A doxycycline for injection and a method for preparing the same

Abstract

The application discloses doxycycline for injection and a preparation method thereof, and belongs to the field of pharmaceutical preparations. The preparation contains doxycycline hydrochloride, polyethylene glycol-poly (caprolactone) and other raw materials, vitamin C phosphate is used to replace vitamin C, and an 'acid phosphatase-GSH' double-reducing agent hierarchical response nano-carrier is constructed. The preparation is prepared through an organic phase, an aqueous phase, homogenization, evaporation, centrifugation and other steps. The problems of poor stability of doxycycline and lack of specificity of release are solved, the storage period is extended to 36 months, the release at an infected site reaches 85% in 48 hours, the release at normal tissue is less than 20%, and precise targeted drug release is realized.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of pharmaceutical preparations, and particularly relates to a kind of doxycycline for injection and a preparation method thereof. BACKGROUND

[0002] Doxycycline, as a second-generation tetracycline broad-spectrum antibiotic, is widely used in the treatment of severe conditions such as respiratory tract infection, skin and soft tissue infection, and sepsis due to its broad antibacterial spectrum and strong tissue penetration. However, its injection preparation has two major defects: first, poor stability, the phenolic hydroxyl and enol groups in the molecule are easily oxidized and degraded by light, temperature and pH, and the potency loss can reach 15% after 30 days of storage at 25℃, and the degradation products can also cause allergies; second, lack of specificity in release, after traditional lyophilized powder injection administration, systemic diffusion occurs, normal tissues are exposed to high levels, which can easily cause gastrointestinal reactions, liver toxicity and bacterial flora imbalance, while the local drug concentration at the infection site is only 30%-50% of the blood drug concentration, which affects the therapeutic effect.

[0003] Although existing nanocarrier delivery technologies attempt to improve, there are obvious limitations. Single response mechanism (such as only pH or glutathione response) is prone to "off-target release", for example, pH-responsive carriers may release prematurely in normal weak acid environments such as tumors, and glutathione-responsive carriers may also leak prematurely in low-concentration glutathione environments in normal tissues such as liver and kidney, with a 72-hour non-specific release rate of up to 30%. At the same time, some polymer carriers (such as PLGA) have poor compatibility with doxycycline, with an encapsulation efficiency often below 60%, and are prone to "burst release" or "insufficient sustained release".

[0004] The unique microenvironment of bacterial infection sites provides a natural signal for precise delivery: first, high concentration of reducing substances, the concentration of glutathione produced by bacterial metabolism reaches 5-10 mmol / L, which is 5-100 times higher than that in normal tissues, and lipopolysaccharide can induce macrophages to secrete reductase to enhance the local reducing potential; second, specific enzymes, the concentration of acid phosphatase secreted by pathogenic bacteria (such as Staphylococcus aureus) in the infection site reaches 10-50 U / mL, which is significantly higher than that in normal tissues; third, weakly acidic environment, lactic acid and other substances produced by bacterial metabolism reduce the local pH to 5.5-6.5, forming a difference with normal tissues.

[0005] Chinese patent CN1868460A discloses a lyophilized preparation of doxycycline hydrochloride for injection and its preparation process, which is composed of active ingredient doxycycline hydrochloride and pharmaceutical excipients vitamin C and glycine, and the weight ratio of each component is doxycycline hydrochloride 50-250, vitamin C 200-1000, and glycine 200-1500. The large amount of antioxidant vitamin C in the patent increases the related substances during the preparation and storage process, and the patent does not involve specific treatment.

[0006] Chinese patent CN101555215A discloses a production process of doxycycline hydrochloride for injection, and the freeze-dried preparation thereof comprises doxycycline hydrochloride, vitamin C, mannitol and sodium pyrosulfite in a weight ratio of 115:380:300:2.9. The patent does not control the related substances of vitamin C, and secondly, the patent does not involve specific treatment.

[0007] Chinese patent CN107638395A discloses a freeze-dried powder of doxycycline hydrochloride for injection and a preparation method thereof. The freeze-dried powder comprises an active ingredient doxycycline hydrochloride, a stabilizer, an excipient and water for injection. The stabilizer is vitamin C and hydrochloric acid cysteine, and the excipient is lactose. The freeze-dried powder prepared by the technical scheme has good forming, full appearance, loose and porous, and good reconstitution, but this patent also does not involve specific treatment.

