Composite bioglass as well as preparation method and application thereof

Abstract

The invention relates to the field of bioglass, in particular to composite bioglass as well as a preparation method and application thereof. The rhBMP-2, the 45S5 bioactive glass doped with the antibacterial ions and the polyethylene glycol-lactide-glycolide polymer are compounded to prepare the slow-release antibacterial bioglass, the rhBMP-2 and the 45S5 bioactive glass doped with the antibacterial ions have a synergistic interaction effect, and compared with homologous growth factors, the slow-release antibacterial bioglass can play a better synergistic effect with the antibacterial ions, so that the slow-release antibacterial bioglass has the advantages that the slow-release antibacterial bioglass is prepared, and the antibacterial activity of the slow-release antibacterial bioglass is improved. Therefore, the antibacterial and osteogenic hydrogel has higher antibacterial property and osteogenic property, and has a longer slow-release effect in a polyethylene glycol-lactide-glycolide polymer. The composite bioglass provided by the invention overcomes pain points generated by oral bone defect treatment in the prior art, improves the treatment effect, reduces the use amount of antibiotics, and reduces the infection risk.

Description

Technical Field

[0001] The present invention relates to the field of bioglass, in particular to composite bioglass and a preparation method and application thereof. Background Art

[0002] Hench invented bioglass (BG) in 1970, and it has been in use for over 50 years. With the continuous advancement of technology and in-depth research, bioglass has demonstrated strong vitality. For example, the addition of antimicrobial ions can enhance its antibacterial properties; various drugs can be added to serve as sustained-release carriers; special treatments can be used to create stronger bioglass scaffolds; and bioglass can be used as coatings for attachment to other medical devices. These advancements have broadened the application scenarios and usage of bioglass.

[0003] In the oral field, bioglass plays an even more important role. It is used in dental implants, root canal treatment, alveolar ridge repair, and even tooth desensitization treatment. The oral environment contains a variety of bacterial flora, and daily cleaning can maintain the balance of the flora and reduce oral problems such as ulcers and inflammation. However, patients who have undergone oral surgery have limited daily cleaning, and their oral flora is unbalanced, which makes them more prone to additional oral problems, especially in the surgical area, which is often supplemented with antibiotic treatment. However, long-term use of antibiotics can lead to problems such as drug resistance. Therefore, there is an urgent need to provide a new and efficient slow-release antibacterial bioglass to reduce the use of antibiotics, accelerate wound healing, reduce the risk of infection at the wound, and alleviate patient pain. Summary of the Invention

[0004] In view of this, the technical problem to be solved by the present invention is a composite bioglass and its preparation method and application. The composite bioglass provided by the present invention has excellent biological activity, antibacterial and osteogenic activity, good sustained-release effect, and is suitable for oral environment.

[0005] The present invention provides a composite bioglass, which comprises: a polyethylene glycol-lactide-glycolide polymer gel, rhBMP-2 doped in the polyethylene glycol-lactide-glycolide polymer gel, and 45S5 bioactive glass doped with antibacterial ions;

[0006] The antibacterial ions are selected from one or more of Ag, Zn, Cu and Ce.

[0007] Specifically, the composite bioactive glass of the present invention is cross-linked by rhBMP-2, polyethylene glycol-polylactide-glycolide polymer gel, and 45S5 bioactive glass doped with antibacterial ions. Among them, the rhBMP-2 is a growth factor; the polyethylene glycol-polylactide-glycolide polymer gel is a hydrogel component; the 45S5 bioactive glass doped with antibacterial ions is a bioactive glass containing antibacterial components, which is synthesized by the polymer template method. Among them, the mass ratio of the rhBMP-2, polyethylene glycol-polylactide-glycolide polymer, and 45S5 bioactive glass doped with antibacterial ions is (0.001~0.003):(2~3):(0.02~0.03), preferably (0.001~0.002):(2.3~2.7):(0.023~0.027), and more preferably 0.0018:2.5:0.025.

[0008] The doping amount of antibacterial ions in the 45S5 bioactive glass doped with antibacterial ions of the present invention is (5~15) wt%, preferably 8 wt%. The antibacterial ions of the present invention are preferably selected from Zn, and the 45S5 bioactive glass doped with Zn has better protein adsorption ability.

