Novel halofuginone derivative, preparation method thereof and application of halofuginone derivative in anticoccidial drugs

Abstract

The invention discloses a novel halofuginone derivative, a preparation method thereof and application of the halofuginone derivative in anticoccidial drugs, and belongs to the field of veterinary drugs. The compound provided by the invention is based on a halofuginone compound as a lead compound, a halofuginone derivative with a novel structure is designed and synthesized by utilizing a computer-aided drug design technology and a skeleton migration principle, a one-pot synthesis method is adopted, the synthesis process is simple, and the used raw materials are simple, easy to obtain, economical and practical. The anticoccidial index (ACI) of the anticoccidial drug prepared from the derivative provided by the invention can reach 180, and the anticoccidial drug has a good anticoccidial effect.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of veterinary medicine, in particular to a new halofuginone derivative, a preparation method thereof and application thereof in anticoccidial drugs. BACKGROUND

[0002] Coccidiosis is a global distribution of intestinal parasitic disease that is extremely harmful, and the morbidity and mortality are extremely high. Coccidiosis is a common parasitic disease in farmed animals, especially poultry, and is extremely difficult to prevent and control. The application of traditional Chinese medicine in the prevention and treatment of animal coccidiosis is a unique research in China. Compared with chemically synthesized drugs, traditional Chinese medicine has less toxic and side effects on the chicken body, and long-term use will not cause the quality of chicken related products to decline or cause drug residues. However, the current application of traditional Chinese medicine in the treatment of coccidia is not satisfactory, mainly in the simple processing method, low technical content; non-medicinal parts are used for medicine, and the effect is difficult to guarantee; unstable drug quality leads to unstable product quality; traditional Chinese medicine westernization technology can fundamentally solve various problems faced by traditional Chinese medicine in the breeding industry. Halofuginone is a typical example of traditional Chinese medicine westernization, but there are still many problems to be solved in the use of halofuginone raw materials and as anticoccidial drugs. Mainly in: 1. All the synthesis processes of the disclosed halofuginone raw materials are complex, low in yield, high in cost, and difficult to treat three wastes, which makes it difficult to scale up production, greatly limiting its wide application. 2. Clinical trial data show that the safe dosage range of halofuginone is narrow, and it can affect the weight gain rate of healthy chicks at the treatment concentration; when the concentration exceeds 1.6 mg / kg of feed, it affects the palatability, reduces the intake, and even refuses to eat; when the concentration exceeds 3 mg / kg, it will inhibit the growth rate of waterfowl (geese, ducks), and even cause animal poisoning and death. Therefore, there is still a lot of room for improvement and upgrading of halofuginone as an anticoccidial drug. The development of new halofuginone derivative anticoccidial drugs is of great significance for the prevention and treatment of coccidiosis in China's livestock and poultry, and solving the problems of drug resistance and drug residues.

[0003] Therefore, the present application is proposed to solve the above technical problems. SUMMARY

[0004] In order to solve the technical problems in the background art, the present application proposes a new halofuginone derivative, a preparation method thereof and application thereof in anticoccidial drugs, which is characterized by designing a new anticoccidial drug molecule, preparing a halofuginone derivative with a novel skeleton, and evaluating and screening its anticoccidial activity.

[0005] In order to achieve the above purpose, the present application adopts the following technical scheme: A new halofuginone derivative, wherein the structure of the new halofuginone derivative is: .

[0006] Preferably, the synthetic route of the new halofuginone derivative is as follows: .

[0007] The application further provides a preparation method of the new halofuginone derivative, comprising the following steps: S1, using one-pot multi-step synthesis method, taking a dry reaction bottle, adding 48.5 g of 3-hydroxy-4-methylpyrrole (0.5 mol), 50.0 g of benzyl chloride (66.1 g, 0.53 mol) in sequence, after the first stirring reaction, adding allyl bromide (60.0 g, 0.5 mol), sodium borohydride (50.0 g, 1.3 mol), passing nitrogen atmosphere for 10-20 min, distilling off the excess raw material, then adding 51.0 g of benzyl chloroformate (0.3 mol), boron trifluoride ether (50 mL), stirring at room temperature for the second time, and finally adding 178.0 g of N-bromosuccinimide (1.0 mol) for the third stirring, detecting that the raw material has disappeared by TLC, stopping stirring, transferring to another dry gourd-shaped bottle, adding silica gel, concentrating under reduced pressure, and purifying by column chromatography to obtain the right pyrrole ring structure (b); S2, taking another dry reaction bottle, adding 100.5 g of 5-chloro-2-nitrobenzoic acid (0.5 mol), then adding 10 g of iron and 15 g of hydrochloric acid (36%, 0.14 mol), stirring in a warm water bath for the first time, distilling off the excess raw material, adding 27.0 g of formamide, stirring in a warm water bath for the second time, adding silica gel after the reaction is completed by TLC detection, concentrating to dryness, and purifying by column chromatography to obtain the left quinazoline derivative (a); S3, adding the above obtained compound (a) and compound (b) into a reaction kettle, adding 56.0 g of potassium tert-butoxide and 42.0 g of potassium carbonate, stirring at room temperature, detecting that the raw material has completely disappeared by TLC, stopping stirring, transferring to another dry gourd-shaped bottle, adding silica gel, concentrating under reduced pressure, and purifying by column chromatography to obtain the halofuginone derivative.

