Valnemulin long-acting compound preparation as well as preparation method and application thereof

Abstract

The invention belongs to the field of medicines, and particularly relates to a valnemulin long-acting compound preparation as well as a preparation method and application thereof. The valnemulin long-acting compound preparation comprises the following components in percentage by mass: 5%-10% of valnemulin, 10%-30% of doxycycline, 0.5%-2.0% of tartaric acid, 10%-20% of an oil phase, 10%-30% of an emulsifier, 2%-10% of a co-emulsifier and the balance of water. The nano-emulsion is prepared from valnemulin, doxycycline and corresponding auxiliary materials, so that the release rate of valnemulin can be effectively delayed, the two medicines can be synchronously eliminated in vivo, and a synergistic antibacterial effect is achieved; moreover, the drug is wrapped in the core of the nanoemulsion, so that the solubility of the valnemulin in water is improved, the doxycycline is prevented from forming an insoluble complex with metal ions in water, and the stability of the drug is improved.

Description

Technical Field

[0001] The present invention belongs to the field of medicine, and particularly relates to a valnemulin long-acting compound preparation, a preparation method thereof and an application thereof. Background Art

[0002] In the process of the development of livestock and poultry farming towards intensification, large-scale and high-efficiency, the requirements for veterinary drugs have also been raised due to the needs and challenges of livestock and poultry disease prevention and control. For example: in high-density farming, the incidence of animal diseases increases, the spread is fast, and the main type is mixed infection. This requires that the use of drugs should achieve the prevention and treatment purposes, and be efficient and broad-spectrum. Therefore, in high-efficiency farming, the improvement of efficiency is the key, which also requires reducing the number of drug administrations and lowering the labor intensity; the rapid development of farming has also brought risks such as food safety and bacterial drug resistance, and the prominent contradiction between disease prevention and control and drug use must be solved.

[0003] In livestock and poultry farming, mixed infections of Gram-positive cocci, Gram-negative bacilli and mycoplasmas are common. However, most drugs have specific antibacterial spectra and are less effective in simultaneously preventing and treating different pathogenic bacteria. Therefore, multiple drugs need to be used in combination in disease prevention and treatment. However, due to the lack of clinical veterinarians in farming, accurate drug combinations are rarely carried out, which leads to a series of problems such as treatment failure, drug waste, increased veterinary drug residues and even rapid induction of bacterial drug resistance.

[0004] In recent years, in the clinical practice of poultry farming, mixed infections of mycoplasmas and bacteria often occur, seriously affecting the healthy development of the poultry farming industry. Valnemulin is a pleuromutilin-class animal-specific antibiotic, which mainly binds to the 50S subunit on the ribosome of pathogenic microorganisms to inhibit protein synthesis and achieve an antibacterial effect. Valnemulin has a strong inhibitory effect on a variety of pathogenic mycoplasmas such as Mycoplasma gallisepticum and Mycoplasma synoviae, and Gram-positive bacteria, but is ineffective against Gram-negative bacteria. Doxycycline is a tetracycline-class antibiotic, and its mechanism of action is to bind to the 30S subunit on the ribosome of pathogenic microorganisms to inhibit the synthesis of pathogenic bacteria proteins and achieve a bactericidal effect. It is mainly used in farming clinics for the prevention and treatment of Gram-negative bacterial infections. Valnemulin and doxycycline are complementary in antibacterial mechanism and antibacterial spectrum and can produce a synergistic effect when combined.

[0005] Pharmacokinetic studies have shown that valnemulin and doxycycline can be rapidly absorbed after oral administration to chickens, with peak times of 1 - 2 h and 2 - 5 h respectively. However, there are significant differences in their elimination rates in chickens: valnemulin has a very fast elimination rate, with an elimination half-life of 2 - 4 h, while doxycycline has a slower elimination rate, with an elimination half-life of 9 - 13 h. Therefore, when the two drugs are used in combination, their elimination rates are inconsistent and it is impossible to implement the same dosing interval, which is also the key problem hindering the development of valnemulin and doxycycline compound preparations. In addition, modern high-efficiency farming models tend to use the easier-to-operate drinking water route for drug administration. However, the solubility of valnemulin in water is difficult to meet the drug use requirements for high-efficiency farming drinking water administration, and doxycycline is sensitive to metal ions in water and has poor stability in water, which is another problem in the development of valnemulin and doxycycline compound preparations.

