Propofol emulsion injection and preparation method thereof

Abstract

This invention discloses a propofol emulsion injection and its preparation method, belonging to the field of biomedical technology. The method involves preparing an oil phase using structural triglycerides, phospholipids, and propofol. The oil phase is then subjected to a first-stage microfluidic emulsification with a first aqueous phase to obtain a propofol proemulsion. This proemulsion is then mixed with a second aqueous phase containing cyclodextrin-loaded phospholipid nanovesicles in a second-stage microfluidic process to obtain a propofol final emulsion. The nanovesicles are loaded with sodium sulfobutyl ether-β-cyclodextrin. This invention can reduce free propofol in the continuous aqueous phase, improve the uniformity of droplet size distribution, and enhance the enzymatic hydrolysis characteristics of the oil phase lipids, making it suitable for the preparation of propofol emulsion injections.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to a propofol emulsion injection and its preparation method. Background Technology

[0002] Propofol is a commonly used intravenous anesthetic in clinical practice, characterized by rapid onset, short duration of action, and quick recovery. Due to its poor water solubility, propofol is currently typically formulated as a fat emulsion injection, using an oil phase to load the propofol for intravenous administration. However, existing propofol fat emulsions usually use natural vegetable oils such as soybean oil as the oil phase, and are primarily formed into droplets through high-speed shearing or high-pressure homogenization. On the one hand, long-term infusion of large amounts of lipids may increase the body's lipid metabolism burden; on the other hand, during emulsification, high-pressure homogenization, and formulation dilution, some propofol may migrate from the oil phase to the continuous aqueous phase, and free propofol in the aqueous phase is a significant factor causing propofol injection irritation and pain. Therefore, how to reduce the level of free propofol in the continuous aqueous phase and improve lipid metabolism characteristics while ensuring the particle size and stability of the propofol emulsion remains a problem to be solved in existing propofol emulsion injections.

[0003] Chinese invention patent application CN108096187A discloses a propofol injection and its preparation method. This technology uses structured triglycerides to replace soybean oil, medium-chain triglycerides, or a mixture thereof in the oil phase, and uses propofol, structured triglycerides, and egg yolk lecithin to form an emulsion injection. This improves the compatibility and irritation issues of traditional propofol fat emulsions from the perspective of oil phase composition. This technology demonstrates that using structured triglycerides as the propofol oil phase has certain advantages; however, it mainly achieves propofol encapsulation through a conventional emulsion system and does not set up an independent secondary capture carrier for free propofol that has migrated to the continuous aqueous phase. Therefore, there is still room for improvement in how to further reduce free propofol in the aqueous phase.

[0004] Chinese invention patent application CN104490780A discloses a method for preparing propofol fat emulsion injection. The method involves spraying an oil phase composed of soybean oil, lecithin, and oleic acid into an aqueous phase. After high-speed shearing to obtain a primary emulsion, the emulsion is then subjected to multiple high-pressure homogenization processes to obtain a propofol fat emulsion with relatively uniform particle size. This technology can reduce droplet size and improve the physical stability of the emulsion through high-pressure homogenization. However, the adjustment of droplet size depends on the strong mechanical action generated by high-speed shearing and high-pressure homogenization. For hydrophobic drugs like propofol, which are mainly distributed in the oil phase, repeated strong shearing and droplet fragmentation processes may still increase the chance of drug redistribution into the continuous aqueous phase. Therefore, relying solely on high-pressure homogenization to control particle size is insufficient to simultaneously address the coordination issue between droplet uniformity and low free propofol levels.

[0005] Chinese invention patent application CN116350579A discloses a clarified propofol injection and its preparation method. This method uses cyclodextrins such as hydroxypropyl-β-cyclodextrin or sulfobutyl ether-β-cyclodextrin to directly encapsulate propofol, thereby reducing free propofol in the aqueous phase and obtaining a clarified injection without the need for a fat emulsion as a carrier. This technology demonstrates that cyclodextrins can be used to reduce free propofol. However, its method of directly dispersing cyclodextrins in a continuous aqueous phase and directly undertaking the solubilization and encapsulation of propofol differs from the system where the oil phase of a fat emulsion primarily carries the drug, with a small amount of propofol migrating to the aqueous phase. Furthermore, the document itself also points out that cyclodextrin-based propofol preparations have issues requiring further optimization, such as higher cyclodextrin dosage, control of free propofol, and storage stability.

[0006] Therefore, although existing technologies have improved propofol injections by changing the type of oil phase, increasing the degree of emulsification homogenization, or using cyclodextrin inclusion, a technical solution that can organically combine "oil phase as the main drug carrier," "secondary capture of free propofol in the aqueous phase," and "mild and controllable droplet formation process" is still lacking. Based on this, it is necessary to provide a new propofol emulsion injection and its preparation method. Summary of the Invention

[0007] To address the shortcomings of the existing technology, the present invention provides a propofol emulsion injection and its preparation method.

[0008] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is as follows: A method for preparing a propofol emulsion injection includes the following steps: Preparation of the oil phase: Propofol, phospholipids and structured triglycerides were mixed and heated to dissolve, thus obtaining the oil phase; Preparation of aqueous phase: Glycerin, disodium edetate and sodium oleate are dissolved in water for injection to obtain a basic aqueous phase. The basic aqueous phase is divided into a first aqueous phase and a second basic aqueous phase. Cyclodextrin-supported phospholipid nanovesicle dispersion is added to the second basic aqueous phase and mixed evenly to obtain the second aqueous phase. The oil phase and the first aqueous phase are introduced into the first-stage microchannel for microfluidic emulsification to obtain propofol primary emulsion. The propofol primary emulsion and the second aqueous phase are introduced into a second-stage microchannel for microfluidic emulsification to obtain the propofol final emulsion. The propofol final emulsion is then filled into containers to obtain the final product.

