Mesoporous hydroxyapatite microspheres regulated by malic acid and a preparation method thereof

Abstract

This invention provides mesoporous hydroxyapatite microspheres regulated by malic acid and their preparation method. The preparation method includes: dissolving a calcium source and a phosphorus source sequentially in deionized water, mixing thoroughly, and then adding nitric acid or sodium hydroxide dropwise to the reaction solution to adjust the pH until clear; adding malic acid of different concentrations as a morphology regulator to the clear reaction solution, followed by the addition of urea; transferring the prepared solution to a hydrothermal reactor and reacting at a set temperature for 15 min–12 h; after the reaction is complete, cooling the solution to room temperature, centrifuging and drying to obtain hydroxyapatite powder samples. This invention regulates the sphericity, particle size, surface micro / nano structure, and crystallinity of hydroxyapatite microspheres by controlling the concentration of the morphology regulator malic acid and adjusting the reaction time and temperature. The hydroxyapatite microspheres prepared by this invention are expected to be applied in clinical fields such as drug controlled-release systems, injectable facial wrinkle removal, and hard tissue repair.

Description

Technical Field

[0001] This invention relates to the field of inorganic biomaterials technology, and more specifically, to a mesoporous hydroxyapatite microsphere regulated by malic acid and its preparation method. Background Technology

[0002] Hydroxyapatite (HAP) is a major inorganic component of human hard tissue and is widely used in biomedical fields such as bone implantation, wound repair, regenerative filling, and drug delivery. HAP's biocompatibility and osteoconductivity with bone tissue give it significant advantages in bone defect repair. Designing HAP particles with specific morphological structures according to the application needs of different fields is crucial for improving its functionality and application performance.

[0003] The morphology and surface nanostructure of HAP particles significantly influence their performance in biomedicine. Porous HAP microspheres, due to their high specific surface area and unique pore structure, can significantly enhance the material's adsorption capacity, interactions with proteins and cells, and its bioactivity, showing strong application potential in drug loading and tissue engineering. Therefore, developing HAP microspheres with tunable pore structure and morphology is crucial for improving their performance in biomedical applications.

[0004] Currently, methods for preparing calcium phosphate microspheres include the sol-gel method, microemulsion method, spray drying method, and template method. The sol-gel and microemulsion methods offer mild synthesis conditions but suffer from low yields and difficulty in precisely controlling the morphology and surface nanostructure of the microspheres. While the spray drying method offers high production efficiency, it struggles to control the morphology and size of the microspheres, resulting in poor particle uniformity. In contrast, the template method provides a relatively simple and effective means of controlling the morphology and surface structure of calcium phosphate particles. Soft template methods, in particular, utilize the interaction between organic molecules and specific sites on the inorganic crystal surface to regulate nucleation and crystal growth by self-assembling templates with specific morphologies in solution, thereby achieving control over the morphology of HAP particles. However, compared to macromolecular template agents, the limited adsorption modes and restricted surface coverage of small molecules result in insufficient stability in controlling crystal morphology. Therefore, developing a stable and efficient hydrothermal synthesis method that combines the advantages of small molecule modulators to achieve precise control over the morphology and structure of HAP microspheres remains a challenge. Summary of the Invention

[0005] In view of the limitations of existing technologies, and to achieve the above objectives, the present invention adopts the following technical solution:

[0006] In a first aspect, this application provides a method for preparing mesoporous hydroxyapatite microspheres regulated by malic acid, comprising the following steps:

[0007] Step S1: Dissolve the calcium source and phosphorus source in deionized water in sequence, mix them evenly, and prepare a reaction solution;

[0008] Step S2: Add nitric acid or sodium hydroxide dropwise to the reaction solution to adjust the pH of the solution until it is clear;

[0009] Step S3: Add malic acid of different concentrations as morphology modifiers to the clear reaction solution, then add urea and stir continuously for 5 to 15 minutes;

[0010] Step S4: Transfer the prepared solution into the reaction vessel and keep it at the set temperature for 15 min-12 h.

