Preparation method and application of antibiotic delivery system

Abstract

The present application proposes a preparation method and application of an antibiotic delivery system, which relates to the technical field of medical excipients. The present application specifically adjusts various pharmaceutical excipients, such as soybean oil, phospholipids, glycerol and other materials, to prepare an emulsion that can be loaded with antibiotics, and uses it as an antibiotic delivery system, so that the antibiotic delivery system can efficiently load antibiotics, significantly improve the bactericidal efficiency, and has good atomization inhalation performance and good drug stability, so that the antibiotic delivery system has a sustained-release effect. Compared with the inhaled antibiotic solution dosage form, the antibiotic-loaded emulsion can be retained in the mouse lung tissue for a longer time after inhalation administration to mice, significantly improving the efficiency of lung drug delivery.

Description

Technical Field

[0001] The present application relates to the technical field of medical excipients, and in particular to a preparation method and application of an antibiotic delivery system. Background Art

[0002] Currently, there is a severe shortage of inhaled antibiotics for clinical use. The only inhaled antibiotic approved for clinical use in China is tobramycin inhalation solution. Furthermore, several antibiotics, such as rifampicin and polymyxin, have been used off-label for inhalation administration. Therefore, the development of antibiotic formulations specifically for inhalation is urgently needed.

[0003] Existing inhaled antibiotics primarily consist of solution-based formulations, and the only marketed nanoformulation for inhalation is amikacin inhalation liposomal suspension. However, solution-based inhaled antibiotics suffer from the disadvantages of rapid bloodstream entry, low lung retention, high dosages, and limited antibacterial efficacy.

[0004] Based on this, a new antibiotic delivery system is urgently needed. Summary of the Invention

[0005] The present application provides a preparation method and application of an antibiotic delivery system to solve the problems of existing inhaled antibiotics such as rapid entry into the blood, low lung retention, large dosage, and insufficient antibacterial efficiency.

[0006] In a first aspect, the present application provides a method for preparing an antibiotic delivery system, the method comprising the following steps:

[0007] Step 1: uniformly mixing 1-100 mg of soybean oil and 1-100 mg of medium-chain triglycerides to prepare an oil phase;

[0008] Step 2: adding 1 to 10 mg of antibiotics, 1 to 12 mg of phospholipids, and 1 to 25 mg of glycerol into a sterile liquid and dissolving them to prepare an aqueous phase, wherein the antibiotics are water-soluble antibiotics;

[0009] Step 3: uniformly mixing the oil phase and the aqueous phase to obtain a mixture, and after ultrasonic treatment, obtaining an initial emulsion, wherein each milliliter of the mixture contains 1 to 100 mg of soybean oil, 1 to 100 mg of medium-chain triglycerides, 1 to 10 mg of antibiotics, 1 to 12 mg of phospholipids, and 1 to 25 mg of glycerol;

[0010] Step 4: The initial emulsion is subjected to high-pressure homogenization to obtain an antibiotic-loaded emulsion as an antibiotic delivery system.

[0011] Optionally, in step 2, the sterile liquid is at least one of water for injection, deionized water, physiological saline, glucose injection, citrate buffer and phosphate buffer.

[0012] Optionally, in step 2, the antibiotic is at least one of polymyxin E sodium methanesulfonate, polymyxin B sulfate, tobramycin and amphotericin B.

[0013] Optionally, in step 3, the power of the ultrasonic treatment is 50 watts to 350 watts, and the ultrasonic treatment time is 1 minute to 20 minutes.

[0014] Optionally, in step 4, the pressure of the high-pressure homogenization treatment is 200 bar to 1200 bar, the temperature of the high-pressure homogenization treatment is 4° C. to 25° C., and the time of the high-pressure homogenization treatment is 2 minutes to 30 minutes.

[0015] Optionally, the particle size of the prepared antibiotic-loaded emulsion is less than 1 micron, preferably less than 300 nanometers, and most preferably less than 100 nanometers.

[0016] In a second aspect, the present application provides an antibiotic drug delivery system, which is prepared by any one of the preparation methods described in the first aspect.

[0017] In a third aspect, the present application proposes a use of the antibiotic delivery system described in the second aspect in the preparation of inhaled antibiotic drugs.

[0018] Optionally, the administration route of the antibiotic delivery system includes at least intravenous administration, subcutaneous administration, and oral administration.

