Hydroxyapatite nanorod with ferromagnetism and preparation method thereof
By adding an appropriate amount of iron and fluorine ions to hydroxyapatite, and preparing nanorods by microwave auxiliary heat method, and then surface modification, the problem of insufficient morphological uniformity and biocompatibility of iron-based hydroxyapatite in the prior art is solved, and ferromagnetic hydroxyapatite nanorods with excellent magnetic properties and good biocompatibility is achieved.
Patent Information
- Application Number
- CN202510383604.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-03-28
- Publication Date
- 2025-06-27
AI Technical Summary
After the existing iron-based hydroxyapatite is incorporated into iron ions, the crystal system is damaged, the morphological uniformity is reduced, and it is difficult to meet the requirements of the field of biomedical medicine.
Hydrophobic ferromagnetic hydroxyapatite nanorods were prepared by adding aqueous solutions of calcium salt, iron salt, phosphate and sodium fluoride to the mixed solution of oleic acid, oleamine and ethanol, and hydrophilic nanorods were obtained by microwave auxiliary heat treatment.
The prepared ferromagnetic hydroxyapatite nanorods have the characteristics of uniform morphology, good biocompatibility and excellent magnetic properties, and are suitable for applications in the field of biomedical medicine.
Smart Images
Figure CN120208173A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of biomaterials, and particularly relates to a ferromagnetic hydroxyapatite nanorod and a preparation method thereof. Background Art
[0002] Iron-based hydroxyapatite has been widely used in the biological field. It can be used for bone tissue repair, as a carrier for drug targeted delivery, and can also play a role in biomedical imaging, antibacterial, and biosensors. With its good biocompatibility, magnetism, and multifunctional characteristics, it has great prospects in the fields of regenerative medicine and personalized treatment. However, when iron ions are incorporated into hydroxyapatite, since the ionic radius of iron ions is 64 pm, which is smaller than that of calcium ions (100 pm), the original hexagonal crystal system of hydroxyapatite is destroyed, and the morphological uniformity of hydroxyapatite will rapidly decline, and even by-products will appear.
[0003] Ideal iron-based hydroxyapatite is required to have a uniform morphology, hydrophilicity that meets the biological and medical fields, and at the same time, it also needs to have good biocompatibility and excellent magnetic properties. Therefore, developing iron-based hydroxyapatite with uniform morphology, good biocompatibility, excellent magnetic properties, and high product purity is of great significance for broadening the application of iron-based hydroxyapatite in the biomedical field. Summary of the Invention
[0004] The technical problem to be solved by the present invention is to provide a preparation method of ferromagnetic hydroxyapatite nanorods in view of the above-mentioned deficiencies of the prior art. The hydroxyapatite nanorods prepared by the method of the present invention have the characteristics of uniform morphology, good biocompatibility, and excellent magnetic properties, and have good application prospects in the biological and medical fields, such as being used in antibacterial or magnetic imaging, or as a bone defect repair material, etc.
[0005] To achieve the above object, on the one hand, the present invention provides a preparation method of ferromagnetic hydroxyapatite nanorods, which includes the following steps:
[0006] S1. Add an aqueous solution of calcium salt, iron salt, phosphate, and sodium fluoride to a mixed solution of oleic acid, oleylamine, and ethanol according to a molar ratio of (Ca + Fe) / P of 1.66, a molar ratio of Fe / (Ca + Fe) of 0.05 - 0.1, and a molar ratio of F / Ca of 0.1 - 0.2, and prepare hydrophobic ferromagnetic hydroxyapatite nanorods through microwave-assisted heating.
[0007] S2. Disperse the obtained hydrophobic ferromagnetic hydroxyapatite nanorods in cyclohexane, and add sodium citrate for surface modification to obtain hydrophilic ferromagnetic hydroxyapatite nanorods.
[0008] As a further preferred technical solution of the present invention, after the reaction in step S1 is completed, it is cooled to room temperature, the precipitate is collected, and it is centrifuged and washed with cyclohexane and absolute ethanol; after the reaction in step S1 is completed, it is centrifuged, washed, and freeze-dried.
[0009] As a further preferred technical solution of the present invention, the calcium salt is Ca(NO3)2, the iron salt is Fe(NO3)3, and the phosphate salt is Na3PO4.
[0010] As a further preferred technical solution of the present invention, the volume ratio of oleic acid, oleic acid, ethanol, and the salt solution is 1:(3 - 4):(9 - 16):(6 - 8), where the salt solution refers to an aqueous solution of calcium salt, iron salt, phosphate salt, and sodium fluoride.