[0011] Step S5: After the reaction is complete, the solution is cooled to room temperature, the precipitate is separated by centrifugation, the precipitate is repeatedly washed with deionized water and ethanol, and then dried at 80°C for 48 hours. After centrifugation and drying, a hydroxyapatite powder sample with uniform morphology and high specific surface area is finally obtained.

[0012] By using a specific morphology modifier—malic acid, an organic small molecule acid—and controlling the concentration of the morphology modifier, the particle size, pore size, and specific surface area of ​​hydroxyapatite can be regulated, resulting in excellent spheroidization.

[0013] Preferably, in step S1, calcium nitrate tetrahydrate and disodium hydrogen phosphate dodecahydrate are added sequentially to 50 mL of deionized water to make the calcium source concentration 0.1 mol / L and the calcium-to-phosphorus molar ratio 1.67, thus obtaining a reaction solution.

[0014] Preferably, in step S2, the pH of the solution is adjusted with nitric acid or sodium hydroxide until the solution is clear, and the pH value of the solution is measured to be 2-3.

[0015] Preferably, in step S3, the molar ratio of urea to calcium source is 0.56:0.1, and the malic acid is L-malic acid, with a molar ratio of (0.05-0.2):0.1 to the calcium source.

[0016] Preferably, in step S3, the concentration of malic acid added is 335 mg - 1.34 g / L.

[0017] Preferably, in step S4, the set temperature is 120℃-180℃. By adjusting the reaction time and temperature, the particle size and pore size of the hydroxyapatite microspheres can be adjusted.

[0018] Preferably, as the reaction time increases, the particles change from amorphous nanoparticles (ACP) to HAP particles, and the particle phase changes; at the same time, as the reaction time prolongs, the crystallinity of the HAP particles increases.

[0019] Preferably, the increase in malic acid concentration leads to a denser surface of hydroxyapatite particles, a smaller pore size, and a corresponding decrease in average particle size. Specifically, the average particle size decreases from 17.90±7.52μm to 11.97±4.79μm, and the pore size decreases from the micrometer level to the nanometer level. When the malic acid concentration is 100mM, larger particles of about 40μm appear.

[0020] Preferably, the hydroxyapatite powder obtained by the preparation method has uniform morphological characteristics and significant specific surface area, which enables the preparation of hydroxyapatite microspheres with different morphologies.

[0021] Secondly, this application also provides mesoporous hydroxyapatite microspheres regulated by malic acid, obtained by the above preparation method.

[0022] The beneficial effects of this invention are as follows:

[0023] This invention proposes a method for preparing mesoporous hydroxyapatite (HAP) microspheres regulated by malic acid. Hydroxyapatite (HAP) particles with uniform morphology and high specific surface area are synthesized via a hydrothermal method. This method aims to address the problems of imprecise morphology control, difficulty in controlling particle size, and low specific surface area in existing HAP preparation techniques. By using malic acid as a small-molecule regulator, this invention can effectively adjust the morphology, size, and surface microstructure of HAP particles, thereby obtaining more uniform HAP microspheres. Compared with existing technologies, the preparation method proposed in this invention has the advantages of simple preparation process and high morphology controllability. It can also achieve highly crystalline HAP particles without relying on complex equipment, making it suitable for the preparation of HAP materials requiring high specific surface area and uniform morphology, laying the foundation for further biomedical research and applications.

[0024] This invention utilizes malic acid as a small molecule regulator to precisely control the morphology of HAP particles by adjusting its concentration. The resulting particles exhibit a uniform spherical nanostructure with a high specific surface area, which is beneficial for further applications, such as drug loading, bone defect filling, or the preparation of composite biomaterials.

[0025] This invention utilizes the regulatory effect of malic acid to produce particles with a porous surface structure, which significantly increases the specific surface area of ​​HAP. This high specific surface area not only enhances the adsorption capacity of the material but also provides it with broad application potential in areas such as drug delivery and bone defect filling.

[0026] This invention employs a hydrothermal synthesis process, which is simple to operate, has mild reaction conditions, requires no complex equipment, and is easy to achieve large-scale preparation. Compared with other complex template methods or spray drying methods, it can achieve precise control of HAP particle morphology while ensuring high crystallinity.

