tannin of condensation type having flavan-2-ol (catechin) shown by formula I [R is group shown by formula II (n is 1-15), 4'-position of R is bonded to 6-position or 8-position of the compound shown by formula I, plural R are bonded to 4'-position, 6'-position and 8'-position and R' shown by formula III is bonded to 6'-position or 8'-position of R], further forming a trimer or a polymer by various bonding manners. Purified tannin of condensation type may be used as the tannin of condensation type and a plant (preferably Parabenzoin trilobum N., Neolitsea aciculata B., of lauraceous plant) or crude drug (e.g. powdered Cinnamon bark or Cinnamomum loureirii Nees) may be used. Chicken coccidiosis, swine coccidiosis and bovine coccidiosis are preferable as the target diseases of the preventive and remedy.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of the Invention The present invention relates to a prophylactic and therapeutic agent for protozoal diseases containing condensed tannins.

PRIOR ART Diseases caused by protozoa include, for example, chicken coccidiosis, chicken leukocytozoonosis, swine and bovine coccidiosis, toxoplasmosis, malaria, piroplasmosis, human malaria, tropical fever, chicken and swine. .

Various protozoal diseases such as trypanosomiasis and leishmaniasis are known to occur in cattle, and conventional preventive and therapeutic agents for these protozoal diseases include sulfa drugs, quinoline derivatives, nucleic acid derivatives, quinazoline derivatives, guanidine derivatives, and 1-folate antagonists. agents, polyether antibiotics, etc. are used.

On the other hand, tannins widely contained in higher plants are classified into hydrolyzable type and condensed type (non-hydrolyzable type) based on their structure, and their chemical properties and physiological activities can be found in Pharmaceutical Journal 103.
125 (1983), but it is not known to have any biological activity against protozoa.

Problems to be Solved by the Invention Chicken coccidiosis, an example of a protozoal disease, is caused by the orally ingested protozoan Eimeria tenella.
tenella), Eimeria necatolic (Ei
meria necatrix), Eimeria acervulna (Eimeria acervultna)
This disease is caused by the proliferation of oocysts in the sporulation stage in the gastrointestinal tract of chickens. Affected chickens excrete diarrhea, mucus, mucus, bloody stool, etc., lose their appetite, and lose their appetite. This results in delayed development, weight loss, reduced egg production, and even death. Chicken leucocytozoonosis is caused by the protozoan Leukocytozoon callaraeri (L).

caulleryi),
Affected chickens exhibit symptoms such as anemia and green stools, which results in delayed growth, weight loss, reduced egg production, and even death.

Furthermore, swine coccidiosis is a disease in which young pigs are generally susceptible to infection after weaning, and the symptoms include clear diarrhea, mainly catarrhal enteritis, and in cases of heavy infection, the wings may be affected. In such cases, anemia, weakness, and loss of appetite are seen, leading to subsequent growth retardation.

As mentioned above, diseases caused by protozoa, particularly sporozoites, are causing great damage to livestock industries such as poultry and pig farming.

Sulfa drugs, quinoline derivatives, nucleic acid derivatives, quinazoline derivatives, which are conventionally used for such protozoal diseases;
Guanidine derivative 1 folate antagonists, polyether antibiotics, and the like are expensive and have a very narrow safety margin, so there are many problems when mixing them with feed and using them in large quantities.

Means for Solving the Problems In view of the above-mentioned circumstances, the present inventors conducted intensive studies to provide an excellent preventive treatment for protozoal diseases, and as a result, unexpectedly discovered that condensed tannins contained in higher plants It was discovered that the present invention showed biological activity against Otsu protozoa, and based on this, further research was conducted and the present invention was completed.

That is, the present invention is a prophylactic and therapeutic agent for protozoal diseases, which contains condensed plastic tannins.

The condensed tannins in the present invention are a group of compounds that have flavan-3-ol (catechin) as a constituent unit and form trimers and even multimers through various bonding modes, as represented by the following general formula (1). It is.

(Left below) [In the formula, n represents an integer from 1 to 15, the 4'-position of R is bonded to the 6- or 8-position of the formula, and the 4'-position of R is bonded to the 6'-position of R. or bonded to the 8' position, and R' indicates bonded to the 6' or 8' position of R.] As the condensed tannin in the present invention, purified condensed tannin may be used; Containing substances such as plants or herbal medicines containing the condensed tannins may be used.
Flowers (seeds, m parts, roots, stems, etc.) can be used fresh, dried, or dried and then crushed or powdered.