[0011] As a further preferred technical solution of the present invention, the aqueous solution concentrations of calcium salt, iron salt, phosphate salt, and sodium fluoride are all 0.05 - 0.5 mol / L.
[0012] As a further preferred technical solution of the present invention, the microwave-assisted heating method uses a microwave solid-liquid synthesizer, the reaction temperature is 80 - 180 °C, the time is 5 - 30 minutes, and the power is 280 - 800 W.
[0013] As a further preferred technical solution of the present invention, the hydrophobic ferromagnetic hydroxyapatite nanorods are dispersed in cyclohexane at a concentration of 0.01 - 0.1 g / mL.
[0014] As a further preferred technical solution of the present invention, the mass of sodium citrate is 1 - 16 times the mass of the hydrophobic ferromagnetic hydroxyapatite nanorods.
[0015] As a further preferred technical solution of the present invention, surface modification is carried out using an aqueous solution of sodium citrate added, and its concentration is 0.02 - 0.1 g / mL.
[0016] As a further preferred technical solution of the present invention, the reaction temperature for surface modification by adding sodium citrate is 10 - 30 °C, and the reaction time is 4 - 8 hours.
[0017] According to another aspect of the present invention, the present invention also provides a ferromagnetic hydroxyapatite nanorod prepared by the above method, the nanorod size of which is 40 - 50 nm, having hydrophilicity, co-doped with iron and fluoride ions, and having characteristics such as uniform morphology, good biocompatibility, and excellent magnetism.
[0018] Compared with the prior art, the present invention has the following advantages:
[0019] 1. The present invention provides a method for preparing ferromagnetic hydroxyapatite nanorods with a size of 40 - 50 nm. The ferromagnetic hydroxyapatite nanorods have a uniform morphology and, after surface modification, exhibit characteristics such as high hydrophilicity, good biocompatibility, and excellent magnetism, and have good application prospects in the field of biomedicine.
[0020] 2. The ferromagnetic hydroxyapatite nanorods of the present invention contain iron ions and fluoride ions doping. Through simple fluoride ions doping, ion radius compensation is achieved to balance the lattice distortion of hydroxyapatite generated by iron ions.
[0021] 3. The preparation method of the ferromagnetic hydroxyapatite nanorods of the present invention is simple, the raw materials are convenient to purchase, the price is appropriate, the production environment is friendly, and it is easy to scale up, and has good market application value. Brief Description of the Drawings
[0022] The present invention will be further described in detail below in conjunction with the drawings and specific embodiments.
[0023] Figure 1 It is the TEM image of the ferromagnetic hydroxyapatite nanorods of Example 1 of the present invention.
[0024] Figure 2 It is the TEM image of the Fe-doped hydroxyapatite of Comparative Example 1 of the present invention.
[0025] Figure 3 It is the TEM image of the ferromagnetic hydroxyapatite nanorods of Comparative Example 2 of the present invention.
[0026] Figure 4 It is the XRD image of the ferromagnetic hydroxyapatite nanorods of Example 1 of the present invention.
[0027] Figure 5 It is the EDS image of the ferromagnetic hydroxyapatite nanorods of Example 1 of the present invention.
[0028] Figure 6 It is the schematic diagram of the cytotoxicity of the ferromagnetic hydroxyapatite nanorods of Example 1 of the present invention to L929.
[0029] Figure 7 It is the M - H diagram of the ferromagnetic hydroxyapatite nanorods of Example 1 and the Fe-doped hydroxyapatite of Comparative Example 1 of the present invention.
[0030] The realization of the object, functional characteristics, and advantages of the present invention will be further described in conjunction with the embodiments with reference to the drawings. Specific Embodiments
[0031] The following will describe the specific embodiments of the present invention in detail with reference to the drawings. It should be understood that the specific embodiments described herein are only used to illustrate and explain the present invention and are not used to limit the present invention.
[0032] Unless otherwise defined, the technical terms used in the following examples have the same meanings as commonly understood by those skilled in the art to which this invention pertains. The test reagents used in the following examples are all conventional biochemical reagents unless otherwise specified; the experimental methods are all conventional methods unless otherwise specified.
