A biodegradable microsphere sustained-release preparation loaded with bupivacaine and a preparation method thereof

Abstract

The present invention discloses a biodegradable microsphere sustained-release preparation loaded with bupivacaine and a preparation method thereof. Using a biodegradable polymer material as a carrier, the sustained-release microspheres loaded with bupivacaine are prepared by an electrostatic spraying method. The mass percentages of the components in the microspheres respectively account for the total weight of the microspheres as follows: polymer material: 50% - 93%; bupivacaine: 7% - 50%. Compared with the traditional complex preparation methods of drug microspheres, the microspheres prepared by the present invention have uniform particle sizes; the encapsulation rate and drug loading rate of the loaded microspheres are relatively high, the encapsulation rate is above 80%, and the drug loading rate can reach 50%; the drug can be slowly released for 72 hours, and the burst release rate is relatively low, and the drug release in the first 4 hours is below 35%; the preparation method is simple and feasible; the reaction conditions require low; it is easy to be prepared and scaled up in the laboratory.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and particularly relates to a biodegradable microsphere sustained-release preparation loaded with bupivacaine and a preparation method thereof. Background Art

[0002] Acute postoperative pain is a common complaint among patients after invasive surgical operations, usually occurring within 48 hours after surgery. It is reported that these operations can cause persistent neuropathic pain, and more than 50% of patients will experience this pain after surgery. Poor control of acute postoperative pain will seriously affect the quality of life and functional recovery of patients, increase the probability of postoperative complications, and prolong the hospital stay of patients. Clinically, opioids are commonly used to achieve postoperative pain management, but serious side effects such as nausea, vomiting, and respiratory depression will occur during the application of such drugs.

[0003] Bupivacaine (Bup) is a type of long-acting amide local anesthetic with a fast onset and strong anesthetic and analgesic effects. It is widely used in spinal anesthesia, epidural anesthesia, brachial plexus block, postoperative pain, and the treatment of some chronic severe cancer pain. However, its half-life is relatively short, resulting in its analgesic effect usually only lasting for 3 to 6 hours. To achieve a continuous local anesthetic analgesic effect, multiple administrations are often required, and high doses may cause cardiac and nerve toxicity, limiting its clinical application in postoperative analgesia. Therefore, the preparation of biodegradable sustained-release microspheres of bupivacaine can effectively improve the problems existing in the clinical application of bupivacaine.

[0004] The first marketed product of bupivacaine is Marcaine, an injection of bupivacaine hydrochloride, which was developed and marketed by Hospira in 1972 for anesthesia and analgesia in surgical operations, oral surgeries, diagnostic and therapeutic procedures, and obstetric operations. Due to its short local anesthetic effect, it is often necessary to extend the analgesic time through methods such as catheters, patient-controlled pumps, and nerve destruction, but this may cause problems such as infection, itching, and lower limb weakness, reducing patient compliance. To solve this problem, Pacira Pharmaceuticals launched a liposomal suspension injection of bupivacaine in 2011, becoming the first long-acting injectable analgesic of bupivacaine directly injected into the surgical site. However, due to the characteristics of liposomes, its encapsulation efficiency and total drug loading are limited, resulting in poor effects in large peripheral nerve trunk blocks. In addition, Posimir, a gel solution of bupivacaine from Durect, was approved by the FDA for marketing in 2021 and is suitable for analgesia 72 hours after arthroscopic subacromial decompression surgery. Although it uses SABER technology and can form a drug depot gel that can be continuously released in the body after injection. However, clinical data show that its analgesic effect basically disappears after 24 hours, showing no significant difference from the placebo.

[0005] Microsphere preparations have become a promising choice for sustained and controlled release preparations due to their controllable release period. Poly(lactic-co-glycolic acid) (PLGA) is a biodegradable organic polymer, and its hydrolysis products are non-toxic to the human body, so it is widely used in drug sustained release carriers. There are various preparation methods for PLGA as a drug carrier, including spray drying, emulsion solvent evaporation method, phase separation method, salting-out method, membrane emulsification method, electrostatic spraying method, etc.