[0006] In summary, how to balance the half-lives of valnemulin and doxycycline, improve their water solubility, and develop new compound preparations is an effective measure to meet the prevention and control needs of mixed infection diseases. Summary of the Invention

[0007] In view of the above technical problems, the present invention provides a long-acting compound preparation of valnemulin, which enables valnemulin and doxycycline to be eliminated at a similar rate, and improves the solubility and stability of doxycycline and valnemulin in water, and can effectively solve the problem of mixed infection diseases in poultry.

[0008] In order to achieve the above object, the technical solution adopted by the present invention is as follows:

[0009] A long-acting compound preparation of valnemulin, comprising the following components and their mass percentages: valnemulin 5% - 10%, doxycycline 10% - 30%, tartaric acid 0.5% - 2.0%, oil phase 10% - 20%, emulsifier 10% - 30%, co-emulsifier 2% - 10%, and the balance being water.

[0010] Valnemulin and doxycycline are prone to form flocculent floating substances after being mixed and dissolved in water. Therefore, the drug-forming property of the two prepared into a compound preparation is poor. In the experimental process of the present invention, it was found that adding an appropriate amount of tartaric acid can significantly improve the compatibility of the two in water and is more conducive to the preparation of a compound preparation. By adding an oil phase, an emulsifier and a co-emulsifier, valnemulin and doxycycline can be encapsulated into nanoemulsions, which not only improves the solubility and stability of the drugs in water, but also can effectively delay the release rate of valnemulin, so that the two drugs achieve synchronous elimination in the body.

[0011] Preferably, the long-acting compound preparation of valnemulin is composed of the following components and their mass percentages: valnemulin 10%, doxycycline 30%, tartaric acid 2.0%, oil phase 20%, emulsifier 10%, co-emulsifier 8%, and the balance being water.

[0012] Preferably, valnemulin is selected from its hydrochloride or tartrate; doxycycline is selected from its hydrochloride.

[0013] Preferably, the mass ratio of valnemulin to doxycycline is 1:3 - 4.

[0014] Preferably, the oil phase is glyceryl triacetate; the emulsifier is Tween - 80, and the co - emulsifier is propylene glycol.

[0015] In the formulation of the nano - emulsion, the inventors found that the oil phase glyceryl triacetate is the main component of the hydrophobic core in the nano - emulsion structure. The formed nano - core can effectively encapsulate valnemulin and doxycycline, achieving a sustained - release and long - acting effect. When Tween - 80 is selected as the emulsifier and propylene glycol as the co - emulsifier, a nano - emulsion with more stable properties can be prepared under the action of high - intensity mechanical force with the oil phase glyceryl triacetate.

[0016] The present invention also provides a preparation method of the long - acting compound preparation of valnemulin, including the following processes:

[0017] S1. Add tartaric acid to water, stir evenly, and then add the emulsifier and co - emulsifier thereto, and continue to mix and stir to make it evenly mixed to obtain mixture I;

[0018] S2. Add valnemulin and doxycycline to mixture I obtained in step S1, stir and mix evenly to completely dissolve the drugs to obtain mixture II;

[0019] S3. Add the oil phase to mixture II obtained in step S2, stir and mix evenly to obtain mixture III;

[0020] S4. Perform high - speed shearing treatment on mixture III obtained in step S3 to obtain the product.

[0021] For the long - acting compound preparation of valnemulin provided by the present invention, after mixing each component evenly, during the high - speed shearing process, due to centrifugal force and shearing force, the drugs, emulsifier, co - emulsifier, oil phase, etc. are fully homogenized and emulsified to form a homogeneous and stable nano - emulsion.

[0022] Preferably, the condition for the uniform stirring in step S1 is to stir at a rotation speed of 300 - 800 rpm for 10 - 15 min; the condition for the continued mixing and stirring is to stir at a rotation speed of 500 - 800 rpm for 15 - 20 min.