[0033] Example 1:
[0034] This example provides a method for preparing Fe- and F-doped ferromagnetic hydroxyapatite nanorods, which includes the following steps:
[0035] Step (1): Mix 4 mL of oleic acid, 1 mL of oleylamine, and 16 mL of ethanol to obtain a mixed solution. Under stirring conditions, sequentially add 6.3 mL of an aqueous Ca(NO3)2 solution (0.25 mol / L), 0.7 mL of an aqueous Fe(NO3)3 solution (0.25 mol / L), 7 mL of an aqueous Na3PO4 solution (0.15 mol / L), and 0.7 mL of an aqueous NaF solution (0.25 mol / L) to the mixed solution, and stir for 15 minutes to obtain reaction system A; place reaction system A in a computer microwave solid-liquid phase synthesizer for reaction, with a reaction temperature of 120 °C, a reaction time of 15 minutes, and a power of the computer microwave solid-liquid phase synthesizer of 300 W; after the reaction ends and cools to room temperature, collect the precipitate, disperse the precipitate in cyclohexane, add anhydrous ethanol, and centrifuge to obtain hydrophobic ferromagnetic hydroxyapatite nanorods.
[0036] Step (2): Disperse 0.3 g of the above-mentioned hydrophobic ferromagnetic hydroxyapatite nanorods in 5 mL of cyclohexane, add 5 mL of a sodium citrate aqueous solution with a concentration of 0.04 g / mL, and stir and react at room temperature for 6 hours; centrifuge the stirred system, collect the precipitate, wash the precipitate with water and anhydrous ethanol, and freeze-dry to obtain hydrophilic ferromagnetic hydroxyapatite nanorods, denoted as: Specimen 1.
[0037] Comparative Example 1:
[0038] As a control experiment for Example 1, the only difference is that sodium fluoride is omitted from the reaction system.
[0039] Step (1): Mix 4 mL of oleic acid, 1 mL of oleylamine, and 16 mL of ethanol to obtain a mixed solution. Under stirring conditions, sequentially add 6.3 mL of Ca(NO3)2 aqueous solution (0.25 mol / L), 0.7 mL of Fe(NO3)3 aqueous solution (0.25 mol / L), and 7 mL of Na3PO4 aqueous solution (0.15 mol / L) to the mixed solution, and stir for 15 minutes to obtain reaction system B. Place reaction system B in a computerized microwave solid-liquid synthesizer, with a reaction temperature of 120 °C, a reaction time of 15 minutes, and a power of the computerized microwave solid-liquid synthesizer of 300 W. After the reaction is completed and cooled to room temperature, collect the precipitate, disperse the precipitate in cyclohexane, add absolute ethanol, and centrifuge to obtain hydrophobic Fe-doped hydroxyapatite.
[0040] Step (2): Disperse 0.3 g of the above-mentioned hydrophobic Fe-doped hydroxyapatite in 5 mL of cyclohexane, add 5 mL of sodium citrate aqueous solution with a concentration of 0.04 g / mL, and stir and react at room temperature for 6 hours. Centrifuge the stirred system, collect the precipitate, wash the precipitate with water and absolute ethanol, and freeze-dry to obtain hydrophilic Fe-doped hydroxyapatite, denoted as Comparative Sample 1.
[0041] Comparative Example 2:
[0042] As a control experiment for Example 1, the only difference is: changing the dosages of Ca(NO3)2, Fe(NO3)3, and NaF.
[0043] Step (1): Mix 4 mL of oleic acid, 1 mL of oleylamine, and 16 mL of ethanol to obtain a mixed solution. Under stirring conditions, sequentially add 5.6 mL of Ca(NO3)2 aqueous solution (0.25 mol / L), 1.4 mL of Fe(NO3)3 aqueous solution (0.25 mol / L), 7 mL of Na3PO4 aqueous solution (0.15 mol / L), and 0.35 mL of NaF aqueous solution (0.25 mol / L), and stir for 15 minutes to obtain reaction system A. Place reaction system A in a computerized microwave solid-liquid synthesizer, with a reaction temperature of 120 °C, a reaction time of 15 minutes, and a power of the computerized microwave solid-liquid synthesizer of 300 W. After the reaction is completed and cooled to room temperature, collect the precipitate, disperse the precipitate in cyclohexane, add absolute ethanol, and centrifuge to obtain hydrophobic ferromagnetic hydroxyapatite.
[0044] Step (2): Disperse 0.3 g of the above-mentioned hydrophobic ferromagnetic hydroxyapatite in 5 mL of cyclohexane, add 5 mL of sodium citrate aqueous solution with a concentration of 0.04 g / mL, and stir and react at room temperature for 6 hours. Centrifuge the stirred system, collect the precipitate, wash the precipitate with water and absolute ethanol, and freeze-dry to obtain hydrophilic ferromagnetic hydroxyapatite nanorods, denoted as Comparative Sample 2.