[0008] Preferably, in the step S1, the first stirring time is 1-1.5 hours, and the second and third stirring times are 30-35 min.

[0009] Preferably, in the step S2, the first and second stirring times are both 30-35 min, and the warm water bath temperature is 40-60°C.

[0010] Preferably, in the step S3, the sufficient stirring time is 2-4 h.

[0011] Preferably, the silica gel mesh size is 100-200 mesh.

[0012] The application also provides the application of the new halofuginone derivative in treating chicken coccidiosis.

[0013] Preferably, the new halofuginone derivative is added into commercial feed for use, and the adding amount is 3-5 mg per kg of feed.

[0014] The application has the following advantages: The application modifies and reconstructs the halofuginone nucleus according to the molecular level pharmacological mechanism of halofuginone, synthesizes a new halofuginone derivative, and proves through animal tests that the anti-coccidial index of the new halofuginone derivative can reach 180, and the new halofuginone derivative has good anti-coccidial effect. BRIEF DESCRIPTION OF DRAWINGS

[0015] Figure 1 is a diagram of the molecular level pharmacological mechanism of the halofuginone derivative Figure 2 is a synthesis reverse analysis diagram of halofuginone DETAILED DESCRIPTION

[0016] The technical solutions in the embodiments of the application will be described clearly and completely below. Obviously, the described embodiments are only a part of the embodiments of the application, rather than all the embodiments. Based on the embodiments in the application, all other embodiments obtained by those skilled in the art without creative labor fall within the protection scope of the application.

[0017] Preparation Example 1 Preparation of the new halofuginone derivative A dry reaction flask was taken and 48.5 g of 3-hydroxy-4-methylpyrrole (0.5 mol) was added followed by 50.0 g of benzyl chloride (66.1 g, 0.53 mol). The reaction was stirred for 1-1.5 h, after which 60.0 g of allyl bromide (0.5 mol) and sodium borohydride (50.0 g, 1.3 mol) were added. Nitrogen gas was passed for 10-20 min, after which the excess starting material was distilled off. Then, 351.0 g of benzyl chloroformate (0.3 mol) and boron trifluoride etherate (50 mL) were added and the reaction was stirred at room temperature for 30-35 min. Finally, 178.0 g of N-bromosuccinimide (1.0 mol) was added and stirred for the third time. TLC was taken to confirm the disappearance of the starting material. The reaction was stopped and transferred to another dry flask. Silica gel (100-200 mesh) was added and concentrated under reduced pressure. The product was purified by column chromatography to obtain the pyrrole ring structure on the right (b). Another dry reaction flask was taken and 100.5 g of 5-chloro-2-nitrobenzoic acid (0.5 mol) was added followed by 10 g of iron and 15 g of hydrochloric acid (36%, 0.14 mol). The reaction was stirred in a water bath at 40-60 °C for 30-35 min, after which the excess starting material was distilled off. Then, 27.0 g of formamide was added and the reaction was stirred in a water bath at 40-60 °C for 30-35 min. TLC was taken to confirm the completion of the reaction. Silica gel (100-200 mesh) was added and concentrated under reduced pressure. The product was purified by column chromatography to obtain the quinazoline derivative on the left (a). Another dry reaction flask was taken and the compounds (a) and (b) obtained above were added to the reaction flask. Then, 56.0 g of potassium tert-butoxide and 42.0 g of potassium carbonate were added and stirred at room temperature for 2-4 h. TLC was taken to confirm the disappearance of the starting material. The reaction was stopped and transferred to another dry flask. Silica gel (100-200 mesh) was added and concentrated under reduced pressure. The product was purified by column chromatography to obtain the halofuginone derivative.