[0008] The prior art cannot simultaneously meet the three demands of "precise release, co-loading of antioxidant stabilizer, and adaptation to infection microenvironment signals", lacks the synergistic response design of the "high glutathione + acid phosphatase" double signals of the infection focus, and is difficult to realize the linkage of "efficient drug release at the infection site + activation of antioxidant", which becomes the research and development motivation of the present application. SUMMARY

[0009] The purpose of the present application is to provide a doxycycline for injection and a preparation method thereof to solve the problems in the background art.

[0010] The core improvement points of the present application are: first, vitamin C phosphate (VC-P) is used to replace vitamin C to protect the easily oxidized group by phosphate bond, the storage period is extended to 24 months, and the inactivation is avoided in advance; second, the "acid phosphatase-GSH" double reducing agent graded response is constructed, the release rate in normal tissues is less than 20%, the disulfide bond is broken due to high concentration of GSH in the infection site, and the release rate reaches 85% in 48 hours, realizing precise targeting.

[0011] The purpose of the present application can be realized by the following technical scheme:

[0012] A doxycycline for injection comprises the following mass parts of raw materials:

[0013] Doxycycline hydrochloride 25-30 parts;

[0014] Polyethylene glycol-poly (caprolactone) (PEG-PCL) 35-45 parts;

[0015] Polyethylene glycol disuccinimidyl succinate (SS-PEG-SS) 10-15 parts;

[0016] Vitamin C phosphate 4-6 parts;

[0017] Mannitol 10-15 parts;

[0018] Further, a preparation method of the injectable doxycycline includes the following steps:

[0019] S1, according to the mass fraction of the above-mentioned raw materials, dichloromethane, polyethylene glycol-polycaprolactone, polyethylene glycol disuccinimidyl butane, stirring at a speed of 300 rpm for 30 minutes, then adding doxycycline hydrochloride, vitamin C phosphate, and ultrasonic treatment at a power of 500 W for 5 minutes to form a uniform organic phase;

[0020] S2, another liquid preparation tank is taken, and injectable water and a polyvinyl alcohol solution with a mass concentration of 1% are added, and stirring is carried out at a speed of 500 rpm for 15 minutes to obtain an aqueous phase;

[0021] S3, the organic phase is dropped into the aqueous phase at a rate of 5 L / h, and stirring is carried out at a speed of 800 rpm during dropping, after dropping, stirring is continued for 10 minutes, then it is transferred to a high-pressure homogenizer, and homogenized at a pressure of 800 bar for 3 times, each cycle for 2 minutes, then it is transferred to a rotary evaporator, and evaporated at 30°C under reduced pressure for 4 hours to remove dichloromethane, and at the same time, polyethylene glycol disuccinimidyl butane and polyethylene glycol-polycaprolactone are crosslinked to obtain a suspension;

[0022] S4, the suspension is transferred to a centrifuge barrel, and centrifuged at a speed of 12000 rpm for 20 minutes at 4°C, the supernatant is discarded, the precipitate is resuspended with sterile PBS buffer, and the centrifugation is repeated for 3 times to obtain a nanoparticle precipitate, then the nanoparticle precipitate is resuspended with injectable water to obtain a nanoparticle suspension;

[0023] S5, mannitol is added to the nanoparticle suspension, and stirring is carried out at a speed of 500 rpm for 20 minutes to form a drug solution, then the drug solution is coarsely filtered through a 0.45 μm polypropylene filter membrane, and finally filtered through a 0.22 μm polyether sulfone sterile filter membrane, and collected in a sterile storage tank to obtain a pre-freeze-drying drug solution;

[0024] S6, the pre-freeze-drying drug solution is filled, semi-capped, pre-frozen, dried, vacuum pressed, and capped to obtain the injectable doxycycline.

[0025] Further, the use amount ratio of dichloromethane to polyethylene glycol-polycaprolactone in S1 is 20 L:5 kg.

[0026] Further, the use amount ratio of injectable water and a polyvinyl alcohol solution with a mass concentration of 1% in S2 to dichloromethane in S1 is 100 L:1 kg:20 L.

[0027] Further, the residual amount of dichloromethane in the suspension in S3 needs to be ≤600 ppm.

[0028] Further, the sterile PBS buffer and the water for injection of S4 and the dichloromethane of S1 are used in a ratio of 100L:50L:20L.

[0029] Advantages of the present application:

[0030] The vitamin C phosphate (VC-P) is used to replace vitamin C, the antioxidant group is protected by the phosphate bond, the storage period of the preparation is prolonged from 6 months of the prior art to 24 months (2-8℃), and the stability of the solution state after being compatible with the infusion is improved from 4 hours to 24 hours, and the core pain points that doxycycline is easily degraded by light and oxidation are solved.

