Method for preparing PDLLA porous microspheres with high porosity and uniform particle size distribution in batches

Abstract

The invention discloses a method for preparing PDLLA porous microspheres with high porosity and uniform particle size distribution in batches, and belongs to the technical field of polymer biomedical material preparation. According to the method, an ultrasonic-assisted double-emulsification system is combined with an inner water phase (an ammonium bicarbonate solution), an oil phase and an outer water phase (a polyvinyl alcohol solution), and a staged curing process is adopted to realize accurate regulation and control of a pore structure and a particle size. Petroleum ether is introduced as a poor solvent to optimize the precipitation uniformity, ultrasonic and mechanical stirring synergistically improve the emulsification uniformity, and staged curing ensures that the solvent evaporation rate is matched with pore formation. The porosity of the prepared PDLLA microspheres reaches 75%-87%, the SPAN value is as low as 0.512, the pore size distribution is uniform, the process expandability is high, and the method is suitable for the biomedical fields of drug carriers, tissue engineering scaffolds and the like. The technical problems of low porosity, wide particle size distribution and complex process are solved, and the method has a remarkable application prospect.

Description

Technical Field

[0001] The present invention relates to a method for batch preparing PDLLA porous microspheres with high porosity and uniform particle size distribution, belonging to the field of biomedical materials. Background Art

[0002] Due to their unique porous structure, high specific surface area, and excellent biocompatibility, polymer porous microspheres have important application values in the fields of drug delivery, tissue engineering scaffolds, biosensors, etc. As a biodegradable polymer material, poly-D,L-lactic acid (PDLLA) and its porous microspheres have attracted much attention in scenarios such as sustained-release drugs and cell loading. Currently, the preparation methods of PDLLA porous microspheres mainly include solvent evaporation method, emulsion polymerization method, suspension polymerization method, etc., but these methods still have significant technical bottlenecks in practical applications.

[0003] In the solvent evaporation method, PDLLA is dissolved in a volatile solvent (such as dichloromethane), a porogen (such as n-dodecane) is added, and a porous structure is formed through emulsification, curing, and solvent evaporation. For example, Patent CN112662005B discloses a method for regulating micropores with a NaOH solution, but the prepared microspheres have sparse surface pores and a wide particle size distribution range (SPAN value > 0.7), resulting in low drug loading efficiency and uncontrollable release behavior. In addition, this method relies on single mechanical stirring during the emulsification process, making it difficult to achieve uniform dispersion of droplets, further exacerbating the performance fluctuations of the microspheres. The emulsion polymerization method and the suspension polymerization method form microspheres through monomer polymerization, and the reaction temperature, initiator concentration, and stirring rate need to be precisely controlled. However, the polymerization reaction kinetics are complex, and poor internal pore connectivity of the microspheres (porosity < 65%) is easily caused by uneven local reaction conditions, and residual monomers may cause biotoxicity problems. For example, the coefficient of variation (CV value) of the average pore size distribution of PDLLA microspheres prepared by the emulsion polymerization method reported in the literature is as high as 30%, significantly limiting their application in the field of precision medicine.

[0004] The core problems of the above technologies include: uncontrollable porosity and pore size distribution: traditional methods rely on physical porogens (such as salting-out agents) or chemical foaming agents, but it is difficult to match the decomposition rate of the porogen with the solvent evaporation rate, resulting in discontinuous pore structures or a high proportion of closed pores. Poor particle size uniformity: it is difficult to achieve uniform shear force during the emulsification process with mechanical stirring or conventional homogenization techniques, resulting in a wide particle size distribution of the microspheres (SPAN value > 0.6), affecting batch-to-batch consistency. Complicated process and difficult to scale up: existing methods require multi-step parameter adjustments (such as staged solvent evaporation, multiple washings), with cumbersome operations and time-consuming processes, making it difficult to meet the requirements of industrial production.

[0005] With the development of precision medicine, the market demand for PDLLA microspheres with high porosity (>75%) and narrow particle size distribution (SPAN value <0.55) is increasing. However, the existing technologies cannot balance porosity and particle size uniformity, and the process stability is insufficient. For example, comparative experiments show that the porosity of microspheres prepared by the traditional solvent evaporation method is only 68±5%, while the particle size distribution span (D90-D10) is as high as 50 microns, seriously restricting their application in high-precision drug carriers. Summary of the Invention

[0006] To address the above problems, the present invention proposes a new process of ultrasonic-assisted double emulsification combined with staged curing. By introducing a poor solvent (petroleum ether) to optimize precipitation kinetics, using the synergistic effect of ultrasonic and mechanical stirring to improve emulsification uniformity, and precisely controlling the curing parameters to achieve directional regulation of the pore structure. This solution aims to break through the technical barrier that it is difficult to synergistically optimize porosity and particle size distribution in the existing technology, and provide an efficient and stable solution for the large-scale preparation of PDLLA porous microspheres. The specific solution is as follows:

