CN106750450B - Preparation method of cross-linked adhesive biomimetic coating containing epoxy phosphorylcholine polymer and dopamine - Google Patents

Abstract

The invention discloses a preparation method of a biomimetic coating containing epoxy phosphorylcholine polymer and dopamine. Acryloyloxyethylphosphorylcholine monomer is synthesized by simple solution radical polymerization to synthesize epoxy-containing phosphorylcholine polymer, and dissolve it with dopamine in polar solvent, and coat it on the surface that needs to be modified. The surface of the material is dried and then placed in a pH=8.5 Tris-HCl aqueous solution for heating treatment, so that the amino groups in the dopamine and the surface of the chitosan film react with the epoxy in the phosphorylcholine polymer while the dopamine is cross-linked and adhered. Relying on the anchoring of epoxy and amino groups on the surface of chitosan film in the phosphorylcholine polymer and the adhesion of dopamine to fix the phosphorylcholine group on the surface of the chitosan film, the cell-like outer film can be prepared. Structured cross-linked adhesive biomimetic coating.

Description

Preparation of Adhesive Biomimetic Coatings Containing Epoxy Phosphorylcholine Polymers and Dopamine Cross-Linking method

technical field

The invention relates to the technical field of material surface science and biomedical polymer materials, in particular to a preparation method of a biomimetic coating containing epoxy phosphorylcholine polymer and dopamine cross-linked and adhered.

Background technique

Chitosan has the advantages of degradability, antibacterial, non-toxic, non-irritating, pH-responsive (Carbohydrate Polymers 2010, 79: 724-730), and has been widely used in biomedicine and other fields. More and more studies have shown that chitosan and its derivative materials can be used for blood purification. Amino groups on chitosan molecules contribute to the adsorption of various toxins in blood, and can be used in blood perfusion materials (Journal of Chemistry of Higher Education 2002, 23: 75-77; Journal of Microencapsulation 1993, 10: 475-486). Chitosan membranes have high dialysis rate, selectivity and strength, and can be used as hemodialysis materials (Journal of Applied Polymer Science 1992, 46: 255-261; 263-269). Although chitosan and its derivatives have many advantages as blood purification materials, there are also problems such as protein adsorption and platelet adhesion, which eventually lead to coagulation and thrombus formation. Therefore, the blood compatibility of chitosan and its derivatives is improved. It is imminent (Applied Surface Science 2005, 241: 485-492; Biomaterials 2002, 23: 2561-2568; Biomaterials 2003, 24: 3213-3220).

Phosphorylcholine (PC) is the hydrophilic end group of lecithin, the basic unit of cell membrane, and the outer functional group in the outer membrane of the cell. And hydrophilic properties, the surface of this structure and composition interacts with the physiological environment not only does not adsorb and deposit proteins, but also does not trigger platelet activation, lead to coagulation and other adverse reactions, and has good biocompatibility. Studies in recent years have shown that the use of phosphorylcholine groups and their polymers to construct a cell-like outer membrane structure on the surface of the material can significantly improve the blood compatibility of the material.

In recent years, the approach of grafting small molecules of phosphorylcholine (Carbohydrate Polymers 2007, 70: 82-88; Biomacromolecules 2007, 8: 3169-3176; Biomacromolecules 2006, 7: 3151-3156; Journal of Applied Polymer Science 2003, 88: 489-493; Polymer International 2003, 52: 81-85; Journal of biomaterials science, Polymer edition 2002, 13: 501-510; Colloids and Surfaces B: Biointerfaces 2009, 71: 268-274) modified chitosan, The blood compatibility of chitosan is significantly improved. However, in these methods, the density of phosphorylcholine groups on the surface of the material is often not high, which limits its application in the field of biomedical material modification and further improvement of blood compatibility.