[0031] In order to resist the active oxygen released by the host immune cells, the bacteria synthesize a large amount of glutathione (GSH), the high-concentration glutathione reacts with the disulfide bond (-S-S-) of the nanometer particle shell, the cross-linked structure is disintegrated, and the cumulative release rate of the loaded doxycycline reaches 85% in 48 hours. While the normal tissues are neutral in pH (without activation of acid phosphatase) and low in GSH concentration (<1mM), the carrier only releases <20% of the drug, realizing the “concentrated release at the infection site and low exposure in normal tissues”.

[0032] When the nanometer particle enters the infection site (such as abscess, pH 5.5-6.0), the acid phosphatase (optimum pH 5.0-6.0) secreted by the bacteria specifically hydrolyzes the phosphate bond of VC-P, and releases free vitamin C (VC) and phosphate ions. DETAILED DESCRIPTION

[0033] The technical solutions in the embodiments of the present application will be clearly and completely described below in combination with the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application. Meanwhile, the raw materials, reagents or devices used in the following embodiments are commercially available or can be obtained by known methods, unless otherwise specified.

[0034] Embodiment 1

[0035] Preparation of doxycycline for injection:

[0036] Firstly, the doxycycline for injection comprises the following raw materials in parts by mass:

[0037] Doxycycline hydrochloride 25 parts;

[0038] Polyethylene glycol-poly (caprolactone) (PEG-PCL) 35 parts;

[0039] Polyethylene glycol disuccinimidyl succinate (SS-PEG-SS) 10 parts;

[0040] Vitamin C phosphate 4 parts;

[0041] Mannitol 10 parts;

[0042] Then, the preparation method of the above-mentioned doxycycline for injection comprises the following steps:

[0043] S1, according to the mass parts of the above-mentioned raw materials, dichloromethane, polyethylene glycol-poly caprolactone, polyethylene glycol disuccinimidyl butane, the amount ratio of dichloromethane and polyethylene glycol-poly caprolactone is 20L:5kg, stirring at a speed of 300rpm for 25 minutes, then adding doxycycline hydrochloride and vitamin C phosphate, ultrasonic treatment at a power of 500W for 5 minutes to form a uniform organic phase;

[0044] S2, another liquid preparation tank is taken, and injection water and a polyvinyl alcohol solution with a mass concentration of 1% are added, the amount ratio of injection water and the polyvinyl alcohol solution with a mass concentration of 1% to the dichloromethane in S1 is 100L:1kg:20L, stirring at a speed of 500rpm for 10 minutes to obtain an aqueous phase;

[0045] S3, the organic phase is dropped into the aqueous phase at a rate of 4L / h, and stirring is carried out at a speed of 800rpm during dropping, after dropping, stirring is continued for 10 minutes, then it is transferred to a high-pressure homogenizer, and homogenized at a pressure of 800bar for 3 times, each cycle for 2 minutes, then it is transferred to a rotary evaporator, and evaporated at 30℃ under reduced pressure for 4 hours to remove dichloromethane, and at the same time, polyethylene glycol disuccinimidyl butane and polyethylene glycol-poly caprolactone are crosslinked to obtain a suspension, the residual amount of dichloromethane in the suspension needs to be ≤600ppm;

[0046] S4, the suspension is transferred to a centrifuge barrel, and centrifuged at a speed of 12000rpm for 20 minutes at 4℃, the supernatant is discarded, and the precipitate is resuspended in sterile PBS buffer, and the centrifugation is repeated for 3 times to remove free drugs and polyvinyl alcohol, and ensure that the residual amount of polyvinyl alcohol is ≤0.1%, then the nanoparticle precipitate is resuspended in injection water, and the amount ratio of sterile PBS buffer, injection water and dichloromethane in S1 is 100L:50L:20L to obtain a nanoparticle suspension;

[0047] S5, mannitol is added to the nanoparticle suspension, and stirred at a speed of 500rpm for 20 minutes until completely dissolved to form a drug solution, then the drug solution is coarsely filtered through a 0.45μm polypropylene filter membrane, and finally filtered through a 0.22μm polyether sulfone sterile filter membrane, and collected in a sterile storage tank to obtain a drug solution before freeze-drying;

[0048] S6, the freeze-dried prodrug liquid is filled into a brown Schlenk bottle, semi-tamped, and then the Schlenk bottle is placed into a freeze dryer, kept at -45℃ for 3 hours, sublimation dried, vacuum tamped, and capped to obtain the doxycycline for injection.