[0045] Example 2:
[0046] This comparative example provides a method for preparing hydroxyapatite nanorods, which includes the following steps:
[0047] Step (1): Mix 4 mL of oleic acid, 2 mL of oleylamine, and 16 mL of ethanol to obtain a mixed solution. Under stirring conditions, sequentially add 6.825 mL of an aqueous Ca(NO3)2 solution (0.25 mol / L), 0.175 mL of an aqueous Fe(NO3)3 solution (0.25 mol / L), 7 mL of an aqueous Na3PO4 solution (0.15 mol / L), and 0.7 mL of an aqueous NaF solution (0.25 mol / L) to the mixed solution, and stir for 15 minutes to obtain reaction system C. Place reaction system C in a computer microwave solid-liquid synthesizer, with a reaction temperature of 100 °C, a reaction time of 10 minutes, and a power of 300 W for the computer microwave solid-liquid synthesizer. After the reaction ends and cools to room temperature, collect the precipitate, disperse the precipitate in cyclohexane, add anhydrous ethanol, and centrifuge to obtain hydrophobic ferromagnetic hydroxyapatite nanorods.
[0048] Step (2): Disperse 0.3 g of the above-mentioned hydrophobic ferromagnetic hydroxyapatite nanorods in 5 mL of cyclohexane, add 5 mL of a sodium citrate aqueous solution with a concentration of 1 g / mL, and stir and react at room temperature for 6 hours. Centrifuge the stirred system, collect the precipitate, wash the precipitate with water and anhydrous ethanol, and freeze-dry to obtain hydrophilic ferromagnetic hydroxyapatite nanorods.
[0049] Through comparative tests, the structure and properties of the ferromagnetic hydroxyapatite nanorods in this example are similar to those in Example 1.
[0050] Performance evaluation:
[0051] Figure 1 is the TEM image of the ferromagnetic hydroxyapatite nanorods in Example 1. According to Figure 1 It can be seen that the ferromagnetic hydroxyapatite has a nanorod-like structure, with a uniform morphology and a size of about 40 - 50 nm.
[0052] Figure 2 is the TEM image of the Fe-doped hydroxyapatite in Comparative Example 1. According to Figure 2 It can be seen that the Fe-doped hydroxyapatite has an irregular nanorod-like structure, with a size of about 50 - 80 nm. Compared with the ferromagnetic hydroxyapatite nanorods in Example 1, a uniform morphology cannot be obtained, and there are by-products of nanoparticles.
[0053] Figure 3 is the TEM image of the ferromagnetic hydroxyapatite in Comparative Example 2. According to Figure 3It can be seen that the ferromagnetic hydroxyapatite has an irregular nanorod structure with a size of about 60-80 nm. Compared with the ferromagnetic hydroxyapatite nanorods in Example 1, a uniform morphology cannot be obtained.
[0054] From the comprehensive comparison of Example 1 with Comparative Examples 1 and 2, it can be seen that appropriate fluoride ion doping can effectively achieve ion radius compensation to balance the lattice distortion of hydroxyapatite generated by iron ions, thereby obtaining a nanorod structure with a uniform morphology.
[0055] Figure 4 is the XRD pattern of the ferromagnetic hydroxyapatite nanorods in Example 1. According to Figure 4 It can be seen that the characteristic peaks of Sample 1 are consistent with the hydroxyapatite standard card (PDF#09-0432), indicating that Sample 1 is hydroxyapatite.
[0056] Figure 5 is the EDS detection result of the ferromagnetic hydroxyapatite nanorods in Example 1. According to Figure 4 It can be seen that Sample 1 contains F and Fe elements, indicating that F elements and magnetic Fe elements have been successfully incorporated into Sample 1.
[0057] Figure 6 is the cytotoxicity result of the ferromagnetic hydroxyapatite nanorods in Example 1 and the Fe-doped hydroxyapatite in Comparative Example 1 on normal cells. The specific test method includes: after sterilizing Sample 1, dispersing it in 1640 complete medium (containing 10% serum and 1% double antibody), extracting it at 37°C for 72 h to obtain sample extracts with concentration gradients of 4, 8, 16, 32, 64, 128, 256, and 512 μg / mL. After sealing, store it in a 4°C refrigerator for later use; in a 96-well plate, culture mouse fibroblasts (L929 cells) with 1640 complete medium until the cells adhere. Then, in the experimental group and the control group (without Sample 1), replace the original medium with the sample extract and 1640 complete medium respectively. After continuing to culture the cells for 24 h and 48 h, add 50 μL of MTT to each well, incubate for 3 h, aspirate the liquid in the wells, add 150 μL of DMSO to each well, measure the absorbance at 490 nm, and calculate the cell survival rate through the following formula: Cell survival rate (%) = [(A_sample - A_blank) / (A_control - A_blank)] × 100%, where A_sample is the absorbance of the sample to be measured, A_blank is the absorbance of the culture medium, and A_control is the absorbance of the control group.