[0019] The present application has the following advantages: The present application proposes a method for preparing and applying an antibiotic delivery system. The present application specifically adjusts various inhalable pharmaceutical excipients, such as soybean oil, phospholipids, and glycerol, to prepare an antibiotic-loaded emulsion, which is used as an antibiotic delivery system. This antibiotic delivery system can efficiently load antibiotics, exhibits good atomization and inhalation performance, and has good drug stability, resulting in a sustained-release effect. Compared with inhaled antibiotic solution formulations, the antibiotics in the antibiotic delivery system can be retained in mouse lung tissue for a longer period of time, significantly improving the efficiency of pulmonary drug delivery. Furthermore, the antibiotic delivery system prepared in the present application can significantly enhance the antibacterial activity of antibiotics compared to free antibiotics. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the following briefly introduces the drawings required for use in the description of the embodiments of the present application. Obviously, the drawings described below are only some embodiments of the present application. For ordinary technicians in this field, other drawings can be obtained based on these drawings without any creative work.

[0021] Figure 1This is a transmission electron micrograph of an antibiotic drug delivery system loaded with polymyxin B sulfate provided in an embodiment of the present application;

[0022] Figure 2 Transmission electron microscopy image of a tobramycin-loaded antibiotic delivery system provided in an embodiment of the present application;

[0023] Figure 3 Transmission electron microscopy image of an antibiotic drug delivery system loaded with amphotericin B provided in an embodiment of the present application;

[0024] Figure 4 Transmission electron microscopy image of an antibiotic drug delivery system loaded with polymyxin E sodium methanesulfonate provided in an embodiment of the present application;

[0025] Figure 5 This is a quantitative detection image of lung tissue of mice 48 hours after inhalation of an antibiotic delivery system loaded with polymyxin B sulfate provided in an embodiment of the present application;

[0026] Figure 6 This is an antibacterial efficacy diagram of an antibiotic delivery system loaded with polymyxin B sulfate against carbapenem-resistant Acinetobacter baumannii provided in an embodiment of the present application. DETAILED DESCRIPTION

[0027] The following will be combined with the drawings in the embodiments of this application to clearly and completely describe the technical solutions in the embodiments of this application. Obviously, the embodiments described are part of the embodiments of this application, not all of them. Based on the embodiments in this application, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of this application.

[0028] Currently, there is a severe shortage of inhaled antibiotics for clinical use. The only inhaled antibiotic approved for clinical use in China is tobramycin inhalation solution. Furthermore, several antibiotics, such as rifampicin and polymyxin, have been used off-label for inhalation administration. Therefore, the development of a dedicated inhaled antibiotic delivery system is urgently needed.

[0029] Existing inhaled antibiotics are primarily solution-based, with the only marketed nanoformulation for inhalation being amikacin liposomal suspension. However, solution-based inhaled antibiotics suffer from the disadvantages of rapid bloodstream entry, low lung retention, high dosages, and limited antibacterial efficacy.

[0030] Based on this, the present application proposes a method for preparing an antibiotic delivery system, which utilizes various inhalable pharmaceutical excipients in the prescription, such as soybean oil, phospholipids, glycerol and other materials, to prepare an antibiotic delivery system that can be loaded with antibiotics.

[0031] In a first aspect of an embodiment of the present application, a method for preparing an antibiotic delivery system is provided, the method comprising the following steps:

[0032] Step 1: uniformly mixing 1-100 mg of soybean oil and 1-100 mg of medium-chain triglycerides to prepare an oil phase;

[0033] Step 2: adding 1 to 10 mg of antibiotics, 1 to 12 mg of phospholipids, and 1 to 25 mg of glycerol into a sterile liquid and dissolving them to prepare an aqueous phase, wherein the antibiotics are water-soluble antibiotics;

[0034] Step 3: uniformly mixing the oil phase and the aqueous phase to obtain a mixture, and after ultrasonic treatment, obtaining an initial emulsion, wherein each milliliter of the mixture contains 1 to 100 mg of soybean oil, 1 to 100 mg of medium-chain triglycerides, 1 to 10 mg of antibiotics, 1 to 12 mg of phospholipids, and 1 to 25 mg of glycerol;

[0035] Step 4: The initial emulsion is subjected to high-pressure homogenization to obtain an antibiotic-loaded emulsion as an antibiotic delivery system.