[0006] The emulsion solvent evaporation method is commonly used but the preparation process is discontinuous and requires controlling many variables; the phase separation method is easily affected by residual solvents and coagulants, and there is insufficient control over the microsphere particle size and uniformity; the salting-out method is cumbersome; the membrane emulsification method has a low drug loading rate. Therefore, in-depth research is needed to develop safe, economical, healthy, and controllable microsphere preparation technologies. Electrostatic spraying technology is a simple method for preparing core-shell structured polymer microspheres. The main equipment includes an injection pump, a liquid feeder, a high-voltage DC power supply, and a collector. A jet is formed by an externally applied electric field force, and micro / nano-scale microspheres are obtained at the receiving end after solvent evaporation. Compared with traditional methods, it theoretically has the following advantages: (1) ultra-micron particle size; (2) narrow size distribution; (3) monodispersity; (4) simple operation and good repeatability; (5) conducive to encapsulation.

[0007] However, up to now, there are still relatively few studies on preparing polymer microspheres loaded with bupivacaine by the electrostatic spraying method. Although there are literature reports on preparing microspheres loaded with levobupivacaine hydrochloride, lidocaine, and acemetacin by the electrospray method, which can achieve a slow release of 30 days in vitro, the microspheres prepared by this method have poor particle size uniformity and low drug loading rate. The theoretical drug loading of levobupivacaine hydrochloride is only 5%. Summary of the Invention

[0008] The purpose of this part is to outline some aspects of the embodiments of the present invention and briefly introduce some preferred embodiments. Some simplifications or omissions may be made in this part, as well as in the abstract and title of the present application, to avoid obscuring the purpose of this part, the abstract, and the title, and such simplifications or omissions shall not be used to limit the scope of the present invention.

[0009] In view of the above and / or problems existing in the prior art, the present invention is proposed.

[0010] Therefore, the purpose of the present invention is to overcome the deficiencies in the prior art and provide a preparation method for a biodegradable microsphere sustained release preparation loaded with bupivacaine.

[0011] To solve the above technical problems, the present invention provides the following technical solution: A preparation method for a biodegradable microsphere sustained release preparation loaded with bupivacaine, including,

[0012] Add the biodegradable polymer material to an organic solvent and stir until completely dissolved to obtain a carrier material. Then add bupivacaine and stir to dissolve it to obtain a homogeneous solution.

[0013] Perform high-voltage electrostatic spraying on the obtained homogeneous solution. Among them, control the flow rate at 0.5 - 1.0 mL / h, the voltage at 6 - 15 kV, the inner diameter of the spraying needle at 0.6 mm, and the receiving distance at 10 - 20 cm.

[0014] Dry the product obtained by high-voltage electrostatic spraying overnight at room temperature to obtain the biodegradable microsphere sustained-release preparation encapsulating bupivacaine.

[0015] As a preferred embodiment of the preparation method of the present invention, wherein: the biodegradable polymer material includes one or more of polylactic acid (PLA), polyglycolic acid (PGA), polycaprolactone (PCL), and poly (lactic-co-glycolic acid) (PLGA).

[0016] As a preferred embodiment of the preparation method of the present invention, wherein: the organic solvent includes one or more of dichloromethane, acetonitrile, chloroform, ethyl acetate, ethyl propionate, propyl acetate, hexafluoroisopropanol, and acetone.

[0017] As a preferred embodiment of the preparation method of the present invention, wherein: the biodegradable polymer material is poly (lactic-co-glycolic acid) (PLGA).

[0018] As a preferred embodiment of the preparation method of the present invention, wherein: the molecular weight of the PLGA is 0.7w - 2.0w.

[0019] As a preferred embodiment of the preparation method of the present invention, wherein: the molar ratio of LA to GA in the PLGA is 75:25 - 50:50.

[0020] As a preferred embodiment of the preparation method of the present invention, wherein: the organic solvent is dichloromethane.

[0021] As a preferred embodiment of the preparation method of the present invention, wherein: the mass volume of the PLGA in the organic solvent is 7% - 20% w / v, and the mass ratio of bupivacaine to the carrier material is 1:1 - 1:10.

