A high-bioavailability tyvalocin dry suspension and its preparation method

Abstract

The present invention relates to the technical field of preparing the antibiotic tylvalosin, and provides a high-bioavailability tylvalosin dry suspension and preparation method. The preparation method comprises 5-20 parts of tylvalosin tartrate, 10-40 parts of a suspending agent, 5-20 parts of a surfactant, 10-20 parts of a carrier dispersing material, and an appropriate amount of purified water. The preparation method comprises the following steps: uniformly dispersing the formulated suspending agent with an appropriate amount of purified water, adjusting the pH to acidic with hydrochloric acid, adding a surfactant dissolved in 95% ethanol, then dissolving the drug in hydrochloric acid at pH 2.0 and adding the surfactant to the suspension, grinding the suspension in a wet nano-grinder to obtain a drug-loaded nanosuspension, adding a carrier dispersing material, and spray drying. The drug of the present invention is suspended in water for a long time at the nanoscale, releasing little drug in simulated gastric fluid but slowly in simulated intestinal fluid, thereby extending the drug's duration of action in the intestine. The surfactant increases the drug's solubility and permeability.

Description

Technical Field

[0001] The present invention relates to the technical field of preparation of tylvalosin antibiotics, and in particular to a tylvalosin dry suspension with high bioavailability and a preparation method thereof. Background Art

[0002] Tylvalosin, a macrolide antibiotic developed by Eco Animal Health Products Ltd. in the UK, is commonly used as its tartrate salt. It is a derivative of tylosin, derived from tylosin through microbial transformation, significantly improving its in vivo antimicrobial activity. Its broad antimicrobial spectrum, high antimicrobial activity, and minimal toxicity and side effects make it a commonly used antibiotic for the prevention and treatment of acute and chronic respiratory and digestive tract diseases. Studies have shown (Liu Li, Preliminary Study on Factors Affecting the Oral Bioavailability of Tylvalosin Tartrate, 2015) that tylvalosin tartrate is rapidly absorbed after a single oral dose in broiler chickens, but its absolute oral bioavailability is low, at only 12.67%. The oral bioavailability of tylvalosin tartrate is primarily influenced by intestinal and hepatic metabolism.

[0003] Dry suspensions combine poorly soluble solid drugs with suitable excipients to form a powder or granules. Upon use, they are dispersed into a suspension by adding water and shaking for oral administration. Compared to premixes or powders, dry suspensions disperse more evenly in water, avoiding the problems of uneven mixing during use, which can lead to suboptimal efficacy or localized toxicity caused by excessive concentrations. Furthermore, dry suspensions have a larger surface area than premixes, resulting in faster absorption, higher bioavailability, and more pronounced efficacy.

[0004] Bentonite is a biocompatible clay mineral with montmorillonite as its primary mineral component, ranging from 40% to 90%. It also contains smaller amounts of kaolinite, allophane, chlorite, opal, and mica. Montmorillonite has a monoclinic lattice structure, consisting of two layers of silicon-oxygen tetrahedrons sandwiched between aluminum-oxygen octahedra. It possesses ion exchange capacity. Bentonite is odorless, extremely fine, and has a variegated earthy appearance. It is insoluble in water and organic solvents. Bentonite is widely used in industrial and agricultural production, serving as a catalyst carrier, desiccant, filter, decolorizer, feed additive, and soil conditioner. In the pharmaceutical industry, it is primarily used as an excipient. Its water absorption, suspension, dispersibility, adhesion, and thixotropy make it an excellent pharmaceutical excipient, capable of emulsification, thickening, suspending, and adsorption. Bentonite is a suspending agent with excellent performance in suspensoids. It has a negative charge after absorbing water and can stop the aggregation of particles. Studies have shown that bentonite is used as a suspending agent for calamine lotion, and compared with CMC-Na and sodium alginate at the same concentration, the results show that the indexes such as its sedimentation volume ratio, Brownian motion of particles, redispersibility, caking situation, and particle size are all better than the latter two. In addition, bentonite, due to its high degree of dispersion and characteristics such as non-uniform electrical distribution, has good adsorptivity and produces stronger selective adsorption to some viruses, pathogens and toxins in the digestive tract, and plays a certain synergistic effect to medicines such as antibiotics and pesticides. However, too strong adsorptivity also causes the drug molecule release degree loaded in the bentonite to be lower, thereby having a negative effect on treatment. Therefore, the present invention screens a kind of surfactant to remove the incomplete shortcoming of drug release, which helps to release the drug from the drug bentonite complex under higher pH conditions.