[0036] Specifically, in order to prepare an emulsion that efficiently loads antibiotics, the preparation method proposed in this application first prepares an oil phase. The oil phase components selected in this application are soybean oil and medium-chain triglycerides (MCT). Soybean oil and medium-chain triglycerides serve as the oil phase of the emulsion to maintain the stability of the emulsion particles, form a large specific surface area, and facilitate drug loading. In addition, positively charged antibiotic molecules can bind to negatively charged emulsion particles through positive and negative charge adsorption, intermolecular forces, and hydrogen bonds, thereby improving drug stability and forming antibiotic nanoaggregates, which help enhance the antibacterial ability of the antibiotics. Furthermore, the unsaturated fatty acids in soybean oil also have a certain antibacterial effect. Therefore, the antibiotic-loaded emulsion prepared from this oil phase, as a drug delivery system, significantly improves its antibacterial ability through a multi-faceted combined mechanism of action compared to free antibiotics.

[0037] Specifically, in step 2, during the preparation of the aqueous phase, the present application adds antibiotics, preferably water-soluble antibiotics, to the aqueous phase to promote complete dissolution of the antibiotic in the aqueous phase and increase the antibiotic loading rate. Components in the aqueous phase are preferably phospholipids and glycerol. Phospholipids act as natural surfactants, reducing oil-water interfacial tension, forming a strong emulsion film, promoting drug absorption, and solubilizing poorly soluble drugs. The resulting phospholipid complex exhibits significantly different physicochemical and biological properties from the original compound, increasing solubility and biomembrane permeability, and playing a significant role in the preparation of aerosol inhalation emulsions. Glycerol, as a co-emulsifier, increases the solubility of the emulsifier, assists the emulsifier in reducing the interfacial tension between the oil and water, enhances the fluidity of the interfacial film, and regulates the HLB value (hydrophilic-lipophilic balance) of the emulsifier, enabling the spontaneous formation of nanoemulsion droplets. The addition of glycerol can increase the antibiotic loading capacity, enhance antibiotic solubility, and expand the range of nanoemulsion formation.

[0038] Among them, the phospholipid can be selected from egg yolk lecithin, dipalmitoylphosphatidylcholine (DPPC), soybean lecithin, etc., preferably egg yolk lecithin.

[0039] In an optional embodiment of the present application, the sterile liquid used in the above step 2 can be any at least one of water for injection, deionized water, physiological saline, glucose injection, citrate buffer and phosphate buffer.

[0040] In an optional embodiment of the present application, the antibiotic used in the above step 2 can be at least one of polymyxin E sodium methanesulfonate and polymyxin B sulfate, tobramycin and amphotericin B, or antibiotics and drugs with equivalent efficacy.

[0041] Specifically, in the implementation of step 3, in order to uniformly mix the oil phase and the aqueous phase, the present application performs ultrasonic treatment on the mixed oil phase and aqueous phase, which can improve the emulsification efficiency, reduce the amount of emulsifier used, and ensure the stability of the emulsion, forming nano-scale droplets, and improving the release characteristics of the antibiotic, thereby increasing the bioavailability and efficacy of the antibiotic. By controlling the ultrasonic process, such as controlling the frequency and time of the ultrasound and adjusting the dispersion of the emulsion droplets, emulsions of different particle sizes can be prepared, achieving precise control, and no excessively high temperatures are generated during the ultrasonic treatment process, which allows for a wider range of applicable antibiotics, such as the loading of heat-sensitive antibiotics.

[0042] In the above step 3, 1 ml of the mixture obtained by mixing the oil phase and the water phase contains 1-100 mg of soybean oil, 1-100 mg of medium-chain triglycerides, 1-10 mg of antibiotics, 1-12 mg of phospholipids and 1-25 mg of glycerol.

[0043] In an optional embodiment of the present application, in the above step 3, the power of the ultrasonic treatment is 50 watts to 350 watts, which can be 50 watts, 100 watts, 150 watts, 200 watts, 250 watts, 300 watts, 350 watts, etc. The ultrasonic treatment time is 1 minute to 20 minutes, which can be 1 minute, 3 minutes, 5 minutes, 7 minutes, 10 minutes, 13 minutes, 15 minutes, 20 minutes, etc.