To this end, the polyanion of methacrylic acid-methacryloyloxyethylphosphorylcholine binary copolymer (PMA) containing phosphorylcholine groups was electrostatically separated with chitosan (polycation) layer by layer. After assembly, a coated surface with a structure mimicking the outer membrane of the cell was obtained (Colloids and Surfaces B: Biointerfaces 2011, 85: 48-55). The experimental results of protein adsorption and platelet adhesion showed that the hemocompatibility of the modified surface was significantly improved. In view of the advantages of this modification method, it will certainly provide technical support for improving the blood compatibility of biomedical materials. However, the polymer coating of the cell-like outer membrane structure combined on the surface of the implanted device by physical adsorption will inevitably dissolve and fall off in the complex environment in vivo. To this end, Lewis and Xu Jianping and others (Biomaterials 2001, 22: 99-111; Biomaterials 2004, 25: 3099-3108; European Polymer Journal 2004, 40: 291-298), respectively, used trimethoxysilicon groups and phosphoryl The polymer coatings of choline groups were investigated. The results show that the trimethoxysilicon group on the polymer molecular chain in the coating will be hydrolyzed and cross-linked when it encounters water, and it can also form a covalent bond with the active group on the surface of the substrate, so that the phosphorylcholine polymer The stability of the coating is significantly improved. It can be seen that the crosslinking between polymers and their reaction with functional groups on the surface of the substrate are the key factors to improve the stability of phosphorylcholine polymer coatings.

However, the cross-linkable groups of the polymer are easily hydrolyzed and cross-linked during the synthesis process, which makes the synthesis process conditions too harsh and difficult to store, resulting in limited application scope.

SUMMARY OF THE INVENTION

In order to solve the above problems, the present invention provides a preparation method of a biomimetic coating containing epoxy phosphoryl choline polymer and dopamine cross-linked and adhered.

To achieve the above object, the technical scheme adopted in the present invention is:

The preparation method of the biomimetic coating containing epoxy phosphorylcholine polymer and dopamine comprises the following steps:

S1, under the protection of nitrogen, carry out radical polymerization reaction of vinyl monomer containing epoxy and vinyl monomer containing phosphoryl choline under the action of initiator to obtain epoxy-containing phosphoryl choline polymer ;

S2, dissolving dopamine and the epoxy-containing phosphorylcholine polymer obtained in step S1 in a polar solvent to obtain a mixed solution, then coating the mixed solution on the surface of the material to be modified, and drying;

S3. The material to be modified after drying in step S2 is placed in a pH=8.5 Tris-HCl aqueous solution, and heat-treated at 45°C to 95°C for 2h to 12h to obtain a cross-linking structure with a cell-like outer membrane structure. Adhesive biomimetic coating.

Preferably, the epoxy-containing vinyl monomer in step S1 is glycidyl methacrylate, and the phosphorylcholine-containing vinyl monomer is 2-methacryloyloxyethylphosphorylcholine .

Preferably, the molar ratio of the phosphorylcholine-containing vinyl monomer to the epoxy-containing vinyl monomer in step S1 is about 9.5:0.5-7:3.

Preferably, the solvent of the radical polymerization reaction in step S1 is obtained by mixing methanol and tetrahydrofuran in a volume ratio of 4:1, the initiator is azobisisobutyronitrile, the reaction temperature is 70°C, the reaction time is 24h, and the Dialysis was performed on a dialysis bag with a molecular weight cut-off of 7000.

Preferably, the concentration of dopamine in the mixed solution in step S2 is 0.1 mg/mL to 0.5 mg/mL, and the concentration of the aldehyde group-containing phosphorylcholine polymer is 0.2 mg/mL to 5 mg/mL.

Preferably, the polar solvent in step S2 is methanol or ethanol.

Preferably, the material to be modified in step S2 is chitosan.