[0005] Phosphatidylcholine (PC) is a biocompatible substance rich in cell membranes. It is an amphiphilic substance composed of a hydrophilic zwitterionic head group and a hydrophobic hydrocarbon chain. Due to its excellent amphiphilicity, PC is often used as an emulsifier, solubilizer or lipid carrier material in preparations. Under physiological conditions, PC is a neutral molecule because the positively charged quaternary amine and the negatively charged phosphate group of the hydrophilic zwitterionic head group have similar charges. When pH < 3, PC becomes positively charged due to the protonation of the negatively charged phosphate group. Due to its pH-dependent charge conversion properties, PC combines with bentonite through electrostatic interaction under acidic conditions.

[0006] After entering the intestine, PC can help the drug dissolve from bentonite. Therefore, the present invention will verify the effect of adding PC on the dissolution of the drug from the complex through experiments. Summary of the Invention

[0007] To solve the problems existing in the background technology, the present invention provides a high-bioavailability tyvalocin dry suspension, which comprises the following components by mass: 5-20 parts of tyvalocin tartrate, 10-40 parts of a suspending agent, 5-20 parts of a surfactant, 10-20 parts of a carrier dispersing material, and 1-10 parts of purified water.

[0008] In a preferred embodiment, the suspending agent is selected from silicates, and the silicate suspending agent is selected from one or more of the following: magnesium aluminum silicate, aluminum silicate, sodium bentonite, calcium bentonite, hydrogen bentonite, and organic bentonite.

[0009] In a preferred embodiment, the surfactant is selected from natural amphiphilic surfactant phosphatidylcholine, including one or both of soybean phosphatidylcholine and egg yolk phosphatidylcholine.

[0010] In a preferred embodiment, the carrier dispersing material is selected from one or more of maltodextrin, corn starch, and microcrystalline cellulose.

[0011] A high-bioavailability tyvalocin dry suspension and a preparation method thereof, comprising the following steps:

[0012] S1: Disperse the formulated suspending agent evenly with an appropriate amount of purified water, and adjust the pH value to 2.0 with 0.1 mol / L hydrochloric acid.

[0013] Obtaining an acidic suspending agent suspension;

[0014] S2: Dissolve the surfactant in a small amount of 95% ethanol, add it to the suspending agent suspension, and stir for 10-20 minutes to obtain a suspending agent-surfactant complex;

[0015] S3: Dissolve the formulated drug in a hydrochloric acid solution at pH 2.0, slowly add it to the suspending agent-surfactant complex, and grind it in a wet nano-mill at 3000 rpm for 3-5 cycles to obtain a drug-loaded nanosuspension;

[0016] S4: Add carrier dispersing material to the drug-loaded nanosuspension and mix well, then spray dry. Set the air inlet temperature to 150-180°C and the air outlet temperature to 80-100°C to obtain the product.

[0017] The beneficial effects achieved by the present invention are:

[0018] To address the problem of low oral bioavailability of conventional tyvalocin tartrate premixes, the present invention provides a nano dry suspension of tyvalocin tartrate and a preparation method. Bentonite is used to efficiently load the drug, and the drug is firmly bound to the bentonite layered structure through electrostatic adsorption. After adding a surfactant and a carrier dispersing material, a nano grinder is used to form a uniform nanocomposite with the drug, suspending agent, and surfactant. The addition of the surfactant phosphatidylcholine optimizes the drug's adsorption and in vitro dissolution rate, accelerating the drug's dissolution rate and making the dissolution more thorough. Furthermore, the surfactant micelles significantly solubilize and promote absorption of the drug, thereby facilitating drug absorption at the target site and significantly improving the drug's oral bioavailability. The dry suspension prepared by the present invention has an appearance of off-white to brown powder with good fluidity. After resuspending in water, a white suspension is obtained. The particle size is 500-800 nm, the polydispersity index (PDI) is 0.23-0.32, the zeta potential is -12.0-15.3 mV, and the drug loading is 12.5-17.6%. The preparation has good hydrophilicity and can be stably suspended in water for more than 3 hours after contact with water. It can be quickly dispersed in water and has good palatability, meeting the requirements of drug administration in drinking water for livestock and poultry, and is easy to use.