[0044] When implementing step 4, in order to further improve the quality of the nanoemulsion, such as stability, production efficiency, and sensory quality, the present application uses high-pressure homogenization to further process the initial nanoemulsion after ultrasonic treatment. High-pressure homogenization can further reduce the size of the emulsion droplets in the initial nanoemulsion, making the emulsion more uniform, thereby improving the physical stability of the emulsion. It can also effectively inactivate microorganisms and enzymes in the emulsion and extend the shelf life of the nanoemulsion; it can further improve the rheological properties of the emulsion, reduce the size of the emulsion droplets, and improve the emulsification performance of the emulsion; atomization inhalation administration has certain requirements for the size of the nanoemulsion. The smaller the size, the more drugs can enter the lungs, increasing the drug retention in the lungs. The use of high-pressure homogenization can more accurately control the characteristics of the emulsion, such as droplet size and distribution. In actual production, when the above-mentioned nanoformulations need to be produced on a large scale, high-pressure homogenization is a good processing method.

[0045] In an optional embodiment of the present application, in the above step 4, the pressure of the high-pressure homogenization treatment is 200 bar to 1200 bar, and can be 200 bar, 400 bar, 600 bar, 800 bar, 1000 bar, 1200 bar, etc. The temperature of the high-pressure homogenization treatment is 4°C to 25°C, and can be 4°C, 8°C, 12°C, 15°C, 18°C, 20°C, 23°C, 25°C, etc. The time of the high-pressure homogenization treatment is 2 minutes to 30 minutes, and can be 2 minutes, 3 minutes, 5 minutes, 7 minutes, 10 minutes, 13 minutes, 15 minutes, 18 minutes, 20 minutes, 23 minutes, 25 minutes, 28 minutes, 30 minutes, etc.

[0046] The antibiotic-loaded emulsion prepared by the above preparation method has a particle size of less than 1 micron, preferably less than 300 nanometers as an antibiotic delivery system, and more preferably less than 100 nanometers as an antibiotic delivery system.

[0047] The preparation method of the antibiotic drug delivery system provided in the first aspect of the embodiment of the present application is to adjust various inhalable pharmaceutical excipients, such as soybean oil, phospholipids, glycerol and other materials in a targeted manner to prepare a nanoemulsion preparation that can load antibiotics as an antibiotic drug delivery system, so that the antibiotic drug delivery system can efficiently load antibiotics and has good atomization inhalation performance and good drug stability, so that the antibiotic drug delivery system has a sustained release effect. Compared with the inhaled antibiotic solution dosage form, the antibiotic drug delivery system can retain the loaded antibiotics in the mouse lung tissue for a longer time, significantly improving the efficiency of lung drug delivery. In addition, the present application first adds water-soluble antibiotics to the aqueous phase and then combines it with oil to improve the loading efficiency of antibiotics. At the same time, the antibiotic drug delivery system prepared by the present application can significantly improve the antibacterial ability of antibiotics compared with free antibiotics.

[0048] Based on the same application concept, in a second aspect of the embodiment of the present application, an antibiotic drug delivery system is proposed, which is prepared by the preparation method described in any one of the above-mentioned first aspects.

[0049] Based on the same application concept, a third aspect of the present application provides a use of the antibiotic delivery system described in the second aspect above in the preparation of an inhaled antibiotic drug. The antibiotic delivery system can be administered via intravenous, subcutaneous, and oral routes. Preferably, intravenous administration is used. More preferably, inhalation administration is used.

[0050] In order to clearly illustrate the antibiotic-loaded emulsion, preparation method and application thereof proposed in the present application, a detailed description will be given below with reference to examples.

[0051] Example 1: Preparation method of an antibiotic delivery system

[0052] Example 1A

[0053] When the loaded antibiotic is polymyxin B sulfate, the preparation method of the antibiotic drug delivery system loaded with polymyxin B sulfate comprises the following steps:

[0054] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0055] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0056] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0057] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0058] Example 1B

[0059] When the loaded antibiotic is tobramycin, the preparation method of the tobramycin-loaded antibiotic drug delivery system comprises the following steps:

[0060] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0061] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol and 3 mg of tobramycin were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and after mixing evenly, an aqueous phase was prepared.

[0062] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of tobramycin was 1 mg / ml.

[0063] Step 4: The initial emulsion was subjected to high-pressure homogenization treatment using a high-pressure homogenizer. The pressure of the high-pressure homogenization treatment was 800 bar, the temperature of the high-pressure homogenization treatment was 4° C., and the time of the high-pressure homogenization treatment was 10 minutes to obtain an antibiotic drug delivery system loaded with tobramycin.