[0049] Example 2

[0050] Preparation of the doxycycline for injection:

[0051] First, the doxycycline for injection includes the following mass parts of raw materials:

[0052] Doxycycline hydrochloride 28 parts;

[0053] Polyethylene glycol-poly (ε-caprolactone) (PEG-PCL) 40 parts;

[0054] Polyethylene glycol disuccinimidyl succinate (SS-PEG-SS) 12 parts;

[0055] Vitamin C phosphate 5 parts;

[0056] Mannitol 12 parts;

[0057] Then, the preparation method of the above doxycycline for injection includes the following steps:

[0058] S1, the above raw materials are weighed according to the mass parts, dichloromethane, polyethylene glycol-poly (ε-caprolactone), and polyethylene glycol disuccinimidyl succinate are added to a liquid preparation tank, the amount ratio of dichloromethane to polyethylene glycol-poly (ε-caprolactone) is 20L:5kg, stirring at a speed of 300rpm for 30 minutes, then adding doxycycline hydrochloride and vitamin C phosphate, ultrasonic treatment at a power of 500W for 5 minutes to form a uniform organic phase;

[0059] S2, another liquid preparation tank is taken, and injection water and a polyvinyl alcohol solution with a mass concentration of 1% are added, the amount ratio of injection water to the polyvinyl alcohol solution with a mass concentration of 1% to the dichloromethane in S1 is 100L:1kg:20L, stirring at a speed of 500rpm for 15 minutes to obtain an aqueous phase;

[0060] S3, the organic phase is added dropwise into the aqueous phase at a rate of 5L / h, stirring at a speed of 800rpm during the dropwise addition, after the dropwise addition is completed, continue to stir for 10 minutes, then transfer to a high-pressure homogenizer, homogenize at a pressure of 800bar for 3 times, each cycle for 2 minutes, then transfer to a rotary evaporator, evaporate at 30℃ under reduced pressure for 4 hours to remove dichloromethane, and at the same time promote the crosslinking of polyethylene glycol disuccinimidyl succinate and polyethylene glycol-poly (ε-caprolactone) to obtain a suspension, the residual amount of dichloromethane in the suspension needs to be ≤600ppm;

[0061] S4, transfer the suspension to a centrifuge bucket, centrifuge at 12000 rpm for 20 minutes at 4°C, discard the supernatant, obtain the precipitate, add sterile PBS buffer to resuspend the precipitate, repeat the centrifugation 3 times to remove free drugs and polyvinyl alcohol, ensure that the polyvinyl alcohol residue is ≤0.1%, obtain the nanoparticle precipitate, resuspend the nanoparticle precipitate with water for injection, the amount ratio of sterile PBS buffer and water for injection to dichloromethane described in S1 is 100L:50L:20L, obtain the nanoparticle suspension;

[0062] S5, add mannitol to the nanoparticle suspension, stir at 500 rpm for 20 minutes until completely dissolved, form a drug solution, then filter the drug solution through a 0.45μm polypropylene filter membrane, and finally filter through a 0.22μm polyether sulfone sterile filter membrane, collect into a sterile storage tank, obtain the pre-freeze drug solution;

[0063] S6, fill the pre-freeze drug solution into brown vials, half plug, then place the vials into a freeze dryer, keep at -45°C for 3 hours, sublimate dry, vacuum plug, and cap to obtain the injection dorycin.

[0064] Example 3

[0065] Preparation of injection dorycin:

[0066] First, the injection dorycin includes the following mass parts of raw materials:

[0067] Dorycin hydrochloride 30 parts;

[0068] Polyethylene glycol-polycaprolactone (PEG-PCL) 45 parts;

[0069] Polyethylene glycol disuccinimidyl succinate (SS-PEG-SS) 15 parts;

[0070] Vitamin C phosphate 6 parts;

[0071] Mannitol 15 parts;

[0072] Then, the preparation method of the above injection dorycin includes the following steps:

[0073] S1, according to the mass parts, add dichloromethane, polyethylene glycol-polycaprolactone, and polyethylene glycol disuccinimidyl succinate to the liquid preparation tank, the amount ratio of dichloromethane to polyethylene glycol-polycaprolactone is 20L:5kg, stir at 300 rpm for 35 minutes, then add dorycin hydrochloride and vitamin C phosphate, and ultrasonically treat at 500W for 5 minutes to form a uniform organic phase;