[0007] A method for batch preparation of PDLLA porous microspheres with high porosity and uniform particle size distribution, comprising the following steps:

[0008] (1) Prepare the inner aqueous phase, oil phase and outer aqueous phase;

[0009] (2) Add the inner aqueous phase to the oil phase at low temperature for primary emulsification to form a primary emulsion;

[0010] (3) Add the primary emulsion to the outer aqueous phase for secondary emulsification to form a double emulsion;

[0011] (4) Cure the double emulsion in stages to form porous microspheres;

[0012] (5) Perform post-treatment to obtain the final product.

[0013] Preferably, the inner aqueous phase is an aqueous ammonium bicarbonate solution with a concentration of 5.2% - 6.4%.

[0014] Preferably, the oil phase contains a mixed solvent of dichloromethane solution of PDLLA and petroleum ether, and the volume ratio of the two is 3:1 - 7:1.

[0015] Preferably, in the primary emulsification step, an ultrasonic homogenizer is used, with an ultrasonic power of 50 - 150 W and a homogenizer rotation speed of 8000 - 16000 rpm.

[0016] Preferably, in the secondary emulsification step, mechanical stirring and a homogenizer are simultaneously turned on, with a mechanical stirring rate of 300 - 600 rpm and a homogenizer rate of 2000 - 4000 rpm.

[0017] Preferably, the staged curing includes: the first stage of curing for 1 - 3 hours under stirring at 20 - 40°C and 600 - 400 rpm; the second stage of curing for 20 - 36 hours under stirring at 45 - 55°C and 300 - 400 rpm.

[0018] Preferably, the post-treatment includes soaking in an alkaline solution, water washing, screening, and freeze-drying.

[0019] Preferably, the alkaline solution is a 0.8% sodium hydroxide solution, and the treatment time is 2 - 4 hours.

[0020] Preferably, the external aqueous phase is an aqueous solution of polyvinyl alcohol (PVA) with a concentration of 1% - 1.8%.

[0021] Preferably, the addition rates of the internal aqueous phase and the external aqueous phase in the primary emulsification and secondary emulsification steps are 40 - 60 ml / min and 60 - 80 ml / min, respectively.

[0022] Specifically, the present invention provides a method for batch preparation of PDLLA porous microspheres with high porosity and uniform particle size distribution. This method is based on an ultrasonic-assisted double-emulsion system. By optimizing the compositions of the internal aqueous phase, oil phase, and external aqueous phase, and combining staged curing and post-treatment processes, dual precise regulation of the pore structure and particle size distribution is achieved. The specific technical solutions include the following steps:

[0023] Phase system preparation

[0024] Internal aqueous phase: An aqueous solution of ammonium bicarbonate (NH4HCO3) with a concentration of 5.2% - 6.4% is used. Ammonium bicarbonate decomposes into ammonia (NH3) and carbon dioxide (CO2) during the curing process of the microspheres, forming a through-porous structure.

[0025] Oil phase: It is composed of a dichloromethane (DCM) solution of PDLLA and petroleum ether, and the volume ratio of the two is 3:1 - 7:1. Petroleum ether, as a poor solvent, regulates the precipitation kinetics of PDLLA to reduce the generation of large or small particles, thereby improving the particle size uniformity.

[0026] External aqueous phase: An aqueous solution of polyvinyl alcohol (PVA) with a concentration of 1% - 1.8% is used to stabilize the interface of the double emulsion droplets and prevent the microspheres from sticking together.

[0027] Primary emulsification process

[0028] Under the condition of an ice-water bath (0 - 5 °C), the inner aqueous phase was added to the oil phase at a rate of 40 - 60 ml / min, and primary emulsification was carried out using an ultrasonic homogenizer. The ultrasonic power was 50 - 150 W, the rotational speed of the homogenizer was 8000 - 16000 rpm, and the emulsification time was 5 - 15 minutes. The low-temperature condition inhibited the premature decomposition of ammonium bicarbonate, while ultrasonic assistance could generate high-frequency shear force, making the droplet size uniform and laying the foundation for the formation of subsequent pore structures.

[0029] Double emulsification process

[0030] The primary emulsion was added to the outer aqueous phase at a rate of 60 - 80 ml / min, while mechanical stirring (300 - 600 rpm) and a homogenizer (2000 - 4000 rpm) were turned on simultaneously. The double emulsification time was 30 - 60 minutes. The synergistic effect of the double stirring system (mechanical stirring promoted the overall emulsion circulation, and the homogenizer provided local high shear force) ensured the uniform dispersion of the double emulsion droplets and further reduced the particle size distribution range.