[0022] Another object of the present invention is to overcome the deficiencies in the prior art and provide a biodegradable microsphere sustained-release preparation encapsulating bupivacaine. The particle size of the microspheres is distributed between 1 μm and 10 μm, and the average particle size is between 2 μm and 5 μm.

[0023] As a preferred embodiment of the biodegradable microsphere sustained-release preparation loaded with bupivacaine according to the present invention, wherein: the microspheres loaded with bupivacaine exhibit a slow release for more than 72 hours.

[0024] Advantages of the present invention:

[0025] (1) The present invention uses the biodegradable material PLGA as the carrier material, which is simple and easy to obtain. The in vitro release rate can be adjusted by adjusting the LA / GA ratio, PLGA molecular weight, and drug loading amount in the carrier material, and the drug release can be controlled for more than 72 hours, solving the problem of its short half-life and improving its clinical application in postoperative analgesia.

[0026] (2) The present invention uses the electrospray method to prepare bupivacaine-loaded sustained-release microspheres. The preparation conditions are controllable, the operation is simple and easy, the drug loading amount and encapsulation rate of the prepared microspheres are relatively high, the administration dose can be reduced, and the drug use safety can be improved. Description of the drawings

[0027] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following will briefly introduce the drawings required for the description of the embodiments. Obviously, the following drawings are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can also be obtained based on these drawings. Among them:

[0028] Figure 1 Schematic diagram of the preparation method of PLGA microspheres loaded with bupivacaine according to the present invention;

[0029] Figure 2 SEM and particle size distribution diagram of PLGA microspheres loaded with bupivacaine prepared in Example 1;

[0030] Figure 3 SEM and particle size distribution diagram of PLGA microspheres loaded with bupivacaine prepared in Example 2;

[0031] Figure 4 SEM and particle size distribution diagram of PLGA microspheres loaded with bupivacaine prepared in Example 3;

[0032] Figure 5 SEM and particle size distribution diagram of PLGA microspheres loaded with bupivacaine prepared in Example 4;

[0033] Figure 6 SEM and particle size distribution diagram of PLGA microspheres loaded with bupivacaine prepared in Example 5;

[0034] Figure 7 SEM and particle size distribution diagram of pure PLGA microspheres prepared in Comparative Example 2;

[0035] Figure 8 PXRD patterns of the bupivacaine-loaded PLGA microspheres prepared in Examples 1-5, pure bupivacaine in Comparative Example 1, and pure PLGA microspheres in Comparative Example 2;

[0036] Figure 9 DSC curves of the bupivacaine-loaded PLGA microspheres prepared in Examples 1-5, pure bupivacaine in Comparative Example 1, and pure PLGA microspheres in Comparative Example 2;

[0037] Figure 10 FTIR spectra of the bupivacaine-loaded PLGA microspheres prepared in Examples 1-5, pure bupivacaine in Comparative Example 1, and pure PLGA microspheres in Comparative Example 2;

[0038] Figure 11 Drug release curves of the bupivacaine-loaded PLGA microspheres prepared in Examples 1-5 and pure bupivacaine in Comparative Example 1.

[0039] Figure 12 Optical microscope images of the pure PLGA microspheres prepared in Comparative Examples 3-9. Detailed Description of the Invention

[0040] To make the above objects, features, and advantages of the present invention more apparent and understandable, the following detailed description of the specific embodiments of the present invention will be given in conjunction with the examples of the specification.

[0041] In the following description, many specific details are set forth to fully understand the present invention. However, the present invention can also be implemented in other ways different from those described herein. Those skilled in the art can make similar generalizations without departing from the connotation of the present invention. Therefore, the present invention is not limited by the specific embodiments disclosed below.

[0042] Secondly, the so-called "one embodiment" or "embodiment" herein refers to a specific feature, structure, or characteristic that can be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places in this specification does not necessarily refer to the same embodiment, nor is it a separate or alternative embodiment that excludes other embodiments.

[0043] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods.

[0044] Unless otherwise specified, the materials, reagents, etc. used in the following examples are all commercially available.

