阅读说明：本技术 硝基噻唑衍生物及其盐酸盐、硫酸盐在制备治疗禽畜肠道感染的药物中的应用 (Application of nitrothiazole derivative, hydrochloride and sulfate thereof in preparation of medicines for treating intestinal infection of livestock and poultry ) 是由 衣云鹏 林树乾 杨世发 殷斌 刘月月 李桂明 赵增成 黄中利 于 2021-09-09 设计创作，主要内容包括：本发明涉及医药化学技术领域,尤其涉及硝基噻唑衍生物及其盐酸盐、硫酸盐在制备治疗禽畜肠道感染的药物中的应用。本发明合成的硝基噻唑衍生物在细胞水平上具有较高生物活性,对禽畜肠道感染尤其鸡、猪、羊、鹿细菌性肠炎体现出较好的生物活性,治疗效果显著,应用价值高。(The invention relates to the technical field of medicinal chemistry, in particular to application of nitrothiazole derivatives, hydrochlorides and sulfates thereof in preparation of medicines for treating intestinal infection of livestock. The nitrothiazole derivative synthesized by the invention has higher biological activity on a cellular level, shows better biological activity on intestinal infection of livestock and poultry, particularly bacterial enteritis of chicken, pig, sheep and deer, and has obvious treatment effect and high application value.)

1. Application of nitrothiazole derivatives, and hydrochlorides and sulfates thereof in preparation of medicines for treating intestinal infections of livestock and poultry.

2. The use as claimed in claim 1, wherein the livestock intestinal infections include infections caused by escherichia coli, and/or salmonella, ileitis caused by lawsonia or infections caused by drug-resistant clostridium welchii.

3. The use as claimed in claim 1, wherein the livestock intestinal infections include chicken infections caused by escherichia coli, and/or salmonella, ileitis of pigs caused by lawsonia, pig infections caused by streptococcus, or infections caused by clostridium welchii resistant to deer.

4. The application of nitrothiazole derivatives, and hydrochloride and sulfate thereof in preparing the medicine for treating the peritonitis of livestock is characterized in that the peritonitis of the livestock is peritonitis caused by haemophilus parasuis infection.

5. Application of nitrothiazole derivatives, hydrochlorides and sulfates thereof in preparation of medicaments for treating livestock lung infection is characterized in that the livestock lung infection is infection caused by streptococcus suis infection.

6. The use according to any one of claims 1 to 5, wherein the nitrothiazole derivatives, the hydrochloride and the sulfate thereof are used in an amount of 1mg/kg of large and medium-sized livestock and 3mg/kg of small livestock.

7. The use according to any one of claims 1 to 5, wherein the nitrothiazole derivative has the formula:the synthetic route is as follows:

8. the use according to claim 7, characterized in that the synthesis of said nitrothiazole derivatives comprises the following operating steps:

a, starting from 2-aminothiazole, carrying out nitration reaction by using sulfuric acid and nitric acid to generate a compound 2 (2-amino-5-nitrothiazole);

diazotization using sodium nitrite reaction and CuSO₄Catalytic downfeedPerforming a sanger reaction to generate a compound 3 (2-bromo-5-nitrothiazole); the reactions in the steps a and b are carried out in water;

c, carrying out nucleophilic substitution reaction on the 2-mercapto-5-amino-1, 3,4 thiadiazole and the compound 3 prepared in the step b under an alkaline condition to generate a compound 4, and thus obtaining the compound.

9. The use according to claim 8, wherein the base used in the basic condition of step c comprises one or more of triethylamine, N-diisopropylethylamine, sodium hydroxide, potassium carbonate, piperidine, N-methylmorpholine and diaminopyridine; the dosage of the alkali is 1-4 times of that of the compound 2-mercapto-5-amino-1, 3,5 thiadiazole; the solvent used in the alkaline condition comprises one or more of methanol, ethanol, tetrahydrofuran and acetone.

10. A pharmaceutical composition comprising, as an active ingredient, a nitrothiazole derivative or a pharmaceutically acceptable salt thereof for use according to claim 7.

Technical Field

The invention relates to the technical field of medicinal chemistry, in particular to application of nitrothiazole derivatives, hydrochlorides and sulfates thereof in preparation of medicines for treating intestinal infection of livestock.