[0023] Preferably, the condition for the uniform stirring and mixing in steps S2 and S3 is to stir at a rotation speed of 500 - 800 rpm for 15 - 25 min.

[0024] Preferably, the high-speed shearing conditions in step S4 are homogenizing and shearing at a rotation speed of 7000 - 9000 rpm for 15 - 20 min.

[0025] The present invention also provides an application of the valnemulin long-acting compound preparation in the preparation of veterinary drugs suitable for high-efficiency breeding modes.

[0026] Compared with the prior art, the present invention has the following technical advantages:

[0027] (1) The present invention is a compound preparation composed of valnemulin and doxycycline. The antibacterial mechanisms and antibacterial spectra of the two are complementary. The combined use of the two can achieve the effects of synergistic antibacterial, expanding the antibacterial spectrum and enhancing the efficacy. And the use of tartaric acid effectively solves the problems of poor solubility and poor compatibility of valnemulin and doxycycline in water, significantly improving the druggability of the valnemulin doxycycline compound preparation;

[0028] (2) The present invention prepares the valnemulin and doxycycline into a compound nanoemulsion, which can effectively delay the release rate of valnemulin, enabling valnemulin and doxycycline to achieve synchronous elimination in the body, and achieving the effect of maximizing the synergistic antibacterial effect;

[0029] (3) The present invention prepares the valnemulin and doxycycline into a compound nanoemulsion, which not only improves the solubility and stability of valnemulin in water, but also avoids the contact of doxycycline with metal ions in water, improving its stability in water;

[0030] (4) The compound nanoemulsion provided by the present invention can be completely dissolved in any proportion in water and can tolerate different water qualities, fully meeting the requirements of modern high-efficiency breeding for drinking water administration. Description of the Drawings

[0031] Figure 1 It is a diagram of the detection results of the sample properties in Example 1 of the present invention;

[0032] Figure 2 It is a diagram of the detection results of the properties of Comparative Example 3 (left) and Comparative Example 4 (right) of the present invention. Detailed Embodiments

[0033] The present invention will be further explained below in conjunction with specific embodiments. However, it should be noted that the following embodiments are only used to explain the present invention and cannot be used to limit the present invention. All technical solutions identical or similar to the present invention are within the protection scope of the present invention. For those not specifying specific techniques or conditions in this embodiment, operations are carried out according to the conventional technical methods and instrument instruction manuals in the art; for reagents or instruments not specifying the manufacturer, they are all conventional products that can be obtained through commercial purchase.

[0034] Example 1 A valnemulin long-acting compound preparation

[0035] The valnemulin long-acting compound preparation includes the following components and their mass percentages: valnemulin 5%, doxycycline 15%, tartaric acid (DL-tartaric acid, purchased from Hunan Er-Kang Pharmaceutical Co., Ltd., the same below) 1.0%, triacetin 15%, Tween-80 20%, propylene glycol 10%, and the balance is water.

[0036] The preparation method of the valnemulin long-acting compound preparation is as follows: Add 1.0 g of tartaric acid to 30 mL of water, stir at 500 rpm for 10 min until evenly mixed, then add 20 g of Tween-80 and 10.0 g of propylene glycol, and stir at 500 rpm for 10 min to make it evenly mixed; continue to add 5.8 g of valnemulin hydrochloride (5.0 g in terms of valnemulin) and 16.3 g of doxycycline hydrochloride (15.0 g in terms of doxycycline), stir at 500 rpm for 20 min to completely dissolve the drug; add 15.0 g of triacetin, stir at 500 rpm for 20 min, and perform high-speed shearing at 8000 rpm for 15 min, and make up the volume to 100 mL with water and stir evenly to obtain the product.

[0037] Example 2 A valnemulin long-acting compound preparation

[0038] The valnemulin long-acting compound preparation includes the following components and their mass percentages: valnemulin 10%, doxycycline 30%, tartaric acid 2.0%, triacetin 20%, Tween-80 10%, propylene glycol 8%, and the balance is water.