[0009] The polyethylene glycol-polylactide-glycolide polymer gel of the present invention is polymerized from polyethylene glycol, lactide, and glycolide according to the mass ratio of (18~22):(34~38):(8~12), preferably polymerized according to the mass ratio of (19~21):(35~37):(9~11); specifically, it is polymerized from polyethylene glycol, lactide, and glycolide to obtain a polyethylene glycol-polylactide-glycolide polymer sol and then solidified into a polyethylene glycol-polylactide-glycolide polymer gel. The polyethylene glycol-polylactide-glycolide polymer gel of the present invention, as a hydrogel component, makes the release curve of the composite bioactive glass more in line with the recovery law. The molecular weight of the polyethylene glycol of the present invention is 2000~6000. In some embodiments of the present invention, the polyethylene glycol is PEG3000.

[0010] In the composite bioactive glass provided by the present invention, the rhBMP-2 can have a synergistic effect with the 45S5 bioactive glass doped with antibacterial ions, and can exert better synergy with antibacterial ions compared with homologous growth factors, so as to have higher antibacterial and osteogenic properties, and at the same time have a longer sustained release effect in the polyethylene glycol-polylactide-glycolide polymer.

[0011] The present invention also provides a preparation method of the composite bioactive glass according to any one of the above technical solutions, including the following steps:

[0012] The 45S5 bioactive glass doped with antibacterial ions and rhBMP-2 are coagulated in a polyethylene glycol-polylactide-glycolide polymer sol to obtain the composite bioactive glass;

[0013] The antibacterial ions are selected from one or more of Ag, Zn, Cu, and Ce.

[0014] Specifically, in the present invention, the 45S5 bioactive glass doped with antibacterial ions and rhBMP-2 are placed in a polyethylene glycol-polylactide-glycolide polymer sol at room temperature, evacuated for (8 - 12) min, and left standing at (35 - 40) °C for sol coagulation to obtain the composite bioactive glass.

[0015] In some embodiments of the present invention, the polyethylene glycol-polylactide-glycolide polymer is dissolved in physiological saline to prepare a (20 - 30) wt% polyethylene glycol-polylactide-glycolide polymer sol; then rhBMP-2 is dispersed in the polyethylene glycol-polylactide-glycolide polymer sol at room temperature to obtain a polyethylene glycol-polylactide-glycolide polymer sol containing rhBMP-2; the 45S5 bioactive glass doped with antibacterial ions is completely immersed in the polyethylene glycol-polylactide-glycolide polymer sol containing rhBMP-2 at room temperature, evacuated for (8 - 12) min, and left standing at (35 - 40) °C for sol coagulation to obtain the composite bioactive glass.

[0016] The mass ratio of rhBMP-2, the polyethylene glycol-polylactide-glycolide polymer sol, and the 45S5 bioactive glass doped with antibacterial ions in the present invention is 1.8 mg : 2.5 g : 0.025 g. The doping amount of the antibacterial ions in the 45S5 bioactive glass doped with antibacterial ions in the present invention is (5 - 15) wt%, preferably 8 wt%. The antibacterial ions in the 45S5 bioactive glass doped with antibacterial ions in the present invention are preferably selected from Zn. The room temperature in the present invention refers to the temperature in the natural state indoors, without relying on external heating or cooling equipment intervention, and is preferably 25 °C.

[0017] The poly(ethylene glycol)-lactide-glycolide polymer sol described in the present invention is obtained by polymerizing poly(ethylene glycol), lactide, and glycolide according to a mass ratio of (18-22):(34-38):(8-12). Specifically, the poly(ethylene glycol)-lactide-glycolide polymer described in the present invention is prepared by the following steps: under an organotin catalyst, poly(ethylene glycol), lactide, and glycolide are subjected to a polymerization reaction to obtain the poly(ethylene glycol)-lactide-glycolide polymer sol. More specifically, poly(ethylene glycol) is placed in a vacuum atmosphere for (2-4) h and argon is replaced once per hour, while maintaining stirring and heating at (120-130)°C at (60-80) r / min; after natural cooling, lactide and glycolide are added to the poly(ethylene glycol) for dissolution, and then an organotin catalyst solution is added continuously. After removing the solvent, a polymerization reaction is carried out to obtain the poly(ethylene glycol)-lactide-glycolide polymer sol.