Background

The breeding industry is often confronted with complicated and various bacterial infections, wherein the bacterial infections caused by bacteria such as escherichia coli, salmonella, pasteurella, clostridium welchii, haemophilus parasuis, staphylococcus aureus, streptococcus and the like are the most serious. Complicated bacterial infections occur frequently in breeding, and particularly intestinal bacterial infections occur frequently. For example, necrotic enteritis in chickens is a bacterial infectious disease of chickens caused by clostridium perfringens, but salmonella pullorum and escherichia coli have been reported to be also complicated in the course of the disease. Clinically, one drug is difficult to effectively control the intestinal infection of animals, so that the development of efficient antibacterial drugs for controlling the intestinal infection of livestock is particularly important.

Thiazoles and their derivatives are important groups in many drugs. The current research shows that the nitrothiazole derivative has the functions of improving the cardiac function and ischemia-reperfusion injury. Dharmalasriram et al, 2004 reported that nitrothiazole-based compounds could act on the ATPase domain of mycobacterial gyrase, thereby inhibiting bacterial growth. The approved drugs heterocyclic nitro compound nitrofuran, metronidazole are used as common drugs for treating intestinal infections. Nitroimidazole compounds such as tinidazole, ornidazole and the like are approved for the treatment of parasitic infections. Megazol is used to treat protozoal infections.

Disclosure of Invention

The nitrothiazole derivative synthesized by the method has higher biological activity on a cell level, shows better biological activity on livestock intestinal infections, particularly bacterial enteritis of chickens, pigs, sheep and deer, has obvious treatment effect and high application value, and solves the problems in the prior art.

The invention is realized by the following technical scheme:

application of nitrothiazole derivatives, and hydrochlorides and sulfates thereof in preparation of medicines for treating intestinal infections of livestock and poultry.

Further, the intestinal infections of the livestock include infections caused by escherichia coli, and/or salmonella, ileitis caused by lawsonia or infections caused by drug-resistant clostridium welchii.

Further, the intestinal infections of the livestock include escherichia coli, and/or chicken infections caused by salmonella, ileitis of pigs caused by lawsonia bacteria, or infections caused by clostridium welchii drug-resistant deer.

Application of nitrothiazole derivatives, hydrochlorides and sulfates thereof in preparation of medicaments for treating livestock peritonitis, wherein the livestock peritonitis is peritonitis caused by haemophilus parasuis infection.

Application of nitrothiazole derivatives, hydrochlorides and sulfates thereof in preparation of medicaments for treating livestock lung infection, wherein the livestock lung infection is infection caused by streptococcus suis infection.

Furthermore, the dosage of the nitrothiazole derivative, the hydrochloride and the sulfate thereof is 1mg/kg for large and medium livestock and 3mg/kg for small livestock.

Further, the structural formula of the nitrothiazole derivative is as follows:the synthetic route is as follows:

further, the synthesis of the nitrothiazole derivative comprises the following operation steps:

a, starting from 2-aminothiazole, carrying out nitration reaction by using sulfuric acid and nitric acid to generate a compound 2 (2-amino-5-nitrothiazole);

diazotization using sodium nitrite reaction and CuSO₄Carrying out Sanger reaction under catalysis to generate a compound 3 (2-bromo-5-nitrothiazole), wherein the reactions of the steps a and b are carried out in water;

c, carrying out nucleophilic substitution reaction on the 2-mercapto-5-amino-1, 3, 4-thiadiazole and the compound 3 prepared in the step b under an alkaline condition to generate a compound 4, and thus obtaining the compound.

Further, the base used in the basic condition of step c comprises one or a mixture of triethylamine, N-diisopropylethylamine, sodium hydroxide, potassium carbonate, piperidine, N-methylmorpholine and diaminopyridine; the dosage of the alkali is 1-4 times of that of the compound 2-mercapto-5-amino-1, 3, 5-thiadiazole; the solvent used in the alkaline condition comprises one or more of methanol, ethanol, tetrahydrofuran and acetone.

Further, the reaction temperature for the synthesis of compound 4 in step c is 0-30 ℃.

A pharmaceutical composition comprising the above nitrothiazole derivative or a pharmaceutically acceptable salt thereof as an active ingredient.

The invention has the beneficial effects that:

the 5- ((5-nitrothiazol-2-yl) thio) -1,3, 4-thiadiazole compound synthesized by the process is simple to operate, and the product yield is remarkably improved. The obtained derivative has high biological activity on a cell level through application research, has obvious effect in the aspect of preparing and treating intestinal infection diseases of livestock and poultry, and can be used for pertinently treating infection caused by escherichia coli, ileitis caused by lawsonia bacteria or infection caused by drug-resistant clostridium welchii and the like.