[0009] Preferably, the preparation method of the cyclodextrin-supported phospholipid nanovesicle dispersion includes: dissolving phospholipids and cholesterol in an organic solvent to obtain a phospholipid solution; dissolving sodium sulfobutyl ether-β-cyclodextrin in water for injection to obtain a cyclodextrin aqueous solution; adding the phospholipid solution to the cyclodextrin aqueous solution to allow the phospholipids to self-assemble into phospholipid nanovesicles loaded with sodium sulfobutyl ether-β-cyclodextrin, and obtaining a nanovesicle dispersion after membrane extrusion; removing free sodium sulfobutyl ether-β-cyclodextrin from the nanovesicle dispersion to obtain the cyclodextrin-supported phospholipid nanovesicle dispersion.

[0010] The interactions between the substances are as follows: In the preparation of cyclodextrin-supported phospholipid nanovesicles, hydrogenated soybean phospholipid and DSPE-PEG-OH are both amphiphilic phospholipid molecules. In an aqueous environment, they spontaneously arrange themselves to form a phospholipid bilayer through hydrophobic interactions. The hydrophobic fatty chains aggregate to form hydrophobic regions inside the bilayer, while the hydrophilic phosphate groups and polyethylene glycol chains face the aqueous phase. The hydrophobic steroidal ring structure of cholesterol inserts between the hydrophobic fatty chains of the phospholipid molecules and binds to the phospholipid molecules through hydrophobic interactions and van der Waals forces, thereby forming a relatively stable phospholipid bilayer.

[0011] Sodium sulfonyl ether-β-cyclodextrin has a hydrophilic outer surface and a relatively hydrophobic ring cavity. During the process of phospholipid molecules self-assembling to form closed nanovesicles, some sodium sulfonyl ether-β-cyclodextrin is loaded into the nanovesicles along with the aqueous phase, and finally cyclodextrin-supported phospholipid nanovesicles are obtained.

[0012] Furthermore, the preparation method of the cyclodextrin-supported phospholipid nanovesicle dispersion is as follows: S1. Add 2-6 parts by weight of hydrogenated soybean lecithin, 0.5-1.5 parts by weight of cholesterol and 0.1-0.5 parts by weight of DSPE-PEG-OH to 80-140 parts by weight of anhydrous ethanol, and stir at 50-70℃ and 400-800 rpm until completely dissolved to obtain a lecithin solution; separately take 160-240 parts by weight of a 5-15% sodium sulfobutyl ether-β-cyclodextrin aqueous solution and heat to 50-70℃. S2. Under stirring conditions of 300-500 rpm, the phospholipid solution is slowly added to an aqueous solution of sodium sulfonyl ether-β-cyclodextrin, and stirred at 50-70℃ for 20-60 min. The resulting dispersion is kept at 50-70℃ and passed sequentially through polycarbonate membranes with pore sizes of 100-400 nm and 50-200 nm. Each type of polycarbonate membrane is extruded 2-5 times to obtain a nanovesicle dispersion. S3. The obtained nanovesicle dispersion is ultrafiltered using an ultrafiltration membrane with a molecular weight cutoff of 50-150 kDa, and replaced with water for injection 2-5 times with equal volumes to remove unencapsulated sodium sulfobutyl ether-β-cyclodextrin in the aqueous phase outside the nanovesicles. The cutoff liquid is adjusted to 150-250 parts by weight with water for injection and filtered through a 0.22 μm sterile filter membrane to obtain a cyclodextrin-supported phospholipid nanovesicle dispersion, which is stored under sterile conditions for later use.

[0013] When the obtained cyclodextrin-supported phospholipid nanovesicles are added to propofol emulsion injection, a small amount of propofol migrating from the triglyceride droplets into the continuous aqueous phase can first be distributed to the hydrophobic region of the phospholipid bilayer and further enter the interior of the nanovesicles. The hydrophobic aromatic ring of propofol can enter the hydrophobic cavity of sulfobutyl ether-β-cyclodextrin, forming a reversible host-guest inclusion complex through hydrophobic interactions, van der Waals forces, and hydrogen bonds, thereby secondary binding of free propofol in the aqueous phase. Therefore, cyclodextrin-supported phospholipid nanovesicles can reduce the content of free propofol in the continuous aqueous phase and improve the system's ability to retain propofol after dilution, thus reducing the risk of injection irritation caused by free propofol.

[0014] Preferably, the first-stage microchannel and / or the second-stage microchannel are flow-focusing microchannels; in the first-stage microfluidic emulsification, the flow ratio of the first aqueous phase to the oil phase is 1.5-2.5:1; in the second-stage microfluidic mixing process, the flow ratio of the second aqueous phase to the propofol pre-emulsion is 1.5-2.5:1; the first-stage microfluidic emulsification and the second-stage microfluidic mixing are carried out continuously without storing the propofol pre-emulsion in between.

[0015] Preferably, by weight, the oil phase comprises 80-120 parts of structured triglycerides, 8-16 parts of phospholipids, and 8-12 parts of propofol; The base aqueous phase used to prepare the first aqueous phase and the second base aqueous phase comprises 18-28 parts of glycerol, 0.3-0.8 parts of disodium edetate, 0.2-0.5 parts of sodium oleate, and 600-800 parts of water for injection; The second aqueous phase also contains 80-240 parts of a cyclodextrin-supported phospholipid nanovesicle dispersion.