[0019] The dry suspension of the present invention has pH-dependent drug release characteristics and sustained release characteristics. Under simulated gastric acid conditions, the total drug release is less than 20%, while under simulated intestinal pH conditions, the drug is slowly and controllably released, which is beneficial to increasing the drug concentration and action time at the absorption site, thereby improving the oral bioavailability of the drug.

[0020] The high-bioavailability tylvalosin dry suspension of the present invention has a bioavailability increased by 2.3 times and 2 times compared with conventional tylvalosin premix and ordinary tylvalosin dry suspension, respectively. The nanoformulation significantly improves the oral absorption efficiency of tylvalosin. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 : Scanning electron micrographs of tylvalosin tartrate nano-dry suspension: bentonite (a), phosphatidylcholine (b), tylvalosin tartrate (c), drug-bentonite-PC spray-dried composite (d), physical mixture of drug, bentonite, and PC;

[0022] Figure 2 : Transmission electron microscopy images of the suspension and PC nanoparticles formed after reconstitution of tyvalocin tartrate nano dry suspension (F3);

[0023] Figure 3 : Comparison of the storage time after reconstitution of Tylvalosin tartrate nano dry suspension (F3) and ordinary dry suspension;

[0024] Figure 4 : Schematic diagram of bentonite-PC-drug complex and drug release mechanism;

[0025] Figure 5 :In vitro drug release results of tyvalosin tartrate nano dry suspension;

[0026] Figure 6 : Pharmacokinetic test results of tartrate tyvalosin nano dry suspension in Example 3. DETAILED DESCRIPTION

[0027] The technical solutions of the present invention will be described clearly and completely below in conjunction with the drawings in the present invention. In addition, the forms of the various structures described in the following embodiments are merely examples. The present invention is not limited to the various structures described in the following embodiments. All other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0028] Example 1

[0029] A tylvalosin tartrate nano-dry suspension with high oral bioavailability, the composition of which is formed by mixing the components by mass is as follows (Formula 1):

[0030] (1) 10g of tartrate tyvalosin,

[0031] (2) 10g sodium bentonite,

[0032] (3) Soybean phosphatidylcholine (PC) 0g,

[0033] (4) 30g maltodextrin,

[0034] (5) Add purified water to 500 ml.

[0035] The preparation steps are as follows:

[0036] (1) Dispersing the formulated amount of sodium bentonite uniformly with an appropriate amount of purified water, and adjusting the pH value to 2.0 with 0.1 mol / L hydrochloric acid to obtain an acidic bentonite suspension;

[0037] (2) Dissolve soybean phosphatidylcholine in a small amount of 95% ethanol, add it to the bentonite suspension, and stir for 10 to 20 minutes to obtain a bentonite-PC complex;

[0038] (3) Dissolve the formulated amount of tylvalosin tartrate API in a pH 2.0 HCl solution, slowly add it to the bentonite-PC composite, and grind it in a wet nano-mill at 3000 rpm for 3-5 cycles to obtain a drug-loaded nanosuspension;

[0039] (4) Add maltodextrin to the drug-loaded nanosuspension and mix well, then spray dry (inlet air temperature 150-180°C, outlet air temperature 80-100°C) to obtain the product.

[0040] Example 2

[0041] In this embodiment, the dry blend components are mixed by mass to form a formulation as follows (Formula 2):

[0042] (1) 20g of tylvalosin tartrate,

[0043] (2) 40g calcium bentonite,

[0044] (3) Soy phosphatidylcholine (PC) 5g,

[0045] (4) 30g microcrystalline cellulose,

[0046] (5) Add purified water to 500 ml.