Drawings

FIG. 1 is a culture diagram of a culture medium for identifying pathogenic bacteria in cecum contents of a died spotted deer in example 7 of the present invention;

FIG. 2 is a graph of the growth of E.coli ATCC-25922 at various concentrations of Compound 4 in accordance with example 8 of the present invention;

FIG. 3 is a graph of the growth of Staphylococcus aureus ATCC-29213 at various concentrations of Compound 4 according to example 8 of the present invention;

FIG. 4 is a photograph of feces before administration to a herd of a test group in example 10 of the present invention;

FIG. 5 is a photograph of day 2 feces of swinery administered to the test group in example 10 of the present invention;

FIG. 6 is a photograph of day 3 feces of the swine herd of the test group of example 10 of the present invention;

FIG. 7 is a photograph of day 2 feces after withdrawal from a herd in test group of example 10 according to the present invention;

FIG. 8 is a photograph of the day 9 of discontinuation of treatment in the swine herd of the test group of example 10 according to the present invention;

FIG. 9 is a photograph of feces of test deer in accordance with example 11 of the present invention before administration;

FIG. 10 is a photograph of day 2 feces of test deer in example 11 of the present invention;

FIG. 11 is a photograph of feces of deer in test group 11 on day 4 after stopping medication;

FIG. 12 shows the results of the detection of Clostridium welchii RAP in the pre-treatment intestinal tract of 11 cases in accordance with the present invention;

FIG. 13 shows the results of detection of Clostridium welchii RAP in the intestine following treatment with the present invention 11;

FIG. 14 shows the results of the examination of fecal flora of young deer at different times of the treatment according to embodiment 11 of the present invention;

FIG. 15 shows the results of the fecal flora test of adult deer performed at various times in accordance with the present invention 11;

FIG. 16 is a photograph of affected sheep anus taken on the first, second and third days in sequence in accordance with example 12 of the present invention;

wherein, A in figure 1 is a drawing of the streaked result of the chromogenic medium of clostridium perfringens koma france; in FIG. 1, B is a drawing showing the streaking results of tryptone-sulfite-cycloserine agar medium.

Detailed Description

In order to clearly explain the technical features of the present invention, the following detailed description of the present invention is provided with reference to the accompanying drawings.

Example 1

The synthesis of nitrothiazole derivatives for treating intestinal infections of livestock comprises the following steps:

1. synthesis of Compound 2:

a250 mL flask was charged with 20g (0.2mol) of Compound 1: 2-aminothiazole, and 30mL of concentrated sulfuric acid and 10mL of nitric acid (40%) were added dropwise under ice bath, and stirred at 15 ℃ overnight. The pH was adjusted to 8 using 1M NaOH, the precipitate was filtered and washed with copious amounts of water. The method comprises the following steps of (1) mixing petroleum ether: ethyl acetate 5:1 as a mobile phase was passed through a silica gel column to obtain 17.1g (yield 59%) of Compound 2.¹H NMR(500MHz,d₆-DMSO)δ8.81(s,1H),8.25(s,1H)。

2. Synthesis of Compound 3:

in a 100mL flask was added 2.88g of Compound 2, 14mL of water was added, 18mL of HBr (40%) was slowly added dropwise in an ice bath, and the mixture was stirred for 30min and cooled to-15 ℃. Slowly dripping 10mLNaNO into the reaction solution₂Mixed solution of copper sulfate and copper sulfate (containing NaNO)₂3g of copper sulfate pentahydrate, 3g) and stirring and reacting for 12 hours at 0 ℃ after the dropwise addition. The temperature is increased to 25 ℃ and stirring is continued for 2 h. The stirring was stopped and the filtration was stopped, washed three times with 10mL of water and the residue was collected. Adding 3mL of ethanol, pulping at 50 ℃ for 3h, filtering again, and washing filter residue with 3mL of cold ethanol. After drying by suction filtration, the sample was dried to obtain 2.7g of a product (yield 65%).

3. Synthesis of Compound 4:

2.67g of 2-mercapto-5-amino-1, 3, 4-thiadiazole is weighed, and solid K is added₂CO₃2.72g and 15mL of ethanol, introducing nitrogen for protection, and stirring at room temperature for 30 min. 33.31 g of the compound obtained above was added thereto, and the mixture was stirred at room temperature overnight. The reaction solution was concentrated to 8mL, and the residue was collected by filtration, washed with 20mL of water, and collected 3 times. Adding petroleum ether: the ethyl acetate 6:1 solution 15mL is beaten for 2h, beaten for 2 times, filtered and washed with petroleum ether. Compound 41.92g (73% yield) was obtained.