[0031] Staged curing

[0032] First-stage curing: Curing was carried out at 20 - 40 °C and a stirring speed of 600 - 400 rpm for 1 - 3 hours to allow the preliminary volatilization of the solvent (DCM) and the preliminary formation of the microsphere surface.

[0033] Second-stage curing: The temperature was raised to 45 - 55 °C, and the stirring speed was reduced to 300 - 400 rpm, and curing was carried out for 20 - 36 hours. Low-speed stirring slowed down the solvent volatilization rate, avoiding the collapse of the microsphere surface. At the same time, high temperature promoted the complete removal of the solvent, forming a smooth and stable porous structure.

[0034] Post-treatment process

[0035] The cured microspheres were soaked in a 0.8% sodium hydroxide solution for 2 - 4 hours to remove the residual ammonium bicarbonate; then they were centrifugally washed with deionized water 6 - 8 times to completely remove the alkaline substances; finally, the final product was obtained through sieving (particle size classification) and freeze-drying.

[0036] Technical key points and innovation

[0037] Synergistic emulsification mechanism of ultrasound and mechanical stirring

[0038] Traditional methods rely on a single stirring method, which is prone to uneven droplet size distribution. In the present invention, the high-frequency vibration of the ultrasonic homogenizer is combined with the macroscopic circulation of mechanical stirring, significantly improving the emulsification efficiency, reducing the coefficient of variation (CV value) of the droplet size to 10% - 18%, and the SPAN value is as low as 0.512 - 0.691.

[0039] Introduction of poor solvent (petroleum ether)

[0040] Petroleum ether, as a non-solvent, regulates the precipitation rate of PDLLA, inhibits local supersaturation, and reduces microsphere aggregation or fragmentation. Experiments show that when the proportion of petroleum ether in the oil phase increases from 0% to 20%, the SPAN value of the microspheres decreases from 0.892 to 0.512 (Example 6 vs Example 1), and the particle size distribution is significantly optimized.

[0041] Staged curing strategy

[0042] The low-temperature curing in the first stage ensures the preliminary stability of the microsphere skeleton, and the high-temperature and low-speed curing in the second stage takes into account both the complete volatilization of the solvent and the morphological integrity. Compared with single-stage curing (Comparative Example 1), the staged curing increases the porosity from 68% to 80%, and the CV value of the pore size distribution decreases from 35% to 10%.

[0043] Inner aqueous phase concentration matching decomposition kinetics

[0044] The concentration of ammonium bicarbonate is controlled at 5.2% - 6.4%, which can not only provide sufficient gas for pore formation but also avoid pore collapse caused by excessive decomposition. When the concentration is lower than 5%, the porosity is less than 70% (Comparative Example 2); when it is higher than 6.4%, cracks appear on the surface of the microspheres (Comparative Example 3).

[0045] Beneficial effects

[0046] High porosity and uniform pore size distribution: Through the synergistic effect of ultrasonic-assisted emulsification and staged curing, the porosity can reach 75% - 87%, and the CV value of the pore size ≤ 18%, significantly superior to traditional methods (porosity < 70%, CV value > 30%).

[0047] Narrow particle size distribution: The SPAN value is controlled at 0.512 - 0.691, meeting the stringent requirements of drug carriers for batch consistency.

[0048] Process scalability: The dual-stirring system and parameter range design are suitable for industrial production, with a single batch feeding amount reaching the kilogram level and a product yield > 90%.

[0049] Excellent biocompatibility: No residual toxic solvents (DCM residue < 0.1 ppm), meeting the medical-grade material standards.

[0050] Coverage scope of the technical solution

[0051] The present invention not only covers the above specific implementation parameters but also enables flexible regulation of microsphere performance by adjusting the following conditions:

[0052] The mass ratio of ammonium bicarbonate in the inner aqueous phase to PDLLA (1:3 - 1:4);

[0053] The ratio of DCM to petroleum ether in the oil phase (3:1 - 7:1);

[0054] Combination of ultrasonic power (50 - 150 W) and homogenizer speed (8000 - 16000 rpm);

[0055] Curing temperature and time gradient (for example, the curing time in the first stage is extended to 5 hours to further reduce the SPAN value). Description of the Drawings

[0056] Figure 1 SEM image of the product of Example 1;

[0057] Figure 2 SEM image of the product of Example 2;

[0058] Figure 3 SEM image of the product of Example 3;

[0059] Figure 4 SEM image of the product of Example 4;

[0060] Figure 5 SEM image of the product of Example 5;

[0061] Figure 6 SEM image of the product of Example 6. Detailed Description of the Invention

[0062] The technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0063] Unless otherwise defined, all professional and scientific terms used herein have the same meaning as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to the described content can be applied to the present invention. The preferred methods and materials described herein are only for illustrative purposes and do not limit the content of this application.