[0018] The molecular weight of the poly(ethylene glycol) described in the present invention is 2000-6000. In certain embodiments of the present invention, the poly(ethylene glycol) is PEG3000. The organotin catalyst described in the present invention is selected from at least one of stannous octoate, dibutyltin dilaurate, tributyltin oxide, and dibutyltin oxide. The temperature of the polymerization reaction described in the present invention is (140-160)°C, and the time of the polymerization reaction is (10-14) h.

[0019] The present invention also provides the use of the composite bioglass described in any of the above technical solutions or the composite glass obtained by the preparation method described in any of the above technical solutions as an oral bone repair material. The composite bioglass provided by the present invention has higher antibacterial properties and osteogenicity, and at the same time has a longer and more regular release curve, and is very suitable for use as an oral bone repair material in the oral environment.

[0020] The present invention provides a composite bioglass, a preparation method thereof, and an application thereof. The present invention uses rhBMP-2, 45S5 bioactive glass doped with antibacterial ions, and poly(ethylene glycol)-lactide-glycolide polymer to prepare a sustained-release antibacterial bioglass. The rhBMP-2 can have a synergistic effect with the 45S5 bioactive glass doped with antibacterial ions, and can exert better synergy with antibacterial ions compared with homologous growth factors, thereby having higher antibacterial properties and osteogenicity, and at the same time having a longer sustained-release effect in the poly(ethylene glycol)-lactide-glycolide polymer. The composite bioglass provided by the present invention overcomes the pain points of the existing technology for the treatment of oral bone defects, improves the treatment effect, reduces the use amount of antibiotics, and reduces the risk of infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 It is a bioactivity result diagram of a bioactive glass doped with silver ions;

[0022] Figure 2 Bioactivity result graph of bioactive glass doped with zinc ions;

[0023] Figure 3 Bioactivity result graph of bioactive glass doped with copper ions;

[0024] Figure 4 Bioactivity result graph of bioactive glass doped with cerium ions;

[0025] Figure 5 Bioactivity result graph of rhBMP-2 / BG described in Example 1 of the present invention;

[0026] Figure 6 Bioactivity result graph of the polymer described in Comparative Example 1 of the present invention;

[0027] Figure 7 Bioactivity result graph of rhBMP-4 / BG described in Comparative Example 2 of the present invention. Detailed implementation manners

[0028] The present invention discloses a composite bioactive glass and its preparation method and application. Those skilled in the art can draw on the content of this article and appropriately improve the process parameters to achieve. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art, and they are all regarded as included in the present invention. The methods and applications of the present invention have been described through preferred embodiments, and those skilled in the art can obviously make changes or appropriate modifications and combinations to the methods and applications in this article without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0029] The following further elaborates on the present invention in conjunction with embodiments:

[0030] Example 1

[0031] Preferred antibacterial bioactive glass:

[0032] Step 1: Select silver (Ag), zinc (Zn), copper (Cu), and cerium (Ce) as doping antibacterial ions, and use the polymer template method to synthesize 45S5 bioactive glass doped with different antibacterial ions, with an ion doping amount of 8 wt% for all; among them, the bioactive glass doped with silver ions is denoted as sample 1, the bioactive glass doped with zinc ions is denoted as sample 2, the bioactive glass doped with copper ions is denoted as sample 3, and the bioactive glass doped with cerium ions is denoted as sample 4;

[0033] Step 2: Perform in vitro culture on the bioactive glass doped with different antibacterial ions obtained in Step 1, and then test its bioactivity and antibacterial properties to optimize an antibacterial bioactive glass. Among them, the bioactivity results of the bioactive glass doped with different antibacterial ions are as Figures 1 to 4 shown Figure 1Biological activity result diagram of silver ion-doped bioactive glass Figure 2 Biological activity result diagram of zinc ion-doped bioactive glass Figure 3 Biological activity result diagram of copper ion-doped bioactive glass Figure 4 Biological activity result diagram of cerium ion-doped bioactive glass. The antibacterial properties of bioactive glasses doped with different antibacterial ions are shown in Table 1:

[0034] Table 1

[0035]

[0036] Example 2

[0037] Preparation of composite human bone morphogenetic protein-2 hydrogel antibacterial bioactive glass (rhBMP-2 / BG):

[0038] (1) Hydrogel preparation

[0039] Place a three-necked flask containing 20.25 g of polyethylene glycol (PEG3000) in an oil bath, set the temperature to 125 °C and the stirring speed to 70 r / min. Evacuate for 3 h and replace with argon, once per hour. After natural cooling, add 36.8 g of lactide and 9.9 g of glycolide respectively. After complete dissolution, add a toluene solution of stannous octoate, evacuate to remove toluene, set the oil bath temperature to 150 °C, and react for 12 h. Evacuate to remove residual raw materials and small molecule products, wash with hot water, and freeze-dry to obtain a polymer, and the obtained polymer is the hydrogel.