[0027] This invention allows for the synthesis of HAP particles of different sizes and structures by controlling the ratio of malic acid to calcium source, meeting the specific material requirements of different application fields. In particular, the adjustability of particle size and morphology greatly improves the applicability of the material in applications such as bone repair materials and drug carriers.

[0028] The preparation method provided by this invention uses a specific morphology regulator, the organic small molecule acid malic acid, and controls the concentration of the morphology regulator to regulate the particle size, pore size, and specific surface area of ​​hydroxyapatite, resulting in excellent spheroidization effect. By adjusting the reaction time and temperature, the particle size and pore size of hydroxyapatite microspheres can be adjusted, enabling the preparation of hydroxyapatite microspheres with different morphologies.

[0029] Other features and advantages of the invention will be set forth in the following description, and will be apparent in part from the description, or may be learned by practicing embodiments of the invention. The objects and other advantages of the invention may be realized and obtained by means of the structures particularly pointed out in the written description, claims, and drawings. Attached Figure Description

[0030] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention and should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can be obtained based on these drawings without creative effort.

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

[0032] Figure 1B The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in Example 1 of this invention;

[0033] Figure 2A This is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 2 of the present invention;

[0034] Figure 2B The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in Example 2 of this invention;

[0035] Figure 2C This is a particle size distribution diagram of the hydroxyapatite microspheres prepared in Example 2 of the present invention;

[0036] Figure 3A This is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 3 of the present invention;

[0037] Figure 3BThe X-ray diffraction pattern of the hydroxyapatite microspheres prepared in Example 3 of this invention;

[0038] Figure 3C This is a particle size distribution diagram of the hydroxyapatite microspheres prepared in Example 3 of the present invention;

[0039] Figure 3D The Fourier transform infrared spectrum of the hydroxyapatite microspheres prepared in Example 3 of this invention;

[0040] Figure 4A This is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 4 of the present invention;

[0041] Figure 4B This is a particle size distribution diagram of the hydroxyapatite microspheres prepared in Example 4 of the present invention;

[0042] Figure 5A This is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 5 of the present invention;

[0043] Figure 5B The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in Example 5 of this invention;

[0044] Figure 5C This is a particle size distribution diagram of the hydroxyapatite microspheres prepared in Example 5 of the present invention;

[0045] Figure 6A This is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 6 of the present invention;

[0046] Figure 6B This is a particle size distribution diagram of the hydroxyapatite microspheres prepared in Example 6 of the present invention;

[0047] Figure 7 The image shown is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 7 of this invention.

[0048] Figure 8 This is a scanning electron microscope image of the hydroxyapatite microspheres prepared in Example 8 of the present invention;

[0049] Figure 9 This is a schematic diagram of the preparation method of hydroxyapatite microspheres regulated by malic acid according to the present invention. Detailed Implementation

[0050] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the claimed invention, but merely to illustrate selected embodiments of the invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention.

[0051] It should be noted that similar reference numerals and letters in the following figures indicate similar items; therefore, once an item is defined in one figure, it does not need to be further defined and explained in subsequent figures. Furthermore, in the description of this invention, terms such as "first," "second," etc., are used only to distinguish descriptions and should not be construed as indicating or implying relative importance.

[0052] Existing methods for preparing calcium phosphate microspheres include the sol-gel method, microemulsion method, spray drying method, and template method. The sol-gel and microemulsion methods offer mild synthesis conditions but have low yields and struggle to precisely control the morphology and surface nanostructure of the microspheres. While the spray drying method offers high production efficiency, it is difficult to control the morphology and size of the microspheres, resulting in poor particle uniformity. This invention provides a technical solution for synthesizing mesoporous hydroxyapatite (HAP) microspheres with uniform morphology and high specific surface area via a hydrothermal method using malic acid as a morphology regulator. The proposed method for preparing malic acid-regulated mesoporous hydroxyapatite microspheres introduces malic acid during the hydrothermal reaction, utilizing its coordination with calcium ions to precisely control the nucleation and growth process of HAP particles, thereby achieving control over particle morphology. Specifically, by adjusting the ratio of malic acid to calcium source, the reaction temperature, and the time, spherical HAP particles with nanoporous structures can be obtained, and both particle size and specific surface area are controllable.

