JP2821912B2 - Animal antibacterial agent - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to the following general formula (I) [Wherein, R represents an alkyl group having 1 to 6 carbon atoms (provided that R is
In the case of a racemic compound, excluding those in which R is a methyl group)], a racemic compound or a 3S compound of a pyrido [1,2,3-de] [1,4] benzoxazine derivative, or a salt or hydrate thereof The present invention relates to a prophylactic / therapeutic agent for animal infectious diseases as an active ingredient.

[Conventional technology and its problems]

In recent years, in the livestock industry, the number of breeding heads per unit area has increased, but on the other hand, the frequency of occurrence of various infectious diseases of animals such as livestock and poultry has increased, the damage has increased year by year, and economic loss has been large. Is in question.

For the prevention or treatment of infectious diseases in animals, antibiotics such as tylosin and oxytetracycline and synthetic antibacterial agents such as oxophosphoric acid and pyromidic acid have conventionally been added to feed and drinking water by oral administration or injection. Methods have been implemented.

However, these antibacterial agents for animals have weak defects such as weak antibacterial activity, narrow antibacterial spectrum, emergence of resistant bacteria, little therapeutic effect, narrow safety margin, side effects, and uneconomical efficiency. It was not satisfactory.

In addition, the pyrido [1,2,
A compound in which R of the 3-de] [1,4] benzoxazine derivative is a methyl group and is in a racemic form is already known as a prophylactic / therapeutic agent for mycoplasma infection in animals (Japanese Patent Application Laid-Open No. 60-1985). No. 184014), however, this compound has a narrow antibacterial spectrum and its antibacterial activity is not always satisfactory.

[Means for solving the problem]

Under such circumstances, the present inventors have conducted intensive studies on a safe animal antibacterial agent which has a wide antibacterial spectrum and is effective against various animal infectious diseases and has low toxicity, and as a result, the pyrido represented by the general formula (I) Among the [1,2,3-de] [1,4] benzoxazine derivatives, the racemic form (excluding those in which R is a methyl group) or the 3S form has excellent preventive and therapeutic effects on animal infectious diseases over a wide range. And found that it is safe with little toxicity.

That is, the present invention relates to a racemic (excluding those in which R is a methyl group) or a 3S-form of the pyrido [1,2,3-de] [1,4] benzoxazine derivative represented by the above formula (I). A bacterium or chicken selected from pig or chicken-derived Salmonella, pig-derived Bordetella, pig-derived Pasteurella, dog-derived Brucella, dog-derived Yersinia, and dog-derived Lactobacillus containing a salt or a hydrate thereof as an active ingredient. It is intended to provide a prophylactic / therapeutic agent for animal infectious diseases caused by derived mycoplasma.

The pyrido [1,2,3-de] [1,
4] benzoxazine derivatives (hereinafter, “compound (I)”
There are several optical isomers, but the racemic or 3S form is used as the preventive / therapeutic agent for animal infectious diseases of the present invention, and the 3S form is preferable because of its high antibacterial activity.

Examples of the compound (I) include the following compounds.

