Low-crystallinity hydroxyapatite microspheres and method for preparing the same

Abstract

The application provides a low-crystallinity hydroxyapatite microsphere for implantation and a preparation method thereof, and the preparation method comprises the following steps: preparing a mixed solution of a calcium source reagent and a phosphorus source reagent, adding inorganic acid, alkali and a buffer into the mixed solution, adding a surfactant, and obtaining a hydroxyapatite micro-particle precipitate through constant-temperature reaction; cleaning the hydroxyapatite micro-particle precipitate, and preparing a hydroxyapatite micro-particle suspension by adding water; and performing spray drying on the hydroxyapatite micro-particle suspension to obtain the hydroxyapatite microsphere; wherein the surfactant is one or more of PEG and PVP. The hydroxyapatite microsphere obtained by the application has stable structure, belongs to spherical large particles, is easy to inject, and is not prone to agglomeration; when the hydroxyapatite microsphere is injected into a human body, the hydroxyapatite microsphere has good supporting effect; and the preparation method of the application is simple, has few process steps, and is easy to realize reaction conditions, and is suitable for large-scale production.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of medical biomaterials, and in particular to a low-crystallinity hydroxyapatite microsphere and a preparation method thereof. BACKGROUND

[0002] Hydroxyapatite is the main inorganic component of human and animal bones. It can realize chemical bond combination with body tissues at the interface, has a certain solubility in the body, can release ions harmless to the body, and can participate in metabolism in the body. As an ideal bone repair material, hydroxyapatite has the advantages of good bone conduction, biological activity, and biocompatibility. Porous hydroxyapatite microspheres have a wide range of applications in the fields of tissue engineering and drug separation due to their low relative density, high specific surface area, and light weight.

[0003] At present, the methods for preparing hydroxyapatite microspheres mainly include hydrothermal method, microemulsion method, and spray drying method. Chinese patent (application number 201580022238.7) discloses a spherical porous hydroxyapatite adsorbent and a method thereof. The method is to first prepare a suspension of hydroxyapatite primary particles, then spray dry the suspension of hydroxyapatite primary particles to obtain hydroxyapatite microspheres, and finally perform a complex screening method to obtain spherical hydroxyapatite. Chinese patent (application number 201710225034.4) discloses a hydroxyapatite microsphere and a preparation method thereof. The method is to first prepare hydroxyapatite primary particles, then filter, wash, and dry the particles, grind the particles into powder, re-prepare an aqueous solution, and finally spray dry. The process is too complex and is not suitable for mass production.

[0004] The current methods for preparing hydroxyapatite microspheres have many problems, such as uneven particle size distribution of the microspheres, low yield, and complex preparation process. The current methods for preparing hydroxyapatite microspheres do not have a process for specifically preparing low-crystallinity hydroxyapatite.

[0005] Therefore, it is urgent to develop a new type of low-crystallinity hydroxyapatite microsphere preparation method that can be industrially produced. SUMMARY

[0006] The purpose of the present application is to provide a hydroxyapatite microsphere that has a uniform particle size distribution and a low crystallinity, is suitable for bone tissue repair and filling, and has a simple preparation process.

[0007] In a first aspect, the present application provides a low-crystallinity hydroxyapatite microsphere preparation method, which comprises the following steps:

[0008] S1: preparing a mixed solution of a calcium source reagent and a phosphorus source reagent;

[0009] S2: adding an inorganic acid, a base, and a buffer to the mixed solution, and then adding a surfactant, and reacting at a constant temperature to obtain hydroxyapatite microparticle precipitation;

[0010] S3: washing the hydroxyapatite particle precipitate and adding water to prepare a hydroxyapatite particle suspension;

[0011] S4: spray drying the hydroxyapatite microparticle suspension, wherein the spray drying parameters are a feed rate of 0.5-5 L / h, a temperature of 120-150° C., and a centrifugal speed of 15,000-30,000 rpm to obtain the hydroxyapatite microspheres;

[0012] Wherein, the surfactant is one or more of PEG and PVP.

[0013] By adopting the above technical solution, by adding PEG and / or PVP in conjunction with specific spray drying conditions, the performance limitations of traditional high-crystallinity hydroxyapatite microspheres are overcome, and the bioactivity of the implant material is improved. In addition, by directly forming in one step through spray drying, steps such as screening and secondary dispersion can be omitted, reducing the number of process steps by more than 50%.

