A chlortetracycline soluble powder and a method for preparing the same

Abstract

The application relates to the technical field of veterinary drugs, and particularly discloses aureomycin soluble powder and a preparation method thereof. The aureomycin soluble powder is composed of the following components in mass percentage: aureomycin phosphate salt 20-55%, chelating agent 0.5-2.5%, penetration promoter 0.5-2.5%, and organic acid carrier in residual amount; the aureomycin phosphate salt is mainly obtained by chemically modifying hydrochloric acid aureomycin with chlorophosphite di-tert-butyl ester, and then reacting with an alkali to form a salt. According to the application, the aureomycin is chemically modified into a phosphate salt, and the chelating agent and the penetration promoter are used in cooperation, so that the key problems of poor water solubility and low absorption rate are fundamentally solved, the solubility is improved by an order of magnitude, the oral bioavailability is significantly improved, and meanwhile, the complicated super-micro pulverization treatment required by a traditional process is avoided.

Description

Technical Field

[0001] This application relates to the field of veterinary drug technology, and more specifically, to a chlortetracycline soluble powder and its preparation method. Background Technology

[0002] Tetracycline antibiotics for veterinary use are commonly used broad-spectrum antibacterial drugs in livestock farming, showing good efficacy against a variety of Gram-positive and Gram-negative bacteria, as well as rickettsiae, mycoplasma, and spirochetes. Among them, chlortetracycline hydrochloride, as one of the representative drugs in this class, primarily inhibits bacterial protein synthesis by specifically binding to the 30S subunit of bacterial ribosomes. Therefore, it is widely used in the prevention and treatment of bacterial diseases and the promotion of growth in poultry such as chickens and pigs.

[0003] Currently, large-scale farms mostly use drinking water to administer medication, dissolving the drug in water before administration via a dosing device. Therefore, the initial solubility and stability of the drug are crucial. Chlortetracycline hydrochloride is only slightly soluble in water, with a solubility of approximately 0.4 g / 100 ml at room temperature. In water with high hardness, it is more prone to precipitation at high concentrations. Once precipitated, even further dilution is difficult to completely dissolve, leading not only to a decrease in effective concentration and reduced efficacy but also potential clogging of the dosing device, causing inconvenience to farm management. Therefore, improving the initial solubility and stability of chlortetracycline hydrochloride is of significant practical importance.

[0004] According to the Biopharmaceutics Classification System (BCS), chlortetracycline hydrochloride belongs to Class IV drugs, meaning it has low solubility and low permeability. To address the low solubility issue, methods such as reducing particle size, salt formation, and preparing solid dispersions or inclusion complexes can be used to improve the dissolution rate. For the low permeability issue, permeability enhancers (such as sodium salicylate and sodium deoxycholate) are often added to improve the drug's membrane permeability. However, most existing methods either have limited effectiveness or are too costly, making them unsuitable for large-scale aquaculture.

[0005] As a basic veterinary chemical, chlortetracycline hydrochloride often plays a synergistic role in combination drug therapy, leading to a continuously growing market demand. However, its low solubility severely restricts the development and practical application of related formulations. Currently, the solubility of commonly available chlortetracycline hydrochloride powder, even with the addition of excipients such as glucose, is only maintained at 4-6g per liter of water. A Chinese patent application, publication number CN110507616A, discloses a chlortetracycline hydrochloride soluble powder and its preparation method. By adding excipients such as urea, anhydrous magnesium sulfate, and citric acid, the solubility is increased to 6-10g per liter of water, and the dissolution rate is also improved. The clinical dosage is: 1-2g per liter of drinking water for chickens; 1-1.5g for weaned piglets, for 7 consecutive days. However, in large-scale farms using 1:50 dosing systems, the actual requirement is a solubility of 50-100g per liter of water in the initial dissolution stage. Currently, no product can meet this high concentration dissolution requirement.

[0006] Most conventional soluble powders are made by simply mixing chlortetracycline hydrochloride with anhydrous sugar, etc. They have poor solubility, long dissolution time, and low bioavailability, making them unsuitable for the actual conditions of high-concentration administration and complex water quality in large-scale aquaculture farms, which greatly limits their clinical application. Summary of the Invention

[0007] To address the aforementioned technical problems, this application provides a chlortetracycline soluble powder and its preparation method. By chemically modifying chlortetracycline hydrochloride with di-tert-butyl chlorite and then forming a salt, its solubility and absorbability in water are greatly improved, and the prepared soluble powder has extremely high stability.