[0039] The preparation process of the valnemulin long-acting compound preparation is as follows: Add 2.0 g of tartaric acid to 30 mL of water, stir at 500 rpm for 10 min until evenly mixed, then add 10.0 g of Tween-80 and 8.0 g of propylene glycol, and stir at 500 rpm for 10 min to make it evenly mixed; continue to add 11.6 g of valnemulin hydrochloride (10.0 g in terms of valnemulin) and 32.6 g of doxycycline hydrochloride (30.0 g in terms of doxycycline), stir at 500 rpm for 20 min to completely dissolve the drug; add 20.0 g of triacetin, stir at 500 rpm for 20 min, and perform high-speed shearing at 8000 rpm for 15 min, and make up the volume to 100 mL with water and stir evenly to obtain the product.

[0040] Example 3 A valnemulin long-acting compound preparation

[0041] The valnemulin long-acting compound preparation includes the following components and their mass percentages: valnemulin 7%, doxycycline 28%, tartaric acid 1.5%, triacetin 15%, Tween-80 20%, propylene glycol 10%, and the balance is water.

[0042] The preparation process of the valnemulin long-acting compound preparation is as follows: Add 1.5 g of tartaric acid to 30 mL of water, stir at 500 rpm for 10 min until evenly mixed, then add 20.0 g of Tween-80 and 10.0 g of propylene glycol, and stir at 500 rpm for 10 min to make it evenly mixed; continue to add 8.1 g of valnemulin hydrochloride (7.0 g in terms of valnemulin) and 30.4 g of doxycycline hydrochloride (28.0 g in terms of doxycycline), stir at 500 rpm for 20 min to completely dissolve the drugs; add 15.0 g of triacetin, stir at 500 rpm for 20 min, and perform high-speed shearing at 8000 rpm for 15 min, then make up the volume to 100 mL with water and stir evenly to obtain the product.

[0043] Comparative Example 1 A valnemulin hydrochloride solution

[0044] Weigh 5.8 g of valnemulin hydrochloride (5.0 g in terms of valnemulin), add it to 75 mL of water, stir at 500 rpm for 10 min until completely dissolved, and make up the volume to 100 mL with water.

[0045] Comparative Example 2 A doxycycline hydrochloride solution

[0046] Weigh 16.3 g of doxycycline hydrochloride (15.0 g in terms of doxycycline), add it to 75 mL of water, stir at 500 rpm for 10 min until completely dissolved, and make up the volume to 100 mL with water.

[0047] Comparative Example 3 A valnemulin-doxycycline compound solution

[0048] The valnemulin-doxycycline compound solution contains 10% valnemulin, 30% doxycycline, and the balance is water.

[0049] The specific preparation process is as follows: Weigh 11.6 g of valnemulin hydrochloride (10.0 g in terms of valnemulin) and 32.6 g of doxycycline hydrochloride (30.0 g in terms of doxycycline), add them to 75 mL of water, stir at 500 rpm for 20 min, and make up the volume to 100 mL with water.

[0050] Comparative Example 4 A valnemulin-doxycycline compound solution

[0051] The valnemulin-doxycycline compound solution contains 10% valnemulin, 30% doxycycline, 2.0% citric acid, and the balance is water.

[0052] The specific preparation process is as follows: Weigh 11.6 g of valnemulin hydrochloride (10.0 g in terms of valnemulin) and 32.6 g of doxycycline hydrochloride (30.0 g in terms of doxycycline), add them to 75 mL of water, then add 2.0 g of citric acid thereto, stir at 500 rpm for 20 min, and make up the volume to 100 mL with water.

[0053] Comparative Example 5 A valnemulin-doxycycline composite solution

[0054] The valnemulin-doxycycline composite solution comprises 10% valnemulin, 30% doxycycline, 2.0% tartaric acid, and the balance water.

[0055] The specific preparation process is as follows: Add 2.0 g of tartaric acid to 75 mL of water, stir at 500 rpm for 10 min until evenly mixed, then add 11.6 g of valnemulin hydrochloride (10.0 g calculated as valnemulin) and 32.6 g of doxycycline hydrochloride (30.0 g calculated as doxycycline), stir at 500 rpm for 20 min to completely dissolve the drugs, and make up the volume to 100 mL with water.