[0047] The preparation steps are as follows:

[0048] (1) Dispersing the formulated amount of calcium bentonite uniformly with an appropriate amount of purified water, and adjusting the pH value to 2.0 with 0.1 mol / L hydrochloric acid to obtain an acidic bentonite suspension;

[0049] (2) Dissolve soybean phosphatidylcholine in a small amount of 95% ethanol, add it to the bentonite suspension, and stir for 10 to 20 minutes to obtain a bentonite-PC complex;

[0050] (3) Dissolve the formulated amount of tylvalosin tartrate API in a pH 2.0 HCl solution, slowly add it to the bentonite-PC composite, and grind it in a wet nano-mill at 3000 rpm for 3-5 cycles to obtain a drug-loaded nanosuspension;

[0051] (4) Add microcrystalline cellulose to the drug-loaded nanosuspension and mix well, then spray dry (inlet air temperature 150-180°C, outlet air temperature 80-100°C) to obtain the product.

[0052] Example 3

[0053] In this embodiment, the dry blend components are mixed by mass to form a formulation as follows (Formula 3):

[0054] (1) 20g of Tylvalosin tartrate,

[0055] (2) 50g sodium bentonite,

[0056] (3) Egg yolk phosphatidylcholine (PC) 10g,

[0057] (4) 30g maltodextrin,

[0058] (5) Add purified water to 500 ml.

[0059] The preparation steps are as follows:

[0060] (1) Dispersing the formulated amount of sodium bentonite uniformly with an appropriate amount of purified water, and adjusting the pH value to 2.0 with 0.1 mol / L hydrochloric acid to obtain an acidic bentonite suspension;

[0061] (2) Dissolve egg yolk phosphatidylcholine in a small amount of 95% ethanol, add it to the bentonite suspension, and stir for 10 to 20 minutes to obtain a bentonite-PC complex;

[0062] (3) Dissolve the formulated amount of tylvalosin tartrate API in a pH 2.0 HCl solution, slowly add it to the bentonite-PC composite, and grind it in a wet nano-mill at 3000 rpm for 3-5 cycles to obtain a drug-loaded nanosuspension;

[0063] (4) Add maltodextrin to the drug-loaded nanosuspension and mix well, then spray dry (inlet air temperature 150-180°C, outlet air temperature 80-100°C) to obtain the product.

[0064] Example 4

[0065] In this embodiment, the dry blend components are mixed by mass to form a formulation as follows (Formula 4):

[0066] (1) 20g of Tylvalosin tartrate,

[0067] (2) 40g of organic bentonite,

[0068] (3) Soy phosphatidylcholine (PC) 15g,

[0069] (4) 30g corn starch,

[0070] (5) Add purified water to 500 ml.

[0071] The preparation steps are as follows:

[0072] (1) Dispersing the formulated amount of organobentonite uniformly with an appropriate amount of purified water, and adjusting the pH value to 2.0 with 0.1 mol / L hydrochloric acid to obtain an acidic bentonite suspension;

[0073] (2) Dissolve soybean phosphatidylcholine in a small amount of 95% ethanol, add it to the bentonite suspension, and stir for 10 to 20 minutes to obtain a bentonite-PC complex;

[0074] (3) Dissolve the formulated amount of tylvalosin tartrate API in a pH 2.0 HCl solution, slowly add it to the bentonite-PC composite, and grind it in a wet nano-mill at 3000 rpm for 3-5 cycles to obtain a drug-loaded nanosuspension;

[0075] (4) Add corn starch to the drug-loaded nanosuspension and mix well, then spray dry (inlet air temperature 150-180°C, outlet air temperature 80-100°C) to obtain the product.