Examples of plants containing condensed tannins used in the present invention include camphor tree (C4nnamoIIlumCa)
Mphora S.), Lourei C.
rii N,), Macbilus Th
umnbergii S,), Kuromoji (Benzoi)
umbellatum R,), Shiromoji (P
arabenzojn trilobum N,)
, Japanese woodpecker (L.

erythrocarpa M,), Tendaiuyaku (
L.

5trychnHolia F,), Ya? -”L Ubashi (L.

glauca B,), Cinnam
omumjaponicum S, ), ? Rubanikai (C, dophnoidesS,), Gekkeiju (
Lauras nobilis L,), Arabian (Parabenzoin praecox N,
), Snuzzle (Cassytha riliform)
is L, ), Neolitseaseri
cea B,), dankorabai (], indcraob
tusiloba B,), Dog Goose (Neolits)
Lauraceae plants such as ea aciculata B,), cinnamomum cassia B,
Magnoliaceae plants such as Rheum palmatum L,
Roim tanguticum (R, tanguticum)
, Polygonum multir
lorum T.), Polygonaceae plants such as Ephedra equisetina (Ephedra equisetina)
B,), I-7 Edora distachia (E, dista
Chya L,) and other plants of the Ephedra family, betel nut (Ar
Palm family plants such as eca catechuL), lingonberry (Vaccinium 'vitis -1da)
ea L,), Ericaceae plants such as Myri
stica iragrans Il,), Myrica rubra (Myrica rubra)
Proteaceae plants such as S, ), Uncaria gambir Rl, Chinonaledgeriana
a M,), Rubiaceae plants such as Rosaceae (Rosa
multiflora T.), Jarimbai (
Rhaphiolepis umbellata
Rosaceae plants such as M.), Kandelia
Candel D.
Among them, Nymphaeaceae plants such as Nelumbo nucifera G, ) are preferred, and among them, Nymphaeaceae plants, Lauraceae plants, Shiromoji, Inugashi, and Dankoubai are preferred.
More preferred are Shiromoji and Inugan.

The herbal medicines containing condensed tannins used in the present invention include whole parts of the above plants or parts containing condensed tannins and secondary processed products thereof, such as bark, cinnamon, Shimoji bark, rhubarb, Heshucho, Ephedra, Hama Eiko, Cowberry,
Meuduzhuang, Yang Meipi, Yingshi, Asianyaku.

Examples include cinchona bark, jarinbai bark, cane bark, lotus capsule, and among these, cinnamon bark, large amount, heshuchodori, and Asenyaku are preferred, and Asenyaku is more preferred.

When administering the above-mentioned plants and herbal medicines as prophylactic and therapeutic agents for protozoal diseases, whole or condensed tannin-containing parts (e.g., bark, 7 m/seed of flower, roots) should be administered.

Administer by mixing 1 type or 2B or more of fresh, dried, or dried and then crushed or powdered (preferably dried and then crushed or powdered) stems, etc.). Bye.

The purified condensed tannin used in the present invention can be produced, for example, by the following method.

Whole or condensed tannin-containing parts of the above-mentioned plants or herbal medicines (e.g., bark, one flower, one seed, one leaf).

Roots, stems, etc.) fresh, dried, or dried and then crushed or powdered (preferably dried and then crushed or powdered) are mixed with alcohol (
e.g., methanol, ethanol, etc.).

Ketones (e.g., acetone, methyl ethyl ketone, etc.),
About 50% of solvents such as chloroform or their combined solvents are used.
Extract using ~10x m for 2 hours to 2 weeks under conditions of room temperature to reflux heating. Concentrate the extract and use polystyrene resin or dextran resin (e.g., Diaion HP-
20, Diaion XAD-2, MCl-gel CHP-
20, Sephadex LH-20, etc.) and condensing the fraction eluted with hydrous alcohol or acetone to obtain condensed tannins.