[0056] Comparative Example 6 A valnemulin composite preparation

[0057] The valnemulin composite preparation comprises the following components and their mass percentages: valnemulin 10%, doxycycline 30%, tartaric acid 2.0%, glyceryl stearate 20%, Tween-80 10%, glycerol 8%, and the balance water.

[0058] The preparation process of the valnemulin composite preparation is as follows: Add 2.0 g of tartaric acid to 30 mL of water, stir at 500 rpm for 10 min until evenly mixed, then add 10.0 g of Tween-80 and 8.0 g of glycerol, stir at 500 rpm for 10 min to make it evenly mixed; continue to add 11.6 g of valnemulin hydrochloride (10.0 g calculated as valnemulin) and 32.6 g of doxycycline hydrochloride (30.0 g calculated as doxycycline), stir at 500 rpm for 20 min to completely dissolve the drugs; add 20.0 g of glyceryl stearate, stir at 500 rpm for 20 min, and perform high-speed shearing at 8000 rpm for 15 min, make up the volume to 100 mL with water, and stir evenly to obtain the product.

[0059] Test Example 1 Detection of sample clarity

[0060] 1. Test samples: The products prepared in Examples 1-3 and Comparative Examples 1-6.

[0061] 2. Test method: Take 30 mL of each of the above product solutions, place them in beakers respectively, perform aseptic treatment throughout the process, and observe the color change of each sample after standing for 24 h.

[0062] 3. Test results: The specific test results are shown in Table 1 below.

[0063] Table 1 Investigation results of the properties of samples from different examples and comparative examples

[0064] Serial number Sample property Property after standing for 24 h Example 1 Yellow clear solution( Figure 1 ) Yellow clear solution Example 2 Yellow clear solution Yellow clear solution Example 3 Yellow clear solution Yellow clear solution Comparative example 1 Colorless clear solution White precipitate precipitates at the bottom Comparative example 2 Yellow clear solution Yellow clear solution Comparative example 3 Yellow turbid solution, with white floating matter on the liquid surface / Comparative example 4 Yellow turbid solution, with white floating matter on the liquid surface / Comparative example 5 Yellow clear solution Yellow clear solution Comparative example 6 Yellow clear solution Yellow clear solution

[0065] As can be seen from Table 1 above, after valnemulin hydrochloride is dissolved in water, white precipitates will precipitate out after standing for 24 hours, indicating that it is unstable in water (Comparative Example 1); after valnemulin hydrochloride and doxycycline hydrochloride are mixed in water, they cannot be completely dissolved, forming a yellow turbid solution with white floating substances on the liquid surface (Comparative Example 3, Figure 2 left figure); after replacing tartaric acid with citric acid, although the solution stability is better than that of Comparative Example 3, it is still much worse than that of the example group, and white floating substances can be clearly seen (Comparative Example 4, Figure 2 right figure); adding an appropriate amount of tartaric acid can significantly improve the solubility and compatibility of valnemulin hydrochloride and doxycycline hydrochloride in water, and the two can be completely dissolved (Comparative Example 5 and Comparative Example 6). After valnemulin hydrochloride and doxycycline hydrochloride are made into a compound nanoemulsion, the sample is a yellow clear solution and is relatively stable, and no precipitation or other situations are seen after long-term placement (Examples 1 to 3).

[0066] Test Example 2 Dilution resistance in water

[0067] Considering the complexity of the water quality in aquaculture clinics, the dilution resistance of Examples 1 to 3 in purified water and standard hard water was investigated respectively to avoid precipitation after drug dilution, which would affect the drug efficacy and block the drinking water pipeline, affecting the use of drugs in aquaculture clinics. At the same time, Comparative Example 1, Comparative Example 2, Comparative Example 5 and Comparative Example 6 were used as control groups for comparison.

[0068] 1. Test samples: The products prepared from the groups of Examples 1-3, the groups of Comparative Examples 1-2 and the groups of Comparative Examples 5-6;

[0069] 2. Test process: Take the samples of Example 1, Example 2, Example 3, Comparative Example 1, Comparative Example 2, Comparative Example 5 and Comparative Example 6, and dissolve them in purified water and standard hard water at ratios of 1:10, 1:100, and 1:500 respectively. Stir well at a speed of 500 rpm and let stand overnight at room temperature to observe the dilution resistance of the sample solutions in purified water and standard hard water.