Example 2

The synthesis of nitrothiazole derivatives for treating intestinal infections of livestock comprises the following steps:

1. the synthesis method of compound 1 and compound 2 was the same as that of compound 2 in example 1.

2. Synthesis of Compound 3:

a100 mL flask was charged with 2.88g of Compound 2, 14mL of water and 2g of NaBr, and the mixture was slowly added dropwise under ice bath to give a concentrated solutionSulfuric acid 9mL, the mixture was stirred for 30 min. Slowly dripping 10mLNaNO into the reaction solution₂Solution (containing NaNO)₂3g) The reaction was stirred at 0 ℃ for 12 h. The temperature is increased to 25 ℃ and stirring is continued for 2 h. The stirring was stopped and the filtration was stopped, washed three times with 10mL of water and the residue was collected. Adding 3mL of ethanol, pulping at 50 ℃ for 3h, filtering again, and washing filter residue with 3mL of cold ethanol. After drying by suction filtration, the sample was dried to obtain 2.40g of a product (yield 58%).

3. Synthesis of Compound 4:

weighing 2.67g of 2-mercapto-5-amino-1, 3, 4-thiadiazole, adding 0.8g of solid NaOH and 15mL of ethanol, introducing nitrogen to protect, and stirring at room temperature for 30 min. 33.31 g of compound was added thereto, and the mixture was stirred at room temperature overnight. The reaction solution was concentrated to 8mL, and the residue was collected by filtration, washed with 20mL of water, and collected 3 times. Adding petroleum ether: the ethyl acetate 6:1 solution 15mL is beaten for 2h, beaten for 2 times, filtered and washed with petroleum ether. Compound 41.76 g (68% yield) was obtained.¹HNMR(500MHz,d₆-DMSO)δ8.74(s,1H),7.94(s,2H)。

Example 3

The synthesis of nitrothiazole derivatives for treating intestinal infections of livestock comprises the following steps:

1. the synthesis of compound 2 was performed as described above for compound 2 in example 1.

2. Synthesis of Compound 3:

in a 100mL flask was added 2.88g of Compound 2, 14mL of water was added, 18mL of hydrobromic acid (40%) was slowly added dropwise under ice bath, and the mixture was stirred for 30 min. Slowly dripping 10mLNaNO into the reaction solution₂Solution (containing NaNO)₂3g) The reaction was stirred at 0 ℃ for 12 h. The temperature is increased to 25 ℃ and stirring is continued for 2 h. The stirring was stopped and the filtration was stopped, washed three times with 10mL of water and the residue was collected. Adding 3mL of ethanol, pulping at 50 ℃ for 3h, filtering again, and washing filter residue with 3mL of cold ethanol. After drying by suction filtration, the sample was dried to obtain 3.1g of a product (yield: 75%).¹H NMR(500MHz,CDCl₃)δ8.34(s,1H)。

3. Synthesis of Compound 4:

2.67g of 2-mercapto-5-amino-1, 3, 4-thiadiazole is weighed, 2.5mL of triethylamine and 35mL of ethanol are added, nitrogen is introduced to protect and the mixture is stirred for 30min at room temperature. 33.31 g of compound was added thereto, and the mixture was stirred at room temperature overnight. The reaction solution was concentrated to 15mL, and the residue was collected by filtration, washed with 20mL of water, and collected 3 times. Adding petroleum ether: the ethyl acetate 6:1 solution 15mL is beaten for 2h, beaten for 2 times, filtered and washed with petroleum ether. 41.24 g (48% yield) of compound was obtained.

Example 4

The synthesis of nitrothiazole derivatives for treating intestinal infections of livestock comprises the following steps:

1. the synthesis of compound 2 was performed as described above for compound 2 in example 1.