The dosage of purified condensed tannins, plants or crude drugs containing condensed tannins in the prophylactic and therapeutic agent for protozoal diseases of the present invention will vary depending on the administration method, purpose of administration, recipient, disease symptoms, target disease, etc. When mixed with feed, insects that account for about 0.1 to 40% by weight of the normal feed are preferable, and in the case of purified condensed tannins, about 0.1 to 10% by weight, more preferably about 0.5 to 3% by weight, In the case of plants or herbal medicines containing condensed tannins, about 2.5 to 40% by weight;
More preferably, the amount is about 4 to 20% by weight.

The purified condensed tannins, plants and herbal medicines containing condensed tannins used in the present invention have extremely low toxicity; for example, the 50% lethal dose (
(Observation for 8 days after administration) was 20g/20g of purified condensed tannin.
kg/day or more, and in plants and herbal medicines containing condensed tannins (for example, Shiromoji, Inugan bark, Nikuduso, Asenyaku, Heshudori, etc.), the amount is 100g/kg/day or more, so the protozoa of the present invention Preventive and therapeutic agents for sexual diseases can be safely administered.

The prophylactic and therapeutic agent for protozoal diseases of the present invention is prepared by diluting purified condensed tannins, the above-mentioned plants or herbal medicines with a solid or liquid carrier, or not diluting them, or stabilizing them by coating, etc. , granules, tablets, liquids, emulsions, pastes, capsules, premixes, injections, etc., or as feed.

It is produced by adding it directly to drinking water or by adding it dispersed in a carrier. As the carrier, any carrier may be used as long as it is physiologically harmless in itself, and it is more preferable to use a carrier that can be used as feed or a component of feed. Examples of solid carriers include lactose, sucrose, starch, wheat flour,
Examples include corn flour, bran, soybean oil cake, fatty rice bran, rapeseed oil meal, tofu meal, cellulose yeast cells, fish meal, groundnut lees, shell powder, calcium carbonate, etc. Examples of liquid carriers include water, Examples include physiological saline and physiologically harmless organic solvents. Other appropriate adjuvants,
For example, appropriate amounts of emulsifiers, dispersants, suspending agents, wetting agents, thickening agents, gelling agents, and solubilizing agents may be added.

In addition, preservatives, bactericidal agents, coloring agents, air fresheners, and antibacterial agents.

Antibiotics, enzyme preparations, lactic acid bacteria preparations, antipyretics, II static drugs.

Anti-inflammatory agents and the like may be blended, and conventionally used prophylactic and therapeutic agents for protozoal diseases may also be blended and used in combination.

Also various vitamins and minerals.

Amino acids and the like may also be blended.

Target diseases of the preventive and therapeutic agent for protozoal diseases of the present invention include:
・For example, chicken coccidiosis, chicken leukocytozoonosis, pig and cow coccidiosis, toxoplasmosis, malaria, piroplasmosis, human malaria, tropical fever, chicken,
Examples include trypanosomiasis and leishmaniasis that occur in pigs and cattle, but among them, chicken coccidiosis, pigs,
Bovine coccidiosis is preferred.

EXAMPLES The present invention will be specifically explained with reference to Examples below, but the present invention is not limited thereto.

Example 1 An efficacy test against coccidia was conducted using chickens. Specifically, each group of 9-day-old white Leghorn male chicks was made up of 3 birds, and 24 hours after the start of the test, 5 x 10' sporulating oocysts of Eimeria tenella per chicken were orally administered to the chickens in all groups except the non-infected control group. was inoculated.

The drug is dried and ground into standard feed (SDL No.
, l: 10% by weight (manufactured by Nippon Compound Feed Co., Ltd.) was added and continuously administered by ad libitum administration for 9 days from 24 hours before infection to 8 days after infection. During the rearing period, the weight of each chick was measured every day and the appearance of bloody stool was observed.Furthermore, on the 8th day after oocyst inoculation, a necropsy was performed to examine the lesion status of the cecum, and the anti-coccidial effect of the administered test drug was determined. . The results are shown in Table 1.

As is clear from the test results in Table 1, the weight gain ratio increased in the non-invention plots compared to the infected plots, and excellent anti-coccidial effects were observed.

As controls, persimmon leaves, whole mugwort plants, and valerian roots, which do not contain condensed tannins, were used.

2) Blood stool index: The amount of blood excreted from the chick's intestines is expressed as the number of blood stains found on the bedding (number of blood stains/chicken).