[0076] Example 5

[0077] Preparation of Tylvalosin Tartrate Ordinary Dry Suspension

[0078] The recipe is as follows (Recipe 5):

[0079] (1) 20g of Tylvalosin tartrate,

[0080] (2) 10g of tartaric acid,

[0081] (3) Hydroxypropyl methylcellulose 5g,

[0082] (4) 4g xanthan gum,

[0083] (5) Silicon dioxide 0.5g

[0084] (6) Add anhydrous glucose to 100g

[0085] The preparation method is as follows:

[0086] The raw materials, suspending agents, cosolvents, dispersants, fillers and other materials are mixed evenly, 30% ethanol is added to granulate, and the mixture is dried at 60°C and crushed into 80-100 meshes using a grinder to obtain the product.

[0087] Example 6

[0088] Evaluation of different formulations of tyvalocin tartrate dry suspension preparations

[0089] The physical and chemical properties of Formulations 1 to 5, including appearance, fluidity, hydration dispersibility, morphology, particle size, potential, and drug loading, were investigated. The results are shown in the following table:

[0090] Table 1 Physicochemical properties of tylvalosin tartrate dry suspension preparations with different formulations

[0091]

[0092] The electron microscope photos of the raw materials and auxiliary materials used in the preparation of tartrate tyvalosin are shown in Figure 1 As shown in Table 1, the Tylvalosin tartrate nano dry suspension has good fluidity and good water dispersibility after reconstitution (see Figure 2 ), record the initial height (H0) of the hydrated dispersible suspension of the nano dry suspension, let it stand for 3 hours, record the final height (H) of the suspension, and use H / H0 to obtain the sedimentation volume ratio F. The larger the F value, the more stable the suspension. Figure 3As shown, the stability of the tyvalosin tartrate nano dry suspension in water after reconstitution is better than that of the ordinary dry suspension, indicating that the nano dry suspension can be administered through drinking water and will not clog the water line during use.

[0093] Example 7

[0094] In vitro drug release of tyvalosin tartrate nanoparticles dry suspension

[0095] The prescribed amount of tylvalosin tartrate nano-dry suspension and tylvalosin premix were subjected to the dissolution and release assay method described in the Chinese Veterinary Pharmacopoeia using 900 ml of 0.1 mol / L hydrochloric acid solution and 900 ml of pH 6.8 phosphate buffer, respectively, at 75 rpm. The solution was filtered at 0, 0.5, 1, 2, 4, 6, 8, and 12 h. An appropriate amount of the filtrate was accurately measured and diluted with mobile phase to a solution containing approximately 100 μg per 1 ml. This served as the test solution. Separately, a precisely weighed tylvalosin tartrate reference standard was diluted with mobile phase to a solution containing approximately 100 μg per 1 ml. This served as the reference standard solution. The dissolution rate of each batch of tylvalosin tartrate nano-dry suspension was determined by HPLC injection. Chromatographic conditions: C18 column, acetonitrile-0.15 mol / L ammonium acetate solution-acetic acid (45:45:10) as mobile phase, detection wavelength 280 nm.

[0096] The results are as follows Figure 5 As shown in Figure 2, tyvalosin is released slowly under conditions of low pH simulated gastric acid, while the release is significantly accelerated under conditions of simulated intestinal fluid. This is because at low pH values, the tertiary amine groups on the tyvalosin molecule are protonated, and the drug interacts strongly with the ions in the bentonite layered molecules, resulting in a low drug release rate. When the pH is raised to neutral conditions, the tertiary amine groups are deprotonated, the polarity of the drug decreases, and the interaction between the drug and the bentonite complex is weakened, thereby increasing drug release. The addition of PC makes the drug dissolve faster and more completely under intestinal pH conditions. The complexation of the drug with phospholipids can improve the dissolution rate on the one hand, and on the other hand, it is beneficial to the transmembrane absorption at the target site, thereby improving the absorption and utilization of the drug (see Figure 4 ).

[0097] Example 8

[0098] Eighteen healthy, 30-day-old piglets were randomly divided into three groups, with six piglets in each group. All piglets were fasted for 12 hours before dosing. Group one received an oral tylvalosin tartrate nanoparticle suspension, group two received an oral tylvalosin tartrate conventional dry suspension, and group three received an oral tylvalosin tartrate premix. All three groups received a single gavage of 25 mg / kg of tylvalosin. Blood samples were collected at predetermined time points after dosing. The collected blood was placed in tubes containing sodium heparin at room temperature for 2 hours and then centrifuged at 3000 rpm for 10 minutes. Plasma was separated, and the supernatant was frozen at -20°C for analysis. Plasma concentrations of tylvalosin tartrate were determined by liquid chromatography-mass spectrometry (LC / MS / MS) according to the literature (Ludi et al., Pharmacokinetic Study of Tylvalosin Tartrate Premix in Piglets). Pharmacokinetic parameters were calculated using DAS 2.0 software.