[0008] In a first aspect, this application provides a chlortetracycline soluble powder, which adopts the following technical solution:

[0009] A chlortetracycline soluble powder is composed of the following components in weight percentage: chlortetracycline phosphate salt 20-55%, chelating agent 0.5-2.5%, penetration enhancer 0.5-2.5%, and organic acid carrier balance;

[0010] The chlortetracycline phosphate salt is prepared by a method comprising the following steps:

[0011] (1) Reaction of chlortetracycline hydrochloride with an organic base yields free chlortetracycline base;

[0012] (2) The free base of chlortetracycline reacts with di-tert-butyl chlorite and is oxidized to generate di-tert-butyl chlortetracycline phosphate;

[0013] (3) Remove the tert-butyl protecting agent from chlortetracycline di-tert-butyl ester to obtain chlortetracycline phosphate;

[0014] (4) React chlortetracycline phosphate with a base to obtain the chlortetracycline phosphate salt.

[0015] More preferably, in step (1), the organic base is selected from at least one of triethylamine, N,N-diisopropylethylamine and pyridine.

[0016] More preferably, in step (2), the oxidant used for oxidation is selected from at least one of hydrogen peroxide, m-chloroperoxybenzoic acid and tert-butylhydrogen peroxide.

[0017] More preferably, in step (3), an acid hydrolysate is used to remove the tert-butyl protecting group, wherein the acid hydrolysate is selected from trifluoroacetic acid, hydrochloric acid, formic acid and trimethylsilane bromide.

[0018] More preferably, in step (4), the base can be an organic base or an inorganic base to suit different formulation requirements, such as solubility, stability, irritation, etc. Specifically, the organic base can be meglumine, arginine or lysine, and the inorganic base can be sodium hydroxide, potassium hydroxide, sodium carbonate or potassium carbonate.

[0019] More preferably, the chelating agent is selected from at least one of disodium ethylenediaminetetraacetate, sodium tripolyphosphate, and sodium hexametaphosphate; preferably, the chelating agent is disodium ethylenediaminetetraacetate.

[0020] The phenolic hydroxyl groups, enol structures, and dimethylamino groups on the chlortetracycline phosphate molecule, as well as the newly introduced phosphate group, readily form complexes with polyvalent cations. Therefore, its bioavailability decreases when the calcium content in feed increases. Chelating agents preferentially bind to polyvalent cations (such as calcium and magnesium) in feed and drinking water, thereby preventing these ions from interacting with chlortetracycline phosphate. Consequently, the amount of absorbable tetracyclines in the gastrointestinal tract increases, improving the bioavailability of chlortetracycline phosphate and consequently increasing the concentration of chlortetracycline phosphate in serum.

[0021] More preferably, the penetration enhancer is selected from at least one of sodium salicylate, sodium deoxycholate, and sodium dodecyl sulfate; preferably, the penetration enhancer is sodium dodecyl sulfate.

[0022] More preferably, the organic acid carrier is selected from at least one of citric acid, tartaric acid, and malic acid; preferably, the organic acid carrier is citric acid.

[0023] Currently, the carriers for soluble powders are generally glucose, sucrose, lactose, or soluble starch. If added to drinking water and not promptly cleaned and disinfected, it can lead to bacterial growth on the waterline. Organic acids not only increase the solubility and stability of chlortetracycline phosphate salts, improving blood drug concentration and bioavailability, but also regulate the acidity of the drinking water, inhibiting the growth of harmful bacteria such as E. coli, effectively preventing the formation of biofilms on the waterline, and reducing the incidence of diarrhea in piglets.

[0024] Secondly, this application provides a method for preparing chlortetracycline soluble powder, using the following technical solution:

[0025] A method for preparing a chlortetracycline soluble powder includes the following steps:

[0026] Chlortetracycline phosphate salt and organic acid carrier are mixed evenly to obtain a premix;

[0027] Add the chelating agent and penetration enhancer to the premix, mix well, and obtain chlortetracycline soluble powder.

[0028] The method for preparing chlortetracycline soluble powder provided in this application can be obtained through a simple mixing process, with low requirements for production equipment, low production cost, and the ability to be mass-produced industrially.