[0014] Optionally, the calcium source reagent is calcium chloride; the phosphorus source reagent is selected from at least one of phosphate ion, monohydrogen phosphate ion or dihydrogen phosphate ion.

[0015] Optionally, the inorganic acid is sulfuric acid or hydrochloric acid, the base is sodium hydroxide, and the buffer is PBS buffer.

[0016] Optionally, the molecular weight of the PEG is 3000-6000, and the molecular weight of the PVP is 20000-50000.

[0017] By adopting the above technical solution and adding PEG and PVP of specific molecular weight, the growth morphology of hydroxyapatite particles can be effectively controlled to form spherical precipitates while reducing their crystallinity.

[0018] Optionally, in step S2, the temperature of the isothermal reaction is 80-90° C., and the reaction time is 2-4 hours.

[0019] Optionally, in step S2, the temperature of the isothermal reaction is 50-60° C., and the reaction time is 18-24 hours.

[0020] By adopting the above technical solution, the reaction temperature (50-60°C) and time (18-24h) are adjusted to inhibit the excessive growth of HA crystal nuclei and reduce the crystallinity.

[0021] In a second aspect, the present invention provides hydroxyapatite microspheres obtained by the preparation method described above, wherein the particle size of the hydroxyapatite microspheres is 25 μm-50 μm.

[0022] By adopting the above technical solution, hydroxyapatite microspheres in this particle size range have good filling effect and cell compatibility in bone repair and tissue engineering applications.

[0023] Optionally, the crystallinity of the hydroxyapatite microspheres is less than 50%.

[0024] By adopting the above technical solution, the degradation rate of low-crystallinity microspheres in the body is significantly improved, increasing the safety of human use; under the same conditions, the smaller the crystallinity, the greater the solubility, the higher the concentration of released calcium and phosphorus, and thus the stronger the reaction performance; and the lower the crystallinity of hydroxyapatite, the larger the specific surface area and the better the biological activity.

[0025] Optionally, the tap density of the hydroxyapatite microspheres is 0.6-1.0 g / mL.

[0026] In a third aspect, the present invention provides a use of the hydroxyapatite microspheres described above in bone tissue repair and filling, a drug sustained-release carrier, or a tissue engineering scaffold.

[0027] The beneficial effects of the present invention are as follows:

[0028] 1. The prepared hydroxyapatite has low crystallinity, breaking through the performance limitations of traditional high-crystallinity hydroxyapatite microspheres, facilitating cell adhesion, proliferation, and tissue ingrowth, thereby improving the bioactivity of implant materials;

[0029] 2. The prepared hydroxyapatite microspheres have uniform particle size, high sphericity and good dispersibility, and have good filling effect and cell compatibility in bone repair and tissue engineering applications;

[0030] 3. The entire preparation method is relatively simple and does not require complicated equipment and operations, which reduces production costs and is conducive to large-scale production. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a scanning electron microscope image of hydroxyapatite microspheres obtained in Example 1 of the present invention;

[0032] Figure 2 This is a scanning electron microscope image of hydroxyapatite microspheres obtained in Example 2 of the present invention;

[0033] Figure 3 This is a scanning electron microscope image of hydroxyapatite microspheres obtained in Comparative Example 1 of the present invention;

[0034] Figure 4 This is a scanning electron microscope image of hydroxyapatite microspheres obtained in Comparative Example 2 of the present invention;

[0035] Figure 5 This is the XRD pattern of hydroxyapatite microspheres obtained in Example 1 of the present invention;

[0036] Figure 6 This is the XRD test pattern of hydroxyapatite microspheres obtained in Comparative Example 2 of the present invention. DETAILED DESCRIPTION

[0037] In order to better illustrate the purpose, technical solutions and advantages of the present invention, the present application will be further described below in conjunction with specific embodiments.

[0038] In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments.

[0039] Example 1

[0040] A method for preparing hydroxyapatite microspheres comprises the following steps:

[0041] S1: Weigh 71 g of disodium hydrogen phosphate and 122 g of calcium chloride dihydrate, dissolve them in 40 L of water, and mix to prepare solution A;

[0042] S2: Add 10 L of 1 mol / L hydrochloric acid to Solution A and stir to mix. Once the reaction solution becomes clear, add 192 g of sodium hydroxide to neutralize the reaction. Then, add PBS buffer and adjust the pH to 7.0. Then, add 2.51 g of PEG (McLean, average Mn 4000) and 0.94 g of PVP (McLean, average molecular weight 24000, K23-27). Maintain the reaction temperature at 50°C for 20 hours, until the reaction is complete and all hydroxyapatite particles are precipitated.