[0070] 3. Test results: The specific test results are shown in Tables 2 to 4 below.

[0071] Table 2 Results of the investigation of dilution resistance in water (1:10)

[0072]

[0073] Table 3 Results of the investigation of dilution resistance in water (1:100)

[0074]

[0075]

[0076] Table 4 Results of the investigation on dilution resistance in water (1:500)

[0077]

[0078] As can be seen from the above Tables 2 to 4, after the samples of Comparative Example 1 were diluted with water and left overnight, turbidity could be observed, indicating that the aqueous solution of valnemulin hydrochloride has poor dilution in water. Although the samples of Comparative Example 2 and Comparative Example 5 can be diluted in purified water in any proportion, turbidity or precipitation will occur after the above samples are diluted in standard hard water, indicating that the above samples have poor tolerance to water quality and are difficult to meet the requirements of the complexity of clinical water for aquaculture; although the oil phase, emulsifier and co-emulsifier were also added in Comparative Example 6, turbidity or precipitation will also occur due to the change in components. Examples 1 to 3 can be dissolved in purified water and standard hard water in any proportion, and no turbidity or precipitation was observed after standing overnight at room temperature, indicating that the preparation of valnemulin hydrochloride and doxycycline hydrochloride into nanoemulsion can improve its stability in water and tolerance to different water qualities.

[0079] Test Example 3 Stability test in water

[0080] 1. Test samples: The long-acting compound preparation of valnemulin prepared from the groups of Examples 1-3;

[0081] 2. Test procedure: According to the method of Test Example 2, the drug contents in the diluents of Examples 1 to 3 at different waters, different times and different dilution ratios were measured respectively, and the changes in the drug contents in the nanoemulsion were observed.

[0082] 3. Test results: The specific test results are shown in Table 5.

[0083] Table 5 Results of the investigation on drug content for stability in water (unit: %)

[0084]

[0085] As can be seen from the results in Table 5 above, after Examples 1 to 3 were diluted in purified water and standard hard water at different dilution concentrations, the drug stability was good, and no obvious degradation reaction of the drug was observed after standing overnight.

[0086] Test Example 4 Drug stability

[0087] According to the requirements of the guiding principles for stability tests in the Chinese Veterinary Pharmacopoeia (2020 Edition), the stability evaluation tests of the valnemulin doxycycline compound nanoemulsion of Examples 1 and 2 were carried out, and Comparative Example 5 and Comparative Example 6 were used as control groups for comparison.

[0088] 4.1 High temperature test

[0089] After the samples of Example 1, Example 2, Example 3, Comparative Example 5 and Comparative Example 6 were sub-packaged and sealed, they were placed at a high temperature of 60 °C for 10 days, and samples were taken on the 0th day and the 10th day respectively to detect the drug content. The results are shown in Table 6 below.

[0090] Table 6 Results of drug content in high temperature test (unit: %)

[0091]

[0092] As can be seen from Table 6 above, after the compound aqueous solution of valnemulin hydrochloride and doxycycline hydrochloride was placed at high temperature for 10 days, the drug contents of valnemulin and doxycycline both decreased by more than 4% (Comparative Example 5); in the nanoemulsion obtained by emulsification with other emulsifiers, the drug contents of valnemulin and doxycycline both decreased by about 3%; while the compound nanoemulsion of valnemulin hydrochloride and doxycycline hydrochloride was placed at the same high temperature for 10 days, the decrease amounts of the two drug contents were both less than 1% (Examples 1 to 3). It can be seen that after the nanoemulsion is prepared according to the formula of the present invention, the drug stability of valnemulin and doxycycline can be effectively improved.

[0093] 4.2 Accelerated test:

[0094] After the samples of Example 1, Example 2, Example 3, Comparative Example 5 and Comparative Example 6 were sub-packaged and sealed, they were placed under the accelerated test conditions of a temperature of 40 °C ± 2 °C and a relative humidity of 75% ± 5% for 6 months, and samples were taken at the 0th, 1st, 3rd and 6th months respectively to detect the drug content. The test results are shown in Table 7 below.