[0064] Example 1C

[0065] When the loaded antibiotic is amphotericin B, the preparation method of the amphotericin B loaded antibiotic drug delivery system comprises the following steps:

[0066] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0067] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of amphotericin B were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed uniformly to prepare an aqueous phase.

[0068] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of amphotericin B was 1 mg / ml.

[0069] Step 4: The initial emulsion was subjected to high-pressure homogenization treatment using a high-pressure homogenizer. The pressure of the high-pressure homogenization treatment was 800 bar, the temperature of the high-pressure homogenization treatment was 4° C., and the time of the high-pressure homogenization treatment was 10 minutes to obtain an antibiotic drug delivery system loaded with amphotericin B.

[0070] Example 1D

[0071] When the loaded antibiotic is polymyxin E sodium methanesulfonate, the preparation method of the antibiotic drug delivery system loaded with polymyxin E sodium methanesulfonate comprises the following steps:

[0072] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0073] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin E sodium methanesulfonate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0074] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin E sodium methanesulfonate was 1 mg / ml.

[0075] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer at a pressure of 800 bar, a temperature of 4° C., and a time of 10 minutes to obtain an antibiotic drug delivery system loaded with polymyxin E sodium methanesulfonate.

[0076] To illustrate the preparation method proposed in the examples of the present application, by specifically adjusting the type of inhalable pharmaceutical excipients, the type and proportion of the added inhalable pharmaceutical excipients, the ratio of the amount of the oil phase and the aqueous phase, as well as the ultrasonic power, homogenization pressure, and time, the antibiotic delivery system prepared can efficiently load antibiotics and has good atomization inhalation performance and high drug stability. The following comparative examples are listed below for comparison with the antibiotic delivery systems prepared by the preparation method proposed in Example 1 (Example 1A-Example 1D).

[0077] Comparative Example 1A

[0078] Comparative Example 1A Compared with Example 1A, the egg yolk lecithin was replaced with soybean lecithin. The remaining operations were the same as those in Example 1A. The specific operations are as follows:

[0079] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0080] Step 2: 3.6 mg of soybean lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0081] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of soy lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0082] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0083] Comparative Example 1B

[0084] Comparative Example 1B is compared with Example 1A, except that egg yolk lecithin is replaced with dipalmitoylphosphatidylcholine. The remaining operations are the same as those in Example 1A. The specific operations are as follows:

[0085] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0086] Step 2: 3.6 mg of DPPC, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0087] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of DPPC was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0088] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0089] Comparative Example 1C

[0090] Comparative Example 1C was compared to Example 1A, except that soybean oil was not added. The remaining operations were the same as those in Example 1A. The specific operations were as follows:

[0091] Step 1: Add 30 mg of medium-chain triglycerides as the oil phase;

[0092] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 1; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0093] Step 3: Add the oil phase from step 1 dropwise to the aqueous phase from step 2 to obtain mixed material 2. Adjust the volume of mixed material 2 to 3 ml with sterile deionized water, vortex mix for 5 minutes, and sonicate in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In mixed material 3, the content of MCT is 10 mg / ml, the content of egg yolk lecithin is 1.2 mg / ml, the content of glycerol is 2.5 mg / ml, and the content of polymyxin B sulfate is 1 mg / ml.

[0094] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0095] Comparative Example 1D

[0096] Comparative Example 1D was performed in the same manner as in Example 1A, except that medium-chain triglycerides were not added. The remaining operations were the same as in Example 1A. The specific operations were as follows:

[0097] Step 1: Add 30 mg of soybean oil as the oil phase;

[0098] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 1; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0099] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 2. The volume of the mixture 2 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0100] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer at a pressure of 800 bar, a temperature of 4° C., and a time of 10 minutes to obtain an antibiotic drug delivery system loaded with polymyxin B sulfate.

[0101] Comparative Example 1E

[0102] Comparative Example 1E was performed in the same manner as in Example 1A, except that egg yolk lecithin was not added. The specific operations were as follows:

[0103] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0104] Step 2: 7.5 mg of glycerol and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0105] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0106] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0107] Comparative Example 1F

[0108] Comparative Example 1F was compared to Example 1A, except that glycerol was not added. The remaining operations were the same as those in Example 1A. The specific operations were as follows:

[0109] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0110] Step 2: 3.6 mg of egg yolk lecithin and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0111] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0112] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer at a pressure of 800 bar, a temperature of 4° C., and a time of 10 minutes to obtain an antibiotic drug delivery system loaded with polymyxin B sulfate.