[0029] In summary, this application has the following beneficial effects:

[0030] (1) This application converts chlortetracycline into phosphate salt and introduces a strongly hydrophilic and highly ionized phosphate group into its molecule, so that the solubility of chlortetracycline phosphate salt in water is increased by more than ten times compared with chlortetracycline hydrochloride after it is made into soluble powder, which fundamentally solves the problem of the dissolution rate limit of active ingredients.

[0031] (2) High solubility ensures that chlortetracycline can quickly reach and maintain a high concentration in the gastrointestinal tract, thus creating a prerequisite for efficient absorption. In addition, as a prodrug, the phosphate salt has a better absorption mechanism than chlortetracycline hydrochloride. Combined with the penetration enhancer in the soluble powder, the high absorption effect is greatly improved.

[0032] (3) Due to the high solubility of chlortetracycline phosphate salts, the formulation process does not require the use of complex and energy-intensive ultra-high-efficiency pulverization processes, which simplifies the production process, reduces costs, and improves product stability. Detailed Implementation

[0033] The following specific examples illustrate the implementation of this application. Those skilled in the art can easily understand other advantages and effects of this application from the content disclosed in this specification. This application can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of this application.

[0034] Furthermore, it should be understood that the one or more method steps mentioned in this application do not preclude the existence of other method steps before or after the combined steps, or the insertion of other method steps between these explicitly mentioned steps, unless otherwise stated. Moreover, unless otherwise stated, the numbering of each method step is merely a convenient tool for identifying each method step, and is not intended to limit the order of the method steps or limit the scope of implementation of this application. Changes or adjustments to their relative relationships, without substantially altering the technical content, shall also be considered as within the scope of implementation of this application.

[0035] Unless otherwise specified, the experimental conditions used in the examples are generally in accordance with conventional conditions in the art or the conditions recommended by the reagent company. Unless otherwise specified, the materials and reagents used in the examples can be purchased commercially.

[0036] Example 1 Preparation of sodium chlortetracycline phosphate

[0037] (1) Place 10g of chlortetracycline hydrochloride (Pucheng Zhengda Biochemical Co., Ltd.) in a dry reaction flask, add 100mL of anhydrous pyridine, and stir under an ice-water bath. Slowly add 3.36mL of triethylamine, and stir for 1 hour at the same temperature to obtain free chlortetracycline base.

[0038] (2) Slowly add 5.37 mL of di-tert-butyl chlorite to the above-mentioned chlortetracycline free base. After the addition is complete, remove the ice bath and stir the reaction at 20-25°C for 4 hours. Place the reaction system back in an ice-water bath and slowly add 4.4 mL of 70% tert-butyl hydrogen peroxide aqueous solution. After the addition is complete, continue stirring the reaction for 1 hour.

[0039] (3) Pour the reaction solution into 200 mL of ice water and extract three times with dichloromethane. Combine the organic phases and wash them successively with 100 mL of 5% citric acid aqueous solution, 100 mL of saturated sodium bicarbonate aqueous solution, and 100 mL of saturated saline solution. Dry the organic phase with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain crude di-tert-butyl chlortetracycline phosphate.

[0040] (4) Dissolve the crude product in 100 mL of dichloromethane, cool in an ice-water bath, and slowly add 12.8 mL of trimethylsilane bromide while stirring. After the addition is complete, remove the ice bath and stir the reaction at 20-25 °C for 3 hours.

[0041] (5) Slowly pour the above reaction solution into 200 mL of ice water. Under ice bath and stirring, adjust the pH of the aqueous phase to 3-4 with 2 mol / L sodium hydroxide aqueous solution. Extract the aqueous phase with a mixed solvent (dichloromethane:isopropanol = 4:1, v / v, total 150 mL × 3). Combine the organic phases, dry with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain chlortetracycline phosphate.

[0042] (6) The obtained chlortetracycline phosphate acid was dissolved in a mixed solvent of 30 mL acetone and 10 mL water and cooled in an ice-water bath. 20 mL of 2 mol / L sodium hydroxide aqueous solution was added dropwise with stirring, controlling the pH of the system to 8.5. After the addition was complete, the solvent was removed by vacuum evaporation, and the residue was dried under vacuum to obtain 7.6 g of pale yellow solid sodium chlortetracycline phosphate.

