Teicoplanin sustained-release preparation for injection and preparation method of teicoplanin sustained-release preparation

Abstract

The invention belongs to the field of biological medicine, and discloses a teicoplanin sustained-release injection as well as a preparation method and application thereof. The injection is composed of a teicoplanin active component and a polylactic acid-glycolic acid copolymer (PLGA) carrier (the molar ratio of lactic acid to glycolic acid is 75: 25-85: 15, and the molecular weight is 10,000-50,000 Da) in a specific proportion, the weight ratio of a medicine to the carrier is 1: 3-1: 5, and sustained release microspheres with the particle size of 10-50 microns are formed. The surface of the microsphere is coated with a composite stable layer containing polyvinyl alcohol (PVA, 0.5-2% w / w) and mannitol (1-5% w / w), a gradient heating curing technology and a freeze-drying protection technology are combined, the burst release effect (the release amount in 24 h is smaller than or equal to 50%) is remarkably reduced, the accumulated release amount in 7 days is larger than or equal to 80%, the viscosity of a dispersion medium after redissolution is controlled to be 10-50 mPa.s (25 DEG C), and the injection needle passing performance is ensured. According to the preparation method, through optimization of a W / O / W emulsification process, low-temperature solvent volatilization and surface coating steps, the drug encapsulation efficiency (greater than 90%) and stability are improved, and the problems that traditional PLGA microspheres are high in burst release, low in encapsulation efficiency and easy to aggregate during storage are solved.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine and provides a teicoplanin sustained-release injection and a preparation method thereof. Background Art

[0002] Teicoplanin is a glycopeptide antibiotic with potent antibacterial activity against Gram-positive bacteria (including methicillin-resistant Staphylococcus aureus MRSA), and is widely used clinically to treat deep tissue infections such as bone and joint infections and infective endocarditis. However, existing teicoplanin preparations are mainly conventional injections, which require multiple doses per day (usually once every 12-24 hours), resulting in poor patient compliance, large fluctuations in blood drug concentrations, and frequent injections may cause local tissue irritation reactions. For chronic infections that require long-term treatment (such as bone and joint infections), existing preparations are difficult to meet the needs of sustained and effective antibacterial effects.

[0003] In recent years, sustained-release microspheres using poly(lactic-co-glycolic acid) (PLGA) as a carrier have been used in attempts to deliver antibiotics in a sustained-release manner. However, existing PLGA-based sustained-release preparations still have significant drawbacks:

[0004] 1. Inadequate control of burst release: Most PLGA microspheres release more than 50% of their drug within 24 hours, failing to meet the requirements of "rapidly reaching effective concentrations and sustained release" in the treatment of deep infections. High burst release may lead to the risk of early toxicity.

[0005] 2. Poor drug encapsulation efficiency and stability: Teicoplanin is a hydrophilic drug. Microspheres prepared by traditional emulsification methods are prone to low encapsulation efficiency (usually <70%) due to drug diffusion, and microsphere aggregation or drug degradation are prone to occur during storage;

[0006] 3. Formulation process defects: Existing microsphere curing processes (such as constant temperature volatilization) easily lead to uneven porosity within the microspheres, affecting the reproducibility of the release curve. The microsphere structure is prone to collapse during the freeze-drying process, resulting in poor dispersibility after reconstitution and high viscosity (>50mPa·s), making it difficult to meet the requirements for injectable needle permeability.

[0007] 4. Limitations of excipient compatibility: Commonly used surfactants (such as single PVA) are difficult to take into account both microsphere stability and burst release control, and lack protection designs for the chemical stability of teicoplanin (such as oxidation or metal ion catalyzed degradation).

[0008] To address the above problems, there is an urgent need to develop a sustained-release injection of teicoplanin with precise release kinetics, high encapsulation rate and good stability. Summary of the Invention

[0009] To address the above issues, we have disclosed a sustained-release injection of teicoplanin and its preparation method. By optimizing the PLGA carrier composition, microsphere structure design, and preparation process, combined with composite stabilizing layer coating technology, the present invention successfully overcomes the bottlenecks of the existing technology and provides a safer and longer-lasting treatment for deep infections caused by MRSA.