[0064] The experimental methods in the following embodiments are all conventional methods unless otherwise specified. The test materials used in the following embodiments are all obtained from commercial channels unless otherwise specified.

[0065] Example 1: Preparation of PDLLA Porous Microspheres with Standard Parameters

[0066] Experimental Steps

[0067] Preparation of the Phase System

[0068] Inner aqueous phase: Dissolve 16 g of ammonium bicarbonate in 250 mL of deionized water and stir until completely dissolved, with a concentration of 6.4% (w / w).

[0069] Oil phase: Add 50 g of PDLLA powder into 600 mL of dichloromethane (DCM), stir magnetically until completely dissolved, and then add 200 mL of petroleum ether (the volume ratio of DCM to petroleum ether is 3:1), and mix evenly.

[0070] External aqueous phase: Add 40 g of polyvinyl alcohol (PVA) powder into 4 L of deionized water, heat to 80 °C and continuously stir for 2 hours, and form a 1% concentration PVA aqueous solution after cooling to room temperature.

[0071] Primary emulsification

[0072] Place the oil phase in an ice-water bath (0 - 5 °C), and the internal aqueous phase is added dropwise to the oil phase at a rate of 60 mL / min through a constant flow pump.

[0073] Turn on the ultrasonic homogenizer (power 100 W, probe diameter 10 mm), set the rotation speed of the homogenizer to 13000 rpm, and continuously emulsify for 5 minutes.

[0074] Secondary emulsification

[0075] Add the primary emulsion to the external aqueous phase at a rate of 80 mL / min.

[0076] At the same time, turn on the mechanical stirrer (rotation speed 300 rpm, blade diameter 50 mm) and the homogenizer (rotation speed 2000 rpm), and continuously perform secondary emulsification for 30 minutes.

[0077] Staged curing

[0078] First-stage curing: Transfer the secondary emulsion to a 30 °C constant temperature water bath and cure at a stirring speed of 600 rpm for 2 hours.

[0079] Second-stage curing: Raise the temperature to 45 °C, reduce the stirring speed to 400 rpm, and continue to cure for 24 hours.

[0080] Post-treatment

[0081] The cured microsphere suspension is collected by vacuum filtration, added with 1.0 L of 0.8% sodium hydroxide solution and soaked for 3 hours to remove residual ammonium bicarbonate.

[0082] Centrifuge and wash 6 times with deionized water (5000 rpm, 10 minutes each time) to remove alkaline substances.

[0083] The washed microspheres are classified through a 200-mesh sieve and freeze-dried for 48 hours to obtain the final product.

[0084] Product characterization

[0085] Particle size distribution: Measured by a laser particle size analyzer (Malvern Mastersizer 3000), D10 = 82 μm, D50 = 90 μm, D90 = 98 μm, SPAN value = 0.512.

[0086] Porosity: Measured by mercury intrusion porosimetry (Micromeritics AutoPore IV), the porosity is 80%.

[0087] Pore size distribution: Analyzed by scanning electron microscopy (SEM, Hitachi SU8010), the pore size range is 5 - 20 μm, CV value = 10%.

[0088] SEM image: The surface of the microspheres shows a uniform porous structure, the pores are interconnected, and there is no obvious collapse or adhesion ( Figure 1 ).

[0089] Example 2: Reduce the ultrasonic power (50 W)

[0090] Experimental procedure

[0091] Phase system preparation

[0092] Inner aqueous phase: Dissolve 16 g of ammonium bicarbonate in 250 mL of deionized water, stir until completely dissolved, and the concentration is 6.4% (w / w).

[0093] Oil phase: Add 50 g of PDLLA powder to 600 mL of dichloromethane (DCM), stir magnetically until completely dissolved, and then add 200 mL of petroleum ether (the volume ratio of DCM to petroleum ether is 3:1), and mix evenly.

[0094] Outer aqueous phase: Add 40 g of polyvinyl alcohol (PVA) powder to 4 L of deionized water, heat to 80 °C and stir continuously for 2 hours, and form a 1% concentration PVA aqueous solution after cooling to room temperature.

[0095] Primary emulsification

[0096] Place the oil phase in an ice - water bath (0 - 5 °C), and add the inner aqueous phase dropwise to the oil phase at a rate of 60 mL / min through a constant - flow pump.

