A composite gel product and its use in the preparation of a cartilage joint lesion repair material

Abstract

The application provides a composite gel product which is designed according to the main components of articular cartilage based on the bionics principle and is made of biological cellulose (BC), polyvinyl alcohol (PVA), an anionic surfactant and chitosan, has mechanical properties, friction characteristics close to articular cartilage and good biocompatibility and degradability and can be used for preparing a cartilage joint injury repair material.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of medical materials, in particular to a composite gel product and application of the composite gel product in preparation of cartilage joint injury repair material. BACKGROUND

[0002] Articular cartilage is a complex and special structure, which covers the bone end constituting the joint, and its function is to provide a low-friction contact surface for the joint within a certain range of activities, and can buffer the impact force during joint movement. The damage of articular cartilage can be caused by acute trauma, or can be a chronic degenerative structure. Articular cartilage has no blood supply, so once the cartilage is damaged, it is often difficult to heal. The current methods for treating cartilage injury are: 1) drug treatment: oral administration of glucosamine and chondroitin to promote the self-recovery of cartilage, while reducing the weight to avoid further damage; 2) bone marrow stimulation method, including cartilage trimming and local injection of autologous blood; 3) cartilage transplantation. Methods 1) and 2) are greatly dependent on the patient's own condition, especially for older patients, the effect is difficult to satisfy. For method 3), it is urgent to find a suitable material as a cartilage substitute. For the articular cartilage of human or animal body, it is mainly composed of water (60-85%), collagen fibers with a diameter of about 100 nanometers (15-22%) and negatively charged polyprotein sugar (4-7%). The collagen fiber network can give the cartilage high tensile strength. Polyprotein sugar is a brush-like molecule with negative charge, and the negative charge comes from the sulfate group on the glycosaminoglycan chain attached to the core of the protein. Polyprotein polysaccharide forms large aggregates with hyaluronic acid, which are trapped in the collagen network, resulting in osmotic pressure against compression load.

[0003] Biocellulose (BC) is a natural cellulose gel material produced by microbial metabolism, which has high purity, high water retention and good mechanical properties and biocompatibility. The chemical formula of polyvinyl alcohol (PVA) is [C2H4O] n , which can form a polyvinyl alcohol hydrogel elastomer by repeated freeze-thaw method, not only has good lubricating effect, but also has certain elasticity and shock absorption capacity. Anionic surfactants can provide fixed negative charge after ionization in solution; and chitosan hydrogel has good biodegradability, safety and biocompatibility, and soft texture, which is a good biomimetic material. SUMMARY

[0004] From the perspective of bionics, according to the main components of the articular cartilage of human or animal body, a composite gel product is designed and prepared, which has good physicochemical properties and can be used to prepare cartilage joint injury repair material.

[0005] The present application provides a composite gel product, which is mainly prepared from a bio-cellulose hydrogel, polyvinyl alcohol, an anionic surfactant and chitosan.

[0006] The anionic surfactant can be various anionic surfactants commonly used, such as anionic polyacrylamide, higher fatty acid salt, triethanolammonium salt and sulfonate.

[0007] The present application also provides use of the composite gel product in preparation of a joint injury repair material.

[0008] The composite gel in the present application is prepared by the following method:

[0009] (1) preparing a bio-cellulose (BC) hydrogel and purifying it;

[0010] (2) preparing a bio-cellulose-polyvinyl alcohol (BC-PVA) composite gel: immersing the BC hydrogel prepared in step (1) into a heated PVA solution, allowing the PVA to diffuse into the three-dimensional space network of the BC hydrogel by incubation and standing, taking it out, quickly freezing and then naturally thawing to prepare the BC-PVA composite gel;

[0011] (3) preparing a composite gel product: dissolving chitosan in an acetic acid solution, adding an anionic surfactant, immersing the BC-PVA composite gel prepared in step (2) into the solution, and then adding a crosslinking agent;

[0012] (4) solidifying the composite gel.

[0013] In the above method, the bio-cellulose hydrogel can be prepared by various known methods, such as using various known bio-cellulose producing bacteria (Gluconacetobacter xylinus, Monascus ruber, etc.), using various known culture media (such as coconut water culture medium, artificial glucose culture medium, molasses culture medium, etc.), using various known fermentation culture methods (such as shallow plate static fermentation method, shaking table dynamic fermentation method, etc.), and determining the specific conditions of fermentation according to the habits of the bio-cellulose producing bacteria (such as 25-30℃, fermentation for 3-9 days).