[0010] The present invention includes the following specific technical solutions:

[0011] A sustained-release injection of teicoplanin, comprising: 1) and 2) components, wherein

[0012] 1) Teicoplanin active ingredient;

[0013] 2) a biodegradable polymer carrier, wherein the carrier is poly(lactic acid-co-glycolic acid) (PLGA) having a molar ratio of lactic acid to glycolic acid of 75:25 to 85:15;

[0014] The weight ratio of 1) to 2) is 1:3 to 1:5;

[0015] The combination of 1) and 2) forms sustained-release microspheres with a particle size of 10-50 μm;

[0016] The surface of the microspheres is coated with a stabilizing layer containing polyvinyl alcohol (PVA) and mannitol, wherein the PVA accounts for 0.5-2% w / w of the total weight of the microspheres, and the mannitol accounts for 1-5% w / w of the total weight of the microspheres;

[0017] The injection has a sustained-release property of releasing ≤50% of the drug within 24 hours and ≥80% of the drug in cumulative amount within 7 days after subcutaneous or intramuscular injection;

[0018] The injection is a freeze-dried powder injection, and the viscosity of the dispersion medium after reconstitution is 10-50 mPa·s, measured at 25°C.

[0019] Furthermore, the molecular weight of the PLGA in the above-mentioned teicoplanin sustained-release injection is 10,000-50,000 Da, and the intrinsic viscosity is 0.2-0.6 dL / g, as measured in chloroform at 25°C.

[0020] The present invention also discloses a method for preparing the sustained-release injection, comprising the following steps:

[0021] a) dissolving PLGA in dichloromethane to form a 5-15% w / v organic phase;

[0022] b) dispersing teicoplanin in an aqueous phase containing 1-3% w / v PVA and sonicating until the particle size is ≤5 μm;

[0023] c) injecting the organic phase into the aqueous phase at a stirring speed of 800-1200 rpm to form a W / O / W emulsion;

[0024] d) curing the microspheres using a gradient temperature ramp process: initially maintaining the temperature at 10-15°C for 2 hours, then increasing the temperature to 30-35°C at a rate of 0.5°C / min and maintaining the temperature for 2-4 hours;

[0025] e) evaporating the solvent at 2-8° C. for 12-24 hours, collecting the microspheres by filtration through a 5-20 μm pore size mesh, washing them three times with deionized water and three times with a 0.1% w / v mannitol aqueous solution, and drying them under vacuum at 25-30° C. until the water content is ≤3%;

[0026] f) immersing the microspheres in an aqueous solution containing 1-5% w / v mannitol and stirring for 30-60 minutes to form a surface coating layer;

[0027] g) adding 2-5% w / w trehalose as a lyoprotectant and then freeze-drying.

[0028] Furthermore, in the method for preparing the above-mentioned teicoplanin sustained-release injection, optionally, in the step b), a metal ion chelating agent is further added, and the metal ion chelating agent accounts for 0.01-0.1% w / w of the weight of the final injection.

[0029] Furthermore, in the preparation method of the above-mentioned teicoplanin sustained-release injection, the metal ion chelating agent is selected from calcium sodium EDTA or sodium citrate.

[0030] Furthermore, in the method for preparing the above-mentioned teicoplanin sustained-release injection, in the step c), the formation of the W / O / W emulsion is carried out under nitrogen protection.

[0031] Furthermore, in the preparation method of the above-mentioned teicoplanin sustained-release injection, the freeze-drying process in step g) is carried out at a temperature of -40°C to -60°C, and the freeze-drying time is not less than 24 hours.

[0032] The present invention also discloses the use of the sustained-release injection in preparing medicine for treating bone and joint infection or infective endocarditis caused by methicillin-resistant Staphylococcus aureus (MRSA).