[0045] Example 1

[0046] A method for preparing bupivacaine-loaded PLGA microspheres, comprising the following steps:

[0047] (1) Add 400 mg of PLGA (LA:GA = 75:25, Mw = 8076) to 4 mL of dichloromethane, stir magnetically until completely dissolved, then add 40 mg of bupivacaine (i.e., Bup:PLGA = 1:10, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0048] (2) Subject the homogeneous solution obtained in (1) to high-voltage electrostatic spraying, controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0049] (3) Dry the product obtained in (2) overnight at room temperature to obtain the bupivacaine-loaded sustained-release microspheres.

[0050] For the schematic diagram of the preparation method of PLGA microspheres loaded with bupivacaine, see Figure 1 .

[0051] Example 2

[0052] A preparation method of PLGA microspheres loaded with bupivacaine, comprising the following steps:

[0053] (1) Add 600 mg of PLGA (LA:GA = 75:25, Mw = 8076) to 3 mL of dichloromethane, stir magnetically until completely dissolved, then add 60 mg of bupivacaine (i.e., Bup:PLGA = 1:10, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0054] (2) Subject the homogeneous solution obtained in (1) to high-voltage electrostatic spraying, controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0055] (3) Dry the product obtained in (2) overnight at room temperature to obtain the bupivacaine-loaded sustained-release microspheres.

[0056] Example 3

[0057] A preparation method of PLGA microspheres loaded with bupivacaine, comprising the following steps:

[0058] (1) Add 800 mg of PLGA (LA:GA = 75:25, Mw = 19422) to 4 mL of dichloromethane, stir magnetically until completely dissolved, then add 80 mg of bupivacaine (i.e., Bup:PLGA = 1:10, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0059] (2) Carry out high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 1.0 mL / h, the voltage at 8.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0060] (3) Dry the product obtained in (2) overnight at room temperature to obtain the bupivacaine-loaded sustained-release microspheres.

[0061] Example 4

[0062] A method for preparing bupivacaine-loaded PLGA microspheres, comprising the following steps:

[0063] (1) Add 400 mg of PLGA (LA:GA = 75:25, Mw = 19422) to 2 mL of dichloromethane, magnetically stir until completely dissolved, then add 80 mg of bupivacaine (i.e., Bup:PLGA = 1:5, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0064] (2) Carry out high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 1.0 mL / h, the voltage at 9.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0065] (3) Dry the product obtained in (2) overnight at room temperature to obtain the bupivacaine-loaded sustained-release microspheres.

[0066] Example 5

[0067] A method for preparing bupivacaine-loaded PLGA microspheres, comprising the following steps:

[0068] (1) Add 400 mg of PLGA (LA:GA = 75:25, Mw = 19422) to 3 mL of dichloromethane, magnetically stir until completely dissolved, then add 400 mg of bupivacaine (i.e., Bup:PLGA = 1:1, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0069] (2) Carry out high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 1.0 mL / h, the voltage at 9.50 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0070] (3) Dry the product obtained in (2) overnight at room temperature to obtain the bupivacaine-loaded sustained-release microspheres.

[0071] Table 1 Prescription composition and quality evaluation of Examples 1-5

[0072]

[0073]

[0074] Comparative Example 1

[0075] Bupivacaine.

[0076] Comparative Example 2

[0077] The pure PLGA microspheres prepared by the electrospray method, the preparation method includes the following steps:

[0078] (1) Add 600 mg of PLGA (LA:GA = 75:25, Mw = 19422) to 3 mL of dichloromethane, and stir magnetically until completely dissolved to obtain a homogeneous solution for use;

[0079] (2) Perform high-voltage electrospray on the homogeneous solution obtained in (1), control the flow rate to be 1.0 mL / h, the voltage to be 9.00 kV, the inner diameter of the spray needle to be 0.6 mm, and the receiving distance from the needle to the aluminum foil to be 10 cm;

[0080] (3) Dry the product obtained in (2) overnight at room temperature to obtain the blank pure PLGA microspheres.