[0043] The product was analyzed by electrospray ionization mass spectrometry (ESI-MS), and two main quasi-molecular ion peaks were observed in positive ion mode, with mass-to-charge ratios (m / z) of [M+Na]. + =625.5 and [M+Na] + =749.8. Among them, the peak at m / z 625.5 corresponds to the theoretically calculated value of disodium chlortetracycline monophosphate [M+Na]. + The ion (molecular weight 602.8 + 23 = 625.8) matches; the peak at m / z 749.8 matches the theoretically calculated tetrasodium chlortetracycline diester [M + Na]. + The ions (molecular weight 726.8 + 23 = 749.8) are consistent. The above results indicate that the product obtained in this example is a mixture of disodium chlortetracycline phosphate monoester and tetrasodium chlortetracycline phosphate diester.

[0044] Example 2 Preparation of chlortetracycline phosphate arginine salt

[0045] (1) Place 10g of chlortetracycline hydrochloride (Pucheng Zhengda Biochemical Co., Ltd.) in a dry reaction flask, add 100mL of anhydrous pyridine, and stir under an ice-water bath. Slowly add 3.36mL of triethylamine, and stir for 1 hour at the same temperature to obtain free chlortetracycline base.

[0046] (2) Slowly add 5.37 mL of di-tert-butyl chlorite to the above-mentioned chlortetracycline free base. After the addition is complete, remove the ice bath and stir the reaction at 20-25°C for 4 hours. Place the reaction system back in an ice-water bath and slowly add 4.4 mL of 70% tert-butyl hydrogen peroxide aqueous solution. After the addition is complete, continue stirring the reaction for 1 hour.

[0047] (3) Pour the reaction solution into 200 mL of ice water and extract three times with dichloromethane. Combine the organic phases and wash them successively with 100 mL of 5% citric acid aqueous solution, 100 mL of saturated sodium bicarbonate aqueous solution, and 100 mL of saturated saline solution. Dry the organic phase with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain crude di-tert-butyl chlortetracycline phosphate.

[0048] (4) Dissolve the crude product in 100 mL of dichloromethane, cool in an ice-water bath, and slowly add 12.8 mL of trimethylsilane bromide while stirring. After the addition is complete, remove the ice bath and stir the reaction at 20-25 °C for 3 hours.

[0049] (5) Slowly pour the above reaction solution into 200 mL of ice water. Under ice bath and stirring, adjust the pH of the aqueous phase to 3-4 with 2 mol / L sodium hydroxide aqueous solution. Extract the aqueous phase with a mixed solvent (dichloromethane:isopropanol = 4:1, v / v, total 150 mL × 3). Combine the organic phases, dry with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain chlortetracycline phosphate.

[0050] (6) Dissolve the obtained chlortetracycline phosphate in a mixed solvent of 30 mL acetone and 10 mL water, and cool in an ice-water bath. Add dropwise an aqueous solution containing 20 mL of L-arginine (containing 6.8 g of L-arginine) while stirring, controlling the pH of the system to 7-7.5. After the addition is complete, evaporate the solvent under reduced pressure, and dry the residue under vacuum to obtain 8.5 g of pale yellow solid chlortetracycline phosphate arginine salt.

[0051] The product was analyzed by electrospray ionization mass spectrometry (ESI-MS), and a major molecular ion peak was observed in positive ion mode, with a mass-to-charge ratio (m / z) of [M+H]. + =908.9, which is consistent with the theoretically calculated value of chlortetracycline phosphate monoester diarginine salt [M+H]. + The ion (molecular weight 907.3) was consistent; simultaneously, a strong signal with a mass-to-charge ratio m / z of approximately 668.8 was observed, which is consistent with the double-charged ion [M+2H] of chlortetraarginine phosphate diester (theoretical molecular weight 1335.7). 2+ The theoretical calculation values ​​are consistent. The above results indicate that the product obtained in this embodiment is a mixture of chlortetracycline monophosphate diarginine salt and chlortetracycline diester tetraarginine salt.

[0052] Example 3: Preparation of chlortetracycline soluble powder

[0053] Formula composition:

[0054] Example 1 prepared a chlortetracycline phosphate sodium salt of 22%, a disodium ethylenediaminetetraacetate of 0.5%, a sodium dodecyl sulfate of 0.5%, and tartaric acid of 77%.

[0055] Preparation steps:

[0056] S1. After crushing the tartaric acid, pass it through an 80-mesh analytical sieve, and then take the sieved raw materials according to the above raw material ratio;

[0057] S2. Add sodium chlortetracycline phosphate to tartaric acid in equal increments and premix for 20 minutes to obtain the premix.