[0095] Table 7 Results of drug content in accelerated test (unit: %)

[0096]

[0097]

[0098] As can be seen from Table 7 above, after the compound aqueous solution of valnemulin hydrochloride and doxycycline hydrochloride was placed under the accelerated test conditions for 6 months, the drug contents of valnemulin and doxycycline both decreased by more than 5% (Comparative Example 5); in the nanoemulsion obtained by emulsification with other emulsifiers, the drug contents of valnemulin and doxycycline both decreased by more than 3%; while the compound nanoemulsion of valnemulin hydrochloride and doxycycline hydrochloride was placed at the same high temperature for 10 days, the decrease amounts of the two drug contents were both less than 2% (Examples 1 to 3). Compared with the compound aqueous solution of valnemulin hydrochloride and doxycycline hydrochloride, the nanoemulsion of valnemulin hydrochloride and doxycycline hydrochloride has better stability under the accelerated test conditions.

[0099] Test Example 5 Pharmacokinetic test

[0100] 1. Test samples: Samples prepared in Example 2, Comparative Examples 1-2, and Comparative Examples 5-6;

[0101] 2. Test animals: Commercial broiler chickens, with a body weight of 1.30 - 1.50 kg, half male and half female, without obvious adverse reactions, a total of 50, randomly divided into 5 groups evenly;

[0102] 3. Test process: Randomly divided into 4 groups: the test group of Example 2, the test group of Comparative Example 1, the test group of Comparative Example 2, the test group of Comparative Example 5, and the test group of Comparative Example 6. Each test group has 10 test animals. Each test group was orally administered by gavage according to the dosing regimen in Table 8.

[0103] Table 8 Dosing regimen for pharmacokinetic experiments

[0104]

[0105]

[0106] Blood samples were collected from each test group before dosing and at 0.25 h, 0.5 h, 1 h, 2 h, 3 h, 4 h, 6 h, 8 h, 10 h, 12 h, 16 h, 18 h, 24 h, and 36 h after dosing. Plasma was separated. After the plasma samples were extracted with 50% acetonitrile, the concentrations of valnemulin and doxycycline in plasma at different time points were determined by high performance liquid chromatography-tandem mass spectrometry. WinNonlin software was used to fit the pharmacokinetic parameters, and the results are shown in Table 9.

[0107] Table 9 Main pharmacokinetic parameters of each test group

[0108]

[0109] The results show that compared with the test group of valnemulin hydrochloride aqueous solution (Comparative Example 1) and the test group of doxycycline hydrochloride aqueous solution (Comparative Example 2), there is no obvious difference in the pharmacokinetic parameters of the test group of valnemulin hydrochloride-doxycycline hydrochloride composite aqueous solution (Comparative Example 5). Compared with the test group of valnemulin hydrochloride aqueous solution (Comparative Example 1), after valnemulin hydrochloride was made into a nanoemulsion (Example 2), the peak time (T max ) and elimination half-life (T 1 / 2β ) of valnemulin in chicken plasma were significantly prolonged, achieving a long-acting effect, and the bioavailability increased by 26.41%. Compared with the test group of doxycycline hydrochloride aqueous solution (Comparative Example 2), after doxycycline hydrochloride was made into a nanoemulsion (Example 1), the peak time (T max ) and the elimination half-life in chickens (T 1 / 2β) was also prolonged, and the bioavailability was increased by 10.62%. Similarly, compared with the test group of valnemulin hydrochloride-doxycycline hydrochloride complex aqueous solution (Comparative Example 5), in the test group of valnemulin hydrochloride-doxycycline hydrochloride compound nanoemulsion (Example 1), the peak time (T max ) and elimination half-life (T 1 / 2β ) of valnemulin and doxycycline in chicken plasma were significantly prolonged, and the bioavailabilities were increased by 28.85% and 15.62% respectively. Compared with Comparative Example 6, the half-life (T 1 / 2β ) of valnemulin in the valnemulin hydrochloride-doxycycline hydrochloride compound nanoemulsion prepared in Example 2 was also significantly prolonged, and the bioavailability was also increased by 17.71%.