[0053] Example 1:

[0054] Reference Figure 9 A schematic diagram of the preparation method of malic acid-regulated hydroxyapatite microspheres is shown. This embodiment provides a method for preparing malic acid-regulated mesoporous hydroxyapatite microspheres as follows:

[0055] S1. Add 1.180g Ca(NO3)2·4H2O and 1.074g Na2HPO4·12H2O to 50mL of deionized water in sequence, so that the calcium-to-phosphorus molar ratio is 1.67. Stir continuously for a period of time, from 5 to 15 minutes.

[0056] S2. Gradually add nitric acid until the solution becomes clear, about 500 μL. Measure the pH of the solution to be 2-3.

[0057] S3. Add 335.2 mg of malic acid (MA) to the solution to make the malic acid concentration 50 mM. Then add 1.68 g of urea and stir continuously for a period of time to obtain the final reaction solution. The stirring time should be 5-15 minutes.

[0058] S4. Transfer the final reaction solution into a hydrothermal reactor and place it in a drying oven that has been heated to 150°C for 3 hours.

[0059] S5. After the reaction is complete, cool the solution to room temperature. Centrifuge at 5000 rpm for 3 min. Wash the precipitate twice each with deionized water and ethanol. Dry the washed precipitate in an oven at 80℃ for 48 hours to obtain hydroxyapatite microspheres.

[0060] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 1A As shown in the figure, it can be seen that the particles collected after the reaction have certain unformed cluster structures, and the particles exhibit a flower-like structure composed of plates.

[0061] The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 1B As shown in the figure, it can be seen that the malic acid-regulated hydroxyapatite microspheres (MA-HA) obtained correspond to the standard diffraction characteristic peaks of pure hydroxyapatite, indicating that the phase composition of the prepared microspheres is hydroxyapatite and no other impurity phases are generated.

[0062] Example 2:

[0063] Reference Figure 9 A schematic diagram of the preparation method of malic acid-regulated hydroxyapatite microspheres is shown. This embodiment provides a method for preparing malic acid-regulated mesoporous hydroxyapatite microspheres as follows:

[0064] S1. Add 1.180g Ca(NO3)2·4H2O and 1.074g Na2HPO4·12H2O to 50mL of deionized water in sequence, so that the calcium-to-phosphorus molar ratio is 1.67. Stir continuously for a period of time, from 5 to 15 minutes.

[0065] S2. Gradually add nitric acid until the solution becomes clear, about 500 μL. Measure the pH of the solution to be 2-3.

[0066] S3. Add 670.4 mg of malic acid to the solution to make the malic acid concentration 100 mM. Then add 1.68 g of urea and stir continuously for 5 to 15 minutes to obtain the final reaction solution.

[0067] S4. Transfer the final reaction solution into a hydrothermal reactor and place it in a drying oven that has been heated to 150°C for 3 hours.

[0068] S5. After the reaction is complete, cool the solution to room temperature. Centrifuge at 5000 rpm for 3 min. Wash the precipitate twice each with deionized water and ethanol. Dry the washed precipitate in an oven at 80℃ for 48 hours to obtain hydroxyapatite microspheres.

[0069] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 2A As shown in the figure, the particles collected after the reaction exhibit a good spherical structure, and nanopores appear on the particle surface. The particle size distribution of the hydroxyapatite microspheres prepared in this example is shown in the figure. Figure 2C As shown in the figure, the particle size distribution ranges from 5 to 30 μm, mainly concentrated around 15 μm, with an average particle size of 15.58 ± 4.65 μm. Compared with Example 1, this indicates that an increased concentration of the regulator is beneficial to the formation of spherical particles.