[0018] Example 1 The halofuginone derivative obtained in Preparation Example 1 was pulverized and passed through a 100 mesh sieve. Three milligrams of the same was added to one kilogram of millet and stirred to mix uniformly.

[0019] Example 2 The halofuginone derivative obtained in Preparation Example 1 was pulverized and passed through a 100 mesh sieve. Four milligrams of the same was added to one kilogram of millet and stirred to mix uniformly.

[0020] Example 3 The halofuginone derivative obtained in Preparation Example 1 was pulverized and passed through a 100 mesh sieve. Five milligrams of the same was added to one kilogram of millet and stirred to mix uniformly.

[0021] Comparative Example 1 Halofuginone was pulverized and passed through a 100 mesh sieve. Three milligrams of the same was added to one kilogram of millet and stirred to mix uniformly.

[0022] Comparative Example 2 Crude pyrrhotone was ground to pass through a 100-mesh sieve, and 4 mg was added to 1 kg of millet food and mixed evenly.

[0023] Comparative Example 3 Crude pyrrhotone was ground to pass through a 100-mesh sieve, and 5 mg was added to 1 kg of millet food and mixed evenly.

[0024] Experimental Example 1 The efficacy evaluation test was performed on the groups of Example 1-3, Comparative Example 1-3, and the infection without drug administration group, and the blank control group (not infected and not administered drugs). The newly hatched yellow-feathered chicken chicks were selected as the test chicks, and were raised in a coccidia-free environment. The chicks were fed with a mixture of feed that did not contain any coccidiostat, and drank clean tap water, and were raised until 12 days old for use. The oocysts were obtained from the ceca of blood-carrying chicken feces and dead chicken ceca from a chicken farm with coccidiosis, and were isolated, purified, and then used to reproduce and rejuvenate in live chickens to obtain 1.0 x 10 5 spores per chicken. The test chickens were weighed and recorded, and were randomly divided into groups, with 20 chickens in each group, and the total weight of each group was adjusted to be substantially consistent. Then, the 12-day-old chickens in the test drug administration groups were fed with the basic feed and the test compound, respectively, and 2 days later, the chickens were artificially infected with coccidia spores, with 1.0 x 10 5 spores per chicken. On the 5th day after infection, 3 chickens were randomly killed from each test group for necropsy to observe the cecal lesions, and the lesions were scored according to the 0-4 lesion scoring system and the lesion value was calculated; on the 7th day after infection, the test ended, and 5 chickens were randomly selected from each group for necropsy, and the oocyst value was calculated according to the number of oocysts in each pair of ceca. Each test chicken was weighed again, and finally the remaining chickens in each group were anesthetized for necropsy, and the relative weight gain, survival rate, lesion value, and oocyst value of each test group were recorded, and the ACI value was calculated.

[0025] Survival rate (%) = number of surviving chicks in each group ÷ total number of chicks in each group x 100%; Relative weight gain rate (%) = weight gain rate of the drug administration group or the infection without drug administration group ÷ weight gain rate of the infection without drug administration group x 100%.

[0026] The lesion score was scored according to the 5-point scoring standard of Johnson et al. according to the severity of the intestinal lesions after necropsy.

[0027] 0 points: no gross lesions; 1 point: very few scattered point hemorrhages in the intestinal wall, no thickening of the intestinal wall, and normal contents; 2 points: a small amount of blood attached to the intestinal contents, thickening of the intestinal wall, and many hemorrhagic lesions; 3 points: multiple blood or intestinal nucleus (blood clots or gray cheese-like banana-shaped mass) in the intestine, intestinal wall thickening and obvious deformation and atrophy; 4 points: the intestine is significantly atrophied, the intestinal wall is extremely thickened, and the intestine contains blood clots or intestinal nucleus. When judging, sometimes the lesions of the two sides of the cecum are not consistent, so the side with the more serious lesion should be used as the standard for judgment.

[0028] Lesion value = average lesion score of each group × 10.

[0029] Oocyst value is converted from the number of oocysts in the cecal contents per gram of feces (OPG).

[0030] ACI = (relative weight gain rate + survival rate) - (lesion value + oocyst value), and the standard for determining the efficacy according to ACI is as follows: ACI < 120, not suitable as an anticoccidial drug; ACI < 160, low anticoccidial efficacy; 160 < ACI < 180, moderate anticoccidial efficacy; ACI > 180, strong anticoccidial efficacy.