[0040] (2) rhBMP-2 / BG preparation

[0041] Dissolve 2.5 g of the polymer prepared in (1) above in 7.5 mL of physiological saline to prepare a 25 wt% sol. At room temperature, add 1.8 mg of rhBMP-2 dry powder to a test tube containing 10 mL of 25 wt% hydrogel, and shake with an oscillator for 1 min to fully disperse rhBMP-2 in the sol. At room temperature, completely immerse 0.025 g of the antibacterial bioactive glass doped with 8 wt% zinc ions obtained in Example 1 in the sol, evacuate for 10 min, and place it in an incubator at 37 °C. Wait for the sol to solidify to obtain rhBMP-2 / BG.

[0042] (3) rhBMP-2 / BG performance testing

[0043] Perform antibacterial testing, biological activity testing, sustained-release effect testing, and osteogenic effect (cell expression) on the rhBMP-2 / BG obtained in (2) above, and the methods are as follows:

[0044] ① Antibacterial testing: Antibacterial ring;

[0045] ②Bioactivity: Refer to "YY / T 0964-2014 Surgical Implants - Bioactive Glass and Glass-Ceramic Materials" for implementation;

[0046] ③Sustained-release effect: Immerse the samples in SBF solution, replace the SBF solution at regular intervals, and measure the ion content in the solution;

[0047] ④Osteogenic effect (cell expression): Seed bone marrow mesenchymal stem cells (BMSCs) on the bottom of a 24-well plate, and embed 5 mg of the scaffold sample in the upper part of the Transwell system. Immerse both the cells and the microspheres in osteogenic induction medium (50 μg / mL ascorbic acid, 10 nM dexamethasone, and 10 mM β-glycerophosphate), and replace the medium every other day. After culturing for 7 days, quantitatively evaluate the expression of osteogenic genes, including Runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), ALP, and Osterix. Use TRIzol reagent (Invitrogen, USA) for cell lysis and total RNA extraction. Subsequently, use a commercial kit (Takara, Japan) to reverse transcribe the extracted RNA into complementary DNA (cDNA) and perform RT-PCR quantitative analysis. The specific primer sequences used are listed in detail in Table 2.

[0048] Table 2

[0049]

[0050] The antibacterial action time of rhBMP-2 / BG obtained in (2) above is 22 d, and the specific test results are shown in Table 3; the action time of the sustained-release effect is 22 d; the test results of the in vitro osteogenic effect are shown in Table 4; the test results of the bioactivity are as Figure 5 shown, Figure 5 This is the bioactivity result graph of rhBMP-2 / BG described in Example 1 of the present invention.

[0051] Table 3

[0052]

[0053] Table 4

[0054]

[0055] Comparative Example 1

[0056] Place a three-necked flask containing 20.25 g of polyethylene glycol in an oil bath, set the temperature to 125 °C and the stirring speed to 70 r / min. Evacuate for 3 h and displace with argon, once per hour. After natural cooling, add 36.8 g of lactide and 9.9 g of glycolide respectively. After complete dissolution, add a toluene solution of stannous octoate, evacuate to remove toluene, set the oil bath temperature to 150 °C, and react for 12 h. Evacuate to remove residual raw materials and small molecule products, wash with hot water, and freeze-dry to obtain a polymer, and the obtained polymer is the hydrogel.

[0057] The polymers obtained above were subjected to antibacterial tests, bioactivity tests, sustained release effect tests, and osteogenic effects (cell expression), and the methods were the same as those in Example 1 and will not be elaborated here.

[0058] The antibacterial action time of the polymers obtained above was 18 d, and the specific test results are shown in Table 5; the action time of the sustained release effect was 18 d; the test results of the in vitro osteogenic effect are shown in Table 6; the test results of the bioactivity are as Figure 6 shown, Figure 6 This is the bioactivity result diagram of the polymer described in Comparative Example 1 of the present invention.