[0113] Comparative Example 1G

[0114] Comparative Example 1G was performed in the same manner as in Example 1A, except that the ultrasonic treatment parameters were changed. The specific operations were as follows:

[0115] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0116] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0117] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 50 watts for 1 minute to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0118] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0119] Comparative Example 1H

[0120] Comparative Example 1H was performed in the same manner as in Example 1A, except that the high-pressure homogenization parameters were changed. The remaining operations were the same as in Example 1A. The specific operations were as follows:

[0121] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0122] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0123] Step 3: The oil phase from step 1 was added dropwise to the aqueous phase from step 2 to obtain a mixture 3. The volume of the mixture 3 was adjusted to 3 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 10 mg / ml, the proportion of MCT was 10 mg / ml, the proportion of egg yolk lecithin was 1.2 mg / ml, the proportion of glycerol was 2.5 mg / ml, and the proportion of polymyxin B sulfate was 1 mg / ml.

[0124] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer at a pressure of 200 bar, a temperature of 4° C., and a time of 2 minutes to obtain an antibiotic drug delivery system loaded with polymyxin B sulfate.

[0125] Comparative Example II

[0126] Comparative Example 1I is compared with Example 1A, except that the proportions of the various substances are changed. The remaining operations are the same as those of Example 1A. The specific operations are as follows:

[0127] Step 1: 30 mg of soybean oil and 30 mg of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0128] Step 2: 3.6 mg of egg yolk lecithin, 7.5 mg of glycerol, and 3 mg of polymyxin B sulfate were mixed to obtain a mixture 2; 2 ml of sterile deionized water was used to dissolve the mixture 2, and the mixture was mixed evenly to prepare an aqueous phase.

[0129] Step 3: The aqueous phase and the oil phase from step 1 were mixed to obtain a mixture 3. The volume of the mixture 3 was adjusted to 30 ml with sterile deionized water, vortexed for 5 minutes, and sonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion. In the mixture 3, the proportion of soybean oil was 1 mg / ml, the proportion of MCT was 1 mg / ml, the proportion of egg yolk lecithin was 0.12 mg / ml, the proportion of glycerol was 0.25 mg / ml, and the proportion of polymyxin B sulfate was 0.1 mg / ml.

[0130] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0131] Comparative Example 1J

[0132] Comparative Example 1J is compared with Example 1A, except that the proportions of the various substances are changed. The remaining operations are the same as those of Example 1A. The specific operations are as follows:

[0133] Step 1: 3 g of soybean oil and 3 g of medium-chain triglycerides were mixed to obtain a mixture 1 as the oil phase;

[0134] Step 2: Mix 360 mg of egg yolk lecithin, 750 mg of glycerol, and 300 mg of polymyxin B sulfate to obtain a mixture; dissolve mixture 2 in 5 ml of sterile deionized water and mix thoroughly to prepare an aqueous phase. In mixture 3, soybean oil accounts for 100 mg / ml, MCT accounts for 100 mg / ml, egg yolk lecithin accounts for 12 mg / ml, glycerol accounts for 25 mg / ml, and polymyxin B sulfate accounts for 10 mg / ml.

[0135] Step 3: The aqueous phase and the oil phase in step 1 were mixed to obtain a mixture 3, and the volume of the mixture 3 was adjusted to 30 ml with sterile deionized water. The mixture was vortex-mixed for 5 minutes, and ultrasonicated in an ice-water bath using a cell disruptor at a power of 200 watts for 10 minutes to obtain an initial emulsion.

[0136] Step 4: The initial emulsion was subjected to high-pressure homogenization using a high-pressure homogenizer. The pressure of the high-pressure homogenization was 800 bar, the temperature of the high-pressure homogenization was 4° C., and the time of the high-pressure homogenization was 10 minutes to obtain an antibiotic delivery system loaded with polymyxin B sulfate.