[0081] Perform structural characterization and performance testing on the Bupivacaine-loaded PLGA microspheres prepared in Examples 1-5 and Comparative Examples 1-2:

[0082] 1. Scanning electron microscope (SEM) characterization:

[0083] The acceleration voltage is 5 kV, and observe the surface morphology of the Bup / PLGA microspheres prepared in Examples 1-5 and the pure PLGA microspheres of Comparative Example 2.

[0084] Figures 2 - 7 For the SEM images and particle size distribution diagrams of the Bup / PLGA microspheres prepared in Examples 1-5 and the pure PLGA microspheres of Comparative Example 2, from Figures 2 - 6 It can be seen from the SEM images in that the Bup / PLGA microspheres prepared under different conditions are all three-dimensional spherical structures with slightly wrinkled surfaces, uniform, round and well-dispersed.

[0085] Use Nano Measurer 1.2 to randomly select 100 microspheres to draw a particle size frequency distribution histogram: From Figures 2 - 7 It can be seen from the particle size distribution diagram in that the particle sizes of the microspheres in Examples 1-5 are distributed between 1.6 μm and 6.4 μm, and the particle sizes of the pure PLGA microspheres in Comparative Example 2 are distributed between 4.8 μm and 10.0 μm. In comparison, the PLGA microspheres in Example 2 have better formability, are round and plump, the particle size between 2.6 μm and 3.4 μm accounts for 88%, the average particle size is 3.03 μm, and the particle size distribution is relatively uniform.

[0086] From Figures 2 - 3It can be seen that increasing the concentration of PLGA in the oil phase can slightly increase the particle size of the microspheres. From Figures 4 - 6 It can be seen that increasing the drug loading amount may reduce the particle size of the microspheres, but the uniformity of the microsphere particle size distribution decreases.

[0087] 2. Powder X-ray diffraction (PXRD) characterization:

[0088] Using Cu-Kα radiation, the scanning diffraction angle range 2θ is 5 - 40°, the scanning speed is 10° / min, the voltage is 40 kV, and the current is 40 mA. Analyze the crystal forms of the Bup / PLGA microspheres prepared in Examples 1 - 5 and Comparative Examples 1 - 2.

[0089] Figure 8 Figure shows the PXRD patterns of the Bup / PLGA microspheres prepared in Examples 1 - 5, pure Bup in Comparative Example 1, and pure PLGA microspheres in Comparative Example 2.

[0090] As can be seen from the figure, Bup has obvious crystal diffraction peaks at 9.93°, 12.61°, 16.16°, 17.77°, 19.99°, 21.47°, 25.44°, and 30.13°. The pure PLGA microspheres in Comparative Example 2 are in an amorphous structure.

[0091] The PXRD patterns of the Bup / PLGA microspheres in Examples 1 - 5 have no obvious diffraction peaks attributed to Bup characteristics. There is only a broad and weak absorption peak near 2θ = 18°, similar to that of the pure PLGA microspheres, indicating that the crystal lattice of Bup is destroyed during the electrospray process, and Bup is highly dispersed in the PLGA matrix in a molecular or amorphous state. The Bup / PLGA microspheres exhibit an amorphous structure similar to that of PLGA. This is because the receiving distance during the electrospray process is relatively short, the solvent evaporation rate is fast, the Bup chains do not have enough time to crystallize, and coupled with the interaction between Bup and PLGA, the crystallization process is inhibited.

[0092] 3. Differential scanning calorimetry (DSC) characterization:

[0093] The heating rate is 10°C / min. Analyze the melting and crystallization behaviors of the Bup / PLGA microspheres prepared in Examples 1 - 5 and Comparative Examples 1 - 2.

[0094] Figure 9DSC analysis charts of the Bup / PLGA microspheres prepared in Examples 1-5, pure Bup in Comparative Example 1, and pure PLGA microspheres in Comparative Example 2. As can be seen from the figure, Bup shows a sharp and narrow melting endothermic peak at 113.73 °C, and the melting enthalpy ΔHm is 113.16 J / g, indicating that Bup has a crystal structure. The pure PLGA microspheres in Comparative Example 2 show a broad and weak melting endothermic peak at 53.08 °C. There is no melting peak attributed to Bup in the Bup / PLGA microspheres of Examples 1-5, but instead shows the same melting trend as PLGA, which also indicates that Bup is completely loaded in PLGA, consistent with the results of PXRD research.