[0110] In the test group of Example 2, compared with doxycycline hydrochloride, the peak time and elimination half-life of valnemulin hydrochloride in vivo were more significantly prolonged after being made into nanoemulsion. The reasons may be: (1) The solubility of doxycycline hydrochloride in water is better than that of valnemulin hydrochloride, and it is more easily dissolved and released in vivo; (2) The hydroxyl group on the four-membered ring of doxycycline hydrochloride is easy to form intramolecular hydrogen bonds, resulting in steric hindrance; while the amino group on the side chain of valnemulin hydrochloride has small steric hindrance, which is more conducive to its close combination with excipients through intermolecular hydrogen bond interaction or van der Waals force, and is conducive to its slow release from the lipophilic nanocore formed by excipients to achieve a longer-lasting goal.

[0111] From the above pharmacokinetic parameters, it can be seen that after making valnemulin and doxycycline into compound nanoemulsion, the elimination half-life of valnemulin in vivo was significantly prolonged, and the elimination half-lives of the two were closer, which was more conducive to the two playing a synergistic antibacterial maximization effect.

[0112] Finally, it should be noted that the above embodiments are only illustrative of the principles, properties and effects of the present invention, and are not used to limit the present invention. Any person familiar with this technology can modify or change the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or changes completed by those with ordinary knowledge in the technical field without departing from the spirit and technical ideas disclosed by the present invention should still be covered by the claims of the present invention.

Claims

1. A valnemulin long-acting compound preparation, characterized in that, It comprises the following components and their mass percentages: valnemulin 5% - 10%, doxycycline 10% - 30%, tartaric acid 0.5% - 2.0%, oil phase 10% - 20%, emulsifier 10% - 30%, co-emulsifier 2% - 10%, and the balance is water.

2. The valnemulin long-acting compound preparation according to claim 1, characterized in that, It is composed of the following components and their mass percentages: valnemulin 10%, doxycycline 30%, tartaric acid 2.0%, oil phase 20%, emulsifier 10%, co-emulsifier 8%, and the balance is water.

3. The valnemulin long-acting compound preparation according to claim 1 or 2, characterized in that, The valnemulin is selected from its hydrochloride or tartrate; the doxycycline is selected from its hydrochloride.

4. The valnemulin long-acting compound preparation according to claim 1 or 2, characterized in that, The mass ratio of valnemulin to doxycycline is 1:3 - 4.

5. The valnemulin long-acting compound preparation according to claim 1 or 2, characterized in that, The oil phase is glyceryl triacetate; the emulsifier is Tween - 80, and the co-emulsifier is propylene glycol.

6. A preparation method of the valnemulin long-acting compound preparation according to any one of claims 1-5, characterized in that, It includes the following process: S1. Add tartaric acid to water, stir evenly, and then add the emulsifier and co-emulsifier thereto, and continue to mix and stir to make it evenly mixed to obtain mixture I. S2. Add valnemulin and doxycycline to the mixture I obtained in step S1, stir and mix evenly to completely dissolve the drugs to obtain mixture II. S3. Add the oil phase to the mixture II obtained in step S2, stir and mix evenly to obtain mixture III. S4. Perform high-speed shearing treatment on the mixture III obtained in step S3 to obtain the product.

7. The preparation method according to claim 6, characterized in that, The condition for the uniform stirring in step S1 is to stir at a speed of 300 - 800 rpm for 10 - 15 min; the condition for the continued mixing and stirring is to stir at a speed of 500 - 800 rpm for 15 - 20 min.

8. The preparation method according to claim 6, characterized in that, The condition for the uniform stirring and mixing in steps S2 and S3 is to stir at a speed of 500 - 800 rpm for 15 - 25 min.

9. The preparation method according to claim 6, wherein The condition for the high-speed shearing in step S4 is to perform homogeneous shearing at a speed of 7000 - 9000 rpm for 15 - 20 min.

10. Use of a valnemulin long-acting compound preparation according to any one of claims 1 - 5 in the preparation of veterinary drugs suitable for high-efficiency breeding modes.

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