Citric acid fentanyl coated PLGA-PEG nano-microsphere and preparation method thereof

Abstract

The invention relates to the technical field of medicines, in particular to citric acid fentanyl-coated PLGA-PEG nano microspheres and a preparation method thereof. The preparation method comprises the following steps: step S1, preparing a citric acid fentanyl citrate solution and a PLGA-PEG solution; step S2, dropwise adding the citric acid fentanyl solution into the PLGA-PEG solution, mixing, and carrying out ultrasonic emulsification to obtain primary emulsion; s3, dropwise adding the primary emulsion into a PVA (Polyvinyl Alcohol) solution, mixing, and carrying out ultrasonic emulsification to obtain multiple emulsion; and step S4, dropwise adding the multiple emulsion into water, stirring to volatilize the organic solvent, and performing centrifugal filtration to obtain the citric acid fentanyl coated PLGA-PEG nano-microspheres. The PLGA-PEG nano-carrier is adopted as a drug carrier to carry the citric acid fentanyl citrate, and compared with an existing oral administration citric acid fentanyl citrate preparation, slow release of the citric acid fentanyl citrate can be achieved.

Description

Technical Field

[0001] The invention belongs to the technical field of medicine, and specifically relates to fentanyl citrate@PLGA-PEG nano-microspheres and a preparation method thereof. Background Art

[0002] Fentanyl citrate is an analgesic that is suitable for severe pain without shock and respiratory depression. It is used in pre-hospital emergency treatment for trauma, surgical anesthesia, and breakthrough cancer pain.

[0003] Currently, oral fentanyl citrate preparations (such as transmucosal lozenges) are Buccal effervescent tablets Cheek Patches Sublingual tablets ) generally releases quickly, but the duration of drug effect is insufficient, with the longest duration being only 2.5-5 hours. In the case of insufficient medical evacuation forces at the disaster accident site, the shorter duration of drug effect will lead to the repeated use of analgesics, increase patient pain, and increase the risk of side effects. Therefore, it cannot meet the needs of disaster relief analgesia when evacuation conditions are insufficient. Summary of the invention

[0004] In view of the above analysis, the present invention aims to provide a fentanyl citrate@PLGA-PEG nanoparticle and a preparation method thereof, so as to solve the problem that the fentanyl citrate preparation for oral use in the prior art cannot achieve sustained drug release.

[0005] On the one hand, an embodiment of the present invention provides a method for preparing fentanyl citrate@PLGA-PEG nanomicrospheres, comprising: step S1, preparing a fentanyl citrate solution and a PLGA-PEG solution; step S2, dropping the fentanyl citrate solution into the PLGA-PEG solution, mixing and then performing ultrasonic emulsification to obtain colostrum; step S3, dropping the colostrum into the PVA solution, mixing and then performing ultrasonic emulsification to obtain a double emulsion; step S4, dropping the double emulsion into water, stirring to volatilize the organic solvent, and then centrifuging it to obtain fentanyl citrate@PLGA-PEG nanomicrospheres.

[0006] Furthermore, in step S1, preparing the PLGA-PEG solution includes: preparing a mixed solution of dichloromethane and acetone, dissolving PLGA-PEG in the mixed solution of dichloromethane and acetone, and obtaining the PLGA-PEG solution.

[0007] Furthermore, the mass percentage of PEG in the used PLGA-PEG is less than or equal to 33%.

[0008] Furthermore, in step S3, when the colostrum is ultrasonically emulsified, the ultrasonic power is 15 to 30 W.

[0009] Furthermore, in step S1, a fentanyl citrate solution is prepared, including: dissolving fentanyl citrate in a 1% PVA solution to obtain a fentanyl citrate solution.

[0010] Furthermore, the volume ratio of the 1% PVA solution to the mixed solution of dichloromethane and acetone is 1:10 to 1:5.

[0011] Furthermore, in step S1, the mass percentage of fentanyl citrate and PLGA-PEG is 5% to 25%.

[0012] Furthermore, in step S1, the mass percentage of PEG in the PLGA-PEG used is 33%; in step S3, the ultrasonic power is 20W when the colostrum is ultrasonically emulsified; in step S1, the volume ratio of 1% PVA solution to the mixed solution of dichloromethane and acetone is 1:7; in step S1, the mass percentage of fentanyl citrate and PLGA-PEG is 25%.