[0033] Compared with the prior art, the present invention has the following outstanding beneficial effects:

[0034] The present invention significantly improves the performance of teicoplanin sustained-release injection through innovative formulation design and process optimization. The specific effects are as follows:

[0035] 1. Precise control of drug release kinetics

[0036] Low burst release effect: By optimizing the hydrophobicity and microsphere structure of the PLGA carrier (lactic acid: glycolic acid = 75:25–85:15), combined with a gradient temperature curing process (initial low-temperature curing at 10–15°C followed by slow temperature increase), the burst release effect is significantly reduced, achieving a release amount of ≤50% within 24 hours, thus avoiding the risk of toxicity caused by excessively high blood drug concentrations in the early stages.

[0037] Long-acting sustained release: The cumulative release amount in 7 days is ≥80%, which meets the demand for sustained and effective antibacterial concentration in the treatment of deep infections. The dosing frequency can be extended to once a week, greatly improving patient compliance.

[0038] 2. High encapsulation efficiency and stability

[0039] Efficient drug loading: After ultrasonically dispersing teicoplanin to a particle size of ≤5μm, the drug encapsulation efficiency is increased to >90% through a W / O / W emulsification process combined with a low-temperature solvent volatilization method, reducing drug waste.

[0040] Enhanced physicochemical stability: The surface of the microspheres is coated with a PVA-mannitol composite layer (PVA 0.5–2% w / w, mannitol 1–5% w / w) to inhibit microsphere aggregation and drug degradation; supplemented with trehalose freeze-drying protectant, the microspheres have an intact structure after freeze-drying, excellent re-dissolution and dispersibility, and extended storage life.

[0041] 3. Advantages of formulation process optimization

[0042] Highly reproducible release curves: The gradient temperature ramp process avoids uneven porosity within the microspheres, ensuring consistent release behavior between batches.

[0043] Excellent injectability: By controlling the microsphere particle size (10–50 μm) and the viscosity after reconstitution (10–50 mPa·s, 25°C), the injection is ensured to pass smoothly through commonly used clinical needles (such as 21–23G), reducing the risk of local irritation.

[0044] 4. Clinical treatment advantages

[0045] Targeting deep infections: After subcutaneous or intramuscular injection, the microspheres form a local drug reservoir and slowly release teicoplanin to the lesions (such as bones, joints or endocardium), thereby increasing the drug concentration at the infection site and reducing systemic exposure side effects.

[0046] Indication expansion: It is particularly suitable for stubborn infections such as bone and joint infections and infective endocarditis caused by MRSA that require long-term treatment, filling the gap where existing preparations cannot achieve weekly administration.

[0047] In summary, the present invention has excellent sustained-release properties, stability and clinical applicability, and provides an efficient, safe and long-acting drug delivery solution for the treatment of deep bacterial infections. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 Particle size D50 (μm) comparison chart;

[0049] Figure 2 Comparison of encapsulation efficiency (%);

[0050] Figure 3 Comparison of 24h burst release (%). DETAILED DESCRIPTION

[0051] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0052] To ensure experimental repeatability and data reliability, the various test methods involved in the present invention are performed according to the following standardized procedures.

[0053] 1. Particle size distribution determination

[0054] Instrument: Malvern Mastersizer 3000 laser particle size analyzer

[0055] Sample processing:

[0056] 10 mg of freeze-dried microspheres were dispersed in 1% w / v Tween-80 aqueous solution (5 mL) and sonicated (40 kHz, 100 W) for 1 min to avoid air bubble interference.

[0057] Test conditions:

[0058] Refractive index of dispersion medium: 1.33 (water);

[0059] Particle refractive index: 1.50 (PLGA);

[0060] Shading rate range: 5-15%;

[0061] Number of measurements: 3 independent parallel experiments, and the average value of D50 (median particle size) was taken.

[0062] 2. Encapsulation Efficiency Determination

[0063] Instrument: Agilent 1260 High Performance Liquid Chromatograph (HPLC)

[0064] Chromatographic conditions:

[0065] Chromatographic column: C18 column (4.6×250 mm, 5 μm);

[0066] Mobile phase: acetonitrile-0.1% phosphoric acid aqueous solution (25:75, v / v);

[0067] Flow rate: 1.0 mL / min;

[0068] Detection wavelength: 280nm;

[0069] Column temperature: 30°C;

[0070] Injection volume: 20 μL.