[0070] The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 2B As shown in the figure, it can be seen that the obtained MA-HA corresponds to the standard diffraction characteristic peaks of pure hydroxyapatite, indicating that the phase composition of the prepared microspheres is hydroxyapatite and no other impurity phases are generated.

[0071] Example 3:

[0072] Reference Figure 9 A schematic diagram of the preparation method of malic acid-regulated hydroxyapatite microspheres is shown. This embodiment provides a method for preparing malic acid-regulated mesoporous hydroxyapatite microspheres as follows:

[0073] S1. Add 1.180g Ca(NO3)2·4H2O and 1.074g Na2HPO4·12H2O to 50mL of deionized water in sequence, so that the calcium-to-phosphorus molar ratio is 1.67, and stir continuously for 5 to 15 minutes.

[0074] S2. Gradually add nitric acid until the solution becomes clear, about 500 μL. Measure the pH of the solution to be 2-3.

[0075] S3. Add 1340.9 mg of malic acid to the solution to make the malic acid concentration 200 mM. Then add 1.68 g of urea and stir continuously for 5 to 15 minutes to obtain the final reaction solution.

[0076] S4. Transfer the final reaction solution into a hydrothermal reactor and place it in a drying oven that has been heated to 150°C for 3 hours.

[0077] S5. After the reaction is complete, cool the solution to room temperature. Centrifuge at 5000 rpm for 3 min. Wash the precipitate twice each with deionized water and ethanol. Dry the washed precipitate in an oven at 80℃ for 48 hours to obtain hydroxyapatite microspheres.

[0078] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 3A As shown in the figure, the particles collected after the reaction exhibit a well-defined spherical structure with nanoporous structures on their surface. The particle size distribution of the hydroxyapatite microspheres prepared in this embodiment is shown in the figure below. Figure 3C As shown in the figure, the particle size distribution ranges from 0 to 25 μm, mainly concentrated around 11 μm, with an average particle size of 11.36 ± 3.49 μm. Compared with Example 2, this indicates that increasing the concentration of the regulator leads to a decrease in particle size.

[0079] The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 3B As shown in the figure, the obtained MA-HA corresponds to the standard diffraction characteristic peaks of pure hydroxyapatite. The Fourier transform infrared spectrum of the hydroxyapatite microspheres prepared in this example is shown below. Figure 3D As shown in the figure, the characteristic absorption peaks of the obtained MA-HA correspond to those of pure hydroxyapatite. This indicates that the microspheres were composed of hydroxyapatite and no other impurities were formed.

[0080] Example 4:

[0081] Reference Figure 9 A schematic diagram of the preparation method of hydroxyapatite microspheres regulated by malic acid is shown. In this embodiment, the reaction time in step S4 of Example 1 is changed to 5h, and the remaining operation steps are the same as in Example 1.

[0082] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 4A As shown in the figure, the particles collected after the reaction have a certain spherical structure, and the surface morphology of the particles is uneven. The particle size distribution diagram of the hydroxyapatite microspheres prepared in this example is shown in the figure. Figure 4BAs shown in the figure, the particle size distribution ranges from 5 to 35 μm, with an average particle size of 14.15 ± 6.84 μm. Compared with Example 1, this demonstrates that reaction time affects particle growth.

[0083] Example 5:

[0084] Reference Figure 9 A schematic diagram of the preparation method of hydroxyapatite microspheres regulated by malic acid is shown. In this embodiment, the reaction time in step S4 of Example 2 is changed to 5h, and the remaining operation steps are the same as in Example 2.

[0085] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 5A As shown in the figure, the particles collected after the reaction have a good spherical structure, but the surface morphology of the particles is uneven. The particle size distribution diagram of the hydroxyapatite microspheres prepared in this example is shown in the figure. Figure 5C As shown in the figure, the particle size distribution ranges from 5 to 45 μm, with an average particle size of 17.90 ± 7.52 μm, indicating a relatively large particle size distribution range. Compared to Example 2, larger microspheres of approximately 40 μm appear, suggesting that reaction time affects particle growth.