[0013] On the other hand, an embodiment of the present invention further provides a fentanyl citrate@PLGA-PEG nanoparticle, which is obtained by the preparation method of the above embodiment, and includes fentanyl citrate and a PLGA-PEG nanocarrier carrying fentanyl citrate.

[0014] Furthermore, the sustained release time of fentanyl citrate@PLGA-PEG nanoparticles exceeded 5 days, the drug release rate in the first 15 minutes of in vitro release was not less than 5.36%, the drug loading rate was not less than 10.68%, and the encapsulation rate was not less than 99%.

[0015] Compared with the prior art, the present invention can achieve at least one of the following beneficial effects:

[0016] 1. The present invention uses PLGA-PEG nanocarrier as a drug carrier to carry fentanyl citrate, which is different from the existing oral fentanyl citrate preparations. It can achieve the sustained release of fentanyl citrate, and provide a reference for the drug development of oral preparations for disaster relief analgesia.

[0017] 2. The present invention adjusts the dosage of fentanyl citrate, the mass percentage of PEG in PLGA-PEG, the ultrasonic power when forming an emulsion, the volume ratio of the internal water phase and the oil phase, etc., so that the drug loading amount, encapsulation rate, sustained release time and rapid drug release percentage of fentanyl citrate @PLGA-PEG can achieve better effects at the same time.

[0018] 3. The fentanyl citrate@PLGA-PEG of the present invention can achieve the following performances: sustained release time exceeds 5 days, the drug release rate in the first 15 minutes of in vitro release is not less than 5.36%, the drug loading rate is not less than 10.68%, and the encapsulation rate is not less than 99%.

[0019] In the present invention, the above-mentioned technical solutions can also be combined with each other to achieve more preferred combination solutions. Other features and advantages of the present invention will be described in the subsequent description, and some advantages can become obvious from the description, or can be understood by practicing the present invention. The purpose and other advantages of the present invention can be achieved and obtained through the contents particularly pointed out in the embodiments of the description and the drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The accompanying drawings are only used to illustrate specific embodiments and are not to be considered as limiting the present invention.

[0021] Figure 1 Schematic diagram of fentanyl citrate@PLGA-PEG microspheres according to Example 1 of the present invention;

[0022] Figure 2 (a) to (i) are the particle size distribution diagrams of the samples of Examples 1 to 9, respectively;

[0023] Figure 3 The standard curve of fentanyl citrate for HPLC detection of encapsulation efficiency;

[0024] Figure 4 The figure is a comparison chart of the percentage of drug released of fentanyl citrate at different time points in Examples 1 to 9; wherein a is a curve chart of the percentage of drug released from 15 min to 12 h, and b is a bar chart of the percentage of drug released at the 15th minute;

[0025] Figure 5 The standard curve for fentanyl citrate concentration in the supernatant of the sustained-release solution was determined by high performance liquid chromatography;

[0026] Figure 6 It is a comparison chart of the percentage of drug released of fentanyl citrate in Examples 1 to 9 on days 1 to 6; wherein a is a curve chart of the percentage of drug released on days 1 to 6, and b is a bar chart of the percentage of drug released on day 5;

[0027] Figure 7 The figure is a schematic diagram of the influence of the four parameters A, B, C, and D on the drug performance of fentanyl citrate in Examples 1 to 9, wherein a is the influence of the four parameters on the particle size; b is the influence of the four parameters on the dispersion index; c is the influence of the four parameters on the drug loading rate; d is the influence of the four parameters on the 15 min drug release rate; and e is the influence of the four parameters on the 5-day drug release rate. DETAILED DESCRIPTION

[0028] The preferred embodiments of the present invention will be described in detail below in conjunction with the accompanying drawings, wherein the accompanying drawings constitute a part of the present invention and are used together with the embodiments of the present invention to illustrate the principles of the present invention, but are not used to limit the scope of the present invention.

[0029] In order to accurately describe the embodiments of the present invention, the relevant professional terms are further explained:

[0030] PLGA-PEG: polylactic acid-co-glycolic acid-polyethylene glycol; wherein PLGA is polylactic acid-co-glycolic acid and PEG is polyethylene glycol.