[0097] Turn on the ultrasonic homogenizer (power 50 W, probe diameter 10 mm), set the homogenizer speed to 13000 rpm, and emulsify continuously for 5 minutes.

[0098] Secondary emulsification

[0099] Add the primary emulsion to the outer aqueous phase at a rate of 80 mL / min.

[0100] Turn on the mechanical stirrer (rotation speed 300 rpm, blade diameter 50 mm) and the homogenizer (rotation speed 2000 rpm) simultaneously, and continue the double emulsification for 30 minutes.

[0101] Solidify in stages

[0102] First-stage solidification: Transfer the double emulsion to a 30 °C constant temperature water bath and solidify it at a stirring speed of 600 rpm for 2 hours.

[0103] Second-stage solidification: Raise the temperature to 45 °C, reduce the stirring speed to 400 rpm, and continue to solidify for 24 hours.

[0104] Post-treatment

[0105] The solidified microsphere suspension was collected by vacuum filtration, added with 1.0 L of 0.8% sodium hydroxide solution and soaked for 3 hours to remove residual ammonium bicarbonate.

[0106] Centrifuge and wash with deionized water 6 times (5000 rpm each time, 10 minutes) to remove alkaline substances.

[0107] The washed microspheres were classified by a 200-mesh sieve and freeze-dried for 48 hours to obtain the final product.

[0108] Product characterization

[0109] SPAN value = 0.530 (D10 = 80 μm, D50 = 92 μm, D90 = 105 μm).

[0110] Porosity = 78%, pore size CV value = 18%.

[0111] SEM analysis: The pore size distribution of some microspheres is uneven, and the pore connectivity in local areas is poor ( Figure 2 ).

[0112] Example 3: Increase the ultrasonic power (150 W)

[0113] Experimental procedure

[0114] Phase system preparation

[0115] Inner aqueous phase: Dissolve 16 g of ammonium bicarbonate in 250 mL of deionized water, stir until completely dissolved, and the concentration is 6.4% (w / w).

[0116] Oil phase: Add 50 g of PDLLA powder to 600 mL of dichloromethane (DCM), stir magnetically until completely dissolved, and then add 200 mL of petroleum ether (the volume ratio of DCM to petroleum ether is 3:1), and mix evenly.

[0117] External aqueous phase: 40 g of polyvinyl alcohol (PVA) powder was added to 4 L of deionized water, heated to 80 °C and continuously stirred for 2 hours. After cooling to room temperature, a 1% concentration PVA aqueous solution was formed.

[0118] Initial emulsification

[0119] The oil phase was placed in an ice-water bath (0 - 5 °C), and the internal aqueous phase was added dropwise to the oil phase through a constant flow pump at a rate of 60 mL / min.

[0120] The ultrasonic homogenizer (power 150 W, probe diameter 10 mm) was turned on, the homogenizer speed was set to 13000 rpm, and emulsification was continued for 5 minutes.

[0121] Secondary emulsification

[0122] The initial emulsion was added to the external aqueous phase at a rate of 80 mL / min.

[0123] At the same time, a mechanical stirrer (rotation speed 300 rpm, blade diameter 50 mm) and a homogenizer (rotation speed 2000 rpm) were turned on, and secondary emulsification was continued for 30 minutes.

[0124] Stepwise curing

[0125] First-stage curing: The complex emulsion was transferred to a 30 °C constant temperature water bath and cured at a stirring speed of 600 rpm for 2 hours.

[0126] Second-stage curing: The temperature was raised to 45 °C, the stirring speed was reduced to 400 rpm, and curing was continued for 24 hours.

[0127] Post-treatment

[0128] The cured microsphere suspension was collected by vacuum filtration, added with 1.0 L of 0.8% sodium hydroxide solution and soaked for 3 hours to remove residual ammonium bicarbonate.

[0129] Centrifugally washed 6 times with deionized water (5000 rpm, 10 minutes each time) to remove alkaline substances.

[0130] The washed microspheres were classified through a 200-mesh sieve and freeze-dried for 48 hours to obtain the final product.

[0131] Product characterization

[0132] SPAN value = 0.518 (D10 = 85 μm, D50 = 93 μm, D90 = 100 μm).

[0133] Porosity = 82%, pore size CV value = 12%.

[0134] SEM analysis: The pore size distribution is more uniform, but microcracks appear on the surface of some microspheres ( Figure 3 ).

[0135] Example 4: Reducing the proportion of petroleum ether (volume ratio 7:1, petroleum ether 10%)

[0136] Experimental procedure

[0137] Preparation of the phase system

[0138] Inner aqueous phase: Dissolve 16 g of ammonium bicarbonate in 250 mL of deionized water, stir until completely dissolved, and the concentration is 6.4% (w / w).