The present invention has the following beneficial effects:

1. In the present invention, the glycidyl methacrylate monomer containing epoxy and the methacryloyloxyethyl phosphorylcholine monomer containing phosphorylcholine hydrophilic group are polymerized by simple solution radicals, A phosphorylcholine polymer containing epoxy was synthesized, dissolved in a polar solvent with dopamine, coated on the surface of the material to be modified, dried and then placed in a pH=8.5 Tris-HCl aqueous solution for heat treatment. Dopamine crosslinks and adheres while the amino group in dopamine and the amino group on the surface of chitosan film react with the epoxy in the phosphorylcholine polymer, relying on the anchor of the reaction between the epoxy in the phosphorylcholine polymer and the amino group on the surface of the chitosan film The phosphorylcholine groups were fixed on the surface of the chitosan film, and the cross-linked adhesion biomimetic coating with the structure of the outer cell membrane could be prepared.

2. The preparation method of the cell-imitation outer layer membrane structure coating of the present invention is simple and convenient to operate, and a new approach is provided for obtaining a stable coating surface with the cell outer layer membrane structure.

3. The imitative cell outer membrane structure coating of the present invention will have broad application prospects in the fields of blood purification, implanted materials in vivo, tissue engineering, sustained drug release and biosensors.

Description of drawings

Fig. 1 is the dynamic contact angle of the chitosan film of the present invention and the modified chitosan film.

Fig. 2 is the surface fine energy spectrogram of the chitosan film of the present invention and the modified chitosan film.

Detailed ways

In order to make the objects and advantages of the present invention more clear, the present invention will be further described in detail below with reference to the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention, but not to limit the present invention.

Example 1

Weigh 16mmol of 2-methacryloyloxyethylphosphorylcholine and 4mmol of glycidyl methacrylate, take 0.1mmol of azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The hydrogen NMR of the polymer was tested with a 400MHz NMR spectrometer with D ₂ O as the solvent. There is no peak at 5-7 ppm, indicating that there is no residual monomer in the obtained copolymer, and the polymer is successfully synthesized. The characteristic peak of -N ⁺ (CH ₃ ) ₃ is at 3.28 ppm, and the characteristic peak at 0.9-2.2 ppm is The polymer composition was calculated from the peaks of methylene and side chain methyl groups on the main chain, and it was known that the polymer composition was basically consistent with the feeding ratio.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 1 mg/mL, and then 0.5 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of the epoxy-containing phosphorylcholine polymer and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution for 5 hours at 80 °C, so that the amino groups in dopamine and the surface of the chitosan film were mixed with phosphorus. Dopamine cross-links at the same time as the epoxy reaction in the acylcholine polymer, and the phosphorylcholine group is fixed on the surface of the chitosan film by the anchoring of the epoxy in the phosphorylcholine polymer and the amino group reaction on the surface of the chitosan film and the adhesion of dopamine. On the surface of the chitosan film, a cross-linked adhesive biomimetic coating with a cell-like outer membrane structure can be prepared.

As shown in Figure 1, the advancing angle and the receding angle of the coated chitosan film are reduced compared with the uncoated chitosan film in this example. , this is because the phosphorylcholine polymer with good hydrophilicity adheres and fixes the phosphorylcholine group on the surface of the chitosan film through the reaction of epoxy with the amino group on the surface of chitosan and the reaction of the amino group in dopamine. , to obtain a surface with a structure that mimics the outer membrane of the cell, so that its hydrophilicity is significantly improved, and the advancing and receding angles are significantly reduced. In addition, the presence of dopamine increased the hydrophilicity of modified chitosan, and both the advancing and receding angles decreased. This indicated that dopamine increased the content of phosphorylcholine groups on the surface of chitosan film.

As shown in Figure 2, compared with the uncoated chitosan film, the modified chitosan film has phosphorylcholine groups on the surface of the coated chitosan film. The characteristic absorption peaks of N and P indicate that the hydrophilic phosphorylcholine groups are immobilized on the surface of the chitosan film. Dopamine crosslinks while the amino group in dopamine and the amino group on the surface of chitosan film react with the epoxy in the phosphorylcholine polymer. The adhesion of the phosphorylcholine groups to the surface of the chitosan film can prepare a surface with a structure that mimics the outer membrane of the cell, so that its hydrophilicity is significantly improved, and the characteristics of N and P on the phosphorylcholine groups appear. absorption peak.