[0031] PLGA-w%PEG: w% indicates the mass percentage of PEG in the composite.

[0032] PVA: polyvinyl alcohol.

[0033] The embodiment of the present invention provides a method for preparing fentanyl citrate@PLGA-PEG nanoparticles, comprising the following steps:

[0034] Step S1, preparing fentanyl citrate solution and PLGA-PEG solution;

[0035] Step S2, adding the fentanyl citrate solution dropwise to the PLGA-PEG solution, mixing and then performing ultrasonic emulsification to obtain colostrum;

[0036] Step S3, adding colostrum dropwise to the PVA solution, mixing and then performing ultrasonic emulsification to obtain a double emulsion;

[0037] Step S4, adding the emulsion dropwise into water, stirring to volatilize the organic solvent, and then centrifuging and filtering the emulsion to obtain fentanyl citrate@PLGA-PEG nanoparticles.

[0038] PLGA is a biodegradable drug controlled release material. The present invention uses PLGA-PEG nanocarrier as a drug carrier to carry fentanyl citrate. Compared with the existing oral fentanyl citrate preparations ( The invention adopts glucose powder, magnesium stearate and the like as auxiliary materials to achieve the sustained release of fentanyl citrate, providing a reference basis and technical support for the research of analgesic drugs for disaster relief. In addition, the present invention adopts the emulsified solvent volatilization method to prepare fentanyl citrate @ PLGA-PEG nano-microspheres, which is classic, stable, reliable and easy to operate.

[0039] In some embodiments of the present invention, in step S1, preparing the PLGA-PEG solution includes: preparing a mixture of dichloromethane and acetone, dissolving PLGA-PEG in the mixture of dichloromethane and acetone, and obtaining the PLGA-PEG solution. The volume ratio of dichloromethane to acetone is: V 二氯甲烷 :V 丙酮 =3:2. When dissolving PLGA-PEG, 3.5 mL of a mixed solution of dichloromethane and acetone can be used to dissolve 100 mg of PLGA-PEG.

[0040] In some embodiments of the present invention, the mass percentage of PEG in the used PLGA-PEG is less than or equal to 33%. The present invention adjusts the mass percentage of PEG in PLGA-PEG to a suitable range, so that fentanyl citrate @PLGA-PEG has excellent sustained-release effect and rapid-release effect, thereby increasing the sustained-release time of fentanyl citrate and achieving rapid release of fentanyl citrate.

[0041] In some embodiments, preparing a fentanyl citrate solution comprises: dissolving fentanyl citrate in a 1% PVA solution to obtain a fentanyl citrate solution.

[0042] In some embodiments, when preparing the fentanyl citrate solution and the PLGA-PEG solution in step S1, the mass percentage of fentanyl citrate and PLGA-PEG is 5% to 25%. The present invention controls the dosage of fentanyl citrate to 5% to 25% of PLGA-PEG, so that the finally obtained fentanyl citrate @PLGA-PEG has a large drug loading, the drug loading is above 2%, and the maximum can reach above 10%; at the same time, the sustained release effect and rapid release effect of fentanyl citrate @PLGA-PEG can also be improved to a certain extent. Among them, the sustained release in the present invention refers to the long-term slow release of fentanyl citrate after use to prolong its action time; rapid release refers to the release of fentanyl citrate in a short time after use.

[0043] In some embodiments, when preparing the fentanyl citrate solution and the PLGA-PEG solution in step S1, the volume ratio of the 1% PVA solution to the mixture of dichloromethane and acetone is 1:10 to 1:5, so that in the colostrum formed after the two are mixed, the volume ratio of the internal water phase to the oil phase (i.e., the mixture of 1% PVA solution and dichloromethane and acetone) is 1:10 to 1:5. The present invention can further improve the sustained-release effect and the rapid-release effect of fentanyl citrate @PLGA-PEG by adjusting the volume ratio of the internal water phase to the oil phase within a suitable range.

[0044] In step S2, the fentanyl citrate solution is added dropwise to the PLGA-PEG solution to obtain an O / W type solution; the solution is ultrasonically emulsified so that the fentanyl citrate solution is evenly dispersed in the PLGA-PEG solution to obtain colostrum. The ultrasonic emulsification can be achieved by using a cell disruptor. The ultrasonic power during ultrasonic emulsification can be 15 to 25 W, for example, 15 W, 18 W, 20 W, 22 W, 25 W.