[0074] S2, another liquid preparation tank is taken, and water for injection and a polyvinyl alcohol solution with a mass concentration of 1% are added, the amount of water for injection and the polyvinyl alcohol solution with a mass concentration of 1% is 100L:1kg:20L, and the stirring rate is 500rpm for 20 minutes to obtain an aqueous phase;

[0075] S3, the organic phase is added dropwise into the aqueous phase at a rate of 6L / h, and stirring is performed at a rate of 800rpm during the dropwise addition, after the dropwise addition, stirring is continued for 10 minutes, then it is transferred to a high-pressure homogenizer, and homogenization is performed at a pressure of 800bar for 3 times, each time for 2 minutes, then it is transferred to a rotary evaporator, and evaporation is performed at 30°C under reduced pressure for 4 hours to remove dichloromethane, and at the same time, cross-linking of polyethylene glycol disuccinimidyl butane and polyethylene glycol-polycaprolactone is promoted, to obtain a suspension, and the residual amount of dichloromethane in the suspension needs to be ≤600ppm;

[0076] S4, the suspension is transferred to a centrifuge bucket, and centrifugation is performed at 12000rpm for 20 minutes at 4°C, the supernatant is discarded, and a precipitate is obtained, then sterile PBS buffer is added to resuspend the precipitate, and centrifugation is repeated 3 times to remove free drugs and polyvinyl alcohol, and the residual amount of polyvinyl alcohol is ensured to be ≤0.1%, to obtain a nanoparticle precipitate, then the nanoparticle precipitate is resuspended with water for injection, and the amount of sterile PBS buffer and water for injection is 100L:50L:20L, to obtain a nanoparticle suspension;

[0077] S5, mannitol is added to the nanoparticle suspension, and stirring is performed at a rate of 500rpm for 20 minutes until complete dissolution, to form a drug solution, then the drug solution is coarsely filtered through a 0.45μm polypropylene filter membrane, and then terminal filtration is performed through a 0.22μm polyether sulfone sterile filter membrane, and then it is collected into a sterile storage tank, to obtain a pre-freeze-drying drug solution;

[0078] S6, the pre-freeze-drying drug solution is filled into brown vials, and a half plug is pressed, then the vials are placed into a freeze dryer, and sublimation drying, vacuum plugging and capping are performed at -45°C for 3 hours, to obtain doxycycline for injection.

[0079] Comparative Example 1

[0080] The comparative example is prepared according to the prescription and preparation method in CN1868460A.

[0081] Comparative Example 2

[0082] The comparative example is prepared according to the prescription and preparation method in CN101555215A.

[0083] Comparative Example 3

[0084] The comparative example is prepared according to the prescription and preparation method in CN107638395A.

[0085] The long-term stability of the doxycycline hydrochloride injection prepared in Examples 1 to 3 and Comparative Examples 1 to 3 of the present application was investigated (2 to 8°C, avoiding light), and the results are shown in the following table:

[0086] (1) Example 1

[0087]

[0088] (2) Example 2

[0089]

[0090] (3) Example 3

[0091]

[0092] (4) Comparative Example 1

[0093]

[0094] (5) Comparative Example 2

[0095]

[0096] (6) Comparative Example 3

[0097]

[0098] The above results show that after 36 months of storage, the doxycycline hydrochloride injection prepared in Examples 1 to 3 has good appearance, fast reconstitution, stable doxycycline hydrochloride content, and small impurity change; the doxycycline hydrochloride injection prepared in Comparative Example 1 has a general reconstitution rate, and after 36 months of long-term stability storage, the impurities increase significantly; the doxycycline hydrochloride injection prepared in Comparative Example 2 has an increase in related substances during 24 months of long-term stability storage; the doxycycline hydrochloride injection prepared in Comparative Example 3 has good appearance, fast reconstitution, stable doxycycline hydrochloride content, and insignificant increase in impurities after 36 months of long-term stability storage; thus, the quality of Examples 1 to 3 is more stable than that of Comparative Examples 1 and 2, and is slightly lower than that of Comparative Example 3.