[0086] The X-ray diffraction pattern of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 5B As shown in the figure, it can be seen that the obtained MA-HA corresponds to the standard diffraction characteristic peaks of pure hydroxyapatite, indicating that the phase composition of the prepared microspheres is hydroxyapatite and no other impurity phases are generated.

[0087] Example 6:

[0088] Reference Figure 9 A schematic diagram of the preparation method of hydroxyapatite microspheres regulated by malic acid is shown. In this embodiment, the reaction time in step S4 of Example 3 is changed to 5h, and the remaining operation steps are the same as in Example 3.

[0089] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 6A As shown in the figure, the particles collected after the reaction have a good spherical structure and a relatively uniform surface morphology. The particle size distribution of the hydroxyapatite microspheres prepared in this example is shown in the figure. Figure 6B As shown in the figure, the particle size distribution ranges from 5 to 25 μm, with an average particle size of 11.97 ± 4.79 μm.

[0090] Example 7:

[0091] Reference Figure 9A schematic diagram of the preparation method of hydroxyapatite microspheres regulated by malic acid is shown. In this embodiment, the reaction time in step S4 of Example 2 is changed to 30 min, and the remaining operation steps are the same as in Example 2.

[0092] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 7 As shown in the figure, a good spherical structure appeared after 30 minutes of reaction, and the particle surface exhibited micron-sized pores.

[0093] Example 8:

[0094] Reference Figure 9 A schematic diagram of the preparation method of hydroxyapatite microspheres regulated by malic acid is shown. In this embodiment, the reaction time in step S4 of Example 2 is changed to 1 hour, and the remaining operation steps are the same as in Example 2.

[0095] The scanning electron microscope image of the hydroxyapatite microspheres prepared in this embodiment is as follows: Figure 8 As shown in the figure, relatively dense microspheres appeared after 1 hour of reaction. Compared with Example 7, this indicates that the hydroxyapatite crystals grow more densely with increasing reaction time. In summary, this invention, through a reasonable and feasible hydrothermal process and with the help of malic acid as a small molecule regulator, achieves effective control over the morphology and structure of hydroxyapatite particles, providing a new solution for the preparation of high-performance biomedical materials.

[0096] It is understood that, as shown in Examples 1 to 3 of this invention, with the increase of malic acid concentration (e.g., from 50 mM to 200 mM), the morphology of the hydroxyapatite microspheres changes from a clustered structure to a more uniform spherical structure, and nanopores appear on the particle surface. This indicates that malic acid, as a morphology modifier, can effectively affect the crystallization morphology and size of hydroxyapatite.

[0097] In this invention, Examples 4 to 8 demonstrate that variations in reaction time significantly affect the morphology and size of hydroxyapatite microspheres. For example, in Example 4 (5 hours reaction time), compared to Example 1 (3 hours), the particle surface morphology was uneven, indicating that a longer reaction time may lead to the growth of particles into a spherical structure. In Example 7 (30 minutes reaction time), compared to Example 8 (1 hour), the crystallinity was poor, while it increased, indicating that extending the reaction time helps improve the crystallinity of hydroxyapatite.

[0098] Understandably, the effect of reaction conditions on the size of hydroxyapatite microspheres is as follows: Compared to Example 2, Example 3 showed a decrease in particle size due to a higher malic acid concentration (200 mM vs. 100 mM), indicating that a higher concentration of malic acid may promote the formation of smaller particles. In Example 5 (reaction time 5 hours), compared to Example 2 (reaction time 3 hours), Example 8 (reaction time 1 hour), and Example 7 (reaction time 30 minutes), the average particle size first decreased and then increased, with the appearance of larger microspheres (approximately 40 μm). This suggests that with increasing reaction time, hydroxyapatite crystals may undergo dissolution and recrystallization, and the longer reaction time leads to an increase in the size of some particles.

[0099] The results of the above examples are shown in Table 1:

[0100] Table 1. Summary table of parameters and results data for each embodiment

[0101]

[0102]

[0103] In practical applications, all embodiments of this invention mention centrifugation and washing with deionized water and ethanol. This helps remove unreacted precursors and byproducts, thereby obtaining pure hydroxyapatite microspheres. The embodiments also mention drying in an 80°C oven for 48 hours, a step crucial for obtaining dry and morphologically stable hydroxyapatite microspheres.