[0058] S3. Add disodium ethylenediaminetetraacetate and sodium dodecyl sulfate in equal increments to the premix obtained in step S2, and continue mixing for 20 minutes.

[0059] Example 4: Preparation of chlortetracycline soluble powder

[0060] Formula composition:

[0061] Example 1 prepared a mixture of 55% sodium chlortetracycline phosphate, 1% disodium ethylenediaminetetraacetate, 2% sodium dodecyl sulfate, and 42% anhydrous citric acid.

[0062] Preparation steps:

[0063] S1. After crushing the anhydrous citric acid, pass it through an 80-mesh analytical sieve, and then take the sieved raw materials according to the above raw material ratio;

[0064] S2. Add chlortetracycline phosphate arginine salt to tartaric acid in equal increments and premix for 20 minutes to obtain the premix.

[0065] S3. Add disodium ethylenediaminetetraacetate and sodium dodecyl sulfate in equal increments to the premix obtained in step S2, and continue mixing for 20 minutes.

[0066] Example 5: Preparation of chlortetracycline soluble powder

[0067] Formula composition:

[0068] Example 2 prepared a chlortetracycline phosphate arginine salt of 22%, disodium ethylenediaminetetraacetate of 0.5%, sodium dodecyl sulfate of 0.5%, and tartaric acid of 77%.

[0069] Preparation steps:

[0070] S1. After crushing the tartaric acid, pass it through an 80-mesh analytical sieve, and then take the sieved raw materials according to the above raw material ratio;

[0071] S2. Add sodium chlortetracycline phosphate to tartaric acid in equal increments and premix for 20 minutes to obtain the premix.

[0072] S3. Add disodium ethylenediaminetetraacetate and sodium dodecyl sulfate in equal increments to the premix obtained in step S2, and continue mixing for 20 minutes.

[0073] Example 6 Preparation of chlortetracycline soluble powder

[0074] Formula composition:

[0075] Example 2 prepared a chlortetracycline phosphate arginine salt 55%, ethylenediaminetetraacetic acid disodium salt 1%, sodium dodecyl sulfate 2%, and anhydrous citric acid 42%.

[0076] Preparation steps:

[0077] S1. After crushing the anhydrous citric acid, pass it through an 80-mesh analytical sieve, and then take the sieved raw materials according to the above raw material ratio;

[0078] S2. Add chlortetracycline phosphate arginine salt to tartaric acid in equal increments and premix for 20 minutes to obtain the premix.

[0079] S3. Add disodium ethylenediaminetetraacetate and sodium dodecyl sulfate in equal increments to the premix obtained in step S2, and continue mixing for 20 minutes.

[0080] Comparative Example 1

[0081] The difference from Example 3 is that this comparative example uses chlortetracycline hydrochloride instead of sodium chlortetracycline phosphate.

[0082] Formula composition:

[0083] Chlortetracycline hydrochloride (Pucheng Zhengda Biochemical Co., Ltd.) 22%, disodium ethylenediaminetetraacetate 0.5%, sodium dodecyl sulfate 0.5%, tartaric acid 77%.

[0084] Preparation steps:

[0085] S1. After crushing the tartaric acid, pass it through an 80-mesh analytical sieve, and then take the sieved raw materials according to the above raw material ratio;

[0086] S2. Add chlortetracycline hydrochloride to tartaric acid in equal increments and premix for 20 minutes to obtain the premix.

[0087] S3. Add disodium ethylenediaminetetraacetate and sodium dodecyl sulfate in equal increments to the premix obtained in step S2, and continue mixing for 20 minutes.

[0088] Comparative Example 2

[0089] The difference from Example 3 is that anhydrous glucose was used as an excipient in this comparative example to prepare the soluble powder.

[0090] Formula composition:

[0091] Example 1 prepared 22% sodium chlortetracycline phosphate and 78% anhydrous glucose.

[0092] Preparation steps:

[0093] S1. After crushing the anhydrous glucose, pass it through an 80-mesh analytical sieve, and then take the sieved raw materials according to the above raw material ratio;

[0094] S2. Add chlortetracycline hydrochloride to anhydrous glucose in equal increments and mix for 20 minutes.

[0095] Performance testing

[0096] I. Solubility determination:

[0097] Test samples: chlortetracycline soluble powder prepared in Examples 3, 5, Comparative Example 1 and Comparative Example 2.