[0016] Preferably, in the method for preparing the cyclodextrin-supported phospholipid nanovesicles, the raw materials for preparing the cyclodextrin-supported phospholipid nanovesicle dispersion include 2-6 parts of hydrogenated soybean phospholipid, 0.5-1.5 parts of cholesterol, and 0.1-0.5 parts of polyethylene glycol-modified phospholipid. The mass concentration of the sulfobutyl ether-β-cyclodextrin sodium aqueous solution is 5%-15%; The membrane extrusion includes sequentially extruding a first polycarbonate membrane with a pore size of 100-400 nm and a second polycarbonate membrane with a pore size of 50-200 nm. Furthermore, the polyethylene glycol-modified phospholipids include DSPE-PEG-OH, DSPE-mPEG, etc., and the average molecular weight of PEG is preferably 1000-5000 g / mol.

[0017] Preferably, the sodium sulfobutyl ether-β-cyclodextrin not loaded in the aqueous phase other than the nanovesicle dispersion is removed by ultrafiltration. The ultrafiltration uses an ultrafiltration membrane with a molecular weight cutoff of 50-150 kDa and is performed with water for injection for 2-5 equal volume replacements.

[0018] Preferably, the oil phase, the first aqueous phase, and each component used to prepare the second aqueous phase are sterilized before microfluidic processing; the cyclodextrin-supported phospholipid nanovesicle dispersion is filtered through a 0.22 μm sterile filter membrane; and the microfluidic emulsification, microfluidic mixing, and filling are all performed under aseptic conditions.

[0019] Preferably, the oil phase preparation is carried out under nitrogen protection and the heating temperature is 50-65℃; the first aqueous phase is adjusted to 50-65℃ before the first-stage microfluidic emulsification.

[0020] Preferably, after the propofol final emulsion is filled, a visible foreign matter inspection and a seal integrity inspection are performed, and unqualified products are rejected before packaging.

[0021] A propofol emulsion injection solution is prepared by the method described above.

[0022] This invention first employs structured triglycerides as the main oil-phase carrier for propofol, allowing propofol to be preferentially distributed and retained within the oil-phase droplets. Simultaneously, the superior enzymatic lipolysis properties of structured triglycerides improve the lipid metabolism characteristics of traditional soybean oil-based emulsions. A first-stage flow-focusing microchannel allows the oil and first aqueous phases to form relatively uniform drug-loaded droplets in a controlled microscale flow field, reducing droplet breakage and propofol migration into the continuous aqueous phase caused by the strong mechanical forces during repeated high-pressure homogenization to form small droplets.

[0023] Building upon this, the present invention introduces cyclodextrin-supported phospholipid nanovesicles into the second aqueous phase and continuously adds them to the propofol primary emulsion via a second-stage microfluidic process. A small amount of propofol migrating from the structural triglyceride droplets into the continuous aqueous phase can be further distributed to the phospholipid nanovesicle membrane and undergo reversible inclusion with the sulfobutyl ether-β-cyclodextrin sodium inside the vesicles. This creates a secondary binding space for free propofol within the cyclodextrin-supported phospholipid nanovesicles, thereby reducing the level of free propofol actually in the continuous aqueous phase and improving the system's ability to retain propofol after dilution.

[0024] Therefore, this invention addresses the problems of lipid metabolism burden, high free propofol in aqueous phase, and difficulty in achieving both droplet uniformity and drug migration under strong mechanical emulsification conditions through a technical approach of "primary drug loading of structural triglycerides, secondary binding of cyclodextrin-loaded phospholipid nanovesicles, and secondary continuous microfluidic staged assembly". This results in a propofol emulsion injection with lower free propofol levels, more uniform particle size distribution, and better lipolysis performance.

[0025] The beneficial effects of this invention are: (1) The present invention uses structured triglycerides as the main oil phase carrier of propofol. While meeting the requirements of propofol oil phase solubility, structured triglycerides have better enzymatic lipolysis characteristics than traditional soybean oil, which is conducive to improving the utilization and clearance rate of lipids, thereby improving the lipid metabolism burden that may exist during the long-term use of traditional soybean oil-based propofol fat emulsion.

[0026] (2) In this invention, cyclodextrin-supported phospholipid nanovesicles are introduced into the second aqueous phase, and sodium sulfobutyl ether-β-cyclodextrin is confined inside the phospholipid nanovesicles. This allows the phospholipid membrane and the internal cyclodextrin to form a secondary propofol binding system, which can further bind propofol that migrates from the structural triglyceride droplets to the continuous aqueous phase. This reduces the amount of free propofol that is actually in the outer aqueous phase and can reduce the degree to which propofol is re-released into the continuous aqueous phase after system dilution, thereby reducing the risk of injection irritation caused by free propofol.

[0027] (3) The present invention adopts a two-stage continuous microfluidic process. First, the oil phase and the first aqueous phase are formed into a propofol primary emulsion with relatively uniform particle size in the first-stage microchannel. Then, a second aqueous phase containing cyclodextrin-supported phospholipid nanovesicles is continuously introduced into the second-stage microchannel. This allows the formation of emulsion droplets and the introduction of the second carrier to be carried out in stages, reducing the migration of propofol to the continuous aqueous phase caused by strong shearing, impact and cavitation in the traditional high-pressure homogenization process. At the same time, it can obtain an emulsion injection with relatively uniform particle size distribution.