(1) 3S-9-fluoro-2,3-dihydro-3-methyl-10- (4-methyl-1-piperazinyl) -7-oxo-7H-pyrido [1,2,3-de] [1, 4) benzoxazine-
6-carboxylic acid hemihydrate (2) 3 (RS) -9-fluoro-2,3-dihydro-3
-Methyl-10- (4-ethyl-1-piperazinyl) -7
-Oxo-7H-pyrido [1,2,3-de] [1,4] benzoxazine-6-carboxylic acid (3) 3 (RS) -9-fluoro-2,3-dihydro-3
-Methyl-10- (4-n-propyl-1-piperazinyl)-
7-oxo-7H-pyrido [1,2,3-de] [1,4] benzoxazine-6-carboxylic acid (4) 3 (RS) -9-fluoro-2,3-dihydro-3
-Methyl-10- (4-iso-propyl-1-piperazinyl)
-7-oxo-7H-pyrido [1,2,3-de] [1,4] benzoxazine-6-carboxylic acid (5) 3 (RS) -9-fluoro-2,3-dihydro-3
-Methyl-10- (4-n-butyl-1-piperazinyl) -7
-Oxo-7H-pyrido [1,2,3-de] [1,4] benzoxazine-6-carboxylic acid (6) 3S-9-fluoro-2,3-dihydro-3-methyl-10- (4 -Ethyl-1-piperazinyl) -7-oxo-7H-pyrido [1,2,3-de] [1,4] benzoxazine-
6-carboxylic acid (7) 3S-9-fluoro-2,3-dihydro-3-methyl-10- (4-n-propyl-1-piperazinyl) -7-oxo-7H-pyrido [1,2,3 -de] [1,4] benzoxazine-6-carboxylic acid (8) 3 (RS) -9-fluoro-2,3-dihydro-3
-Methyl-10- (4-ethyl-1-piperazinyl) -7
-Oxo-7H-pyrido [1,2,3-de] [1,4] benzoxazine-6-carboxylic acid hydrochloride hemihydrate Of the above exemplified compounds, compound (6) includes gram-positive bacteria and the like. It has excellent antibacterial activity against a wide range of bacteria and has a high blood concentration when administered to animals (for example, when 10 mg / kg is orally administered to a dog, the blood concentration after 1 hour is about 6 μg / ml. Was particularly preferred.

Compound (I) used in the present invention can also be used as an acid addition salt due to the basicity of piperazinyl at the 10-position substituent. Examples of the acid addition salt include hydrochloride, hydrobromide, and iodine. Inorganic salts such as hydrides and sulfates, or organic salts such as acetate, methanesulfonate, citrate, benzenesulfonate and lactate are exemplified.

The carboxyl group at the 6-position may be in the form of a salt, for example, an alkali metal salt such as a sodium salt or a potassium salt, an alkaline earth metal salt such as a magnesium salt or a calcium salt, an ammonium salt, a triethylamine salt or the like. Organic salts are included.

Further, the compound (I) or a salt thereof can be used in the form of a hydrate.

The method for producing the compound (I) is described, for example, as a racemic form in JP-A-57-46986 or JP-A-58-72589, and as a 3S form, as described in JP-A-62-252790. A method can be used.

As a method for administering compound (I) or a salt or hydrate thereof (hereinafter, referred to as “active compound (I)”) to an animal, a method of orally administering directly or by mixing it in a feed, or a solution After that, a method of orally administering directly or by adding to drinking water or feed, and a method of administering by injection are further exemplified.

The dose at this time varies depending on the purpose of administration (prevention or treatment), the type and size of the animal to be treated, the type of infected pathogen, the degree of infection, and the like.
1 to 200 mg, preferably 5 to 1 kg of animal body weight per day
~ 100mg. Administration can be performed once or several times a day. This dose is only a rough guide, and it is also possible to administer an amount outside the above range according to the age, body weight, medical condition, etc. of the animal. The period of administration is not particularly limited, but a sufficient effect can be usually obtained by administration for about 1 to 10 days, and intermittent administration is also possible.

Preparations for administering the active compound (I) to animals can be suitably prepared as powders, fine granules, soluble powders, syrups, solutions or injections by techniques usually used in this field. The typical formulation examples are described below.

Formulation Example 1 (powder for mixing feed): Active compound (I) 1 to 10 parts by weight Corn starch 98.5 to 89.5 parts by weight Light silica silicic acid 0.5 part by weight Total 100 parts by weight Formulation Example 2 (soluble powder for mixing feed or drinking water) ): 1 to 10 parts by weight of active compound (I) (water-soluble) lactose 99 to 90 parts by weight Total 100 parts by weight Formulation Example 3 (solution): 1 to 10 parts by weight of active compound (I) acetic acid or water Oxidation 5 to 20 parts by weight Sodium paraoxybenzoic acid 0.1 part by weight Ethyl purified water 69.9 to 93.9 parts by weight Total 100 parts by weight [Effect of the Invention] The preventive / therapeutic agent provided by the present invention causes infectious diseases in animals. Has a wide range of antibacterial activity against various bacteria, for example, Escherichia, Salmonella, Pasteurella, Haemophilus, Bordetella, Staphylococcus, Mycoplasma, etc. Demonstrate . Therefore, it is useful in preventing, treating and treating infectious diseases of cattle, pigs, birds, dogs, cats and the like. To illustrate specific disease names, in cattle, colibacillosis, salmonellosis, hemorrhagic sepsis, mycoplasma infection, bovine lung disease, mastitis, etc., and in pigs, colibacillosis, salmonellosis, pasteurellosis, hemophilus infection, atrophic disease Rhinitis, exudative epidermitis,
In birds, such as mycoplasma infection, birds, colibacillosis, chick dysentery, chicken paratyphoidosis, poultry cholera, infectious coryza, staphylococci, mycoplasma infection, etc. Examples include cystitis and the like, and in cats, exudative pleurisy, cystitis, chronic rhinitis, haemophilus infection, kitten diarrhea, mycoplasma infection and the like.