[0099] The results of the system suitability test showed that the chromatographic peaks of tyvalosin were well separated, and the precision and recovery rate met the detection requirements. Figure 6 As shown in Figure 2, oral tartrate premix was rapidly absorbed, reaching an average maximum plasma concentration (Cmax) of 243.2±165.1 ng·mL in 1.61 h. -1 The average area under the concentration-time curve (AUC) was 2788.2±1024.3ng / mL·h. The average peak time of the ordinary dry suspension was delayed to 2.48h, which was related to the sustained release effect of the drug. Its Cmax and AUC were 235.0±189.4ng·mL -1 and 3177.4±1933.5ng / mL·h. In comparison, the absorption peak of the nanosuspension group was slow, with Tmax reaching 6.63h, which is related to the pH-dependent release of the drug in the body, showing a good correlation between in vitro and in vivo drug release. The Cmax and AUC of the tartrate tyvalosin nano dry suspension group were 258.6±133.5ng·mL -1 and 6410.5±2490.0ng / mL·h.

[0100] Compared with the conventional premix and conventional dry suspension, the nanodry suspension group showed significantly enhanced area under the drug-time curve (AUC) and mean residence time (MRT), indicating a prolonged duration of drug action, slower clearance, and enhanced absorption. Compared with the conventional premix, the relative bioavailability of the nanodry suspension and conventional dry suspension was 229% and 113%, respectively, indicating that dry suspensions formulated with bentonite-PC as a carrier can significantly enhance oral bioavailability. The sustained-release capacity and PC-mediated drug enhancement facilitate higher drug absorption. Tylvalosin is a time-dependent drug, with a prolonged duration of action, which is beneficial for its antibacterial activity in vivo. Encapsulating this drug in lipid-containing nanocarriers can increase its blood circulation time, thereby enhancing therapeutic efficacy.

[0101] Table 2 Pharmacokinetic parameters

[0102]

[0103] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A high bioavailability tyvalocin dry suspension, characterized in that: It comprises the following components by mass: (1) 5-20 parts of tylvalosin tartrate, (2) 10-40 parts of suspending agent, (3) 5-20 parts of surfactant, (4) 10-20 parts of carrier dispersion material, (5) 1-10 parts of purified water; The suspending agent is selected from one or more of the following: sodium bentonite, calcium bentonite, and organic bentonite; The surfactant is selected from natural amphiphilic surfactant phosphatidylcholine, including one or two of soybean phosphatidylcholine and egg yolk phosphatidylcholine.

2. The high-bioavailability tyvalocin dry suspension according to claim 1, characterized in that: The carrier dispersing material is selected from one or more of maltodextrin, corn starch and microcrystalline cellulose.

3. The method for preparing a high-bioavailability tyvalocin dry suspension according to claim 1 or 2, wherein: It includes the following steps: S1: Disperse the formulated suspending agent evenly with an appropriate amount of purified water, and adjust the pH to 2.0 with 0.1 mol / L hydrochloric acid to obtain an acidic suspending agent suspension; S2: Dissolve the surfactant in a small amount of 95% ethanol, add it to the suspending agent suspension, and stir for 10-20 minutes to obtain a suspending agent-surfactant complex; S3: Dissolve the formulated drug in a hydrochloric acid solution at pH 2.0, slowly add it to the suspending agent-surfactant complex, and grind it in a wet nano-mill at 3000 rpm for 3-5 cycles to obtain a drug-loaded nanosuspension; S4: Add carrier dispersing material to the drug-loaded nanosuspension and mix well, then spray dry. Set the air inlet temperature to 150-180°C and the air outlet temperature to 80-100°C to obtain the product.

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