[0045] In step S3, colostrum is uniformly added to the PVA solution, mixed evenly, and then ultrasonically emulsified to obtain a double emulsion. The PVA solution is a PVA solution with a mass fraction of 5%. In addition, during the dropping process, the dropping can be stirred, for example, by stirring and mixing by magnetic stirring, and the speed of the magnetic stirring can be 300 to 500 rpm to avoid stirring too fast and splashing the solution.

[0046] In this embodiment, the PVA solution is used as a dispersant or emulsifier to disperse the raw materials into the continuous phase to prevent the microsphere droplets from sticking and merging; at the same time, the microsphere droplets are evenly distributed by uniform dripping and stirring.

[0047] In some embodiments, when the colostrum is ultrasonically emulsified in step S3, the ultrasonic power is 15 to 30 W. The present invention controls the ultrasonic power when preparing the emulsion, so that fentanyl citrate @PLGA-PEG has excellent sustained release effect, rapid release effect and high drug loading, thereby increasing the sustained release time of fentanyl citrate and achieving rapid release of fentanyl citrate.

[0048] In step S4, after the ultrasonic emulsification in step S3 is completed, the emulsion is immediately dripped into pure water at a uniform speed, and volatilized for a certain period of time, such as 4 hours, under magnetic stirring, so that the organic solvent in the emulsion is volatilized, and the spherical material is precipitated in the water, and finally solidified by filtering and drying to form fentanyl citrate@PLGA-PEG microspheres, such as Figure 1 The speed of magnetic stirring can be 300-500 rpm, for example, 300 rpm, 400 rpm, 500 rpm, to ensure uniform dispersion of the drug in the emulsion during the volatilization process, avoid too slow stirring speed, resulting in uneven distribution of the particle size of the obtained nano-microspheres, and avoid too fast stirring, resulting in splashing of the solution.

[0049] Specifically, vacuum freeze drying can be used to dry the drug to avoid drug ineffectiveness.

[0050] In some embodiments, by simultaneously controlling the above parameters, i.e., the mass percentage of PEG in PLGA-PEG, the mass percentage of fentanyl citrate and PLGA-PEG, the volume ratio of 1% PVA solution to the mixture of dichloromethane and acetone, and the ultrasonic power when preparing the double emulsion, the drug loading, encapsulation efficiency, sustained release time and rapid drug release percentage of fentanyl citrate @PLGA-PEG can be improved.

[0051] In some embodiments, when the preparation process of fentanyl citrate @PLGA-PEG meets the following conditions, that is, the mass percentage of PEG in PLGA-PEG is 33%, the volume ratio of 1% PVA solution to the mixed solution of dichloromethane and acetone in step S1 is 1:7, the dosage percentage of fentanyl citrate and PLGA-PEG is 25%, and the ultrasonic power is 20W when the colostrum is ultrasonically emulsified in step S3. The drug loading, encapsulation rate, sustained release time and rapid drug release percentage of the fentanyl citrate @PLGA-PEG finally obtained can achieve good effects at the same time, and its sustained release time exceeds 5 days, the drug release rate in the first 15 minutes of in vitro release is not less than 5.36%, the drug loading rate is not less than 10.68%, and the encapsulation rate is not less than 99%.

[0052] In addition, an embodiment of the present invention further provides a fentanyl citrate@PLGA-PEG nanoparticle, which can be obtained by the preparation method of the above embodiment, and includes fentanyl citrate and a PLGA-PEG nanocarrier carrying fentanyl citrate.

[0053] The present invention adopts PLGA-PEG nanocarrier as a drug carrier to carry fentanyl citrate. Compared with the existing fentanyl citrate preparation for oral use, the present invention can achieve sustained release of fentanyl citrate, so that it can meet the needs of disaster relief analgesics under insufficient medical evacuation conditions.

[0054] In some embodiments, after testing, the sustained release time of fentanyl citrate @PLGA-PEG exceeds 5 days, the drug release rate in the first 15 minutes of in vitro release is not less than 5.36%, the drug loading rate is not less than 10.68%, and the encapsulation rate is not less than 99%, indicating that PLGA-PEG nanocarriers are suitable for loading fentanyl citrate.