[0014] In the above method, the thickness of the bio-cellulose hydrogel can be determined according to the needs, but is preferably 0.3-0.8mm; more preferably 0.5mm; and it can be controlled by various known methods, for example, the bio-cellulose hydrogel can be prepared into the above thickness of hydrogel by controlling the fermentation time or mechanical cutting;

[0015] In the above method, the PVA is preferably PVA with medium polymerization degree, and more preferably PVA with a molecular weight of about 146000; and the concentration of PVA in the aqueous solution is preferably 25-40%, more preferably 35%;

[0016] In the above method, the temperature of the heated PVA solution is preferably 120-140℃, more preferably 135℃;

[0017] In the above method, the time for the BC hydrogel to stand after being immersed in the PVA solution is preferably 20-30h, more preferably 24h;

[0018] In the above method, the rapid freezing is at -60 to -90℃ for 20-40 minutes, more preferably at -78℃ for 30 minutes;

[0019] In the above method, the concentration of the chitosan in the solution is preferably 1-3%, more preferably 2%;

[0020] In the above method, the concentration of the anionic surfactant in the solution is preferably 1-3%, more preferably 2%;

[0021] In the above method, the cross-linking agent can be various conventional cross-linking agents, and is preferably glutaraldehyde, which has a concentration of 1-2% in the solution;

[0022] In the above method, a photoinitiator is also added in step 3), which is preferably I2959, and has a concentration of 1-2% in the solution;

[0023] In the above method, the time for the BC-PVA hydrogel to stand is preferably 20-30h, more preferably 24h;

[0024] In the above method, the solidification includes UV irradiation and heating treatment; the UV irradiation is UV irradiation for 20-40 minutes, preferably 30 minutes; the heating treatment is oven heating at 50-70℃ for 2-6h, preferably oven heating at 60℃ for 3h;

[0025] In the above method, a step of rinsing the solidified composite gel product with PBS buffer and then immersing it in PBS buffer for preservation is further included.

[0026] In the above, unless otherwise specified, all are weight percentages.

[0027] The composite gel product prepared by the method has mechanical properties, friction characteristics close to those of articular cartilage, and good biocompatibility and degradability, and has better lubricity and more excellent wear resistance than PVA, and can be used to prepare joint damage repair materials. DETAILED DESCRIPTION

[0028] The application will be further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the application, and should not limit the scope of the application. After reading the application, those skilled in the art can make various forms of changes on the basis of the application, which should fall within the scope of the application.

[0029] Example 1:

[0030] The bio-cellulose hydrogel was prepared by the static culture method with coconut water medium, and cultured at 28°C for 7 days. After washing and purifying with deionized water and weak alkali solution, the bio-cellulose hydrogel was cut into 0.5mm thick.

[0031] The PVA with molecular weight of about 146000 was added into deionized water to prepare a PVA aqueous solution with concentration of 35wt%, and heated to 135°C. The BC hydrogel was immersed in the heated PVA aqueous solution for 24h. The gel was taken out, frozen at -78°C for 30min, and naturally thawed to prepare the BC-PVA composite gel.

[0032] After 1g of chitosan was dissolved in 50ml of acetic acid solution (2%), 1g of anionic polyacrylamide was added, and the BC-PVA composite gel was immersed in the above solution. Then, 0.5g of glutaraldehyde and 0.5g of I2959 were added, and the immersion was maintained for 24h.

[0033] After the composite gel product was taken out, it was irradiated with ultraviolet light for 30min, and then heated in an oven at 60°C for 3h. After washing with PBS buffer, it was immersed in PBS buffer for storage.

[0034] Example 2:

[0035] The bio-cellulose hydrogel was prepared by the static culture method with molasses medium, and cultured at 26°C for 3 days to a thickness of 0.4mm. Then, it was washed and purified with deionized water and weak alkali solution.