[0098] Test method based on: Chinese Veterinary Pharmacopoeia 2020 Edition, Part I, General Regulations 0113 Soluble Powder [Solubility] Unless otherwise specified, take an appropriate amount of the test sample, place it in a Nessler tube, add water to make a 50 ml solution (concentration is twice the high dose concentration used in clinical applications), invert 10 times at 25℃±2℃, the test sample should be completely dissolved, let stand for 30 minutes, and there should be no turbidity or precipitation.

[0099] Test method: Referring to the solubility test method in the Veterinary Pharmacopoeia, the solubility test method in this test is as follows: Prepare distilled water at 25℃±2℃ for later use. Take a 100ml Nessler colorimetric tube, add the prepared distilled water to about 50ml, accurately weigh each test sample using a 0.01% balance, pour it into the colorimetric tube, add the prepared distilled water to make up to 100ml, start timing, shake vigorously up and down for 30 seconds, and let it stand in a water bath at 25℃±2℃. Repeat this process, shaking vigorously for 30 seconds every 5 minutes, and observe the solubility within 30 minutes. If there are no visible solute particles, it is considered to be completely dissolved.

[0100] At the beginning of the experiment, based on experience, a certain mass of the test sample was weighed and its solubility was tested. If it dissolved completely, a mass +0.1g of the test sample was weighed again; if it did not dissolve completely, a mass -0.1g of the test sample was weighed again, and the solubility was tested again. This continued until a 0.1g increase or decrease resulted in either incomplete or complete dissolution of the solution. At this point, the mass of the test sample added for complete dissolution was considered the critical solubility point; the mass added for incomplete dissolution was considered the critical insoluble point.

[0101] Experimental results:

[0102] Table 1 Solubility of chlortetracycline soluble powder

[0103]

[0104] As can be seen from the test results in Table 1, the solubility of the chlortetracycline soluble powder prepared in this application is significantly higher, reaching 11.3-20.5 g / 100 mL, which far exceeds the requirements of a 1:50 dosing device.

[0105] II. Determination of dissolution rate:

[0106] Test samples: chlortetracycline soluble powder prepared in Examples 3, 5, Comparative Example 1 and Comparative Example 2.

[0107] Experimental Method: Prepare distilled water at 25℃±2℃. Accurately measure 100mL of distilled water and pour it into an Erlenmeyer flask. Weigh 5.0g of chlortetracycline soluble powder from Examples 3, 5, Comparative Example 1, and Comparative Example 2. Invert the flask 10 times and let it stand in a water bath at 25℃±2℃. Repeat this inversion process 10 times every 30 seconds, observing the dissolution. If no visible solute particles are observed, it is considered completely dissolved. Start timing simultaneously and stop timing when the solution is completely clear. Record the time as the dissolution rate at that solubility.

[0108] Experimental results:

[0109] Table 2 Dissolution rate of chlortetracycline soluble powder

[0110]

[0111] As can be seen from the test results in Table 2, the dissolution rate of the chlortetracycline soluble powder prepared in Examples 3 and 5 is significantly faster than that in Comparative Examples 1 and 2. This indicates that preparing chlortetracycline hydrochloride into chlortetracycline phosphate salt can significantly improve the dissolution rate of chlortetracycline, thereby reducing the waiting time during application and improving the convenience of drinking water use.

[0112] III. Stability Testing:

[0113] Test samples: chlortetracycline soluble powder prepared in Example 3 and Comparative Example 1.

[0114] Test water: pure water, soft water (hardness standard <150mg / L), hard water (hardness standard 250mg / L), tap water, and well water. Hard water preparation process: Weigh 1.670g of calcium chloride and 0.952g of magnesium chloride, dissolve and dilute with deionized water to 1.0L, which is 2500mg / L hard water. When using, take 1.0L and dilute to 10.0L, which is 250mg / L hard water.

[0115] Experimental method: Take 0.2g of each test sample, accurately weigh it, dissolve it in water of different qualities, and make up to 100mL with water of the corresponding quality. At 0, 4, 8, 12 and 24h after dissolution, accurately take 20μL of the aqueous solution, filter it through a 0.45μm filter membrane, inject it into the liquid chromatograph, record the chromatogram, and calculate the content of chlortetracycline phosphate or chlortetracycline hydrochloride in the solution according to the external standard method.