[0095] 4. FTIR Characterization:

[0096] In the ATR-FTIR mode, the test wavelength range is 4000-500 cm -1 , and the characteristic functional groups of the Bup / PLGA microspheres prepared in Examples 1-5 and Comparative Examples 1-2 are analyzed.

[0097] Figure 10 FTIR analysis charts of the Bup / PLGA microspheres prepared in Examples 1-5, pure Bup in Comparative Example 1, and pure PLGA microspheres in Comparative Example 2. As can be seen from the figure, in the PLGA structure, the strong absorption peak at 1746 cm -1 corresponds to the C=O stretching vibration, and 2996 cm -1 , 2947 cm -1 correspond to the C-H stretching vibration, and 1082 cm -1 is the C-O stretching vibration peak. In the Bup structure, the strong absorption peak at 1651 cm -1 corresponds to the C=O stretching vibration on the amide, 3171 cm -1 is the N-H stretching vibration, 2931 cm -1 corresponds to the C-H stretching vibration, 1525 cm -1 and 1230 cm -1 are the coupling peaks of the C-N stretching vibration and the N-H bending vibration, and 768 cm -1 is the bending vibration peak of N-H. For the Bup / PLGA microspheres of Examples 1-5, the absorption peaks of the characteristic functional groups of PLGA still exist and the positions are almost unchanged, indicating that Bup is successfully loaded in PLGA, and the characteristic peaks of Bup begin to appear with the increase of the drug loading amount.

[0098] In vitro Sustained Release Performance Test:

[0099] 1. Chromatographic Parameters and Conditions

[0100] Agilent ZORBAX-C18 (5 μm, 250 mm*4.6 mm) was used as the chromatographic column, phosphate buffer solution (0.015 mol / L NaH2PO4 solution (adjusted to pH 7.2 with 1 M NaOH)) was used as the mobile phase, the column temperature was 35 °C, the detection wavelength was 215 nm, the flow rate was 1.0 mL / min, and the injection volume was 10 μL.

[0101] 2. Determination of drug loading and encapsulation efficiency

[0102] Accurately weigh 10 mg of microspheres into a 50 mL volumetric flask, add 2 mL of dichloromethane and ultrasonicate for 30 min to destroy the microsphere structure, remove the solvent with nitrogen, add methanol to dissolve the precipitate, and ultrasonicate for another 30 min. The solution was filtered through a 0.22 μm filter membrane and then analyzed by HPLC injection. The drug loading (LC) and encapsulation efficiency (EE) were calculated by the following formula:

[0103] Drug loading (%) = weight of Bup in microsphere sample / total weight of microsphere

[0104] Encapsulation efficiency (%) = actual drug loading percentage / theoretical drug loading percentage (the percentage of Bup in the total solid weight of the raw material)

[0105] 3. In vitro release experiment

[0106] Use pH 7.4 phosphate buffer solution with 0.1% sodium dodecyl sulfate (SDS) as the dissolution medium. Accurately weigh 3 mg of Bup and an appropriate amount of Bup / PLGA microspheres (approximately equivalent to 3 mg of Bup) in a 50 mL centrifuge tube, add 50 mL of dissolution medium, and oscillate at a constant temperature of 37.0±0.5°C and 100 r / min. Take samples of 0.5 mL at regular intervals and add an equal volume of fresh medium. According to the "Chromatographic Parameters and Conditions", perform sample injection and determination, and calculate the cumulative drug release percentage according to the following formula to draw the Bup release curve in the microspheres.

[0107] Cumulative drug release rate (%) = released Bup mass / theoretical Bup content in microspheres × 100%

[0108] Figure 11 The drug release curves of Bup / PLGA microspheres prepared in Examples 1 to 5 and pure Bup in Comparative Example 1 are as follows:

[0109] As can be seen from the figure, the Bup / PLGA microspheres prepared in Examples 1 to 5 released rapidly within 48 hours and then entered a slow release phase.