Example 2

Weigh 14mmol of 2-methacryloyloxyethylphosphorylcholine and 6mmol of glycidyl methacrylate, take 0.1mmol of azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 2 mg/mL, and then 1 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of epoxy-containing phosphorylcholine polymer and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution for 12 hours at 45 °C, so that the amino groups in dopamine and the surface of chitosan film were mixed with phosphorus. Dopamine cross-links while epoxy reaction in acylcholine polymer, and the phosphorylcholine group is fixed on the shell by the reaction anchoring of epoxy and amino groups on the surface of chitosan film in phosphorylcholine polymer and the adhesion of dopamine On the surface of the glycan film, a cross-linked adhesive biomimetic coating with a cell-like outer membrane structure can be prepared.

Example 3

Weigh 19mmol of 2-methacryloyloxyethylphosphorylcholine and 1mmol of glycidyl methacrylate, take 0.1mmol of azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 5 mg/mL, and then 0.8 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of epoxy-containing phosphorylcholine polymer and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution at 95 °C for 2 hours, so that the amino groups in dopamine and the surface of chitosan film were mixed with phosphorus. Dopamine cross-links while epoxy reaction in acylcholine polymer, and the phosphorylcholine group is fixed on the shell by the reaction anchoring of epoxy and amino groups on the surface of chitosan film in phosphorylcholine polymer and the adhesion of dopamine On the surface of the glycan film, a cross-linked adhesive biomimetic coating with a cell-like outer membrane structure can be prepared.

Example 4

Weigh 15mmol of 2-methacryloyloxyethylphosphorylcholine and 5mmol of glycidyl methacrylate, take 0.1mmol of azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 4 mg/mL, and then 0.6 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of epoxy-containing phosphorylcholine polymer and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution for 4 hours at 60 °C, so that the amino groups in dopamine and the surface of chitosan film were mixed with phosphorus. Dopamine cross-links while epoxy reaction in acylcholine polymer, and the phosphorylcholine group is fixed on the shell by the reaction anchoring of epoxy and amino groups on the surface of chitosan film in phosphorylcholine polymer and the adhesion of dopamine On the surface of the glycan film, a cross-linked adhesive biomimetic coating with a cell-like outer membrane structure can be prepared.

Example 5

Weigh 17mmol 2-methacryloyloxyethylphosphorylcholine and 3mmol glycidyl methacrylate, use 0.1mmol azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 3 mg/mL, and then 0.4 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of the epoxy-containing phosphorylcholine polymer and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution for 6 hours at 70 °C, so that the amino groups in dopamine and the surface of the chitosan film were mixed with phosphorus. Dopamine cross-links while epoxy reaction in acylcholine polymer, and the phosphorylcholine group is fixed on the shell by the reaction anchoring of epoxy and amino groups on the surface of chitosan film in phosphorylcholine polymer and the adhesion of dopamine On the surface of the glycan film, a cross-linked adhesive biomimetic coating with a cell-like outer membrane structure can be prepared.

Example 6

Weigh 18mmol of 2-methacryloyloxyethylphosphorylcholine and 2mmol of glycidyl methacrylate, take 0.1mmol of azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 1 mg/mL, and then 0.2 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of the epoxy-containing phosphorylcholine polymer and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution for 8 hours at 80 °C, so that the amino groups in dopamine and the surface of chitosan film were mixed with phosphorus. Dopamine cross-links during the epoxy reaction in the acylcholine polymer, and the phosphorylcholine group is fixed to the chitosan by the anchoring of the epoxy in the phosphorylcholine polymer to the amino group on the surface of the chitosan film and the adhesion of dopamine. On the surface of the sugar film, a cross-linked adhesive biomimetic coating with a structure that mimics the outer membrane of a cell can be prepared.

Example 7

Weigh 14mmol of 2-methacryloyloxyethylphosphorylcholine and 6mmol of glycidyl methacrylate, take 0.1mmol of azobisisobutyronitrile as initiator, methanol and tetrahydrofuran as solvent, under nitrogen protection, 70 The polymerization reaction was carried out at °C for 24 h, dialyzed after the reaction, and then freeze-dried at -50 °C to obtain an epoxy-containing phosphorylcholine polymer.