[0031] The grouping and administration are shown in Table 1 as follows: Table 1 Group Dosing situation Dosing cycle Example 1 3 mg / kg D12-21 Example 2 4 mg / kg D12-21 Example 3 5 mg / kg D12-21 Comparative Example 1 3 mg / kg D12-21 Comparative Example 2 4 mg / kg D12-21 Comparative Example 3 5 mg / kg D12-21 Infection without administration group / / Blank control group / / The relative weight gain, survival rate, lesion value and oocyst value, and ACI value are shown in Table 2 as follows: Table 2 Group Relative weight gain rate (%) Survival rate (%) Oocyst value Lesion value ACI Blank control group 100 100 0 0 / Infection without administration group 57.2 80 40 47 / Example 1 90.5 100 5 1.5 184 Example 2 90.7 100 5 1.1 184.6 Example 3 91.6 100 5 1.2 185.4 Comparative Example 1 89.3 100 5 1.4 182.9 Comparative Example 2 84.9 95 5 1.9 173 Comparative Example 3 82.5 95 5 2.0 170.5 As can be seen from Table 2, when the addition amount of the oridonin derivative in the chicken feed is 3-5 mg per kg of feed, the ACI index is greater than 180, and the oridonin derivative has excellent anticoccidial effect; with the increase of the drug dose, no damage is caused to the chicken intestinal tract, and the growth rate of the chicken is not affected, and the chicken's food intake is not inhibited, indicating that the oridonin derivative has good anticoccidial effect.

[0032] The above is only the preferred specific embodiment of the present application, but the protection scope of the present application is not limited thereto, and any person skilled in the art can make equivalent replacement or change according to the technical solution and the inventive concept of the present application within the technical range disclosed by the present application, which should be covered within the protection scope of the present application.

Claims

1. A novel halofentrine derivative, characterized in that, The structural formula of the novel halofuginone derivative is: 。 2. The novel halofuginone derivative according to claim 1, characterized by the fact that, The synthetic route of the novel halofuginone derivative is: 。 3. A method for preparing a novel halofuginone derivative according to claim 1 or 2, characterized by, The method comprises the following steps: S1, using one-pot multi-step synthesis method, taking a dry reaction bottle, adding 48.5g of 3-hydroxy-4-methylpyrrole (0.5mol), 50.0g of benzyl chloride (66.1g, 0.53mol) in sequence, after the first stirring reaction, adding allyl bromide (60.0g, 0.5mol), sodium borohydride (50.0g, 1.3mol), passing nitrogen atmosphere for 10-20min, distilling off the excess raw material, then adding 51.0g of benzyl chloroformate (0.3mol), boron trifluoride ether (50mL), stirring at room temperature for the second time, finally adding 178.0g of N-bromosuccinimide (1.0mol) for the third time, detecting that the raw material has disappeared by TLC, stopping stirring, transferring to another dry gourd-shaped bottle, adding silica gel, concentrating under reduced pressure, and purifying by column chromatography to obtain the right pyrrole ring structure (b); S2, taking another dry reaction bottle, adding 100.5g of 5-chloro-2-nitrobenzoic acid (0.5mol), then adding 10g of iron and 15g of hydrochloric acid (36%, 0.14mol), stirring in a warm water bath for the first time, distilling off the excess raw material, adding 27.0g of formamide, stirring in a warm water bath for the second time, adding silica gel after the reaction is completed by TLC detection, concentrating to dryness, and purifying by column chromatography to obtain the left quinazoline derivative (a); S3, adding the obtained compound (a) and compound (b) into a reaction kettle, adding 56.0g of potassium tert-butoxide and 42.0g of potassium carbonate, stirring at room temperature until the raw material completely disappears by TLC detection, stopping stirring, transferring to another dry gourd-shaped bottle, adding silica gel, concentrating under reduced pressure, and purifying by column chromatography to obtain the halofuginone derivative.

4. The method for preparing a novel chrysanthemumone derivative according to claim 3, characterized in that, In the step S1, the first stirring time is 1-1.5h, and the second and third stirring times are 30-35min.

5. The method for preparing a novel chrysanthemumone derivative according to claim 3, characterized in that, In the step S2, the first and second stirring times are both 30-35min, and the temperature of the warm water bath is 40-60℃.

6. A process for the preparation of a novel halofuginone derivative according to claim 3, characterized by, In the step S3, the sufficient stirring time is 2-4h.

7. The method for preparing a novel chrysanthemumone derivative according to claim 3, characterized in that, The mesh number of the silica gel is 100-200.

8. A novel halofuginone derivative according to claim 1 or 2 for use in the treatment of chicken coccidiosis.

9. Use according to claim 8, characterized in that, The novel halofuginone derivative is added to commercial feed for use, and the addition amount is 3-5mg per kilogram of feed.

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