[0110] In Comparative Example 1, more than 66% of the Bup raw material drug was released within 4 hours; in Examples 1 and 2, the Bup / PLGA microspheres with a dosage of about 9% of PLGA (LA:GA = 75:25, Mw = 8076) achieved almost similar release to the raw material drug Bup within 48 hours, and the cumulative release was greater than 80%, indicating that the drug-loaded microspheres had an obvious short-term sustained-release effect.

[0111] When using PLGA with a higher molecular weight (LA:GA = 75:25, Mw = 19422), the prepared drug-loaded microspheres could be slowly released within 72 hours. When the drug loadings were 9%, 16%, and 49% respectively, the cumulative drug releases of the prepared drug-loaded microspheres within 72 hours were 34.35%, 46.78%, and 67.42% respectively, and the drug release was slow without an initial burst phenomenon.

[0112] Thus, the greater the dosage, the faster the drug release rate and the higher the cumulative release rate. From the experimental results, the Bup / PLGA microspheres prepared by the electrospray technique could achieve a sustained-release effect on the loaded Bup without an initial burst phenomenon.

[0113] Comparative Example 3

[0114] This comparative example provides a preparation method for PLGA microspheres loaded with bupivacaine by the electrospray method, including the following steps:

[0115] (1) Add 140 mg of PLGA (LA:GA = 75:25, Mw = 7526) to 2 mL of dichloromethane, stir magnetically until completely dissolved, then add 17.5 mg of bupivacaine (i.e., Bup:PLGA = 1:8, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0116] (2) Perform high-voltage electrospray on the homogeneous solution obtained in (1), control the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0117] (3) Dry the product obtained in (2) overnight at room temperature to obtain the bupivacaine-loaded sustained-release microspheres.

[0118] Comparative Example 4

[0119] This comparative example provides a preparation method for PLGA microspheres loaded with bupivacaine by the electrospray method, including the following steps:

[0120] (1) Add 350 mg of PLGA (LA:GA = 50:50, Mw = 7038) to 5 mL of dichloromethane, stir magnetically until completely dissolved, then add 43.75 mg of bupivacaine (i.e., Bup:PLGA = 1:8, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0121] (2) Perform high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0122] (3) Dry the product obtained in (2) at room temperature overnight to obtain the bupivacaine-loaded sustained-release microspheres.

[0123] Comparative Example 5

[0124] This comparative example provides a preparation method of PLGA microspheres loaded with bupivacaine by electrostatic spraying, including the following steps:

[0125] (1) Add 400 mg of PLGA (LA:GA = 50:50, Mw = 7038) to 4 mL of dichloromethane, stir magnetically until completely dissolved, then add 40.0 mg of bupivacaine (i.e., Bup:PLGA = 1:10, w / w), and stir until completely dissolved to obtain a homogeneous solution for use;

[0126] (2) Perform high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0127] (3) Dry the product obtained in (2) at room temperature overnight to obtain the bupivacaine-loaded sustained-release microspheres.

[0128] Table 2 Formulation composition and quality evaluation of Comparative Examples 3 - 5

[0129]

[0130] Use an optical microscope to observe the morphology of the PLGA microspheres loaded with bupivacaine prepared in Comparative Examples 3 - 5. It can be seen that Figure 12 when the PLGA concentration is low, in addition to spherical shapes, there are also strip-shaped substances in the electrosprayed products, and the particle size uniformity is poor.

[0131] Comparative Example 6

[0132] This comparative example provides a method for preparing pure PLGA microspheres by electrostatic spraying, including the following steps:

[0133] (1) Add 100 mg of PLGA (LA:GA = 75:25, Mw = 7526) to 1 mL of dichloromethane, stir magnetically until completely dissolved to obtain a homogeneous solution for use;

[0134] (2) Perform high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0135] (3) Dry the product obtained in (2) overnight at room temperature to obtain the blank pure PLGA microspheres.