[0055] The following specific examples further illustrate the fentanyl citrate@PLGA-PEG nanoparticles and the preparation method thereof.

[0056] Example 1

[0057] The specific preparation process of fentanyl citrate@PLGA-PEG is as follows.

[0058] Step S1: preparing fentanyl citrate solution and PLGA-PEG solution.

[0059] Preparation of 5% PVA solution: Weigh 2 g of PVA and place it in a beaker, add 40 mL of pure water, seal it with tin foil and rubber band, place it in a 100°C water bath and heat it, swell it for 2 hours, and then obtain a 5% PVA solution.

[0060] The 10 mL 5% PVA that has been swollen was placed in three centrifuge tubes, and a magnetic bar was added. The tubes were placed on a magnetic stirrer at a rotation speed of 600 rpm for later use.

[0061] Take 50mL of pure water in a beaker and set aside.

[0062] Preparation of PLGA-PEG solution: Weigh 100 mg of PLGA-33% PEG for use. Prepare 15 mL of a mixture of dichloromethane and acetone. 二氯甲烷 :V 丙酮 =3:2, add V to PLGA-33% PEG 二氯甲烷 :V 丙酮 =3:2 mixed solution 3.5mL, and dissolve it.

[0063] Preparation of fentanyl citrate solution: weigh 25 mg of fentanyl citrate and add 0.35 mL of 1% PVA solution (obtained by diluting the 5% PVA solution prepared above).

[0064] Step S2: preparing colostrum.

[0065] The fentanyl citrate solution was added dropwise to the PLGA-33% PEG solution, and the obtained O / W solution was placed in a cell disruptor and ultrasonicated for 5 times at an ultrasonic power of 20 W to obtain colostrum.

[0066] Step S3: preparing a double emulsion.

[0067] The colostrum was uniformly added dropwise to the 5% PVA solution on the magnetic stirrer. After mixing evenly, the centrifuge tube and the magnet were removed. The colostrum was ultrasonicated for 2 minutes using a cell crusher with an ultrasonic power of 30W.

[0068] Place the magnet in a beaker containing 50 mL of pure water, place the beaker in a magnetic stirrer and adjust the speed to 400 rpm. Immediately after the ultrasound is finished, drop the emulsion into the beaker containing 50 mL of pure water at a uniform speed, and after the emulsion is added, volatilize it at 400 rpm on a magnetic stirrer for 4 hours.

[0069] Step S4: Collecting microspheres.

[0070] After 4 hours of volatilization, remove the beaker from the magnetic stirrer, filter each sample into a centrifuge tube using a 0.45 μm filter, centrifuge for 1 hour at a speed of 18000G, and save the supernatant for later use. Add 10 mL of pure water to the centrifuge tube to dissolve the precipitate with the help of an ultrasonic cleaner, and centrifuge again for 1 hour (speed 18000G), and save the supernatant for later use.

[0071] The precipitate in the centrifuge tube was quickly frozen with liquid nitrogen, and immediately placed in a vacuum freeze dryer for drying (temperature of -51°C, pressure of 0.035 mbar), and taken out after 24 hours to obtain fentanyl citrate@PLGA-PEG. Figure 1 and stored in a -20°C refrigerator until use.

[0072] Embodiments 2 to 9

[0073] The preparation processes of Examples 2 to 9 are basically similar to the preparation process of Example 1, except for the following four parameters: A, dosage of fentanyl citrate (mg); B, mass percentage of PEG in PLGA-PEG (X%); C, ultrasonic power when forming emulsion (W); D, volume ratio of internal aqueous phase and oil phase (mL:mL).

[0074] The settings of the parameters in Examples 1 to 9 are shown in Table 1. Examples 2 to 9 were used to prepare fentanyl citrate@PLGA-PEG microspheres according to the setting parameters shown in Table 1, respectively.

[0075] Table 1 Settings of parameters in Examples 1 to 9

[0076]

[0077] The performance of the fentanyl citrate@PLGA-PEG microspheres in Examples 1 to 9 was tested:

[0078] (1) Detection of drug loading rate

[0079] The drug loading rate of the samples was detected by high performance liquid chromatography, and the calculation formula of the drug loading rate is as follows.