〔Example〕

 Next, the present invention will be described in more detail with reference to examples.

The test compound numbers of the active compounds (I) used in the examples are as shown in Table 1 below.

Example 1 In Vitro Antibacterial Activity Measurement Test for Various Animal-Derived Pathogens (Part 1): Each test strain shown in Table 2 was cultured overnight in a heart infusion broth medium (manufactured by Eiken Chemical Co., Ltd.).
This culture solution (the number of bacteria is about 10 ⁸ cells / ml) is diluted 100-fold, a fixed amount (about 0.05 ml) is taken, and a Mueller-Hinton medium (Eiken) to which the test compound (1) at each concentration step has been added in advance. (Chemical Co., Ltd.) and inoculated on a plate, and cultured at 37 ° C. for 18 to 24 hours. After culturing, the minimum concentration of the test compound that inhibited the growth of the inoculum, that is, the minimum inhibitory concentration (MIC), was measured. Table 2 shows the results.

As is clear from Table 2, the compound (1) of the present invention
The racemic known compound (OFLX) (JP-A-60-184014)
No. 3) showed higher antibacterial activity.

Example 2 In Vitro Antibacterial Activity Measurement Test for Various Animal-Derived Pathogens (Part 2): The minimum growth inhibitory concentration (MI) of compounds (1) to (8) against various animal-derived pathogens was determined in the same manner as in Example 1.
C) was measured. The results are shown in Table 3 below.

As is clear from Table 3, the compounds (1) to
(8) showed higher antibacterial activity than oxophosphoric acid which is currently widely used as an antibacterial agent for animals in an in vitro antibacterial activity test.

Example 3 In Vitro Antibacterial Activity Measurement Test for Various Animal-Derived Pathogenic Bacteria (Part 3): Minimum growth inhibitory concentrations of compounds (1), (2) and (6) against various animal-derived pathogens by the same method as in Example 1. (MIC) was measured. The results are shown in Table 4 below.

Example 4 Antibacterial activity against chicken-derived Mycoplasma garicepticum: Each test strain of chicken-derived Mycoplasma garicepticum was cultured in Frey's liquid medium for mycoplasma for 3 days, and the culture solution (the number of cells was about 10 ⁸ cells / ml) of the ^10-2 diluted solution was inoculated into Frey's liquid medium for Mycoplasma, which was previously added with the test compound at each concentration step.
Cultured for 5 days. During this period, observe the color tone of the liquid medium daily, and when the color tone of the liquid medium not inoculated with the test compound set as a control changes to red, the minimum concentration at which the color tone of the liquid medium containing the test compound remains yellow MI
C. The results are shown in Table 5 below.

Example 5 Effect of Oral Administration on Chicken Escherichia coli Infected Chicken: Escherichia coli, a causative fungus of chicken Escherichia coli isolated from diseased chickens
Cory was cultured overnight in trypticase soy broth (BBL), and 0.2 ml of this culture was injected into the trachea of chickens averaging about 80 g using a 1 ml disposable syringe to artificially infect the chicken. 2 hours after infection, test compound (1)
Feeds mixed and added to 50, 25, and 12.5 ppm concentrations were fed for 5 days and allowed to freely ingest. After completion of the administration, the animals were allowed to freely ingest the drug-free feed for an additional 5 days, and then the surviving chickens were sacrificed and dissected, and the degree of lesion in the air sac was observed.
Escherichia Cory in lungs and air sacs was searched.
Table 6 shows the results. As is clear from the results, the compound (1) of the present invention exerted an excellent effect on chicken Escherichia coli infection.