[0136] Comparative Example 7

[0137] This comparative example provides a method for preparing pure PLGA microspheres by electrostatic spraying, which includes the following steps:

[0138] (1) Add 100 mg of PLGA (LA:GA = 75:25, Mw = 7526) to 1 mL of hexafluoroisopropanol, and stir magnetically until completely dissolved to obtain a homogeneous solution for use;

[0139] (2) Perform high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0140] (3) Dry the product obtained in (2) overnight at room temperature to obtain the blank pure PLGA microspheres.

[0141] Comparative Example 8

[0142] This comparative example provides a method for preparing pure PLGA microspheres by electrostatic spraying, which includes the following steps:

[0143] (1) Add 100 mg of PLGA (LA:GA = 50:50, Mw = 7038) to 1 mL of hexafluoroisopropanol, and stir magnetically until completely dissolved to obtain a homogeneous solution for use;

[0144] (2) Perform high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spray needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm;

[0145] (3) Dry the product obtained in (2) overnight at room temperature to obtain the blank pure PLGA microspheres.

[0146] Comparative Example 9

[0147] This comparative example provides a method for preparing pure PLGA microspheres by electrostatic spraying, which includes the following steps:

[0148] (1) Add 100 mg of PLGA (LA:GA = 75:25, Mw = 7526) to 1 mL of a mixed solvent (dichloromethane:hexafluoroisopropanol = 3:1, v / v), and stir magnetically until completely dissolved to obtain a homogeneous solution for use.

[0149] (2) Perform high-voltage electrostatic spraying on the homogeneous solution obtained in (1), controlling the flow rate at 0.5 mL / h, the voltage at 10.00 kV, the inner diameter of the spraying needle at 0.6 mm, and the receiving distance from the needle to the aluminum foil at 10 cm.

[0150] (3) Dry the product obtained in (2) at room temperature overnight to obtain the blank pure PLGA microspheres.

[0151] Table 3 Formulation composition and quality evaluation of Comparative Examples 6-9

[0152]

[0153]

[0154] Comparative Examples 6-9 investigated the effect of the solvent on the morphology of the electrosprayed microspheres. The morphology of the PLGA microspheres loaded with bupivacaine prepared in Comparative Examples 6-9 was observed by optical microscopy. As Figure 12 can be seen, when pure dichloromethane, hexafluoroisopropanol, and the mixed solvent are used as the solvent, the microsphere morphology is better.

[0155] It should be noted that the above examples are only used to illustrate the technical solutions of the present invention and are not restrictive. Although the present invention has been described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present invention can be modified or equivalently replaced without departing from the spirit and scope of the technical solutions of the present invention, and they should all be covered within the scope of the present invention.

Claims

1. A preparation method of a biodegradable microsphere sustained-release preparation loaded with bupivacaine, characterized in that: including, adding a biodegradable polymer material into an organic solvent, stirring until completely dissolved to obtain a carrier material, adding bupivacaine and stirring to dissolve to obtain a homogeneous solution, wherein the biodegradable polymer material is poly(lactic-co-glycolic acid) (PLGA), the molecular weight of the PLGA is 8076 - 19422, and the molar ratio of LA to GA in the PLGA is 75:25; performing high-voltage electrostatic spraying on the obtained homogeneous solution, wherein the flow rate is controlled at 0.5 - 1.0 mL / h, the voltage is 6 - 15 kV, the inner diameter of the spraying needle is 0.6 mm, and the receiving distance is 10 - 20 cm; drying the product obtained by high-voltage electrostatic spraying overnight at room temperature to obtain the biodegradable microsphere sustained-release preparation loaded with bupivacaine; wherein, the organic solvent is dichloromethane; the mass-volume of the PLGA in the organic solvent is 20% w / v, and the mass ratio of bupivacaine to the carrier material is 1:

10.

2. The biodegradable microsphere sustained-release preparation loaded with bupivacaine prepared by the preparation method according to claim 1.

3. The biodegradable microsphere sustained-release preparation loaded with bupivacaine according to claim 2, wherein: The microspheres loaded with bupivacaine achieve a slow release for more than 72 h.

Images