[0028] (4) The present invention improves the lipid hydrolysis characteristics while taking into account the control of free propofol and the uniformity of emulsion dispersion through the synergistic combination of structured triglyceride drug-loaded droplets, cyclodextrin-loaded phospholipid nanovesicles and secondary continuous microfluidic process, so that the resulting propofol emulsion injection has good comprehensive performance. Detailed Implementation

[0029] The invention will be further described in detail below with reference to specific embodiments, but it should not be construed as limiting the scope of the invention to the following embodiments.

[0030] The raw materials described in this application are partially described; all other raw materials not described are commercially available. The preparation method of the structured triglyceride is as follows: 64 parts by weight of refined soybean oil and 36 parts by weight of medium-chain triglycerides are mixed and heated to 80°C under nitrogen protection, and stirred for 30 min. Sodium methoxide (0.2% by weight of the total oil mass) is added, and the mixture is stirred at 80°C for 1 h to allow random transesterification of medium-chain fatty acids and long-chain fatty acids between triglyceride molecules. After the reaction is complete, citric acid aqueous solution is added to neutralize the catalyst, and the mixture is allowed to stand for separation. The aqueous phase is removed, and the resulting oil phase is washed with water and dehydrated under reduced pressure. Residual low-molecular-weight substances are removed under vacuum, and the mixture is filtered to obtain the structured triglyceride.

[0031] Hydrogenated soybean lecithin was purchased from Aivito (Shanghai) Pharmaceutical Technology Co., Ltd., model number: N01003.

[0032] DSPE-PEG-OH was purchased from Xi'an Kaixin Biotechnology Co., Ltd. Sodium sulfobutyl ether-β-cyclodextrin was purchased from Shandong Tongwang Biotechnology Co., Ltd., with a water solubility of >50 g / 100 mL.

[0033] Example 1 A method for preparing a propofol emulsion injection includes the following steps: (1) Preparation of oil phase: Under nitrogen protection, 100 parts by weight of structured triglyceride was heated to 55°C, 12 parts by weight of egg yolk lecithin was added and stirred until dissolved; 10 parts by weight of propofol were added and stirred evenly to obtain the oil phase; (2) Preparation of aqueous phase: Under nitrogen protection, 22.5 parts by weight of glycerol, 0.5 parts by weight of disodium edetate and 0.3 parts by weight of sodium oleate were added to 697.9 parts by weight of water for injection and stirred until completely dissolved. 219.5 parts by weight of the solution were taken as the first aqueous phase. 156.8 parts by weight of the pre-prepared cyclodextrin-supported phospholipid nanovesicle dispersion were added to the remaining 501.7 parts by weight of the basic aqueous phase and mixed evenly to obtain 658.5 parts by weight of the second aqueous phase. (3) Dual carrier second-stage microfluidic mixing: The oil phase and the first aqueous phase obtained in step (1) are adjusted to 55°C respectively; the oil phase and the first aqueous phase are respectively introduced into the first-stage flow focusing microchannel, and the flow ratio of the first aqueous phase to the oil phase is controlled to be 1.8:1. Microfluidic emulsification is continuously carried out to obtain propofol primary emulsion. The obtained propofol pre-emulsion was continuously introduced into a second-stage flow-focusing microchannel, while a second aqueous phase was introduced simultaneously. The flow ratio of the second aqueous phase to the propofol pre-emulsion was controlled at 1.93:1. The second-stage microfluidic mixing was continuously performed, and the resulting emulsion was collected to obtain the propofol final emulsion. The first-stage microfluidic emulsification and the second-stage microfluidic mixing were performed continuously without storing the propofol pre-emulsion in between. (4) Sterilization and aseptic filling: The oil phase, the first aqueous phase and the components used to prepare the second aqueous phase were sterilized. The cyclodextrin-supported phospholipid nanovesicle dispersion was filtered through a 0.22 μm sterile filter membrane. Under aseptic conditions, a sterilized microfluidic system was used to prepare propofol final emulsion according to step (3). The obtained final emulsion was aseptically filled into 20 mL glass bottles and sealed to obtain propofol emulsion injection. (5) Inspection and packaging: After filling, the product is inspected for visible foreign matter and seal integrity. Unqualified products are rejected and qualified products are packaged to obtain propofol emulsion injection.

[0034] The preparation method of the cyclodextrin-supported phospholipid nanovesicle dispersion is as follows: S1. Add 4 parts by weight of hydrogenated soybean phospholipid, 1 part by weight of cholesterol and 0.2 parts by weight of DSPE-PEG-OH to 100 parts by weight of anhydrous ethanol, and stir at 60°C and 600 rpm until completely dissolved to obtain a phospholipid solution; separately take 200 parts by weight of a 10% sodium sulfobutyl ether-β-cyclodextrin aqueous solution and heat it to 60°C. S2. Under stirring at 400 rpm, the phospholipid solution is slowly added to an aqueous solution of sodium sulfonyl ether-β-cyclodextrin. The mixture is stirred at 60°C for 30 min. The resulting dispersion is kept at 60°C and then passed through polycarbonate membranes with pore sizes of 200 nm and 100 nm in sequence. Each type of polycarbonate membrane is extruded three times to obtain a nanovesicle dispersion. S3. The obtained nanovesicle dispersion was ultrafiltered using an ultrafiltration membrane with a molecular weight cutoff of 100 kDa, and then replaced three times with water for injection at equal volumes to remove unencapsulated sodium sulfobutyl ether-β-cyclodextrin in the aqueous phase outside the nanovesicles. The cutoff liquid was adjusted to 200 parts by weight with water for injection and filtered through a 0.22 μm sterile filter membrane to obtain a cyclodextrin-supported phospholipid nanovesicle dispersion, which was stored under sterile conditions for later use.