2. One-pot synthesis of compound 3:

a250 mL flask was charged with 20g (0.2mol) of Compound 2, and 30mL of concentrated sulfuric acid and 10mL of nitric acid (40%) were added dropwise under ice bath, and stirred at 15 ℃ overnight. The reaction solution was cooled to 0 ℃ and 30mL of NaBr saturated solution was added. 60mL (containing NaNO) is added dropwise₂18g copper sulfate pentahydrate 18g) NaNO₂And copper sulfate mixed solution, and stirring and reacting for 12 hours at 0 ℃ after the dropwise addition. The temperature is increased to 25 ℃ and stirring is continued for 2 h. The stirring was stopped and the filtration was stopped, washed three times with 10mL of water and the residue was collected. Adding 3mL of ethanol, pulping at 50 ℃ for 3h, filtering again, and washing filter residue with 3mL of cold ethanol. After drying by suction filtration, the sample was dried to obtain 17.3g of a product (yield 42%).

3. Synthesis of Compound 4:

2.67g of 2-mercapto-5-amino-1, 3, 4-thiadiazole is weighed, and solid K is added₂CO₃2.72g and 15mL of ethanol, and stirring at room temperature for 30min under the protection of nitrogen. 33.31 g of compound was added thereto, and the mixture was stirred at room temperature overnight. The reaction solution was concentrated to 8mL, and the residue was collected by filtration, washed with 20mL of water, and collected 3 times. Adding petroleum ether: the ethyl acetate 6:1 solution 15mL is beaten for 2h, beaten for 2 times, filtered and washed with petroleum ether. Compound 41.92g (73% yield) was obtained.

Example 5

The synthesis of nitrothiazole derivatives for treating intestinal infections of livestock comprises the following steps:

1. the synthesis method of compound 2 is the same as that of compound 2 in the previous example 1;

2. effect of changes in Synthesis conditions for Compound 3 on its yield

A100 mL flask was charged with 2.88g (0.2mol) of Compound 2, various amounts of hydrobromic acid (40%) were added at 0 deg.C, and various amounts of n-pentanol and 3g, 10mL of NaNO were added dropwise at different temperatures₂After 4 hours of reaction, 15L was stirred overnight. The temperature is increased to 25 ℃ and stirring is continued for 2 h. The stirring was stopped and the filtration was stopped, three times with 50mL water and the residue was collected. 10mL of ethanol was added, 80L of the mixture was recrystallized, and the residue was washed with 10mL of ethanol. After suction filtration and drying, compound 3 was obtained, the results are shown in table 1.

3. Synthesis of Compound 4 the compound 4 of the preceding examples can be synthesized by any of the methods described for Compound 4.

TABLE 1

In the synthesis of the compound 3 in the synthetic route, copper sulfate is removed and n-amyl alcohol is adopted for substitution. As can be seen from the results in Table 1, different amounts of hydrobromic acid added and different amounts of n-pentanol used at different temperatures have a significant effect on the yield. Since the yield of compound 3 most significantly affects the yield of compound 4 in the synthetic route, the synthesis was performed under the synthesis conditions of No. 3 in the present synthetic route.

The compound 4 synthesized in the embodiment 1 of the invention is selected for subsequent determination and application test.

EXAMPLE 6 determination of minimum inhibitory concentration of Compound 4

Escherichia coli, Staphylococcus aureus, Salmonella paratyphi, Streptococcus pneumoniae, methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant Staphylococcus epidermidis (MRSE), enterococcus faecalis, vancomycin-resistant enterococcus faecium (VRE), Haemophilus parasuis, and Streptococcus suis were resuscitated using blood plates. Inoculating to nutrient broth for 12-24h after recovery, diluting to 10 with MH broth⁵CFU/mL. The minimum inhibitory concentration of each type of bacteria was determined using broth dilution.

Firstly, compound 4 is dissolved in DMSO, sterile organic filter membrane is used for filtration, different volumes are taken and added into bacteria-bearing broth to prepare solutions with different concentrations, the broth is cultured for 16-24h, and the turbidity degree judgment result of the broth is observed, and the result is shown in Table 2.

TABLE 2 antibacterial Activity of different bacteria

Note:_aTSB medium was supplemented with 1% NAD chocolate agar.

Table 2 shows the minimum inhibitory concentrations of compound 4 of the present invention corresponding to each of the above-mentioned pathogenic bacteria, and the minimum inhibitory concentrations of different bacteria are determined as follows: the compound 4 has good effect on gram-positive bacteria, general effect on gram-negative bacteria and poor inhibition effect on bacillus subtilis and pseudomonas aeruginosa. In comprehensive consideration, the compound 4 can be used as an intestinal flora regulator to inhibit the dysbacteriosis of intestinal part.