[0137] Example 2: Performance Characterization of Antibiotic Delivery System

[0138] First, the morphology of the antibiotic drug delivery system prepared in Example 1A, Example 1B, Example 1C, and Example 1D was characterized. Figures 1 to 4 The prepared antibiotic delivery system was morphologically characterized. The characterization results are shown in Figure 1 , Figure 1 This is a transmission electron microscopy morphology of an antibiotic drug delivery system loaded with polymyxin B sulfate provided in an embodiment of the present application; Figure 2 This is a transmission electron microscopy morphology image of a tobramycin-loaded antibiotic delivery system provided in an embodiment of the present application; Figure 3 This is a transmission electron microscopy image of an antibiotic drug delivery system loaded with amphotericin B provided in an embodiment of the present application; Figure 4 This is a transmission electron microscopy image of a polymyxin sodium methanesulfonate-loaded antibiotic delivery system provided in an example of this application. The antibiotic delivery system has a uniform, spherical morphology. The experimental method used in this example to characterize the performance of the polymyxin-loaded nanoemulsion is the same as that commonly used in the art and will not be repeated here.

[0139] The antibiotic drug delivery systems prepared in Examples 1A to 1D and Comparative Examples 1A to 1J were then characterized for performance. The encapsulation efficiency, drug loading, particle size, PDI (Polymer Dispersion Index), potential, and stability were tested, respectively. The test results are shown in Table 1.

[0140] Table 1 Performance characterization results of the antibiotic delivery systems prepared in Example 1A-Example 1D and Comparative Example 1A-Comparative Example 1J

[0141]

[0142] The results in Table 1 indicate that the antibiotic delivery system prepared using the preparation method of the present application, as an inhalable emulsion, exhibits a high antibiotic loading rate, good aerosol inhalation performance, and excellent stability. The ratio of soybean oil, MCT, and glycerol in the antibiotic delivery system, as well as the ultrasonic power and duration, influence the encapsulation efficiency and drug loading. Higher ultrasonic power can reduce the emulsion's particle size, reaching the nanometer scale. However, excessive ultrasonic power can affect the physical stability of the nanoemulsion, leading to precipitation during storage. An appropriate homogenization pressure and a longer homogenization time contribute to the formation of an emulsion with uniform particle size. Furthermore, it can be seen that the ratio of each substance in the antibiotic delivery system is a key factor in the successful preparation of the delivery system. When the ratio of each substance in the antibiotic delivery system is too low (Comparative Example 1I), the resulting emulsion is unstable and prone to turbid precipitation, making it unsuccessful. When the ratio of each substance in the antibiotic delivery system is higher (Comparative Example 1J), the emulsion exhibits a small amount of precipitation after four months of storage at 4°C, but regains clarity and translucency after shaking and mixing. In general, it is difficult to obtain a drug-loaded emulsion with better stability when a higher proportion of excipients is used.

[0143] Therefore, in the preparation method of the present application, the optimal parameter ratio is the one in Example 1A through the reasonable design and ratio of various parameters. Specifically, in the prepared antibiotic delivery system, the proportion of soybean oil is 10 mg / ml, the proportion of MCT is 10 mg / ml, the proportion of egg yolk lecithin is 1.2 mg / ml, the proportion of glycerol is 2.5 mg / ml, and the proportion of polymyxin B sulfate is 1 mg / ml. The vortex mixing time is 5 minutes, and the cell disruptor is used for ice-water bath ultrasound. The ultrasonic treatment power is 200 watts, the ultrasonic treatment time is 10 minutes, the pressure of the high-pressure homogenization treatment is 800 bar, the temperature of the high-pressure homogenization treatment is 4°C, and the high-pressure homogenization treatment time is 10 minutes. An antibiotic delivery system loaded with polymyxin B sulfate is obtained, which can be stored at 4 degrees Celsius for 6 months with good stability, a dispersion index of 0.153, and an encapsulation efficiency of more than 99%.

[0144] Example 3: Characterization of drug delivery efficiency and drug retention of antibiotic delivery systems

[0145] An antibiotic drug delivery system encapsulating Cy5-labeled polymyxin B sulfate was prepared using the preparation method of Example 1A. Cy5-labeled polymyxin B sulfate was used as the control group. An equal amount of polymyxin B sulfate was quantitatively administered to mice using a mouse micro-atomizer inhalation device. 48 hours after administration, the distribution of fluorescence signals in each group of mice was observed using a small animal in vivo imaging device to analyze the drug delivery efficiency and drug retention in lung tissue. Figure 5 It can be seen that ( Figure 5 The left side shows the fluorescence signal distribution results of isolated mouse organs. Figure 5 On the right are the quantitative results of the fluorescence signals of free polymyxin B sulfate and the polymyxin B sulfate delivery system in mouse lung tissue). The polymyxin B sulfate antibiotic delivery system can significantly enhance the accumulation of antibiotics in mouse lung tissue and prolong the exposure time of antibiotics in mouse lung tissue.