[0035] Example 2 The process is basically the same as in Example 1, except that in step (2), under nitrogen protection, 22.5 parts by weight of glycerol, 0.5 parts by weight of disodium edetate and 0.3 parts by weight of sodium oleate are added to 760.6 parts by weight of water for injection and stirred until completely dissolved to obtain 783.9 parts by weight of the basic aqueous phase; 219.5 parts by weight of this phase are taken as the first aqueous phase, and 94.1 parts by weight of the pre-prepared cyclodextrin-supported phospholipid nanovesicle dispersion is added to the remaining 564.4 parts by weight of the basic aqueous phase and mixed evenly to obtain 658.5 parts by weight of the second aqueous phase; in the second aqueous phase, the mass concentration of phospholipid contained in the cyclodextrin-supported phospholipid nanovesicles is 0.3% based on the total mass of hydrogenated soybean phospholipids and DSPE-PEG2000-OH; other steps and parameters are the same as in Example 1.

[0036] Example 3 The process is basically the same as in Example 1, except that in step (2), under nitrogen protection, 22.5 parts by weight of glycerol, 0.5 parts by weight of disodium edetate and 0.3 parts by weight of sodium oleate are added to 635.2 parts by weight of water for injection and stirred until completely dissolved to obtain 658.5 parts by weight of the basic aqueous phase; 219.5 parts by weight of the basic aqueous phase are taken as the first aqueous phase, and 219.5 parts by weight of the pre-prepared cyclodextrin-supported phospholipid nanovesicle dispersion is added to the remaining 439.0 parts by weight of the basic aqueous phase and mixed evenly to obtain 658.5 parts by weight of the second aqueous phase; in the second aqueous phase, the mass concentration of phospholipid contained in the cyclodextrin-supported phospholipid nanovesicles is 0.7% based on the total mass of hydrogenated soybean phospholipids and DSPE-PEG2000-OH; other steps and parameters are the same as in Example 1.

[0037] Example 4 The preparation of the cyclodextrin-supported phospholipid nanovesicle dispersion is basically the same as that of Example 1, except that in step S1 of the preparation of the cyclodextrin-supported phospholipid nanovesicle dispersion, 200 parts by weight of 5% sodium sulfonyl ether-β-cyclodextrin aqueous solution are used instead of 200 parts by weight of 10% sodium sulfonyl ether-β-cyclodextrin aqueous solution in Example 1; the remaining preparation steps of the cyclodextrin-supported phospholipid nanovesicle dispersion and the amount of the dispersion added in step (2) are the same as those of Example 1; other steps and parameters are the same as those of Example 1.

[0038] Example 5 The preparation of the cyclodextrin-supported phospholipid nanovesicle dispersion is basically the same as that in Example 1, except that in step S1 of the preparation of the cyclodextrin-supported phospholipid nanovesicle dispersion, 200 parts by weight of 15% sodium sulfonyl ether-β-cyclodextrin aqueous solution are used instead of 200 parts by weight of 10% sodium sulfonyl ether-β-cyclodextrin aqueous solution in Example 1; the remaining preparation steps of the cyclodextrin-supported phospholipid nanovesicle dispersion and the amount of the dispersion added in step (2) are the same as those in Example 1; other steps and parameters are the same as those in Example 1.

[0039] Comparative Example 1 The process is basically the same as in Example 1, except that in step (1), 100 parts by weight of soybean oil for injection is used instead of 100 parts by weight of structured triglycerides; the other steps and parameters are the same as in Example 1.

[0040] Comparative Example 2 The process is basically the same as in Example 1, except that: in step (2), no cyclodextrin-supported phospholipid nanovesicle dispersion is added; step (2) is adjusted as follows: under nitrogen protection, 22.5 parts by weight of glycerol, 0.5 parts by weight of disodium edetate and 0.3 parts by weight of sodium oleate are added to 854.7 parts by weight of water for injection and stirred until completely dissolved to obtain 878.0 parts by weight of basic aqueous phase; 219.5 parts by weight of this phase are taken as the first aqueous phase and the remaining 658.5 parts by weight of the basic aqueous phase are directly used as the second aqueous phase; other steps and parameters are the same as in Example 1.

[0041] Comparative Example 3 The process is basically the same as in Example 1, except that: in step (2), no cyclodextrin-supported phospholipid nanovesicle dispersion is added, but a 10% sodium sulfonyl ether-β-cyclodextrin aqueous solution is directly added to the second basic aqueous phase; specifically: under nitrogen protection, 22.5 parts by weight of glycerol, 0.5 parts by weight of disodium edetate and 0.3 parts by weight of sodium oleate are added to 697.9 parts by weight of water for injection and stirred until completely dissolved to obtain 721.2 parts by weight of basic aqueous phase; 219.5 parts by weight of this phase are taken as the first aqueous phase, and 156.8 parts by weight of a 10% sodium sulfonyl ether-β-cyclodextrin aqueous solution is directly added to the remaining 501.7 parts by weight of basic aqueous phase and mixed evenly to obtain 658.5 parts by weight of the second aqueous phase; other steps and parameters are the same as in Example 1.