[0036] The PVA with molecular weight of about 146000 was added into deionized water to prepare a PVA aqueous solution with concentration of 30wt%, and heated to 120°C. The BC hydrogel was immersed in the heated PVA aqueous solution for 30h. The gel was taken out, frozen at -60°C for 40min, and naturally thawed to prepare the BC-PVA composite gel.

[0037] After 1.2g of chitosan was dissolved in 50ml of acetic acid solution (2%), 0.6g of anionic polyacrylamide was added, and the BC-PVA composite gel was immersed in the above solution. Then, 0.6g of glutaraldehyde and 0.6g of I2959 were added, and the immersion was maintained for 20h.

[0038] After the composite gel product was taken out, it was irradiated with ultraviolet light for 25min, and then heated in an oven at 50°C for 5h. After washing with PBS buffer, it was immersed in PBS buffer for storage.

[0039] Example 3:

[0040] The bio-cellulose hydrogel is prepared by a dynamic culture method on a shaker, a glucose culture medium is artificially configured, and the culture is carried out at 29°C for 7 days, and then the bio-cellulose hydrogel is washed and purified by using deionized water and a weak alkali solution, and then the bio-cellulose hydrogel is cut into pieces with a thickness of 0.7 mm;

[0041] The PVA with a molecular weight of about 146000 is selected, deionized water is added, and a PVA aqueous solution with a concentration of 40wt% is prepared, the PVA aqueous solution is heated to 140°C, the BC hydrogel is immersed in the heated PVA aqueous solution, and the immersion is maintained for 20 hours; the gel is taken out, frozen at -90°C for 20 minutes, and naturally thawed, and the BC-PVA composite gel is prepared;

[0042] After 1.5g of chitosan is dissolved in 50ml of an acetic acid solution (2%), 0.8g of anionic polyacrylamide is added, the solution is uniformly stirred, the BC-PVA composite gel is immersed in the solution, and then 0.8g of glutaraldehyde and 0.8g of I2959 are added, and the immersion is maintained for 30 hours;

[0043] After the composite gel product is taken out, the product is irradiated with ultraviolet light for 20 minutes, and then the product is heated in an oven at 70°C for 2 hours, and then the product is washed with a PBS buffer and immersed in the PBS buffer for storage.

[0044] Experimental example:

[0045] Experimental object: the composite gel product prepared in Examples 1-3 is cut into square pieces with a diameter of 2cm as an experimental example group, and a conventional PVA hydrogel with the same thickness and size and pig joint cartilage with the same thickness and size are used as Comparative Examples 1 and 2, respectively.

[0046] 1. Mechanical property experiment

[0047] The mechanical experiment is carried out on the experimental example group, the Comparative Example 1 and 2, and the tensile strength, tensile modulus, compressive strength, compressive modulus and elastic modulus are measured, the test conditions are as follows: the probe rate is 500mm / min, the temperature is room temperature, and the humidity is 82%, the experiment is carried out for 20 times, and the average value is taken. The results are shown in the following table:

[0048] Tensile strength (MPa) Tensile modulus (MPa) Compressive strength (MPa) Compressive modulus (MPa) Elastic modulus (MPa) Example 1 24.36 223.43 27.84 13.78 0.74 Example 2 21.67 206.72 25.47 11.56 0.78 Example 3 18.33 188.27 21.83 10.12 0.82 PVA hydrogel 6.43 40.33 22.42 3.78 0.58 Porcine articular cartilage 25.82 220.38 33.81 13.24 0.77

[0049] As can be seen from the above table, the composite gel product prepared in the application is closer to pig joint cartilage in mechanical properties, especially the composite gel product prepared in Example 1, which is basically at the same level as pig joint cartilage in tensile strength, tensile modulus, compressive strength, compressive modulus and elastic modulus; and the PVA hydrogel cannot reach the mechanical properties of pig joint cartilage in each index.

[0050] Friction experiment

[0051] Each of the example groups and the comparative example groups was placed in a 0.15 m PBS buffer, and a friction experiment was performed using a rotating pin disc, 105 rotations, 1 MPa pressure, to measure the friction coefficient and the maximum wear depth. The results are shown in the following table:

[0052] Coefficient of friction Maximum wear depth (mm) Example 1 0.05 0.31 Example 2 0.06 0.35 Example 3 0.06 0.37 PVA hydrogel 0.17 1.23 Porcine articular cartilage 0.11 0.48

[0053] As can be seen from the above table, the composite gel product prepared in the present application has a lower friction coefficient than pig joint cartilage, and has better lubricity. The maximum wear depth is also significantly less than that of pig joint cartilage, and the product has better wear resistance. The friction coefficient of PVA hydrogel is too large, and the wear resistance is not enough.