[0071] Sample processing:

[0072] Determination of total drug dosage: accurately weigh 10 mg of microspheres, dissolve in 1 mL of dimethyl sulfoxide (DMSO), vortex until completely dissolved, filter through a 0.22 μm filter, and dilute to the linear range of HPLC detection (0.1-1.0 mg / mL).

[0073] Determination of free drug content: The washing liquid (deionized water and mannitol solution) from the microsphere preparation process was combined, concentrated, and then the amount of unencapsulated drug was detected by HPLC.

[0074] The encapsulation efficiency E was calculated by the following formula:

[0075]

[0076] 3. Determination of burst release and cumulative release

[0077] Release medium: pH 7.4 phosphate buffer (containing 0.1% w / v SDS, simulating physiological conditions)

[0078] Instrument: RCZ-8M drug dissolution apparatus (paddle method, 100 rpm)

[0079] Test steps:

[0080] 50 mg of microspheres were accurately weighed, placed in a dialysis bag (molecular weight cut-off 10 kDa), and immersed in 900 mL of release medium;

[0081] At a constant temperature of 37 ± 0.5 °C, 5 mL of samples were collected at 0.5, 1, 2, 4, 8, 12, 24 h, and daily thereafter (while adding an equal volume of fresh medium at the same temperature);

[0082] The samples were filtered through a 0.22 μm filter membrane and then the teicoplanin concentration was determined by HPLC, and the cumulative release rate was calculated.

[0083] 4. Determination of reconstitution viscosity

[0084] Instrument: Brookfield DV2T Rotational Viscometer (compatible with LV-4 rotor)

[0085] Sample processing:

[0086] Take 100 mg of freeze-dried microspheres, add 2 mL of water for injection, vortex for 1 min, and let it stand for 5 min to form a uniform suspension.

[0087] Test conditions:

[0088] Temperature: 25±0.1℃ (water bath temperature control);

[0089] Speed: 60rpm;

[0090] Measurement time: 3min, take the steady-state viscosity value;

[0091] The measurements were repeated three times and the average value was taken.

[0092] 5. Storage stability test

[0093] Accelerated test conditions:

[0094] Temperature: 40±2℃;

[0095] Humidity: 75 ± 5% RH;

[0096] Packaging: sealed with semi-permeable aluminum-plastic composite film;

[0097] Time: 0, 1, 2, 3 months.

[0098] Detection indicators:

[0099] Appearance: Visually inspect the microspheres for color changes, agglomeration, or aggregation;

[0100] Encapsulation efficiency: determined by the above HPLC method;

[0101] Redissolution performance: Evaluate dispersion uniformity according to the viscosity measurement method;

[0102] Microbial limits: Check according to Article 1105 of the General Rules of Part IV of the 2020 edition of the Chinese Pharmacopoeia.

[0103] Example 1

[0104] Preparation of Teicoplanin Sustained-Release Injection

[0105] Step a: 4.0 g of PLGA with a molecular weight of 30,000 Da and a lactic acid:glycolic acid molar ratio of 80:20 (Suzhou Beike Nanotechnology Co., Ltd.) was weighed and dissolved in 40 mL of dichloromethane to form a 10% w / v organic phase.

[0106] Step b: 1.0 g of teicoplanin API (Wuhan Dingxintong Pharmaceutical Co., Ltd.) was dispersed in 100 mL of aqueous phase containing 2% w / v PVA (pharmaceutical grade PVA88 CP2020, Xi'an Tianzheng Pharmaceutical Excipients Co., Ltd.) and sonicated for 30 min (power 200 W, frequency 40 kHz) to obtain a uniform suspension with a particle size of ≤3 μm.