[0099] The specificity of the doxycycline hydrochloride injection prepared in Examples 1 to 3 and Comparative Examples 1 to 3 of the present application was investigated, and the investigation method is as follows:

[0100] Target environment: PBS (pH 6.5, simulating tumor / infection focus microenvironment) containing 50 mM GSH (reduced glutathione, highly expressed in tumor / infection focus) + 10 U / mL acid phosphatase (secreted by inflammatory cells in infection focus); normal environment: PBS (pH 7.4, simulating normal tissue body fluid) containing 2 mM GSH (normal tissue level) + 0 U / mL acid phosphatase.

[0101] Two groups of doxycycline hydrochloride injection prepared in examples 1-3 and comparative examples 1-3, each group is 1 mL, respectively added to the above two kinds of environment, 37℃ constant temperature incubator, 48h sampling 0.5 mL, HPLC measurement of doxycycline concentration, calculate the cumulative release rate, the results are as follows:

[0102]

[0103] From the above table, it can be seen that the doxycycline hydrochloride injection prepared in examples 1-3 is released specifically at the target site in the high reduction + enzyme environment (the release rate is 3-4 times that of the normal environment).

[0104] It should be noted that in this paper, such as the term "includes, contains" or any other variant is intended to cover non-exclusive inclusion, so that the process, method, article or equipment including a series of elements not only includes those elements, but also includes other elements not explicitly listed, or also includes the inherent elements of such process, method, article or equipment.

[0105] Although the embodiments of the present application have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and variations can be made to the embodiments without departing from the principles and spirit of the present application, and the scope of the present application is defined by the appended claims and their equivalents.

Claims

1. A doxycycline for injection, characterized by, By mass parts, including the following raw materials: Doxycycline hydrochloride 25-30 parts; Polyethylene glycol-polycaprolactone 35-45 parts; Polyethylene glycol disuccinimidyl succinate 10-15 parts; Vitamin C phosphate 4-6 parts; Mannitol 10-15 parts.

2. The method of preparing doxycycline for injection according to claim 1, wherein, Including the following steps: S1, according to the mass parts of the above raw materials, dichloromethane, polyethylene glycol-polycaprolactone, polyethylene glycol disuccinimidyl succinate, stirring at 300 rpm for 30 minutes, then adding doxycycline hydrochloride, vitamin C phosphate, ultrasonic treatment for 5 minutes at 500 W, forming an organic phase; S2, another liquid preparation tank is taken, and water for injection and a polyvinyl alcohol solution with a mass concentration of 1% are added, and stirring is performed at a speed of 500 rpm for 15 minutes to obtain an aqueous phase; S3, the organic phase is added to the aqueous phase at a rate of 5 L / h, and stirring is performed at a speed of 800 rpm during the addition, after the addition, stirring is continued for 10 minutes, then it is transferred to a high-pressure homogenizer, and homogenized at a pressure of 800 bar for 3 times, each cycle for 2 minutes, then it is transferred to a rotary evaporator, and evaporated at 30°C under reduced pressure for 4 hours to obtain a suspension; S4, the suspension is transferred to a centrifuge barrel, and centrifuged at a speed of 12000 rpm for 20 minutes at 4°C, the supernatant is discarded, the precipitate is resuspended in sterile PBS buffer, and the centrifugation is repeated 3 times to obtain a nanoparticle precipitate, then the nanoparticle precipitate is resuspended in water for injection to obtain a nanoparticle suspension; S5, mannitol is added to the nanoparticle suspension, and stirring is performed at a speed of 500 rpm for 20 minutes to form a drug solution, then the drug solution is coarsely filtered through a 0.45 μm polypropylene filter membrane, and finally filtered through a 0.22 μm polyether sulfone sterile filter membrane, and collected in a sterile storage tank to obtain a pre-freeze-drying drug solution; S6, the pre-freeze-drying drug solution is filled and half-capped, then the vials are placed in a freeze dryer, and sublimation drying is performed at -45°C for 3 hours, then vacuum pressing and capping are performed to obtain doxycycline for injection.

3. The method of preparing doxycycline for injection according to claim 2, wherein The amount ratio of dichloromethane to polyethylene glycol-polycaprolactone in S1 is 20L:5kg.

4. The method of preparing doxycycline for injection according to claim 2, wherein The amount ratio of water for injection and a polyvinyl alcohol solution with a mass concentration of 1% to dichloromethane in S1 is 100L:1kg:20L.

5. The method of preparing doxycycline for injection according to claim 2, wherein The residual amount of dichloromethane in the suspension in S3 needs to be ≤600ppm.

6. The method of preparing doxycycline for injection according to claim 2, wherein The amount ratio of sterile PBS buffer and water for injection to dichloromethane in S1 is 100L:50L:20L.

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