[0043] S3: After the precipitate was washed with purified water six times, an appropriate amount of purified water was added to obtain a hydroxyapatite microparticle suspension B with a concentration of 10 g / L.

[0044] S4: spray drying the suspension B using a centrifugal spray drying device. The spraying process parameters are as follows: the feed flow rate of the suspension is 2 L / h, the rate of the spray drying centrifuge is 18,000 rpm, and the spray drying temperature is 120°C.

[0045] The hydroxyapatite microspheres prepared in Example 1 had a particle size of 25-50 μm, a D50 of 35.06 μm, a tap density of 0.73 g / mL, and a crystallinity of 22%. They were large spherical particles with low crystallinity, a stable spherical structure, high sphericity (roundness ≥ 0.9), and good dispersibility (agglomeration index ≤ 0.1), making them suitable for subcutaneous injection filling.

[0046] Example 2

[0047] A method for preparing hydroxyapatite microspheres comprises the following steps:

[0048] S1: Weigh 71 g of disodium hydrogen phosphate and 122 g of calcium chloride dihydrate, dissolve them in 40 L of water, and mix to prepare solution A;

[0049] S2: Add 10 L of 1 mol / L hydrochloric acid to Solution A and stir to mix. Once the reaction solution becomes clear, add 192 g of sodium hydroxide to neutralize the reaction. Then, add PBS buffer and adjust the pH to 7.0. Then, add 2.51 g of PEG (McLean, average Mn 4000) and 0.94 g of PVP (McLean, average molecular weight 24000, K23-27). Maintain the reaction temperature at 80°C for 3 hours, until the reaction is complete and all hydroxyapatite particles are precipitated.

[0050] S3: After the precipitate was washed with purified water six times, an appropriate amount of purified water was added to obtain a hydroxyapatite microparticle suspension B with a concentration of 10 g / L.

[0051] S4: The suspension B is spray-dried using a centrifugal spray drying device. The spraying process parameters are as follows: the feed flow rate of the suspension is 4 L / h, the rate of the spray drying centrifuge is 25,000 rpm, and the spray drying temperature is 150°C.

[0052] The particle size of the hydroxyapatite microspheres prepared in Example 2 is 25-50 μm, D 50 The particle size is 31.15 μm, the tap density is 0.79 g / mL, and the crystallinity is 30%. It is a large spherical particle with low crystallinity and a stable spherical structure.

[0053] Comparative Example 1

[0054] A method for preparing hydroxyapatite microspheres comprises the following steps:

[0055] S1: Weigh 71 g of disodium hydrogen phosphate and 122 g of calcium chloride dihydrate, dissolve them in 40 L of water, and mix to prepare solution A;

[0056] S2: Add 10 L of 1 mol / L hydrochloric acid to Solution A and stir. Once the reaction solution becomes clear, add 192 g of sodium hydroxide to neutralize the reaction. Then, add PBS buffer and adjust the pH to 7.0. Maintain the reaction temperature at 80°C for 3 hours, until the reaction is complete and all hydroxyapatite particles are precipitated.

[0057] S3: After the precipitate was washed with purified water six times, an appropriate amount of purified water was added to obtain a hydroxyapatite microparticle suspension B with a concentration of 10 g / L.

[0058] S4: The suspension B is spray-dried using a centrifugal spray drying device. The spraying process parameters are as follows: the feed flow rate of the suspension is 4 L / h, the rate of the spray drying centrifuge is 25,000 rpm, and the spray drying temperature is 150°C.

[0059] The particle size of the hydroxyapatite microspheres prepared in this comparative example 1 is 5-30 μm, D 50 The particle size is 15.18 μm, the tap density is 1.2 g / ml, the crystallinity is 65%, and it is spherical with a wide particle size distribution range.

[0060] Comparative Example 2

[0061] A method for preparing hydroxyapatite microspheres comprises the following steps:

[0062] S1: Weigh 71 g of disodium hydrogen phosphate and 122 g of calcium chloride dihydrate, dissolve them in 40 L of water, and mix to prepare solution A;

[0063] S2: Add 10 L of 1 mol / L hydrochloric acid to Solution A and stir. Once the reaction solution becomes clear, add 192 g of sodium hydroxide to neutralize the reaction. Then, add PBS buffer and adjust the pH to 7.0. Maintain the reaction temperature at 80°C for 3 hours, until the reaction is complete and all hydroxyapatite particles are precipitated.