[0116] Experimental results:

[0117] Table 3. Content stability of chlortetracycline soluble powder in different water qualities

[0118]

[0119] As can be seen from the test results in Table 3, during the 24-hour observation period after dissolution, the labeled content of the chlortetracycline soluble powder sample prepared in Example 3 of this application did not change significantly after dissolving in different water qualities and remained relatively stable after 24 hours. However, the chlortetracycline soluble powder sample prepared in Comparative Example 1 showed precipitation and sedimentation after dissolution in hard water, tap water, and well water, and the labeled content also decreased significantly over time. This indicates that although both Example 3 and Comparative Example 1 formulations contain chelating agents, the stability of the soluble powder using chlortetracycline phosphate salt as the active ingredient in this application is far superior to that using chlortetracycline hydrochloride as the active ingredient.

[0120] IV. Comparative Test of Bioavailability of Chlortetracycline Soluble Powder in Pigs

[0121] Test samples: chlortetracycline soluble powder prepared in Example 3 and Comparative Example 1.

[0122] Experimental Methods: Ten weaned piglets, weighing 7.5 ± 0.5 kg, were selected for the experiment and fed a complete diet (calcium content 1.0%) without any antibiotics. They were fasted for 12 hours before administration and randomly divided into two experimental groups, A and B, with five piglets in each group, based on similar weights. Group A received the chlortetracycline soluble powder obtained in Example 3, while Group B received the chlortetracycline soluble powder obtained in Comparative Example 1, administered orally. The powder was dissolved in an appropriate amount of physiological saline before administration. The dosage for both groups was 0.15 g / kg BW. At 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 4.0, 6.0, 8.0, 10.0, and 24.0 hours after administration, 10 mL of blood was collected from the anterior vena cava. The blood drug concentration was analyzed by HPLC, and the pharmacokinetic parameters after administration were calculated using a non-compartmental model in WinNonlin software.

[0123] Experimental results:

[0124] Table 4. Pharmacokinetic parameters of chlortetracycline soluble powder

[0125]

[0126] The data in the table above show that the time to peak concentration (Tmax) in group A was significantly faster than that in group B (P < 0.05), and the area under the curve (AUC0-t) and peak concentration (Cmax) in group A were both significantly greater than those in group B (P < 0.05). This indicates that chlortetracycline hydrochloride, after chemical modification with phosphate groups, has good absorption in vivo. Under the same dietary calcium content, the bioavailability of Example 3 of this application was increased by more than 2 times.

[0127] Compared with chlortetracycline hydrochloride, the chlortetracycline phosphate salt of this application is chemically modified, which can significantly improve solubility, avoid metal ion chelation and gastric acid degradation, and achieve targeted release by intestinal enzymatic hydrolysis. This can significantly improve the oral bioavailability of chlortetracycline, obtain a better blood drug concentration-time curve, and thus achieve stronger and more stable efficacy.

[0128] V. Application Test of Chlortetracycline Soluble Powder in the Treatment of Bronchopneumonia Caused by Susceptible Escherichia coli in Chickens

[0129] Test samples: chlortetracycline soluble powder prepared in Example 3, Comparative Example 1, and Comparative Example 2.

[0130] Experimental animals: 150 broiler chickens of uniform weight at 20 days of age with respiratory diseases were selected. Clinical symptoms included lethargy, loss of appetite, reduced intake, weight loss, runny nose, sneezing, watery eyes, sinusitis, facial edema, coughing, difficulty breathing, and death. The chickens were randomly divided into 3 groups of 50 each.

[0131] Group administration:

[0132] (1) The first group of drugs was the chlortetracycline soluble powder prepared in Example 3. Each 2.0g was mixed with 1L of water for drinking. After drinking, the patient could drink water freely. This was done for three consecutive days.

[0133] (2) The second group was given the chlortetracycline soluble powder prepared in Comparative Example 1. Each 2.0g was diluted with 1L of water for drinking. After drinking, the patient could drink water freely. This treatment was continued for three days.

[0134] (3) The third group was given the chlortetracycline soluble powder prepared in Comparative Example 2. Each 2.0g was diluted with 1L of water for drinking. After drinking, the patient could drink water freely. This treatment was continued for three days.

[0135] Efficacy evaluation indicators:

[0136] The following indicators were all calculated based on data collected one week after medication.

[0137] (1) Number of deaths: refers to the number of chickens that died during the experiment due to worsening respiratory symptoms.