[0054] As can be seen from the above experimental results, the composite gel product prepared in the present application has properties close to joint cartilage, and can be used to prepare joint damage repair materials.

[0055] The above examples only describe the preferred embodiments of the present application, and do not limit the scope of the present application. Various modifications and improvements to the technical solutions of the present application made by ordinary engineering technicians in the art without departing from the design spirit of the present application shall fall within the protection scope defined by the claims of the present application.

Claims

1. A method for preparing a composite gel product, characterized in that: This composite gel is mainly prepared from bio-cellulose hydrogel, polyvinyl alcohol, anionic surfactant, and chitosan; the preparation method is as follows: (1) Prepare bio-cellulose hydrogels and clean and purify them; (2) Preparation of biocellulose polyvinyl alcohol BC-PVA composite gel: Immerse the biocellulose hydrogel prepared in step (1) into a heated PVA solution, keep it warm and stand so that PVA diffuses into the three-dimensional spatial network of the biocellulose hydrogel, take it out, freeze it quickly and then thaw it naturally to make BC-PVA composite gel. (3) Preparation of composite gel products: Chitosan is dissolved in acetic acid solution, anionic surfactant is added, the BC-PVA composite gel prepared in step (2) is immersed in it, and then a crosslinking agent is added; (4) Curing the composite gel; The PVA mentioned therein is PVA with a molecular weight of 146,000, and the concentration of PVA in the aqueous solution is 25-40%; the temperature of the heated PVA solution is 120-140℃; the BC hydrogel is immersed in the PVA solution and kept at a constant temperature for 20-30 hours; the rapid freezing is carried out at -60 to -90℃ for 20-40 minutes. The concentration of chitosan in the solution is 1-3%; the concentration of the anionic surfactant in the solution is 1-3%, and the anionic surfactant is anionic polyacrylamide.

2. The method for preparing the composite gel product according to claim 1, characterized in that: The thickness of the biocellulose hydrogel prepared in step (1) is 0.3-0.8 mm.

3. The method for preparing the composite gel product according to claim 2, characterized in that: The thickness of the biocellulose hydrogel prepared in step (1) is 0.5 mm.

4. The method for preparing the composite gel product according to claim 1, characterized in that: The concentration of PVA in the aqueous solution is 35%; the temperature of the heated PVA solution is 135°C; the BC hydrogel is immersed in the PVA solution and kept at a constant temperature for 24 hours; the rapid freezing is performed at -78°C for 30 minutes.

5. The method for preparing the composite gel product according to claim 1, characterized in that: The concentration of chitosan in the solution is 2%; the concentration of the anionic surfactant in the solution is 2%.

6. The method for preparing the composite gel product according to claim 1, characterized in that: The crosslinking agent is glutaraldehyde, and its concentration in solution is 1-2%.

7. The method for preparing the composite gel product according to claim 1, characterized in that: In step (3) of the method, a photoinitiator is also added; it is I2959, and its concentration in the solution is 1-2%.

8. The method for preparing the composite gel product according to claim 1, characterized in that: The immersion time of the BC-PVA hydrogel in the method is 20-30 hours; the curing includes ultraviolet irradiation and heat treatment; the ultraviolet irradiation is 20-40 minutes; the heat treatment is heating in an oven at 50-70°C for 2-6 hours.

9. The method for preparing the composite gel product according to claim 8, characterized in that: The immersion time of BC-PVA hydrogel in the method is 24 hours; the ultraviolet irradiation is 30 minutes; and the heat treatment is 3 hours in a 60°C oven.

10. The method for preparing the composite gel product according to claim 1, characterized in that: It also includes the step of rinsing the cured composite gel product with PBS buffer and then immersing it in PBS buffer for storage.

11. A composite gel product, characterized in that... Prepared by the method described in any one of claims 1-10.

12. The use of the composite gel product as described in claim 11 in the preparation of joint injury repair materials.