[0059] Table 5

[0060]

[0061] Table 6

[0062]

[0063] Comparative Example 2

[0064] Place a three-necked flask containing 20.25 g of polyethylene glycol in an oil bath, set the temperature to 125 °C and the stirring speed to 70 r / min. Evacuate for 3 h and displace with argon, once per hour. After natural cooling, add 36.8 g of lactide and 9.9 g of glycolide respectively. After complete dissolution, add a toluene solution of stannous octoate, evacuate to remove toluene, set the oil bath temperature to 150 °C, and react for 12 h. Evacuate to remove residual raw materials and small molecule products, wash with hot water, and freeze-dry to obtain a polymer, and the obtained polymer is the hydrogel.

[0065] Dissolve 2.5 g of the polymer prepared above in 7.5 mL of physiological saline to prepare a 25 wt% sol. At room temperature, add 1.8 mg of rhBMP-4 dry powder to a test tube containing 10 mL of 25 wt% hydrogel, and shake with an oscillator for 1 min to fully disperse rhBMP-4 in the sol. Immerse the antibacterial bioactive glass completely in the sol at room temperature, evacuate for 10 min, and place it in an incubator at 37 °C. Wait for the sol to solidify to obtain rhBMP-4 / BG.

[0066] The antibacterial test, bioactivity test, sustained-release effect test, and osteogenic effect (cell expression) of the obtained rhBMP-4 / BG were carried out in the same manner as in Example 1 and will not be elaborated here.

[0067] The antibacterial action time of the obtained rhBMP-4 / BG was 21 d, and the specific test results are shown in Table 7; the action time of the sustained-release effect was 21 d; the test results of the in vitro osteogenic effect are shown in Table 8; the test results of the bioactivity are as Figure 7 shown Figure 7 This is the bioactivity result diagram of the rhBMP-4 / BG described in Comparative Example 2 of the present invention.

[0068] Table 7

[0069]

[0070] Table 8

[0071]

[0072] As described above, the above are only the preferred specific embodiments of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art within the technical scope disclosed by the present invention, according to the technical solution and inventive concept of the present invention, making equivalent substitutions or changes should be covered within the protection scope of the present invention.

Claims

1. A composite bioglass, characterized in that, It includes: Polyethylene glycol-lactide-glycolide polymer gel, rhBMP-2 doped in the polyethylene glycol-lactide-glycolide polymer gel, and 45S5 bioactive glass doped with antibacterial ions; The antibacterial ions are selected from one or more of Ag, Zn, Cu and Ce.

2. The composite bioglass according to claim 1, wherein The mass ratio of the rhBMP-2, the polyethylene glycol-lactide-glycolide polymer gel and the 45S5 bioactive glass doped with antibacterial ions is (0.001-0.003): (2-3): (0.02-0.03).

3. The composite bioactive glass according to claim 1, characterized in that, The doping amount of the antibacterial ions is (5-15) wt%.

4. The composite bioglass according to claim 1, characterized in that, The polyethylene glycol-lactide-glycolide polymer gel is obtained by polymerizing polyethylene glycol, lactide and glycolide in a mass ratio of (18-22): (34-38): (8-12).

5. A method for preparing composite bioglass, characterized in that: The following steps are involved: solidifying 45S5 bioactive glass doped with antibacterial ions and rhBMP-2 in a polyethylene glycol-lactide-glycolide polymer sol to obtain the composite bioglass; The antibacterial ions are selected from one or more of Ag, Zn, Cu and Ce.

6. The preparation method according to claim 5, characterized in that, The mass ratio of the rhBMP-2, the polyethylene glycol-lactide-glycolide polymer sol and the 45S5 bioactive glass doped with antibacterial ions is (0.001-0.003): (2-3): (0.02-0.03).

7. The preparation method according to claim 5, wherein The doping amount of the antibacterial ions is (5-15) wt%.

8. The preparation method according to claim 5, characterized in that, The polyethylene glycol-lactide-glycolide polymer sol is obtained by polymerizing polyethylene glycol, lactide and glycolide in a mass ratio of (18-22): (34-38): (8-12).

9. The preparation method according to claim 5, characterized in that, The polyethylene glycol-lactide-glycolide polymer sol is prepared by the following steps: Polyethylene glycol, lactide and glycolide are polymerized in the presence of an organic tin catalyst to obtain the polyethylene glycol-lactide-glycolide polymer sol.

10. Use of the composite bioglass according to any one of claims 1 to 4 or the composite glass obtained by the preparation method according to any one of claims 5 to 9 as an oral bone repair material.

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