[0058] Through Figure 6It can be seen that after culturing L929 cells with the leaching solution of Specimen 1 for 24 h or 48 h, the survival rates of L929 cells cultured with leaching solutions of different concentrations of Specimen 1 are all greater than 100%. Similarly, Comparative Specimen 1 was used as a control group for the comparative experiment. The experimental results show that Comparative Specimen 1 does not have the ability to promote cell proliferation, while Specimen 1 has good cell proliferation promoting ability, which benefits from the incorporation of F element and the uniform rod-like morphology. In order to further prove the beneficial technical effects of the present invention, a large number of experiments were carried out by changing the incorporation amount of F element, and it was found that when the molar ratio of F / Ca in the system is less than 0.1 (such as Comparative Example 2), although the F element is successfully incorporated and has biocompatibility, the ability to promote cell proliferation is almost absent, which is related to the incorporation amount of F element and the morphology and size of apatite; when the molar ratio of F / Ca in the system is greater than 0.2, the biocompatibility will deteriorate significantly, which is caused by the excessive incorporation of F element. If applied to the human body, the excessive fluoride ions in the human body will damage teeth to form dental fluorosis, cause bone lesions to trigger skeletal fluorosis, and will also affect organs such as the kidneys and liver, as well as the nervous system and endocrine system, interfering with the normal functions of the human body.
[0059] Figure 7 M-H diagrams of ferromagnetic hydroxyapatite nanorods of Example 1 and Fe-doped hydroxyapatite of Comparative Example 1. According to Figure 7 It can be seen that Specimen 1 passes through the origin of the coordinate axis, showing superparamagnetic properties, and has a higher magnetic moment compared with the comparative specimen, indicating that Specimen 1 has more excellent magnetic properties.
[0060] Although the specific embodiments of the present invention have been described above, those skilled in the art should understand that these are only examples, and various changes or modifications can be made to this embodiment without departing from the principle and essence of the present invention. The protection scope of the present invention is only defined by the appended claims.
Claims
1. A method for preparing ferromagnetic hydroxyapatite nanorods, characterized in that: The following steps are involved: S1. Into a mixed solution of oleic acid, oleylamine and ethanol, an aqueous solution of calcium salt, iron salt, phosphate and sodium fluoride is added at a molar ratio of (Ca+Fe) / P of 1.66, a molar ratio of Fe / (Ca+Fe) of 0.05-0.1, and a molar ratio of F / Ca of 0.1-0.2, and hydrophobic ferromagnetic hydroxyapatite nanorods are prepared by microwave assisted heating; S2. Dispersing the obtained hydrophobic ferromagnetic hydroxyapatite nanorods in cyclohexane, adding sodium citrate for surface modification, and obtaining hydrophilic ferromagnetic hydroxyapatite nanorods.
2. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: Calcium salt is Ca(NO3)2, iron salt is Fe(NO3)3, and phosphate is Na3PO4.
3. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: The volume ratio of oleic acid, oleic acid, ethanol and salt solution is 1:(3-4):(9-16):(6-8), wherein the salt solution refers to an aqueous solution of calcium salt, iron salt, phosphate and sodium fluoride.
4. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: The concentrations of the aqueous solutions of calcium salt, iron salt, phosphate and sodium fluoride are all 0.05-0.5 mol / L.
5. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: The microwave-assisted heating method uses a microwave solid-liquid phase synthesizer with a reaction temperature of 80-180°C, a reaction time of 5-30 minutes, and a power of 280-800W.
6. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: Hydrophobic ferromagnetic hydroxyapatite nanorods are dispersed in cyclohexane at a concentration of 0.01-0.1 g / mL.
7. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: The mass of sodium citrate is 1-16 times the mass of the hydrophobic ferromagnetic hydroxyapatite nanorods.
8. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: The surface modification is performed by adding an aqueous solution of sodium citrate, the concentration of which is 0.02-0.1 g / mL.
9. The method for preparing ferromagnetic hydroxyapatite nanorods according to claim 1, characterized in that: The reaction temperature for adding sodium citrate for surface modification is 10-30°C and the reaction time is 4-8 hours.
10. Hydroxyapatite nanorods having ferromagnetism prepared by the method according to any one of claims 1 to 9.