[0042] Comparative Example 4 The method is basically the same as in Example 1, except that a blank phospholipid nanovesicle dispersion without sodium sulfobutyl ether-β-cyclodextrin is used instead of the cyclodextrin-supported phospholipid nanovesicle dispersion in Example 1.

[0043] The preparation method of the blank phospholipid nanovesicle dispersion is as follows: S1. Add 4 parts by weight of hydrogenated soybean phospholipids, 1 part by weight of cholesterol and 0.2 parts by weight of DSPE-PEG2000-OH to 100 parts by weight of anhydrous ethanol, and stir at 60°C and 600 rpm until completely dissolved to obtain a phospholipid solution; separately take 200 parts by weight of water for injection and heat to 60°C. S2. Under stirring at 400 rpm, the phospholipid solution was slowly added to water for injection and stirred at 60°C for 30 min. Then, the temperature of the resulting dispersion was kept at 60°C and passed through polycarbonate membranes with pore sizes of 200 nm and 100 nm in sequence. Each type of polycarbonate membrane was extruded 3 times to obtain a blank nanovesicle dispersion. S3. The obtained blank nanovesicle dispersion was ultrafiltered using an ultrafiltration membrane with a molecular weight cutoff of 100 kDa, and then replaced three times with water for injection at equal volumes. The cutoff liquid was adjusted to 200 parts by weight with water for injection and filtered through a 0.22 μm sterile filter membrane to obtain the blank phospholipid nanovesicle dispersion.

[0044] In step (2), 156.8 parts by weight of the blank phospholipid nanovesicle dispersion are added, and the amount of other raw materials and preparation steps are the same as in Example 1.

[0045] Comparative Example 5 The process is basically the same as in Example 1, except that step (3) does not use the first-stage microfluidic emulsification and second-stage microfluidic mixing method, but instead uses a single-stage microfluidic method to prepare the propofol final emulsion. Specifically, the oil phase, the first aqueous phase, and the second aqueous phase are prepared according to the method in Example 1. The first aqueous phase and the second aqueous phase are continuously merged before entering the microchannel to form a full aqueous phase, and immediately after merging, they are introduced into a single-stage flow-focusing microchannel. At the same time, the oil phase is introduced into the flow-focusing microchannel, and the mass flow ratio of the full aqueous phase to the oil phase is controlled to be 7.20:1. Microfluidic emulsification is continuously performed, and the resulting emulsion is collected to obtain the propofol final emulsion. Other steps and parameters are the same as in Example 1.

[0046] Comparative Example 6 The process is basically the same as in Example 1, except that step (3) does not use two-stage continuous microfluidic control, but instead uses high-speed shear pre-emulsification combined with high-pressure homogenization to prepare the propofol final emulsion. Specifically, the oil phase obtained in step (1) is mixed with the first aqueous phase and the second aqueous phase obtained in step (2), and sheared at 8000 rpm for 5 min at 55°C to obtain a pre-emulsion. The obtained pre-emulsion is then homogenized under high pressure at 60 MPa for a total of 3 times to obtain the propofol final emulsion. Other steps and parameters are the same as in Example 1.

[0047] Test Example 1 Aqueous phase free propofol content test: The aqueous phase free propofol content of the propofol emulsion injections prepared in Examples 1-5 and Comparative Examples 1-6 was tested.

[0048] The separation of free propofol in the aqueous phase was performed using centrifugal ultrafiltration. 1.0 mL of the propofol emulsion injection to be tested was added to a regenerated cellulose ultrafiltration tube with a molecular weight cutoff of 1 kDa. The solution was centrifuged at 25°C and 3000 × g for 10 min, and the filtrate was collected. The ultrafiltration membrane used was able to retain emulsion droplets, phospholipid nanovesicles, and sulfobutyl ether-β-cyclodextrin sodium-propofol conjugates, while free propofol could pass through the ultrafiltration membrane. The recovery rate of the ultrafiltration process was verified using a propofol standard solution before testing, and the recovery rate was controlled at 95%-105%.

[0049] The propofol content in the filtrate was determined by HPLC according to Chapter 0512 of the 2025 edition of the Pharmacopoeia of the People's Republic of China, Part IV. The chromatographic conditions were as follows: a C18 column (4.6 mm × 150 mm, 5 μm particle size); acetonitrile-water (70:30 v / v); flow rate 1.0 mL / min; column temperature 30℃; detection wavelength 270 nm; injection volume 20 μL. A series of standard solutions were prepared using propofol reference standard, and a standard curve was established with peak area as the ordinate and propofol concentration as the abscissa. The concentration of free propofol in the filtrate was calculated based on the standard curve.

[0050] Further evaluation of the changes in free propofol under simulated intravenous administration dilution conditions was conducted. 1.0 mL of the propofol emulsion injection to be tested was added to 9.0 mL of 0.9% sodium chloride solution preheated to 37°C. After mixing, the solution was maintained at 37°C for 5 min. The concentration of free propofol was determined according to the above-described centrifugation, ultrafiltration, and high-performance liquid chromatography methods. The obtained concentration was multiplied by a dilution factor of 10 to obtain the free propofol concentration after a 10-fold dilution. The test results are shown in Table 1.

[0051] Table 1. Results of free propofol test in each group of propofol emulsion injections

[0052] Test Example 2 Z-mean particle size and PDI test of the final emulsion dispersion system: The average particle size and polydispersity index (PDI) of the propofol emulsion injections obtained in Examples 1-5 and Comparative Examples 1-6 were determined according to the dynamic light scattering method in General Chapter 0982 of the 2025 edition of the Pharmacopoeia of the People's Republic of China, Part IV, for determination of particle size and particle size distribution.