The epoxy-containing phosphorylcholine polymer prepared in this example was prepared into a methanol solution of 2 mL and 0.2 mg/mL, and then 0.3 g of dopamine was added to mix uniformly. Then the mixed solution of the above-mentioned epoxy-containing phosphorylcholine polymer and dopamine was drop-coated on the surface of chitosan. The mixed solution of phosphorylcholine polymer containing epoxy and dopamine was dripped and dried, and then placed in a pH=8.5 Tris-HCl aqueous solution for 10 hours at 50 °C, so that the amino groups in dopamine and the surface of the chitosan film were mixed with phosphorus. Dopamine cross-links while epoxy reaction in acylcholine polymer, and the phosphorylcholine group is fixed on the shell by the reaction anchoring of epoxy and amino groups on the surface of chitosan film in phosphorylcholine polymer and the adhesion of dopamine On the surface of the glycan film, a cross-linked adhesive biomimetic coating with a cell-like outer membrane structure can be prepared.

The above are only the preferred embodiments of the present invention. It should be pointed out that for those skilled in the art, without departing from the principles of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be It is regarded as the protection scope of the present invention.

Claims (6)

1. the preparation method that contains epoxy phosphoryl choline polymer and dopamine cross-linked adhesion biomimetic coating, is characterized in that, comprises the steps: S1, under the protection of nitrogen, carry out radical polymerization reaction of vinyl monomer containing epoxy and vinyl monomer containing phosphoryl choline under the action of initiator to obtain epoxy-containing phosphoryl choline polymer ; S2, dissolving dopamine and the epoxy-containing phosphorylcholine polymer obtained in step S1 in a polar solvent to obtain a mixed solution, then coating the mixed solution on the surface of the material to be modified, and drying; S3. The material to be modified after drying in step S2 is placed in a pH=8.5 Tris-HCl aqueous solution, and heat-treated at 45°C to 95°C for 2h to 12h to obtain a cross-linking structure with a cell-like outer membrane structure. Adhesive biomimetic coating; The epoxy-containing vinyl monomer in step S1 is glycidyl methacrylate, and the phosphorylcholine-containing vinyl monomer is 2-methacryloyloxyethylphosphorylcholine. 2. the preparation method of containing epoxy phosphorylcholine polymer and dopamine cross-linked adhesion biomimetic coating according to claim 1, is characterized in that, described in step S1, the vinyl monomer containing phosphorylcholine The molar ratio to the epoxy-containing vinyl monomer is 9.5:0.5-7:3. 3. the preparation method that contains epoxy phosphorylcholine polymer and dopamine cross-linking adhesion biomimetic coating according to claim 1, is characterized in that, the solvent of radical polymerization described in step S1 is methanol and tetrahydrofuran It is obtained by mixing in a volume ratio of 4:1, the initiator is azobisisobutyronitrile, the reaction temperature is 70°C, the reaction time is 24h, and the dialysis bag with molecular weight cut-off of 7000 is used for dialysis. 4. the preparation method of containing epoxy phosphorylcholine polymer and dopamine cross-linked adhesion biomimetic coating according to claim 1, is characterized in that, the concentration of dopamine in the mixed solution described in step S2 is 0.1mg/ mL to 0.5 mg/mL, and the concentration of the epoxy-containing phosphorylcholine polymer is 0.2 mg/mL to 5 mg/mL. 5 . The method for preparing a biomimetic coating containing epoxy phosphorylcholine polymer and dopamine cross-linking adhesion according to claim 1 , wherein the polar solvent in step S2 is methanol or ethanol. 6 . 6 . The method for preparing a biomimetic coating containing epoxy phosphorylcholine polymer and dopamine cross-linking and adhering according to claim 1 , wherein the material to be modified in step S2 is chitosan. 7 .

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