[0107] Step c: Under nitrogen protection, the organic phase was slowly injected into the aqueous phase at a stirring speed of 1000 rpm to form a W / O / W emulsion.

[0108] Step d: Gradient temperature curing: maintain 12°C for the initial 2 hours, then increase the temperature to 32°C at a rate of 0.5°C / min and maintain for 3 hours.

[0109] Step e: evaporate the solvent at 4° C. for 18 h, filter through a 15 μm mesh, wash three times with deionized water and 0.1% mannitol aqueous solution, and vacuum dry (28° C., 24 h) to a water content of 2.5%.

[0110] Step f: Immerse the microspheres in a 3% w / v mannitol solution and stir for 45 minutes to form a surface coating layer.

[0111] Step g: add 3% w / w trehalose, freeze-dry at -50°C for 28 hours to obtain a lyophilized powder injection.

[0112] Key parameter tests:

[0113] Particle size distribution: D50 = 28 μm (measured by laser particle size analyzer);

[0114] Encapsulation efficiency: 93.2% (determined by HPLC);

[0115] Burst release: 42% of the 24h release, 85% of the 7-day cumulative release;

[0116] Reconstituted viscosity: 35 mPa·s (25°C, rotary viscometer);

[0117] Storage stability: 3 months at 40°C / 75% RH, encapsulation efficiency decreased by <5%, with no visible aggregation.

[0118] Example 2

[0119] High PLGA ratio formulation (drug: carrier = 1:5)

[0120] Step a: 5.0 g of PLGA (molecular weight 50,000 Da) with a ratio of lactic acid to glycolic acid of 85:15 was selected and dissolved in 50 mL of dichloromethane (10% w / v).

[0121] Step b: 1.0 g of teicoplanin was dispersed in a 1% w / v PVA aqueous phase, and the particle size was ≤4 μm after ultrasonication.

[0122] Step c: The other steps were the same as those in Example 1. The final microsphere particle size D50 was 45 μm, the encapsulation efficiency was 91.5%, the release amount in 24 hours was 38%, and the release amount in 7 days was 88%.

[0123] Example 3

[0124] Low PLGA ratio formulation (drug:carrier=1:3)

[0125] Step a: 3.0 g of PLGA (molecular weight 10,000 Da) with a ratio of lactic acid to glycolic acid of 75:25 was selected and dissolved in 20 mL of dichloromethane (15% w / v).

[0126] Step b: 1.0 g of teicoplanin was dispersed in a 3% w / v PVA aqueous phase, and 0.05% sodium calcium EDTA was added.

[0127] Step c: In step d, the temperature was raised to 35° C. and maintained for 4 h. The final microsphere particle size D50 was 12 μm, the encapsulation efficiency was 89.7%, the release amount in 24 h was 47%, and the release amount in 7 days was 82%.

[0128] Comparative Example 1

[0129] Comparative Example 1: PLGA molar ratio out of range (lactic acid: glycolic acid = 70:30)

[0130] Preparation method: except that the molar ratio of PLGA was adjusted to 70:30, other procedures were the same as in Example 1.

[0131] Results: The burst release in 24 hours was 65%, the cumulative release in 7 days was only 72%, and the particle size of the microspheres was uneven (D50=50μm, span>2).

[0132] Comparative Example 2

[0133] Drug-to-carrier ratio out of range (1:2)

[0134] Preparation method: 2.0 g of PLGA and 1.0 g of teicoplanin were mixed (1:2), and the rest was the same as in Example 1.

[0135] Results: The encapsulation efficiency was only 68%, the 24h release was 58%, the microspheres aggregated severely after freeze-drying, and the reconstituted viscosity was >80 mPa·s.

[0136] Comparative Example 3

[0137] No gradient heating process is used

[0138] Preparation method: Step d was changed to curing at a constant temperature of 30° C. for 6 h, and the rest was the same as in Example 1.

[0139] Results: The porosity of the microspheres varied greatly, and the 7-day release fluctuated between batches (75%-88%), with a 24-hour burst release of 52%.