[0053] Each sample was appropriately diluted with a 0.9% sodium chloride solution filtered through a 0.22 μm filter membrane to ensure the scattering intensity was within the effective detection range of the instrument. The samples were then tested using a dynamic light scattering particle size analyzer at 25℃. Each sample was measured in triplicate, and the Z-mean particle size and PDI were recorded and averaged. The results are shown in Table 2.

[0054] Table 2. Particle size and PDI test results of propofol emulsion injections in each group.

[0055] Test Example 3 Lipoprotein lipase-mediated in vitro lipolysis test: To evaluate the differences in the hydrolysis of emulsion droplets formed by structural triglycerides and soybean oil by lipoprotein lipase, Example 1 and Comparative Example 1 were selected for in vitro lipolysis experiments.

[0056] Take the propofol emulsion injections obtained in Example 1 and Comparative Example 1 respectively, and adjust the sample addition amount using a triglyceride mass meter so that the triglyceride concentration in the reaction system is 10 mg / mL.

[0057] Prepare a 50 mmol / L Tris-HCl buffer solution with a pH of 8.0, containing 2 mmol / L calcium chloride and 0.5% fatty acid-free bovine serum albumin. Add the emulsion to be tested to the buffer solution, pre-equilibrate at 37°C for 10 min, then add lipoprotein lipase, controlling the addition to 2 U of lipoprotein lipase per 100 mg of triglycerides, and perform enzymatic digestion at 37°C.

[0058] Samples were taken at 10 min, 20 min, and 30 min of reaction, and 4 times the volume of isopropanol was immediately added to terminate the enzymatic hydrolysis. The supernatant was collected by centrifugation, and the free fatty acid content in the reaction system was determined using a free fatty acid enzyme assay kit. The free fatty acid content of the sample at 0 min was then subtracted to obtain the net release of free fatty acids during lipolysis. The results are shown in Table 3.

[0059] Table 3. Results of in vitro lipolysis tests for Example 1 and Comparative Example 1

[0060] The test results of Examples 1-5 show that the present invention, by combining structured triglycerides, cyclodextrin-supported phospholipid nanovesicles, and a two-stage continuous microfluidic emulsification process, can effectively reduce free propofol in the continuous aqueous phase while maintaining a small droplet size and narrow particle size distribution of the propofol emulsion injection. In Example 2, reducing the amount of cyclodextrin-supported phospholipid nanovesicles decreased the ability to capture free propofol in the aqueous phase. In Example 3, increasing the amount of nanovesicles further reduced free propofol, but the increased amount of dispersed phase in the system reduced the overall particle size distribution uniformity, indicating that higher nanovesicle amounts are not necessarily more beneficial. In Example 4, reducing the concentration of sodium sulfobutyl ether-β-cyclodextrin decreased the binding ability of nanovesicles to propofol migrating to the aqueous phase. In Example 5, further increasing the concentration of sodium sulfobutyl ether-β-cyclodextrin resulted in a limited reduction in free propofol, while the particle size distribution uniformity also decreased.

[0061] Compared to Comparative Example 1, Example 1 only replaced the structured triglyceride with soybean oil. Using the structured triglyceride to form the oil phase resulted in faster release of free fatty acids under the same lipoprotein lipase activity, indicating that it is more prone to enzymatic lipolysis, which is beneficial for improving lipid utilization and clearance. In contrast, the lipolysis rate decreased after using soybean oil. Therefore, the structured triglyceride not only serves as an oil phase carrier for propofol but also mitigates the potential lipid metabolism burden that may exist during long-term use of traditional soybean oil-based propofol fat emulsions.

[0062] Compared with Comparative Examples 2-4, Example 1 shows that cyclodextrin-supported phospholipid nanovesicles are an important factor in reducing free propofol in the aqueous phase. In Comparative Example 2, completely eliminating the nanovesicles resulted in a lack of a carrier for the secondary binding of propofol migrating from oil droplets to the continuous aqueous phase, leading to a significant increase in free propofol. In Comparative Example 4, using blank nanovesicles without sodium sulfobutyl ether-β-cyclodextrin, the distribution of propofol by the phospholipid membrane alone was insufficient for effective capture, indicating that sodium sulfobutyl ether-β-cyclodextrin played a crucial role. In Comparative Example 3, although sodium sulfobutyl ether-β-cyclodextrin could bind some propofol initially after being directly added to the aqueous phase, the free propofol increased significantly after dilution, indicating that the binding equilibrium of cyclodextrin-propofol directly dispersed in the external aqueous phase is more easily affected by system dilution. This invention, by internally loading sodium sulfobutyl ether-β-cyclodextrin into phospholipid nanovesicles, forms a relatively independent secondary binding space, thereby improving the retention capacity of propofol under dilution conditions.

[0063] Compared with Comparative Example 5, Comparative Example 5 changed the two-stage continuous microfluidic control to a single-stage microfluidic control, that is, the oil phase directly emulsifies with all the aqueous phase in one go. This requires the oil and water phases to complete the formation of droplets, the introduction of cyclodextrin-supported nanovesicles and the dilution of the system simultaneously in a single microchannel. This is not conducive to the graded control of the droplet formation process, resulting in a wider droplet size distribution and an increase in free propofol in the aqueous phase.