Example 7 separation of Clostridium welchii and determination of minimum inhibitory concentration of Compound 4

The cecal contents of the died sika deer are taken, streaked to a chromogenic medium (figure 1A) of Clostridium komarjia perfringens France and a tryptone-sulfite-cycloserine agar medium (TSC) (figure 1B), and anaerobically cultured at 37 ℃ for 48h, and an obvious orange colony is observed and is identified as clostridium welchii (figure 1).

The agar dilution method was used to prepare a medium containing compound 4 at a concentration of 64ug/mL, 32ug/mL, 16ug/mL, 8ug/mL, 4ug/mL, 2ug/mL, 1ug/mL, 0.5ug/mL, 0.25ug/mL, or 0.125ug/mL, respectively. Coating with Clostridium welchii from different sources, and performing anaerobic culture at 37 deg.C for 48 h. Sources of Clostridium welchii include cattle, sheep, pigs, chickens, seafood and bald irises. All clostridium welchii were from clinical isolates and used after identification on selective media.

TABLE 3 antibacterial Activity of different bacteria

Note: a, the strain is separated from a deer farm infected by clostridium welchii, and drug resistance detection finds that the strain is resistant to dimetridazole.

The results in tables 2 and 3 show that the compound 4 has better effect on clostridium welchii from different sources, and shows that the compound has the effect of inhibiting clostridium welchii infection of different animals. The MIC performance of the compound 4 to clostridium welchii of large-scale animals and medium-scale animals is 4 mug/mL, and the effect is good. The performance is best for rabbits and bald irisory source clostridium welchii of 0.5 mu g/mL. Has poor effect on chicken-derived clostridium welchii, and is 16 mug/mL.

The results of the combined examples 6 and 7 show that: the compound 4 has better bacteriostatic effect on common aerobic bacteria, anaerobic bacteria and facultative anaerobic bacteria. The compound 4 has therapeutic value on intestinal infection of common livestock, and the compound 4 is used in a dosage of not more than 1mg/kg for large and medium-sized livestock and in a dosage of 3mg/kg for small animals by comprehensively considering the clinical use of the compound 4 as an intestinal flora regulator.

EXAMPLE 8 time kill curves of Compound 4 against Staphylococcus aureus and Escherichia coli

Escherichia coli ATCC25922(MIC of 8. mu.g/mL) and Staphylococcus aureus ATCC29213(MIC of 0.5. mu.g/mL) were thawed using blood plates, inoculated into LB broth for 14h, and diluted to the corresponding concentrations (10) using MH broth⁶Left and right), the corresponding compounds were added to final concentrations of 0, 1 MIC, 2 MIC, 4 MIC, 8 MIC. Samples were taken at 0h, 2h, 4h, 8h, 12h, 24h and dilution plates were used to record the number of single colonies.

The results show that: the compound 4 has obvious effect of inhibiting the growth of bacteria.

EXAMPLE 9 use of Compound 4 in the preparation of a medicament for the treatment of enteric infections in livestock and poultry

In order to examine the treatment effect of the compound 4 on mixed infection diarrhea caused by various bacteria, the compound 4 prepared in the synthetic route example 1 of the invention is selected to carry out treatment protection research on naturally-occurring white feather broilers. After the infection is determined to be the infection caused by salmonella and escherichia coli together.

The compound 4 prepared in the example 1, metronidazole and amoxicillin are respectively used for treatment at different dosages. The therapeutic effect is shown in table 4 below.

TABLE 4 in vivo bacteriostatic Activity of Compound 4, Metronidazole and Amoxicillin on diarrhea caused by multiple bacteria

The data in table 4 show that the in vivo antibacterial activity of the compound 4 of the invention on the mixed interference diarrhea caused by multiple bacteria is significantly higher than that of metronidazole and amoxicillin at a low dose, and when the dose of 0.75mg/kg is used, the number of the broiler chickens surviving after the compound 4 of the invention in the example 1 acts is 30, but no broiler chickens survive after the metronidazole and amoxicillin at the same dose are treated; and with the increase of the dose of the compound 4, the antibacterial activity of the compound is gradually increased, which shows that compared with metronidazole and amoxicillin with the same increased dose, the number of the treated and surviving meat is obviously increased, and at the dose of 1.5mg/kg, the number of the surviving meat chicken acted by the compound 4 in the embodiment 1 reaches 100, while the metronidazole and the amoxicillin with the same dose have no effect on mixed infection, and the number of the surviving meat chicken is still 0.