[0140] Comparative Example 4

[0141] No trehalose freeze-drying protective agent added

[0142] Preparation method: Omit the trehalose in step g and directly freeze-dry. Other steps are the same as in Example 1.

[0143] Results: The collapse rate of the microspheres after freeze-drying was >30%, the re-dissolution and dispersibility were poor, and the encapsulation efficiency decreased by 12% after storage for one month.

[0144] Analysis example

[0145] The test results of the embodiments and comparative examples are summarized in Table 1 and Figure 1-3 shown.

[0146] Table 1 Test results

[0147]

[0148]

[0149] Note:

[0150] 1. Items marked with “*”: indicate key parameters that deviate from the technical solution of the present invention;

[0151] 2. Storage stability: Accelerated test conditions were 40°C / 75% RH. The examples were tested for 3 months, and the comparative examples were tested for 1 month. (Due to time constraints, the comparative examples were tested for only 1 month, but this was sufficient to observe performance degradation trends.)

[0152] 3. Reconstituted viscosity: The clinically acceptable range is 10-50 mPa·s (>50 mPa·s may cause injection blockage);

[0153] 4. 7-day release of Comparative Example 3: Due to defects in the curing process, there was fluctuation between batches (±8%), which did not meet the quality control standard (RSD < 5%).

[0154] As can be seen from the test results in Table 1 above, the present invention systematically solves the long-standing technical problems of PLGA-based teicoplanin sustained-release preparations (high burst release, low encapsulation efficiency, and poor stability) through a combination of key parameter optimization and process innovation.

[0155] 1. Synergistic Control of PLGA Composition and Drug Carrier Ratio (Example 1 vs. Comparative Examples 1-2)

[0156] Example 1: Using PLGA (molecular weight 30 kDa) with a lactic acid:glycolic acid ratio of 80:20 and a drug:carrier ratio of 1:4, combined with a gradient temperature increase process, a burst release of 42% in 24 hours and a release of 85% in 7 days were achieved, with an encapsulation efficiency of 93.2%.

[0157] Comparative Example 1 (PLGA molar ratio out of range): When the PLGA molar ratio was adjusted to 70:30 (the glycolic acid ratio was too high), the hydrophilicity of PLGA increased, resulting in a surge in burst release to 65% (24 hours), and insufficient cumulative release (72% over 7 days). This demonstrates that a lactic acid ratio of ≥75% is required to maintain sufficient hydrophobicity and inhibit early drug diffusion.

[0158] Comparative Example 2 (drug-carrier ratio 1:2): Reducing the PLGA ratio resulted in excessive drug loading, with an encapsulation efficiency of only 68%, a burst release of 58%, and excessive viscosity after reconstitution. This validated a drug:carrier ratio of 1:3–1:5 as the critical window for balancing drug loading and sustained release.

[0159] Summarize:

[0160] In the prior art, the composition and drug ratio of PLGA are typically set empirically (e.g., a conventional 50:50 or random ratio). The present invention experimentally discovered that the synergistic effect of a lactic acid ratio of 75–85% and a drug carrier ratio of 1:3–1:5 can precisely control the balance between drug diffusion resistance and PLGA degradation rate without sacrificing drug loading.

[0161] 2. The Core Role of the Gradient Temperature Curing Process (Example 1 vs. Comparative Example 3)

[0162] Example 1: Using an initial low-temperature curing at 12°C and a slow temperature increase to 32°C, the pores inside the microspheres are evenly distributed, and the release difference between batches is less than 5% (7-day release amount is 85±2%).

[0163] Comparative Example 3 (Constant Temperature Curing): Curing at a constant temperature of 30°C resulted in excessively rapid solvent evaporation. A hard shell on the microspheres hindered the escape of the internal solvent, forming irregular pores. This resulted in large batch-to-batch release fluctuations (75–88% over 7 days) and a burst release of 52%. This demonstrates that a gradual temperature increase is essential for controlling microsphere structural uniformity.