[0064] Compared to Comparative Example 6, although high-pressure homogenization can further reduce the Z-mean particle size of the final emulsion dispersion system through stronger mechanical action, the uniformity of the particle size distribution of the resulting system is reduced, and the free propofol in the aqueous phase is significantly increased. This is because the strong shearing, impact, and cavitation effects during high-pressure homogenization break up large emulsion droplets and also easily increase the chance of propofol migrating from the oil phase to the continuous aqueous phase. In contrast, the present invention uses a continuous microfluidic approach to gradually form and regulate emulsion droplets in a microscale flow field, without the need for repeated application of strong mechanical force, thus achieving both particle size uniformity and low free propofol levels.

Claims

1. A method for preparing a propofol emulsion injection, characterized in that, Includes the following steps: Preparation of the oil phase: Propofol, phospholipids and structured triglycerides were mixed and heated to dissolve, thus obtaining the oil phase; Preparation of aqueous phase: Glycerin, disodium edetate and sodium oleate are dissolved in water for injection to obtain a basic aqueous phase. The basic aqueous phase is divided into a first aqueous phase and a second basic aqueous phase. Cyclodextrin-supported phospholipid nanovesicle dispersion is added to the second basic aqueous phase and mixed evenly to obtain the second aqueous phase. The oil phase and the first aqueous phase are introduced into the first-stage microchannel for microfluidic emulsification to obtain propofol primary emulsion. The propofol primary emulsion and the second aqueous phase are introduced into a second-stage microchannel for microfluidic emulsification to obtain the propofol final emulsion. The propofol final emulsion is then filled into vials to obtain propofol emulsion injection.

2. The preparation method according to claim 1, characterized in that, The preparation method of the cyclodextrin-supported phospholipid nanovesicles includes: dissolving phospholipids and cholesterol in an organic solvent to obtain a phospholipid solution; dissolving sodium sulfobutyl ether-β-cyclodextrin in water for injection to obtain a cyclodextrin aqueous solution; adding the phospholipid solution to the cyclodextrin aqueous solution to allow the phospholipids to self-assemble into phospholipid nanovesicles loaded with sodium sulfobutyl ether-β-cyclodextrin, and obtaining a nanovesicle dispersion after membrane extrusion; removing free sodium sulfobutyl ether-β-cyclodextrin from the nanovesicle dispersion to obtain the cyclodextrin-supported phospholipid nanovesicle dispersion.

3. The preparation method according to claim 1, characterized in that, The first-stage microchannel and / or the second-stage microchannel are flow-focusing microchannels; in the first-stage microfluidic emulsification, the flow ratio of the first aqueous phase to the oil phase is 1.5-2.5:1; in the second-stage microfluidic mixing process, the flow ratio of the second aqueous phase to the propofol pre-emulsion is 1.5-2.5:1; the first-stage microfluidic emulsification and the second-stage microfluidic mixing are carried out continuously without storing the propofol pre-emulsion in between.

4. The preparation method according to claim 1, characterized in that, The oil phase comprises, by weight, 80-120 parts of structured triglycerides, 8-16 parts of phospholipids and 8-12 parts of propofol; The base aqueous phase used to prepare the first aqueous phase and the second base aqueous phase comprises 18-28 parts of glycerol, 0.3-0.8 parts of disodium edetate, 0.2-0.5 parts of sodium oleate, and 600-800 parts of water for injection; The second aqueous phase also contains 80-240 parts of a cyclodextrin-supported phospholipid nanovesicle dispersion.

5. The preparation method according to claim 2, characterized in that, The raw materials for preparing the cyclodextrin-supported phospholipid nanovesicle dispersion, by weight, include 2-6 parts of hydrogenated soybean phospholipid, 0.5-1.5 parts of cholesterol, and 0.1-0.5 parts of polyethylene glycol-modified phospholipid. The mass concentration of the sulfobutyl ether-β-cyclodextrin sodium aqueous solution is 5%-15%; The membrane extrusion includes sequentially extruding a first polycarbonate membrane with a pore size of 100-400 nm and a second polycarbonate membrane with a pore size of 50-200 nm; The PEGylated phospholipids include DSPE-PEG-OH, DSPE-mPEG, etc., and the average molecular weight of PEG is preferably 1000-5000 g / mol.

6. The preparation method according to claim 2, characterized in that, The sodium sulfobutyl ether-β-cyclodextrin that was not encapsulated in the aqueous phase, excluding the nanovesicle dispersion, was removed by ultrafiltration. The ultrafiltration used an ultrafiltration membrane with a molecular weight cutoff of 50-150 kDa and was performed with 2-5 equal-volume replacements using water for injection.

7. The preparation method according to claim 1, characterized in that, The oil phase, the first aqueous phase, and each component used to prepare the second aqueous phase are sterilized before microfluidic processing; the cyclodextrin-supported phospholipid nanovesicle dispersion is filtered through a 0.22 μm sterile filter membrane; the microfluidic emulsification, microfluidic mixing, and filling are all performed under aseptic conditions.

8. The preparation method according to claim 1, characterized in that, The oil phase preparation is carried out under nitrogen protection and the heating temperature is 50-65℃; the first aqueous phase is adjusted to 50-65℃ before the first-stage microfluidic emulsification.

9. The preparation method according to claim 1, characterized in that, After the propofol final emulsion is filled, it is inspected for visible foreign matter and seal integrity. Unqualified products are rejected before packaging.

10. A propofol emulsion injection, characterized in that, It is prepared by the preparation method according to any one of claims 1-9.

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