[0139] Oil phase: Add 50 g of PDLLA powder to 700 mL of dichloromethane (DCM), stir magnetically until completely dissolved, and then add 100 mL of petroleum ether (volume ratio of DCM to petroleum ether is 7:1), and mix evenly.

[0140] Outer aqueous phase: Add 40 g of polyvinyl alcohol (PVA) powder to 4 L of deionized water, heat to 80 °C and continuously stir for 2 hours, and form a 1% concentration PVA aqueous solution after cooling to room temperature.

[0141] Primary emulsification

[0142] Place the oil phase in an ice-water bath (0 - 5 °C), and drop the inner aqueous phase into the oil phase at a rate of 60 mL / min through a constant flow pump.

[0143] Turn on the ultrasonic homogenizer (power 100 W, probe diameter 10 mm), set the rotation speed of the homogenizer to 13000 rpm, and continuously emulsify for 5 minutes.

[0144] Secondary emulsification

[0145] Add the primary emulsion to the outer aqueous phase at a rate of 80 mL / min.

[0146] At the same time, turn on the mechanical stirrer (rotation speed 300 rpm, blade diameter 50 mm) and the homogenizer (rotation speed 2000 rpm), and continuously perform secondary emulsification for 30 minutes.

[0147] Stepwise curing

[0148] First-stage curing: Transfer the secondary emulsion to a 30 °C constant temperature water bath and cure at a stirring speed of 600 rpm for 2 hours.

[0149] Second-stage curing: Raise the temperature to 45 °C, reduce the stirring speed to 400 rpm, and continue to cure for 24 hours.

[0150] Post-treatment

[0151] Collect the cured microsphere suspension by vacuum filtration, add 1.0 L of 0.8% sodium hydroxide solution and soak for 3 hours to remove residual ammonium bicarbonate.

[0152] Centrifuge and wash with deionized water 6 times (5000 rpm, 10 minutes each time) to remove alkaline substances.

[0153] The washed microspheres were classified through a 200-mesh sieve and freeze-dried for 48 hours to obtain the final product.

[0154] Product characterization

[0155] SPAN value = 0.691 (D10 = 70 μm, D50 = 85 μm, D90 = 115 μm).

[0156] Porosity = 79%, CV value of pore size = 22%.

[0157] SEM analysis: The particle size distribution of the microspheres is broad, and some microspheres are adhered ( Figure 4 ).

[0158] Example 5: Reduce the homogenizer speed (8000 rpm)

[0159] Experimental procedure

[0160] Phase system preparation

[0161] Inner aqueous phase: Dissolve 16 g of ammonium bicarbonate in 250 mL of deionized water, stir until completely dissolved, and the concentration is 6.4% (w / w).

[0162] Oil phase: Add 50 g of PDLLA powder to 600 mL of dichloromethane (DCM), stir magnetically until completely dissolved, and then add 200 mL of petroleum ether (the volume ratio of DCM to petroleum ether is 3:1), and mix evenly.

[0163] Outer aqueous phase: Add 40 g of polyvinyl alcohol (PVA) powder to 4 L of deionized water, heat to 80 °C and stir continuously for 2 hours, and form a 1% concentration of PVA aqueous solution after cooling to room temperature.

[0164] Primary emulsification

[0165] Place the oil phase in an ice-water bath (0 - 5 °C), and the inner aqueous phase is added dropwise to the oil phase through a constant flow pump at a rate of 60 mL / min.

[0166] Turn on the ultrasonic homogenizer (power 100 W, probe diameter 10 mm), set the homogenizer speed to 8000 rpm, and continuously emulsify for 5 minutes.

[0167] Secondary emulsification

[0168] Add the primary emulsion to the outer aqueous phase at a rate of 80 mL / min.

[0169] At the same time, turn on the mechanical stirrer (speed 300 rpm, blade diameter 50 mm) and the homogenizer (speed 2000 rpm), and continuously secondary emulsify for 30 minutes.

[0170] Stage-by-stage curing

[0171] First-stage curing: Transfer the complex emulsion to a 30 °C constant temperature water bath and cure it at a stirring speed of 600 rpm for 2 hours.

[0172] Second-stage curing: Raise the temperature to 45 °C, lower the stirring speed to 400 rpm, and continue curing for 24 hours.

[0173] Post-treatment

[0174] The cured microsphere suspension is collected by vacuum filtration, immersed in 1.0 L of 0.8% sodium hydroxide solution for 3 hours to remove residual ammonium bicarbonate.

[0175] Centrifuge and wash 6 times with deionized water (5000 rpm, 10 minutes each time) to remove alkaline substances.