[0064] S3: After the precipitate was washed with purified water six times, an appropriate amount of purified water was added to obtain a hydroxyapatite microparticle suspension B with a concentration of 10 g / L.

[0065] S4: The suspension B is spray-dried using a centrifugal spray drying device. The spraying process parameters are as follows: the feed flow rate of the suspension is 4 L / h, the rate of the spray drying centrifuge is 30,000 rpm, and the spray drying temperature is 300°C.

[0066] The particle size of the hydroxyapatite microspheres prepared in this comparative example 2 is 3-30 μm, D 50The particle size is 15.05 μm, the tap density is 1.8 g / mL, the crystallinity is 90%, some microspheres are not spherical, and the particle size distribution range is wide.

[0067] Performance testing

[0068] 1. According to the method of GB / T 23101.3-2023 Hydroxyapatite for surgical implants Part 3: Chemical analysis and characterization of crystalline phase and phase purity, the crystallinity of the microspheres of Example 1 and Comparative Example 2 was tested respectively. The results are as follows: Figure 5 and Figure 6 As shown in the figure, it can be clearly seen that Figure 5 The diffraction peak width increases, and Figure 6 The diffraction peaks are basically sharp, indicating that the crystallinity of the microspheres in Example 1 is significantly lower than that in Comparative Example 2.

[0069] 2. The microspheres obtained in Example 1-2 and Comparative Example 1-2 were detected using GeminiSEM 300. The scanning electron microscope images obtained were as follows: Figures 1-4 As shown in the figure, Figure 3 and Figure 4 The microspheres are small and unevenly distributed. Figure 4 There are also some that are not spherical, indicating that the microspheres prepared in Comparative Examples 1-2 are not suitable for the application of this application; Figure 1 and Figure 2 The microspheres have appropriate particle size, uniform distribution and high sphericity, making them very suitable for subcutaneous injection filling or bone repair.

[0070] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A method for preparing hydroxyapatite microspheres with low crystallinity, characterized in that: The steps include: S1: preparing a mixed solution of a calcium source reagent and a phosphorus source reagent; S2: adding an inorganic acid, a base, and a buffer to the mixed solution, and then adding a surfactant, and reacting at a constant temperature to obtain hydroxyapatite microparticle precipitation; S3: washing the hydroxyapatite particle precipitate and adding water to prepare a hydroxyapatite particle suspension; S4: spray drying the hydroxyapatite microparticle suspension, wherein the spray drying parameters are a feed rate of 0.5-5 L / h, a temperature of 120-150° C., and a centrifugal speed of 15,000-30,000 rpm to obtain the hydroxyapatite microspheres; Wherein, the surfactants are PEG and PVP, the molecular weight of the PEG is 3000-6000, and the molecular weight of the PVP is 20000-50000.

2. The method for preparing hydroxyapatite microspheres according to claim 1, wherein: The calcium source reagent is calcium chloride; the phosphorus source reagent is selected from at least one of phosphate ion, monohydrogen phosphate ion or dihydrogen phosphate ion.

3. The method for preparing hydroxyapatite microspheres according to claim 1, wherein: The inorganic acid is sulfuric acid or hydrochloric acid, the base is sodium hydroxide, and the buffer is PBS buffer.

4. The method for preparing hydroxyapatite microspheres according to claim 1, wherein: In step S2, the temperature of the isothermal reaction is 80-90° C., and the reaction time is 2-4 hours.

5. The method for preparing hydroxyapatite microspheres according to claim 1, wherein: In step S2, the temperature of the isothermal reaction is 50-60° C., and the reaction time is 18-24 hours.

6. Hydroxyapatite microspheres obtained by the method for preparing hydroxyapatite microspheres according to any one of claims 1 to 5, characterized in that: The particle size of the hydroxyapatite microspheres is 25 μm-50 μm.

7. The hydroxyapatite microspheres according to claim 6, characterized in that: The crystallinity of the hydroxyapatite microspheres is less than 50%.

8. The hydroxyapatite microspheres according to claim 6, characterized in that: The tap density of the hydroxyapatite microspheres is 0.6-1.0 g / mL.

9. Use of the hydroxyapatite microspheres according to any one of claims 6 to 8 in bone tissue repair and filling, drug sustained-release carriers or tissue engineering scaffolds.

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