[0164] Summarize:

[0165] Traditional PLGA microsphere curing processes typically utilize a constant temperature process. However, this invention pioneers a gradient temperature strategy, slowly forming a dense shell at a low temperature before gradually increasing the temperature to promote gentle evaporation of the internal solvent, thus preventing pore collapse or cracking. This process requires a specific PLGA molecular weight (10k–50kDa) and solvent system (dichloromethane) to be effective.

[0166] 3. Synergistic Effect of Composite Stabilizing Layer and Freeze-Drying Protection (Example 1 vs. Comparative Example 4)

[0167] Example 1: The surface of the microspheres was coated with PVA (2%) + mannitol (3%) and protected by freeze-drying with trehalose. After 3 months of storage, the encapsulation efficiency of the microspheres decreased by less than 5%, and the re-dissolution and dispersibility were excellent (viscosity 35 mPa·s).

[0168] Comparative Example 4 (no trehalose): The microsphere collapse rate after freeze-drying was >30%, the encapsulation efficiency decreased by 12% after one month of storage, and the re-dispersion and dispersibility were poor, indicating that trehalose inhibited ice crystal damage through glass transition and maintained the integrity of the microspheres together with the PVA-mannitol layer.

[0169] Summarize:

[0170] In the prior art, PVA is often used alone as an emulsifier, but the present invention innovatively introduces mannitol as a stabilizer. Its hydroxyl group forms a hydrogen bond network with PVA, enhancing the mechanical strength of the microsphere surface. At the same time, the low-temperature protection effect of trehalose and the surface coating form a double barrier, solving the industry problems of microsphere structure collapse and poor storage stability during the freeze-drying process.

[0171] 4. Added Value of Metal Ion Chelating Agents (Example 3)

[0172] Example 3: In step b, 0.05% EDTA calcium sodium was added, and the encapsulation efficiency was increased to 89.7% (compared to <85% when no addition was made). 3+ 、Cu 2+ ), inhibiting the oxidative degradation of teicoplanin and prolonging the shelf life of the preparation.

[0173] Existing technology drawbacks: Traditional PLGA microspheres ignore the chemical stability of the drug, especially the sensitivity of teicoplanin to metal ions. The present invention uses a chelating agent + low-temperature process + inert gas protection (nitrogen environment in step c) to form a multi-stability guarantee mechanism.

[0174] Animal experiment test examples

[0175] Animal Models:

[0176] SD rats (200-250 g, purchased from Nanjing Zhishu Yantu Biotechnology Co., Ltd.) were selected to establish a MRSA bone and joint infection model (MRSA bacterial suspension was injected into the tibia, 48 hours after infection).

[0177] Grouping and dosing:

[0178] Experimental group: 10 mice, subcutaneously injected with the sustained-release microspheres prepared in Example 1 (teicoplanin dose 20 mg / kg);

[0179] Control group: 10 rats, intravenously injected with conventional teicoplanin injection (daily administration, total dose 20 mg / kg / day × 7 days, Zhejiang Hisun Pharmaceutical Co., Ltd.);

[0180] Blank group: 5 rats, injected with normal saline.

[0181] Detection indicators:

[0182] Blood concentration: Blood samples were collected at 0.5 h, 1 h, 6 h, 12 h, 24 h, and daily until the 7th day after administration, and teicoplanin concentration was determined by HPLC.

[0183] Tissue drug concentration: On the 7th day, animals were sacrificed and infected bones and joints, heart (target tissue for simulating endocarditis), and liver (representative of systemic exposure) were homogenized and tested for drug concentration.

[0184] Antibacterial effect: Take the tissue homogenate from the infected area, spread it on the MH agar plate, and calculate the colony count (CFU / g).

[0185] Summary of experimental results:

[0186] The experimental group had stable blood drug concentrations, with peak concentrations (Cmax) <10 μg / mL (control group Cmax >30 μg / mL), and systemic exposure was reduced;

[0187] The drug concentration in infected bone and joint tissues was maintained at >5 μg / g (MIC = 5 times of 1 μg / mL) until day 7;

[0188] The colony count decreased to <10 3 CFU / g (the control group fluctuated due to intermittent administration, and the number of colonies repeatedly increased to >10 5 CFU / g).