[0080]

[0081] The drug loading rate test results of Examples 1 to 9 are shown in Table 3.

[0082] (2) Detection of particle size

[0083] The precipitate obtained by centrifugation before drying in step S4 of each embodiment was taken as a sample, which was respectively recorded as PLGA E1 to 9. 2 mL of pure water was added to the sample to fully dissolve it to obtain a sample solution; the sample solution was filtered using a 0.45 μm filter, and dropped into a cuvette, and the cuvette to be tested was placed in a particle size analyzer to measure the particle size.

[0084] The particle size test results of Examples 1 to 9 are shown in Tables 3 and Figure 2 As shown in Table 3, the particle size of the samples is between 144 and 231 nm; and the particle size distribution of the samples of each embodiment is relatively concentrated, and the particle size is relatively uniform. In addition, the dispersion index PDI of the samples of each embodiment is also tested, as shown in Table 3, which also shows that the particle size distribution of the samples of each embodiment is relatively uniform.

[0085] (3) Detection of encapsulation efficiency

[0086] Take 10 mg of fentanyl citrate @PLGA-PEG microspheres from each example and place them in a centrifuge tube, add 10 mL of pure water to dissolve them. It was observed that the freeze-dried materials, especially the samples of Examples 3, 6, and 9, were in the form of flakes and insoluble in water. Use a 0.45 μm filter to filter the sample aqueous solution, take 9 mL of each sample and place it in a retention tube, and centrifuge for 15 minutes (3000 rpm). Take the lower supernatant of the lower layer of the retention tube for later use.

[0087] Preparation V 甲醇 :V 水 =7:3 mixed solution, weigh 5 mg of fentanyl citrate, add 5 mL of the above prepared mixed solution to fentanyl citrate to obtain 1 mg / mL fentanyl citrate standard solution. Dilute the 1 mg / mL fentanyl citrate standard solution to concentrations of 100 μg / mL, 80 μg / mL, 60 μg / mL, 40 μg / mL, and 20 μg / mL for use. Use 500 mg of anhydrous sodium sulfate, 1000 mg of ammonium acetate and 500 mL of water to prepare 500 mL of buffered saline solution, use a pH meter to detect its pH value and adjust it to 6.3±0.1 with glacial acetic acid for use.

[0088] Turn on the high performance liquid chromatograph, and place the mobile phase in the order of A methanol, B acetonitrile, C pure water, and D buffered saline solution. After the bubbles are discharged, rinse the chromatographic column with methanol and water in a proportional gradient, and continue to rinse the chromatographic column with a ratio of methanol: acetonitrile: buffered saline = 1:4:5. After the column pressure is stable, start injection. Transfer the fentanyl citrate standard solution and the lower supernatant in the interception tube to the injection bottle respectively, and inject each injection bottle 3 times to obtain the peak area. Use the standard solution concentration as the X-axis and the peak area percentage as the Y-axis to draw a calibration curve, such as Figure 3 As shown. According to this method, the mass of fentanyl citrate in the supernatant under the interception tube of each embodiment is measured and the encapsulation rate is calculated. The encapsulation rate calculation formula is:

[0089]

[0090] Since the release peak areas of the samples in each example were all less than the detection limit of the standard curve, the calculated drug concentrations were all negative values, as shown in Table 2. Therefore, the encapsulation rates of the samples in each example determined by this method were all 100%.

[0091] Table 2 Concentration of fentanyl citrate in the supernatant of Examples 1 to 9

[0092]

[0093] (4) Testing the rapid release performance of drugs

[0094] Take the fentanyl citrate @ PLGA-PEG microspheres of Examples 1 to 9, respectively record them as PLGA E1 to 9, add artificial gastric juice to dilute them into a solution of 12 mg / mL, and after fully dissolving, use centrifuge tubes to divide them into (E1-E9) × 8 centrifuge tubes, and each tube has 1 mL of solution for standby use. Take the time of the subpackaging as time 0, set the centrifugation time points of 15min, 30min, 1h, 2h, 4h, 6h, 8h, 10h, and 12h for a total of 8 time points. Take out 1 sample solution (E1-E9) at T0, centrifuge for 15min (14000rpm), put it in a -20°C refrigerator for standby use, and the other 7 samples are placed in a 37°C constant temperature environment, centrifuged in sequence according to the set time points, and all supernatants are stored in a -20°C refrigerator after centrifugation. The peak area in the supernatant is obtained by high performance liquid chromatography, and the drug release percentage at each time point is calculated by the difference method.