[0138] (2) Effective number: refers to the number of chickens whose respiratory symptoms have been reduced or disappeared, and whose mental state and feed intake have improved or returned to normal after medication.

[0139] (3) Number of cured chickens: refers to the number of chickens whose respiratory symptoms disappeared and whose mental state and feed intake returned to normal after medication.

[0140] (4) Cure rate: refers to the percentage of chickens cured out of the total number of chickens in the group.

[0141] Table 5. Therapeutic effect of chlortetracycline soluble powder on respiratory diseases in broilers.

[0142]

[0143] Table 5 shows that the chlortetracycline soluble powder prepared in Example 3 of this application achieved a cure rate of 96% for bronchopneumonia in broilers, which is significantly better than the therapeutic effects of chlortetracycline soluble powders in Comparative Examples 1 and 2, indicating that the chlortetracycline hydrochloride soluble powder provided in this application has a better therapeutic effect.

[0144] The above description is merely a preferred embodiment of this application and does not constitute any limitation on this application in any form or substance. It should be noted that those skilled in the art can make several improvements and additions without departing from the method of this application, and these improvements and additions should also be considered within the scope of protection of this application. Any modifications, alterations, and equivalent changes made by those skilled in the art based on the above-disclosed technical content without departing from the spirit and scope of this application are equivalent embodiments of this application; furthermore, any modifications, alterations, and evolutions made to the above embodiments based on the essential technology of this application still fall within the scope of the technical solution of this application.

Claims

1. A soluble powder of chlortetracycline, characterized in that, It is composed of the following components by mass percentage: chlortetracycline phosphate salt 20-55%, chelating agent 0.5-2.5%, penetration enhancer 0.5-2.5%, and organic acid carrier balance; The chlortetracycline phosphate salt is prepared by a method comprising the following steps: (1) Reaction of chlortetracycline hydrochloride with an organic base yields free chlortetracycline base; (2) The free base of chlortetracycline reacts with di-tert-butyl chlorite and is oxidized to generate di-tert-butyl chlortetracycline phosphate; (3) Remove the tert-butyl protecting group from di-tert-butyl chlortetracycline phosphate to obtain chlortetracycline phosphate; (4) React chlortetracycline phosphate with a base to obtain the chlortetracycline phosphate salt; In step (2), the oxidant used for oxidation is tert-butyl hydrogen peroxide; In step (3), the tert-butyl protecting group is removed using an acid hydrolysate, wherein the acid hydrolysate is trimethylsilane bromide; The organic acid carrier is selected from at least one of citric acid and tartaric acid.

2. The chlortetracycline soluble powder according to claim 1, characterized in that, In step (1), the organic base is selected from at least one of triethylamine, N,N-diisopropylethylamine and pyridine.

3. The chlortetracycline soluble powder according to claim 1, characterized in that, In step (4), the base is selected from at least one of meglumine, arginine, lysine, sodium hydroxide, potassium hydroxide, sodium carbonate, and potassium carbonate.

4. The chlortetracycline soluble powder according to claim 1, characterized in that, The chelating agent is selected from at least one of disodium ethylenediaminetetraacetate, sodium tripolyphosphate, and sodium hexametaphosphate.

5. The chlortetracycline soluble powder according to claim 1, characterized in that, The penetration enhancer is selected from at least one of sodium salicylate, sodium deoxycholate, and sodium dodecyl sulfate.

6. A method for preparing chlortetracycline soluble powder according to any one of claims 1-5, characterized in that, Includes the following steps: Chlortetracycline phosphate salt and organic acid carrier are mixed evenly to obtain a premix; Add the chelating agent and penetration enhancer to the premix, mix well, and obtain chlortetracycline soluble powder.

7. The method for preparing chlortetracycline soluble powder according to claim 6, characterized in that, The chlortetracycline phosphate salt is prepared by a method comprising the following steps: (1) Reaction of chlortetracycline hydrochloride with an organic base yields free chlortetracycline base; (2) The free base of chlortetracycline reacts with di-tert-butyl chlorite and is oxidized to generate di-tert-butyl chlortetracycline phosphate; (3) Remove the tert-butyl protecting group from di-tert-butyl chlortetracycline phosphate to obtain chlortetracycline phosphate; (4) React chlortetracycline phosphate with a base to obtain the chlortetracycline phosphate salt.

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