Example 6 Effect of Oral Administration on Mycoplasma garicepticum Artificially Infected Chickens (Part 1): Mycoplasma garicepticum, a causative fungus of chicken respiratory mycoplasma disease isolated from diseased chickens, was used for mycoplasma
The cells were cultured in Frey's liquid medium for 3 days, and 0.2 ml of the culture solution was injected into the trachea of chickens having an average body weight of about 80 g using a 1 ml disposable syringe to artificially infect the chickens. From the day after the infection, a feed containing the test compound (1) at a concentration of 100, 75, or 50 ppm was fed for 5 days and fed freely. After dosing,
After a free intake of the drug-free feed for an additional 5 days, surviving chickens were sacrificed and dissected, the degree of lesion in the air sac was observed, and then Mycoplasma galicepticum in the suborbital sinus, trachea, lungs, and air sac was searched. Table 7 shows the results. As is clear from the results, the compound of the present invention (1)
Exerted an excellent effect on chicken mycoplasma galisepticum infection.

Example 7 Effect of Oral Administration on Mycoplasma garicepticum Artificially Infected Chickens (Part 2): The oral administration effect of test compounds (2) and (3) on Mycoplasma galisepticum artificially infected chickens was examined in the same manner as in Example 6. . Table 8 shows the results. From this example, it was revealed that the compounds (2) and (3) of the present invention have an excellent effect on chicken mycoplasma galisepticum infection.

Example 8 Acute toxicity by intravenous administration to mice: Representative compounds (2) and (3) of test compounds were intravenously injected into mice, and compound (2) was 200 mg / kg.
(10 per group), compound (3) is also 200 mg / kg (6 per group)
In, no deaths, LD ₅₀ values are considered further.

(Reference example)

Next, the compound (5) of the present invention used in the test example,
The production examples of (8) and (7) are shown for reference.

Production Example A (test compound number (8)) 3 (RS) -9-fluoro-2,3-
Dihydro-3-methyl-10- (4-ethyl-1-piperazinyl) -7-oxo-7H-pyrido [1,2,3-de] [1,
4] 2.0 g of benzoxazine-6-carboxylic acid and concentrated hydrochloric acid
Add 2.0ml and heat to dissolve. The excess ethanol was concentrated under reduced pressure, and the precipitated crystals were collected by filtration, washed with ethanol, and recrystallized from aqueous ethanol to give 3 (RS) -9-fluoro-2,3- showing a melting point of 285 to 290 ° C (decomposition). Dihydro-
3-methyl-10- (4-ethyl-1-piperazinyl)-
1.3 g of 7-oxo-7H- [1,2,3-de] [1,4] benzoxazine-6-carboxylic acid hydrochloride hemihydrate was obtained.

Elemental analysis: as C ₁₉ H ₂₂ O ₄ N ₃ F.HCl.0.5H ₂ O Theoretical: C54.22, H5.75, N9.98 (%) Actual: C54.20, H5.79, N9. 85 (%) Production Example B (test compound number (5)) 3 (RS) -9-fluoro-2,3-dihydro-3-methyl-10- (1-piperazinyl) -7-oxo-7H-pyrido [ 1.5 g of anhydrous sodium carbonate was added to a mixture of 2.0 g of dimethylformamide of 2.0 g of 1,2,3-de] [1,4] benzoxazine-6-carboxylate hydrate and 10 ml of water, followed by stirring at room temperature for 10 minutes. -Butyl bromide (2.0 g) was added, and the mixture was stirred at the same temperature for 1 hour. Thereafter, 1.0 g of n-butyl bromide and 1.0 g of triethylamine are added, and stirring is continued at 60 to 70 ° C. for 5.5 hours. After the reaction, the mixture is concentrated, water is added, the mixture is acidified with hydrochloric acid under ice-cooling, then returned to weak basicity (pH 8 to 9) with aqueous ammonia, and extracted with chloroform. The aqueous layer is neutralized with acetic acid and extracted with chloroform. The chloroform layers are combined, washed with water, dried (Na ₂ SO ₄ ) and evaporated. 5 ml of concentrated hydrochloric acid, 40 ml of tetrahydrofuran and 20 ml of chloroform were added to the residue,
Stir at 70 ° C. for 4 hours. After the reaction, the solvent was distilled off, water was added, and aqueous ammonia was added under ice-cooling to make the mixture weakly basic (pH 8 to 10).
9) Extract with chloroform. The aqueous layer is neutralized with acetic acid and extracted with chloroform. The chloroform layers are combined, washed with water, dried (Na ₂ SO ₄ ) and the solvent is distilled off. Activated carbon is added to the residue and recrystallized from chloroform-acetonylyl. Then, if recrystallization is repeated from a concentrated ammonia water-methanol-water system, 3 (RS) -9-fluoro-2,3-dihydro-3-methyl-10- (4-n showing a melting point of 241 to 244 ° C. (decomposition). -Butyl-1-piperazinyl) -7-oxo-7H-pyrido [1,2,
3-de] [1,4] benzoxazine-6-carboxylic acid 300mg
was gotten.