EXAMPLE 10 use of Compound 4 in the preparation of a medicament for the treatment of ileitis in pigs caused by Lawsonia

In order to examine the therapeutic effect of compound 4 on lawsonia-induced porcine ileitis, compound 4 of example 1 of the present invention was selected for therapeutic protection studies in naturally occurring pigs.

1 unit of 854 pigs (38 days old) are stored in a certain fattening field of Taian city, Shandong, 6 months at the beginning of 2021, and lean and weak pigs and diarrhea pigs selected from 4 units, 5 units and 6 units of the pigs are mainly used; after the swinery is transferred into 1 unit, partial pigs are only continuously diluted for several days, the proportion of the diluted pigs accounts for about 20 percent of the number of the pigs in the stockings, and the excrement is mostly in a porridge shape, a water sample, a gray color or a cement sample and a small amount of green/yellow green samples; feces are stained on the buttocks of seriously ill pigs, the anus is red and swollen, the hair of the diarrhea pigs is thick and messy, and the body surfaces of partial pigs are pale; the big group of pigs have better spirit and no other abnormalities; large groups of pigs drink water and dose amoxicillin, colistin, doxycycline, florfenicol powder and other medicaments, the effect is not ideal, and only new diarrhea pigs are added every day; the individual pig is alternately treated by taking drugs such as mequindox, enrofloxacin, florfenicol, lincomycin injection and the like, and has certain effect on treating part of the diarrhea pigs. The phenomenon of diarrhea relapse only after a few healed and recovered pigs are treated for several days/week. And then sampling and inspecting the excrement, wherein the result of detection shows that PPE and PED pathogens are positive in the excrement sample, and the ileitis of the Lawsonia bacteria pig is diagnosed.

The treatment mode comprises the following steps: 30 pigs with severe diarrhea are selected from a swinery, are intensively and uniformly placed in a test bar, anus swabs are randomly collected from 3 pigs with severe diarrhea, relevant marks are made, and samples are sent for inspection before medication. Feeding test pigs after fasting for half a day by uniformly mixing about 8 kg of feed and test medicaments by adopting an artificial stepwise dilution method; in order to ensure that all pigs have equal chances to eat, 3 additional feeding trays are additionally arranged in the fence and matched with the feeding grooves in the fence for use. To ensure the objectivity of the test, no drug was added to the water during the test. Compound 4 was administered daily to each pig as 60mg for 3 consecutive days and swabs of the anus of sick pigs were collected daily for censorship and the results are shown in table 5.

TABLE 5 follow-up effects of drug treatment on diarrhea pigs

With reference to FIGS. 4-7, the pre-treatment feces of the herds of the test group of FIG. 4; feces on day 2 of administration; feces after the 3 rd day of administration; day 2 after drug withdrawal. Most of the diarrhea pigs are recovered after the intervention of the drug, the feces are formed or normal, and the red and swollen phenomenon of the anus of part of the pigs is eliminated. The data collected from table 2 was analyzed for the significant effect of the drug use; and finally, the diarrhea phenomenon still exists in 3 incured pigs, the intestinal mucosa inflammation is serious, and the intestinal tract repair can take more time than other pigs, so that daily clinical observation and tracking are further carried out to find that all pig feces are normal on the 9 th day after the medicine is stopped.

Before treatment, 30 selected pigs have serious diarrhea phenomenon, thick and messy hair, and most pigs have 'back exposure', and the regular health state of the treated pigs is obvious by combining the picture 8. The result shows that the compound 4 has the characteristics of quick response and quick recovery compared with the existing florfenicol, enrofloxacin and other common porcine ileitis treatment medicines.

EXAMPLE 11 use of Compound 4 in the preparation of a medicament for the treatment of infection caused by C.welchii resistant deer

The experiment is carried out in a certain farm in east Shandong. 100 breeding deer are introduced into the breeding base, bacterial infection of breeding deer groups is caused by feed pollution in the breeding process, the bacterial infection is confirmed to be clostridium welchii infection through veterinarian identification, deer bloody stool is found after the infection, and the deer is finally dehydrated and died, and sporadic deer still die after the dimetridazole is used for a long time.

Bloody stool appears in early deer and rumination stops in later period. Blepharorrhage, tearing. Late death, debridement, peritoneal hemorrhage, hemorrhage and necrosis of intestinal tract, and massive necrosis and edema of kidney. PCR amplification is carried out by using the clostridium welchii specific primer, an obvious amplification strip is obtained, and the clostridium welchii alpha toxin is detected.