[0095] The calculated percentage of fentanyl citrate released at each time point is as follows: Figure 4 As shown. Figure 4 It can be seen that the fentanyl citrate in each embodiment can be released quickly, wherein the release rate in the first 15 minutes can reach more than 3.6%, and most of them can reach more than 5%.

[0096] (5) Detection of drug sustained-release performance

[0097] Take the weighed fentanyl citrate@PLGA-PEG microspheres in Examples 1 to 9, dilute them with physiological saline to a 6 mg / mL solution according to the mass, and use centrifuge tubes to divide the samples into (E1-E9)×6 centrifuge tubes, with 1 mL of solution in each tube for later use.

[0098] The time of packaging was set as time 0, and the centrifugation time points were set to 1d, 2d, 3d, 4d, 5d, and 6h, a total of 6 time points. At T0, one sample solution (E1-E9) was taken out, centrifuged for 15min (14000rpm), and placed in a -20℃ refrigerator for standby use. The other 5 samples were placed in a constant temperature environment of 37℃ and centrifuged at the same time point every day. After centrifugation, all supernatants were stored in a -20℃ refrigerator for testing.

[0099] Weigh 5 mg of fentanyl citrate and add 5 mL of normal saline to fentanyl citrate to obtain a 1 mg / mL fentanyl citrate standard solution. Dilute the 1 mg / mL fentanyl citrate standard solution to concentrations of 100 μg / mL, 80 μg / mL, 60 μg / mL, 40 μg / mL, and 20 μg / mL for later use. Use 500 mg of anhydrous sodium sulfate, 1000 mg of ammonium acetate, and 500 mL of water to prepare 500 mL of buffered saline solution, use a pH meter to detect its pH value, and adjust it to 6.3 ± 0.1 with glacial acetic acid for later use.

[0100] Turn on the high performance liquid chromatograph, and place the mobile phase in the order of A methanol, B acetonitrile, C pure water, and D buffered saline solution. After the bubbles are discharged, rinse the chromatographic column with methanol and water in a proportional gradient. Continue to rinse the chromatographic column with a ratio of methanol: acetonitrile: buffered saline = 1:4:5. Start injection after the column pressure stabilizes. Transfer the fentanyl citrate standard solution and the supernatant obtained from the sustained release experiment to the injection bottle. Inject each injection bottle 3 times to obtain the peak area. Use the standard solution concentration as the X-axis and the peak area percentage as the Y-axis to draw a calibration curve, such as Figure 5 shown.

[0101] The drug concentration in the supernatant of the sustained-release solution was calculated according to the standard curve, and the percentage of drug release at the same time point every day during the experiment was calculated using the differential method. The calculation results are shown in Table 3 and Figure 6 As shown. Figure 6 It can be seen that the sustained release effect of fentanyl citrate in each example is excellent, and the drug release rate has not reached 100% on the 6th day, indicating that the sustained release time of the drug is not less than 6 days (the release time has exceeded 5 days).

[0102] Table 3 Performance comparison of fentanyl citrate nanoparticles in various embodiments

[0103]

[0104] According to the above test results, the influence of different parameters on each performance is analyzed. Figure 7 The graph shows the influence of different parameters on drug particle size, dispersibility index, drug loading rate, 15-min release rate, and 5-day release rate.

[0105] like Figure 7 As shown in a, the concentration of fentanyl citrate and the volume ratio of the internal water phase to the oil phase have little effect on the particle size. In comparison, when 0% PEG and 30W power are used for preparation, the particle size becomes larger.

[0106] like Figure 7 As shown in b, when 0% PEG and 30W power are used for preparation, the particle size distribution becomes smaller, but when 0% PEG is used, the drug solubility is too low and is not used.

[0107] like Figure 7 As shown in c, the nanoparticles formed under the conditions of 25 mg fentanyl citrate dosage, 25% PEG in PLGA-PEG, 20 W ultrasonic power, and a ratio of 0.5 / 3.5 mL of inner water phase to outer oil phase have a larger drug loading capacity and are more suitable for drug loading.