[0104] In summary, the morphology, size, and crystallinity of hydroxyapatite microspheres can be precisely controlled by adjusting the malic acid concentration and reaction time. These factors are crucial for the final application performance of the material, such as in biomedicine, drug delivery, and bone defect filling. These examples allow for the optimization of synthesis conditions to obtain hydroxyapatite microspheres with specific properties.

[0105] To achieve precise control of malic acid concentration and reaction time in experiments, the following methods can be used:

[0106] 1. Use precise measuring equipment: In the experiment, weigh an appropriate amount of malic acid powder directly onto the analytical balance according to the required amount of malic acid. Ensure accuracy during the weighing process and avoid the influence of wind and vibration of the balance.

[0107] 2. Temperature Control: Reaction temperature has a significant impact on the rate of chemical reactions. In experiments, high-throughput reaction devices with individual temperature control and monitoring functions can be used to precisely control the reaction temperature, thereby controlling the reaction time. High-throughput reaction devices are often automated, automatically adjusting the temperature, reducing manual intervention, and improving the repeatability and reliability of experiments.

[0108] 3. Time Control: Use a timer or a device with time control function to precisely control the reaction time. For example, in some experiments, it may be necessary to sample or add reagents at specific time points; in this case, a timer can be used to ensure the accuracy of the operation.

[0109] 4. Monitoring of color changes: In the experiment, the precipitated calcium phosphate crystals in the solution can be removed by adjusting the pH value.

[0110] This is because calcium phosphate dissolves under acidic conditions but precipitates under alkaline conditions.

[0111] In summary, malic acid was used as a morphology modifier in the preparation of mesoporous hydroxyapatite microspheres. Specifically, in the examples, different concentrations of malic acid were added to control the morphology and size of the mesoporous hydroxyapatite microspheres. This method is not common in existing technologies. The addition of malic acid not only affects the morphology of the microspheres but also their surface structure and pore size. Furthermore, urea also had a synergistic effect; urea was added simultaneously with malic acid, and the molar ratio of urea to calcium source was 0.56:0.1. This synergistic effect may have a significant impact on the formation mechanism of the microspheres. This is rare in previous studies; at the same time, the temperature and time of the hydrothermal reaction were precisely controlled in the examples, such as reacting at 150°C for 5 hours. The optimization of these conditions helps to obtain the ideal microsphere morphology and size. It is also mentioned that centrifugation was carried out at a speed of 5000 rpm and repeated washing with deionized water and ethanol was performed. This step helps to remove impurities and improve the purity and surface properties of the microspheres. In addition, during the drying process, the examples mentioned drying in an oven at 80°C for 48 hours. These precise drying conditions help to control the final morphology and physical properties of the microspheres.

[0112] Compared with other existing technologies, such as the method for preparing porous hydroxyapatite microspheres, which mentions using sodium citrate as a template agent to regulate the growth of hydroxyapatite crystals and obtain microspheres with a porous surface structure, this differs from the use of malic acid as a regulator in this embodiment. Malic acid has a unique structure with two carboxyl groups and one hydroxyl group. The presence of the two carboxyl groups provides a more stable binding mode in regulating the nucleation and growth of HAP crystals, which helps to form a uniform, mesoporous structure with controllable pore size. At the same time, the hydroxyl group in malic acid may also affect the surface chemical properties of HAP, improving its biocompatibility. Malic acid is relatively hydrophilic and easily dispersed under hydrothermal conditions, providing a more uniform regulation effect.

[0113] In addition, malic acid is naturally found in many fruits and is a common intermediate product in human metabolism. It has good biocompatibility and safety, making it suitable for the development of biomedical materials. L-malic acid, on the other hand, is a naturally occurring form that does not produce harmful byproducts in the human metabolic pathway, making it even more suitable for the application of biomedical materials.