[0189] Conclusion: The sustained-release microspheres prepared in the present invention can form a local drug reservoir, maintain effective concentration in the infection site, and reduce systemic side effects.

[0190] The above are only a few preferred embodiments of the present invention, and their description is relatively specific and detailed, but it should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art may make various modifications and improvements without departing from the scope of the present invention, and such modifications and improvements are within the scope of protection of the present invention.

Claims

1. A teicoplanin sustained-release injection, characterized in that: Contains: 1) and 2) components, wherein 1) Teicoplanin active ingredient; 2) a biodegradable polymer carrier, wherein the carrier is poly(lactic acid-co-glycolic acid) (PLGA) having a molar ratio of lactic acid to glycolic acid of 75:25 to 85:15; The weight ratio of 1) to 2) is 1:3 to 1:5; The combination of 1) and 2) forms sustained-release microspheres with a particle size of 10-50 μm; The surface of the microspheres is coated with a stabilizing layer containing polyvinyl alcohol (PVA) and mannitol, wherein PVA accounts for 0.5-2% w / w of the total weight of the microspheres, and mannitol accounts for 1-5% w / w of the total weight of the microspheres; The injection has a sustained-release property of releasing ≤50% of the drug within 24 hours and ≥80% of the drug in cumulative amount within 7 days after subcutaneous or intramuscular injection; The injection is a freeze-dried powder injection, and the viscosity of the dispersion medium after reconstitution is 10-50 mPa•s, measured at 25°C.

2. The teicoplanin sustained-release injection according to claim 1, characterized in that: The PLGA has a molecular weight of 10,000-50,000 Da and an intrinsic viscosity of 0.2-0.6 dL / g, as measured in chloroform at 25°C.

3. A method for preparing the sustained-release injection according to any one of claims 1 to 2, characterized in that: The following steps are involved: a) dissolving PLGA in dichloromethane to form a 5-15% w / v organic phase; b) dispersing teicoplanin in an aqueous phase containing 1-3% w / v PVA and sonicating until the particle size is ≤5 μm; c) injecting the organic phase into the aqueous phase with stirring at 800-1200 rpm to form a W / O / W emulsion; d) Curing the microspheres using a gradient temperature ramp process: initially maintaining the temperature at 10-15°C for 2 hours, then increasing the temperature at 0.5°C / min to 30-35°C and maintaining the temperature for 2-4 hours; e) evaporating the solvent at 2-8°C for 12-24 hours, collecting the microspheres by filtration through a 5-20 μm pore size mesh, washing them three times with deionized water, three times with a 0.1% w / v mannitol aqueous solution, and drying them under vacuum at 25-30°C until the water content is ≤3%; f) immersing the microspheres in an aqueous solution containing 1-5% w / v mannitol and stirring for 30-60 minutes to form a surface coating; g) Add 2-5% w / w trehalose as a lyoprotectant and then freeze-dry.

4. The method for preparing the teicoplanin sustained-release injection according to claim 3, characterized in that: Optionally, in step b), a metal ion chelating agent is further added, and the metal ion chelating agent accounts for 0.01-0.1% w / w of the weight of the final injection.

5. The method for preparing the teicoplanin sustained-release injection according to claim 4, characterized in that: The metal ion chelating agent is selected from calcium sodium EDTA or sodium citrate.

6. The method for preparing the teicoplanin sustained-release injection according to claim 3, characterized in that: In the step c), the formation of the W / O / W emulsion is carried out under nitrogen protection.

7. The method for preparing the teicoplanin sustained-release injection according to claim 3, characterized in that: The freeze-drying process in step g) is carried out at a temperature of -40°C to -60°C, and the freeze-drying time is not less than 24 hours.

8. Use of the sustained-release injection according to any one of claims 1 to 2 in the preparation of a medicament for treating bone and joint infection or infective endocarditis caused by methicillin-resistant Staphylococcus aureus (MRSA).

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