[0146] Example 4: Investigation of the antibacterial activity of the polymyxin B sulfate-loaded antibiotic delivery system against carbapenem-resistant Acinetobacter baumannii

[0147] The preparation method of Example 1A was used to prepare an antibiotic drug delivery system loaded with polymyxin B sulfate, and carbapenem-resistant Acinetobacter baumannii was cultured. The same concentration of polymyxin B sulfate was set in two groups of experiments, and the groups were divided into: control group (Control), free polymyxin B sulfate (PMB group), and polymyxin B sulfate antibiotic drug delivery system group (PMB-NEs group). Acinetobacter baumannii (bacterial count was 1.0×10 5 cfu / ml) for 12 h, and the bacterial growth of the two groups after drug treatment was observed. Figure 6 The results show that the control group has more Acinetobacter baumannii colonies, followed by the PMB group, and the PMB-NEs group has no Acinetobacter baumannii colonies. It can be seen that the PMB-NEs group has the best bactericidal effect compared with the control group (Control) and the PMB group, and can achieve the effect of sterile growth, indicating that the antibiotic delivery system loaded with polymyxin B sulfate can significantly improve the bactericidal effect.

[0148] The above is a detailed introduction to the preparation method and application of an antibiotic delivery system provided by the present application. Specific examples are used herein to illustrate the principles and implementation methods of the present application. The description of the above embodiments is only used to help understand the method and core ideas of the present application. At the same time, for those skilled in the art, according to the ideas of the present application, there may be changes in the specific implementation methods and application scopes. In summary, the content of this specification should not be understood as limiting the present application.

Claims

1. A method for preparing an antibiotic drug delivery system, characterized in that: The method comprises the following steps: Step 1: Mix 30 mg of soybean oil and 30 mg of medium-chain triglycerides to prepare an oil phase; Step 2: adding 3 mg of antibiotics, 3.6 mg of egg yolk lecithin, and 7.5 mg of glycerol into a sterile liquid and dissolving them to prepare an aqueous phase, wherein the antibiotic is a water-soluble antibiotic, and the antibiotic is at least one of polymyxin E sodium methanesulfonate, polymyxin B sulfate, tobramycin, and amphotericin B; Step 3: uniformly mixing the oil phase and the aqueous phase to obtain a mixture, and ultrasonically treating the mixture to obtain an initial emulsion, wherein each milliliter of the mixture contains 10 mg of soybean oil, 10 mg of medium-chain triglycerides, 1 mg of antibiotics, 1.2 mg of phospholipids, and 2.5 mg of glycerol, and the ultrasonic treatment power is 200 watts to 350 watts, and the ultrasonic treatment time is 10 minutes to 20 minutes; Step 4: The initial emulsion is subjected to high-pressure homogenization treatment to obtain an antibiotic-loaded emulsion as an antibiotic delivery system, and the antibiotic delivery system is used as a nebulized inhalation antibiotic. The pressure of the high-pressure homogenization treatment is 800 bar to 1200 bar, the temperature of the high-pressure homogenization treatment is 4°C to 25°C, and the time of the high-pressure homogenization treatment is 10 minutes to 30 minutes.

2. The method for preparing the antibiotic drug delivery system according to claim 1, characterized in that: In step 2, the sterile liquid is at least one of water for injection, deionized water, physiological saline, glucose injection, citrate buffer and phosphate buffer.

3. The method for preparing the antibiotic drug delivery system according to claim 1, characterized in that: The particle size range of the prepared antibiotic-loaded emulsion is less than 1 micron.

4. An antibiotic delivery system, characterized in that: The antibiotic drug delivery system is prepared by the preparation method according to any one of claims 1 to 3.

5. Use of the antibiotic delivery system according to claim 4 in the preparation of inhaled antibiotic drugs.

6. The use according to claim 5, characterized in that The administration routes of the antibiotic delivery system include at least intravenous administration, subcutaneous administration, and oral administration.

Images