3) Cecal lesion, -1 normal, +: mild, 廿: moderate, t:
Severe 1 book: Extremely severe lesion. The numbers indicate losses.

Example 2 The anti-coccidial effect of the condensed tannin-containing crude drug was determined in the same manner as in Example I. The results were as shown in Table 2.

(Margins below) Table 2 Example 3 Condensed tannins from Shimoji bark were produced by the following method.

8 kg of dried and pulverized Shimoji bark was extracted with 70% acetone 81 at room temperature for one week. The extract was filtered and the solvent was distilled off under reduced pressure to obtain 45.86 kg of an aqueous solution. This was passed through a Diaiont IP-20 column (3 (H) and eluted with H,090Q, then 30% acetone (150C).
45.6 g of anticoccidial active fraction was obtained in the fraction eluted with.

In the same way, Tendo, Nashu Tori, Ephedra, Kippi, Nikuduso.

Anticoccidial active fraction was obtained from Asenyaku.

These anticoccidial active fractions are n-butanol-
A red color appears due to the hydrochloric acid reaction, and (2R,3S) appears due to the decomposition reaction with benzyl mercaptan.

4S)-4-benzylthioflavan-3,3',4'.

Since 5.7-pentaol was obtained together with catechin or epicatechin, it was confirmed that it was a condensed tannin.

The anti-coccidiosis effects of Shimoji bark and Tendo integrated tannin were determined in the same manner as in Example 1.

The results were as shown in Table 3.

The hydrolyzed tannins used as controls were geraniin obtained by the method described in Pharmaceutical Journal 99, 505 (1979) and commercially available Japanese Pharmacopoeia tannic acid (manufactured by Wako Pure Chemical Industries, Ltd.).

Table 3 Example 4 Purification and structure determination of Shimoji bark tannin were carried out as follows.

Condensed tannin of Shimoji bark obtained in Example 3 84.2
g by silica gel column chromatography (φ5x5
5 cm) and ethyl acetate-acetone-water (80:5
After fractionation with a mixed solvent (0:20 upper layer), 45.6 g of brown powder (1-7) was obtained from both fractions eluted with 50% methanol. The ice crystals took on a red color due to hydrochloric acid-butanol. In addition, 500 mg of ice crystals were dissolved in 30 ml of ethanol, 1 d of acetic acid and 2 ml of benzyl mercaptan were added thereto, and the reaction solution was reacted under reflux heating for 15 hours.The reaction solution was subjected to high performance liquid chromatography and gas chromatography. Purified condensed tannin (I-7)
) has a structure in which (-)-epicatechin is the upper unit, and the lower unit is (-)-epicatechin and (+)-rich, with a composition of 3 = 5, and the average degree of condensation is 6.12. It was confirmed.

The anti-coccidial effect of the purified condensed tannin (1-7) was determined in the same manner as in Example 1. The results were as shown in Table 4.

Table 4 Example 5 Corn, North Sea meal, soybean oil cake, powdered refined beef tallow,
Standard feed for broiler fattening 5DI3-N made from wheat flour, alfalfa meal, vitamins, and minerals.
The prophylactic and therapeutic agent for protozoal diseases of the present invention can be produced by mixing and blending 150 g of purified condensed tannin (1-7) with 10 kg of o.1 (manufactured by Japan Compound Feed Co., Ltd.).

Example 6 Corn, North Sea meal, soybean oil cake, powdered refined beef tallow,
Standard feed for broiler fattening using wheat flour, alfalfa meal, vitamins, and minerals 5DB-No.
, l (manufactured by Nippon Compound Feed Co., Ltd.) 40 Asenyaku per 10 kg
By mixing and blending 0 g of the protozoal diseases, the prophylactic and therapeutic agent for protozoal diseases of the present invention can be produced.

Example 7 A premix of the preventive and therapeutic agent for protozoal diseases of the present invention can be prepared by mixing 1 kg of soybean oil cake with 400 g of Asenyaku. This premix preparation can be diluted 10 times, mixed into feed, and administered as the preventive and therapeutic agent of the present invention.

Effects of the Invention The prophylactic and therapeutic agent for protozoal diseases of the present invention exhibits safe and economically excellent effects against protozoan diseases, and is particularly advantageous in livestock farming.

Claims (1)

[Claims] A preventive treatment agent for protozoal diseases containing condensed tannins.