[0108] like Figure 7 As shown in (d), the PEG ratio in PLGA-PEG and the ultrasonic power have the greatest influence on the drug release rate. Among them, under 25% PEG and 20W ultrasonic power, the release rate of fentanyl citrate is the highest in the first 15 minutes, indicating that this condition is more suitable for rapid release.

[0109] like Figure 7 As shown in Figure e, the PEG ratio in PLGA-PEG and the ultrasonic power have the greatest influence on the drug release rate. Among them, under 33% PEG and 20W ultrasonic power, the release rate of fentanyl citrate is the highest on the 5th day, indicating that this condition is more suitable for sustained release.

[0110] In order to make the drug of the embodiment of the present invention meet the performance indicators of drug loading rate, encapsulation rate, rapid release and sustained release at the same time, a dosage of 25 mg of fentanyl citrate, PLGA-20% PEG, an ultrasonic power of 20 W and a volume ratio of the inner water and outer oil phase of 0.5 / 3.5 mL can be used, so that the sustained release time of fentanyl citrate @ PLGA-PEG is not less than 6 days (the release time has exceeded 5 days), the in vitro release is not less than 5.36% in the first 15 minutes, the drug loading rate is not less than 10.68%, and the encapsulation rate is not less than 99%.

[0111] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions that can be easily conceived by any technician familiar with the technical field within the technical scope disclosed by the present invention should be covered within the protection scope of the present invention.

Claims

1. A method for preparing fentanyl citrate@PLGA-PEG nanoparticles, It is characterized in that include: Step S1, preparing fentanyl citrate solution and PLGA-PEG solution; Step S2, adding the fentanyl citrate solution dropwise to the PLGA-PEG solution, mixing and then performing ultrasonic emulsification to obtain colostrum; Step S3, adding the colostrum dropwise to the PVA solution, mixing and then performing ultrasonic emulsification to obtain a double emulsion; Step S4, adding the double emulsion dropwise into water, stirring to volatilize the organic solvent, and then centrifuging and filtering the water to obtain the fentanyl citrate@PLGA-PEG nanoparticles.

2. The preparation method according to claim 1, It is characterized in that In step S1, preparing the PLGA-PEG solution comprises: A mixed solution of dichloromethane and acetone is prepared, and PLGA-PEG is dissolved in the mixed solution of dichloromethane and acetone to obtain the PLGA-PEG solution.

3. The preparation method according to claim 2, It is characterized in that The mass percentage of PEG in the PLGA-PEG used is less than or equal to 33%.

4. The preparation method according to claim 1, It is characterized in that In step S3, when the colostrum is ultrasonically emulsified, the ultrasonic power is 15-30W.

5. The preparation method according to claim 2, It is characterized in that In step S1, preparing a fentanyl citrate solution comprises: Fentanyl citrate is dissolved in 1% PVA solution to obtain the fentanyl citrate solution.

6. The preparation method according to claim 5, It is characterized in that The volume ratio of the 1% PVA solution to the mixed solution of dichloromethane and acetone is 1:10 to 1:

5.

7. The preparation method according to claim 5, It is characterized in that In step S1, the mass percentage of fentanyl citrate and PLGA-PEG is 5% to 25%.

8. The preparation method according to any one of claims 1 to 7, It is characterized in that In step S1, the mass percentage of PEG in the PLGA-PEG used is 33%; In step S3, the ultrasonic power is 20W when the colostrum is ultrasonically emulsified; In step S1, the volume ratio of 1% PVA solution to the mixed solution of dichloromethane and acetone is 1:7; In step S1, the mass percentage of fentanyl citrate and PLGA-PEG is 25%.

9. A fentanyl citrate@PLGA-PEG nanoparticle, It is characterized in that The preparation method according to any one of claims 1 to 8 is used to obtain the fentanyl citrate and the PLGA-PEG nanocarrier carrying the fentanyl citrate.

10. The fentanyl citrate@PLGA-PEG nanoparticles according to claim 9, It is characterized in that The sustained release time of the fentanyl citrate@PLGA-PEG nano-microspheres exceeds 5 days, the drug release rate in the first 15 minutes of in vitro release is not less than 5.36%, the drug loading rate is not less than 10.68%, and the encapsulation rate is not less than 99%.

Images