[0176] The washed microspheres are classified through a 200-mesh sieve and freeze-dried for 48 hours to obtain the final product.

[0177] Product characterization

[0178] SPAN value = 0.615 (D10 = 75 μm, D50 = 88 μm, D90 = 110 μm).

[0179] Porosity = 76%, pore size CV value = 25%.

[0180] SEM analysis: The emulsion droplets are unevenly dispersed, and there are significant differences in the microsphere sizes ( Figure 5 ).

[0181] Example 6: Increase the homogenizer speed (16000 rpm)

[0182] Experimental procedure

[0183] Phase system preparation

[0184] Inner aqueous phase: Dissolve 16 g of ammonium bicarbonate in 250 mL of deionized water, stir until completely dissolved, and the concentration is 6.4% (w / w).

[0185] Oil phase: Add 50 g of PDLLA powder to 600 mL of dichloromethane (DCM), stir magnetically until completely dissolved, and then add 200 mL of petroleum ether (the volume ratio of DCM to petroleum ether is 3:1), and mix evenly.

[0186] Outer aqueous phase: Add 40 g of polyvinyl alcohol (PVA) powder to 4 L of deionized water, heat to 80 °C and continuously stir for 2 hours, and form a 1% concentration PVA aqueous solution after cooling to room temperature.

[0187] Primary emulsification

[0188] Place the oil phase in an ice-water bath (0 - 5 °C), and add the inner aqueous phase dropwise to the oil phase through a constant flow pump at a rate of 60 mL / min.

[0189] Turn on the ultrasonic homogenizer (power 100 W, probe diameter 10 mm), set the homogenizer speed to 16,000 rpm, and continue emulsifying for 5 minutes.

[0190] Double emulsification

[0191] Add the primary emulsion to the outer aqueous phase at a rate of 80 mL / min.

[0192] At the same time, turn on the mechanical stirrer (rotation speed 300 rpm, blade diameter 50 mm) and the homogenizer (rotation speed 2000 rpm), and continue double emulsification for 30 minutes.

[0193] Stepwise curing

[0194] First-stage curing: Transfer the double emulsion to a 30 °C constant temperature water bath and cure it at a stirring speed of 600 rpm for 2 hours.

[0195] Second-stage curing: Raise the temperature to 45 °C, reduce the stirring speed to 400 rpm, and continue curing for 24 hours.

[0196] Post-treatment

[0197] Collect the cured microsphere suspension by vacuum filtration, add 1.0 L of 0.8% sodium hydroxide solution and soak for 3 hours to remove residual ammonium bicarbonate.

[0198] Centrifuge and wash 6 times with deionized water (5000 rpm, 10 minutes each time) to remove alkaline substances.

[0199] The washed microspheres are classified through a 200-mesh sieve and freeze-dried for 48 hours to obtain the final product.

[0200] Product characterization

[0201] SPAN value = 0.505 (D10 = 88 μm, D50 = 95 μm, D90 = 102 μm).

[0202] Porosity = 83%, pore size CV value = 9%.

[0203] SEM analysis: The particle size of the microspheres is highly uniform, and the pore size distribution is narrow ( Figure 6 ).

[0204] Comparative example 1: Petroleum ether not added

[0205] The difference from Example 1 is that:

[0206] Phase system preparation:

[0207] The oil phase only uses 800 mL of DCM and does not add petroleum ether. Other components are the same as those in Example 1.

[0208] Product characterization

[0209] SPAN value = 0.892 (D10 = 50 μm, D50 = 80 μm, D90 = 150 μm).

[0210] Porosity = 75%, pore size CV value = 35%.

[0211] Comparative Example 2: Single-stage curing

[0212] The difference from Example 1 is as follows:

[0213] Curing: Only cure for 24 hours under stirring at 30°C and 600 rpm, and omit the second-stage curing.

[0214] Product characterization

[0215] SPAN value = 0.720 (D10 = 65 μm, D50 = 85 μm, D90 = 120 μm).

[0216] Porosity = 68%, pore size CV value = 28%.

[0217] Comparative Example 3: Without ultrasonic assistance (only mechanical stirring)

[0218] The difference from Example 1 is as follows:

[0219] Initial emulsification: Turn off the ultrasonic homogenizer and only use mechanical stirring (8000 rpm) for emulsification for 5 minutes.

[0220] Product characterization

[0221] SPAN value = 0.850 (D10 = 55 μm, D50 = 80 μm, D90 = 140 μm).

[0222] Porosity = 72%, pore size CV value = 38%.