[0114] This application selects a higher temperature and a shorter reaction time, parameters that significantly affect the addition of malic acid and its performance in the reaction: higher temperatures generally increase the reaction rate because higher temperatures increase the speed of molecular motion and the frequency of collisions, thereby accelerating the reaction process. In this application, selecting higher temperature and shorter reaction time conditions allows the desired reaction conversion rate to be achieved in a shorter time, thereby improving production efficiency. Therefore, at higher temperatures, even with a shorter reaction time, it is possible to achieve reaction efficiencies similar to or better than those achieved with lower temperature and longer reaction times. The thermal decomposition temperature of malic acid is 140℃, and this application uses a temperature of 150℃. The principle for controlling the effective role of malic acid is as follows: ① The hydrothermal reaction is carried out under high pressure, which to some extent inhibits the thermal decomposition of malic acid. Therefore, selecting 150℃, which is slightly higher than the decomposition temperature of malic acid, ensures both the reaction rate and the effectiveness of malic acid. ② Malic acid has a large number of hydroxyl groups, good hydrophilicity, and good dispersibility, providing good material dispersion conditions for rapid reaction. At higher temperatures, the crystal growth rate may be accelerated, but the shorter reaction time may also result in smaller crystals or affect the integrity of the crystals. In this embodiment, the hydroxyapatite particles regulated by malic acid exhibit dense, lamellar stacking on their surface, resulting in a superior surface micro / nano structure. The mesoporous surface enhances drug loading and sustained release; the micron-sized structure makes them ideal for applications such as bone defect filling.

[0115] This invention employs a reaction system in which macroporous spherical structures appear after 30 minutes, and mesoporous microspheres with good crystallinity and uniform morphology are obtained after 5 hours. This invention provides a simple, time-saving, and efficient method for preparing mesoporous hydroxyapatite microspheres with good crystallinity and uniform morphology, which can be applied in various biomedical fields.

[0116] In summary, this invention demonstrates creativity in the selection of morphology modifiers, precise control of reaction conditions, and refinement of post-processing. These factors work together to help obtain mesoporous hydroxyapatite microspheres with specific morphology and size, thus playing an important role in biomedicine and other fields.

[0117] It should be noted that the specific methods by which each module performs operations in the system described in the above embodiments have been described in detail in the embodiments related to the method, and will not be elaborated here.

[0118] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

[0119] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A method for preparing mesoporous hydroxyapatite microspheres regulated by malic acid, characterized in that, Includes the following steps: Step S1: Dissolve the calcium source and phosphorus source in deionized water in sequence, mix them evenly, and prepare a reaction solution; Step S2: Add nitric acid or sodium hydroxide dropwise to the reaction solution to adjust the pH of the solution until it is clear; Step S3: Add malic acid of different concentrations as morphology modifiers to the clear reaction solution, followed by the addition of urea; Step S4: Transfer the prepared solution into the reaction vessel and keep it at the set temperature for 3-5 hours. Step S5: After the reaction is complete, cool the solution to room temperature, centrifuge and dry to obtain hydroxyapatite powder sample; In step S3, the molar ratio of urea to calcium source is 0.56:0.1, and the malic acid is L-malic acid, with a molar ratio of (0.1-0.2):0.1 to the calcium source. In step S4, the set temperature is 120℃-180℃.

2. The method for preparing mesoporous hydroxyapatite microspheres regulated by malic acid according to claim 1, characterized in that, In step S1, calcium nitrate tetrahydrate and disodium hydrogen phosphate dodecahydrate are added sequentially to 50 mL of deionized water to make the calcium source concentration 0.1 mol / L and the calcium-to-phosphorus molar ratio 1.67, thus preparing a reaction solution.

3. The method for preparing mesoporous hydroxyapatite microspheres regulated by malic acid according to claim 1, characterized in that, In step S2, the pH of the solution is adjusted with nitric acid or sodium hydroxide until the solution is clear, and the pH value of the solution is measured to be 2-3.

4. A mesoporous hydroxyapatite microsphere regulated by malic acid, characterized in that, Obtained by the preparation method described in any one of claims 1-3.

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