Before the experiment, 21 live deer were kept in the deer farm. In the experiment, 4 deer are randomly selected as a control group, wherein 3 deer die after the dimetridazole is stopped for 3 days, and 1 deer dies after 5 days. Selecting 16 deer as experimental group, encapsulating the medicine according to average weight of deer, using green grass to wrap the capsule for food calling, and administering 4140mg compound to each deer every day for 3 days, and observing for 11 days. The results are given in table 6 below:

TABLE 6 statistical Table for treatment of C.welchii infected deer herd

As can be seen from Table 6, when the compound 4 of example 1 of the present invention is used in low dose, the phenomena of diarrhea in deer are completely improved after 2 days of medication, no death of deer occurs in the period, and the deer is completely cured after 3 days of medication, so the effect is obvious. Has a targeted and obvious curative effect on infection diseases caused by drug-resistant clostridium welchii of deer.

As can be seen from the accompanying figures 9-11, the feces of infected deer caused by drug-resistant Clostridium welchii are taken 2 days before administration and 4 days after withdrawal. Stopping taking the medicine for 3-4 days, realizing complete cure without relapse, and increasing the average food intake of deer herds by 30%. FIGS. 12-13 show the detection of the RAP of Clostridium welchii in the pre-and post-treatment intestinal tracts, which indicates infection with Clostridium welchii before treatment and does not detect Clostridium welchii after treatment.

Intestinal flora detection was performed on fecal samples collected from the diseased deer on the first day of administration (Z1), the third day of administration (Z2), the first day of withdrawal (Z3), and the 20 days of withdrawal (Z4), fig. 14. The results show that the number of bacteria of the genus clostridium is effectively reduced during the treatment period and the number of other bacterial species is increased.

When compound 4 was fed to non-diseased young deer and fecal samples were analyzed for flora (fig. 15) on the first day of drug administration (Z5), the first day of withdrawal (Z6) and the 20 th day of withdrawal (Z7), compound 4 showed less effect on bacteroides and greater effect on firmicutes and proteus bacteria.

EXAMPLE 12 use of Compound 4 in the treatment of ovine dysentery caused by Clostridium welchii

In order to investigate the scrapie caused by clostridium welchii by compound 4, compound 4 of example 1 of the present invention was selected for therapeutic protection studies in naturally-occurring sheep.

The experiment is carried out by selecting the sheep infected with the clostridium welchii in a certain farm in Shandong tobacco terrace city, the disease remission of the sheep group is still not seen after the sheep group is used for 10 days before the experiment, and the sheep dysentery caused by the clostridium welchii infection is diagnosed by a veterinarian.

The treatment mode comprises the following steps: selecting 4 diseased sheep, fasting the test sheep for half a day, and uniformly mixing about 2 kg of feed and the test medicine by adopting an artificial stepwise dilution method and feeding; to ensure the objectivity of the test, no drug was added to the water during the test. Using the drug for 3 consecutive days, 60mg of compound 4 per day per sheep, and observing the fecal status of the flocks of sheep for three days as shown in fig. 16; the three panels in figure 16 show the condition of the anal diarrhea in sheep on the first day of administration, the second day of administration and the third day of administration in sequence.

The experimental results are as follows: after 3 days, the sheep stops diarrhea, the anus is clean, the excrement is granular, and the diarrhea is not repeated after continuously observing for 14 days.

EXAMPLE 13 use of Compound 4 in the treatment of Swine septicemia caused by Streptococcus suis

In order to examine the swine septicemia caused by streptococcus suis by compound 4, the synthetic compound 4 of the present invention was selected for therapeutic protection studies in naturally-occurring swine.

The treatment mode comprises the following steps: selecting 10 sick pigs, dividing the sick pigs into two groups, fasting the test pigs for half a day, and uniformly mixing about 1 kg of feed and test medicines by adopting an artificial stepwise dilution method and feeding the test pigs; to ensure the objectivity of the test, no drug was added to the water during the test. Compound 4 was administered to each pig at 20mg for 3 consecutive days and pig survival was observed for three days as shown in table 7.

TABLE 7

After 3 days of drug administration, observation was continued for 10 days, and two pigs finally survived in the treatment group, while all the control groups died. The test result shows that the compound 4 has certain treatment effect on the streptococcus suis infection.

The above-described embodiments should not be construed as limiting the scope of the invention, and any alternative modifications or alterations to the embodiments of the present invention will be apparent to those skilled in the art.

The present invention is not described in detail, but is known to those skilled in the art.