Elemental analysis value: C ₂₁ H ₂₆ O ₄ N ₃ F Theoretical value: C62.51, H6.50, N10.41 (%) Actual measurement value: C62.42, H6.56, N10.39 (%) Production example C (Test compound number (7)) 3S-(−)-9,10-difluoro-2,3-dihydro-3-
Methyl-7-oxo-7H-pyrido [1,2,3-de] [1,4]
A solution of 23.1 g of N- (n-propyl) piperazine in 8 ml of dimethylsulfoxide is added dropwise to a suspension of 19.9 g of benzoxazine-6-carboxylate boron difluorochelate in 48 ml of dimethylsulfoxide with stirring under water cooling, and then the mixture is stirred at room temperature for 24 hours. . After completion of the reaction, 120 ml of water was added under ice-cooling and stirring, and the precipitated crystals were collected by filtration, washed with water and dried to obtain 21.3 g of yellow powder crystals. 29.1 g of triethylamine and 200 acetonitrile
Then, 200 ml of water and 200 ml of water are added, and the mixture is refluxed with stirring for 6 hours. The reaction solution was concentrated to dryness, the residue was dissolved in chloroform, washed with water, dried, (Na ₂ SO ₄ ) and the solvent was distilled off, and then repeatedly recrystallized from concentrated aqueous ammonia-ethanol to give a melting point of 219 to 222 ° C (decomposition). 3S-(-)-9-fluoro-2,3-dihydro-3-methyl-10- [4-n-propyl-1-piperazinyl) 7-oxo-7H-pyrido [1,2,3-de [1. 4] benzoxazine-6-carboxylic acid 10.1 g was obtained.

Elemental analysis: as C ₂₀ H ₂₄ O ₄ N ₃ F Theoretical: C61.68, H6.21, N10.79 (%) Actual: C61.45, H6.35, N10.73 (%)

──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Tadayoshi Miura 1-16-13 Kita-Kasai, Edogawa-ku, Tokyo Inside the Daiichi Pharmaceutical Central Research Laboratory (72) Inventor Toshiyuki Yoshioka 1-1-16-13 Kita-Kasai, Edogawa-ku, Tokyo JP-A-62-252790 (JP, A) JP-A-1-165589 (JP, A) European Publication 206283 (EP, A1) Chem, Pharm. Ball. , Vol. 32 (No. 12), p. 4907-4913 (1984) (58) Fields investigated (Int. Cl. ⁶ , DB name) C07D 498/06 A61K 31/535 CA (STN) REGISTRY (STN)

Claims (1)

(57) [Claims] 1. The following general formula (I) [Wherein, R represents an alkyl group having 1 to 6 carbon atoms (provided that R is
In the case of a racemic compound, excluding those in which R is a methyl group)], a racemic compound or a 3S compound of a pyrido [1,2,3-de] [1,4] benzoxazine derivative, or a salt or hydrate thereof Prevention of animal infectious diseases due to bacteria or chicken-derived mycoplasma selected from the group consisting of pig or chicken-derived Salmonella, pig-derived Bordetella, pig-derived Pasteurella, dog-derived Brucella, dog-derived Yersinia, and dog-derived Lactobacillus. -Therapeutic agents.