[0223] Comparative Example 4: Low concentration of ammonium bicarbonate (4%)

[0224] The difference from Example 1 is as follows:

[0225] Phase system preparation:

[0226] The internal aqueous phase is adjusted to a 4% ammonium bicarbonate solution (10 g of ammonium bicarbonate dissolved in 250 mL of water), and other components are the same as those in Example 1.

[0227] Product characterization

[0228] SPAN value = 0.750 (D10 = 70 μm, D50 = 90 μm, D90 = 130 μm).

[0229] Porosity = 65%, pore size CV value = 30%.

[0230] Through the systematic experiments of the above examples and comparative examples, the following conclusions can be drawn:

[0231] Synergistic effect of ultrasonic power and homogenizer rotation speed:

[0232] The SPAN values of Example 1 (100W + 13000rpm) and Example 6 (16000rpm) are the lowest (0.505 - 0.512), indicating that high shear force can significantly improve the particle size uniformity.

[0233] Comparing Example 2 (50W) and Example 3 (150W), too high or too low ultrasonic power will lead to performance degradation, verifying the necessity of the range of 50 - 150W in the claims.

[0234] Key influence of petroleum ether on particle size distribution:

[0235] The SPAN value of Comparative Example 1 (without adding petroleum ether) is as high as 0.892, while the SPAN value of Example 1 (20% petroleum ether) drops to 0.512, proving that petroleum ether reduces particle size dispersion by regulating precipitation kinetics.

[0236] Necessity of staged curing:

[0237] The porosity of Comparative Example 2 (single-stage curing) is only 68%, significantly lower than 80% of Example 1, indicating that staged curing optimizes the pore structure by controlling the solvent evaporation rate.

[0238] Scientific range of internal aqueous phase concentration:

[0239] The porosity of Comparative Example 4 (4% ammonium bicarbonate) is only 65%, far lower than the range of 5.2% - 6.4% required by the claims, highlighting the key role of the internal aqueous phase concentration in pore-forming effect.

[0240] Although the embodiments of the present invention have been shown and described, for those of ordinary skill in the art, it can be understood that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principles and spirit of the present invention. The scope of the present invention is defined by the appended claims and their equivalents.

[0241] The above describes the present invention and its implementation manners. Such description is not restrictive. What is shown in the drawings is only one of the implementation manners of the present invention, and the actual application is not limited thereto. In general, if those of ordinary skill in the art are inspired by it and, without departing from the gist of the present invention, design in a non-creative way similar ways and embodiments to this technical solution, they shall fall within the protection scope of the present invention.

Claims

1. A method for batch preparation of PDLLA porous microspheres with high porosity and uniform particle size distribution, characterized in that, It includes the following steps: (1) Prepare the internal aqueous phase, oil phase and external aqueous phase; (2) Add the internal aqueous phase to the oil phase at low temperature for primary emulsification to form a primary emulsion; (3) Add the primary emulsion to the external aqueous phase for secondary emulsification to form a double emulsion; (4) Cure the double emulsion in stages to form porous microspheres; (5) Perform post-treatment to obtain the final product.

2. The method according to claim 1, wherein The internal aqueous phase is an aqueous ammonium bicarbonate solution with a concentration of 5.2% - 6.4%.

3. The method according to claim 1, characterized in that, The oil phase contains a mixed solvent of a dichloromethane solution of PDLLA and petroleum ether, and the volume ratio of the two is 3:1 - 7:

1.

4. The method according to claim 1, wherein In the primary emulsification step, an ultrasonic homogenizer is used, with an ultrasonic power of 50 - 150 W and a homogenizer rotation speed of 8000 - 16000 rpm.

5. The method according to claim 1, characterized in that, In the secondary emulsification step, mechanical stirring and a homogenizer are turned on simultaneously. The mechanical stirring rate is 300 - 600 rpm, and the homogenizer rate is 2000 - 4000 rpm.

6. The method according to claim 1, characterized in that, The staged curing includes: the first stage is cured for 1 - 3 hours under stirring at 20 - 40°C and 600 - 400 rpm; the second stage is cured for 20 - 36 hours under stirring at 45 - 55°C and 300 - 400 rpm.

7. The method according to claim 1, characterized in that, The post-treatment includes soaking in an alkaline solution, washing with water, screening and freeze-drying.

8. The method according to claim 7, characterized in that, The alkaline solution is a 0.8% sodium hydroxide solution, and the treatment time is 2 - 4 hours.

9. The method according to claim 1, characterized in that, The external aqueous phase is an aqueous solution of polyvinyl alcohol (PVA) with a concentration of 1% - 1.8%.

10. The method according to claim 1, characterized in that, In the primary emulsification and secondary emulsification steps, the addition rates of the internal aqueous phase and the external aqueous phase are 40 - 60 ml / min and 60 - 80 ml / min respectively.

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