Succinic acid polymer and preparation method and derivative thereof, adhesive hydrogel and preparation method and application thereof

Abstract

The invention belongs to the field of material science and clinical medicine, and particularly relates to a succinic acid polymer and a preparation method and derivative thereof, adhesive hydrogel and a preparation method and application thereof. According to the specific technical scheme, the succinic acid polymer is provided, and the succinic acid acrylate polymer is prepared by modifying the succinic acid polymer through acryloyl chloride; further uniformly mixing a succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate and acrylic acid N-hydroxysuccinimide ester with a photoinitiator phenyl-2, 4, 6-trimethylbenzoyl lithium phosphite, and reacting under the irradiation of ultraviolet light to obtain the photoinitiator. The three-dimensional network structure hydrogel with strong wet adhesion, low swelling rate and high toughness is prepared through a photopolymerization reaction. The hydrogel has good biocompatibility, an in-vivo rabbit model proves that an orificium fistulae can be completely sealed through single use or combined suture, a safe and efficient novel biological material is provided for anastomotic fistula treatment, and the hydrogel has important clinical transformation potential.

Description

Technical Field

[0001] This application relates to the fields of materials science and clinical medicine, and specifically relates to succinic acid polymers and their preparation methods and derivatives, adhesive hydrogels and their preparation methods and applications. Background Art

[0002] Anastomotic leakage is currently the most common complication after all digestive tract tumor surgeries. Among them, the incidence of anastomotic leakage after esophageal cancer surgery is about 10 - 20%, which seriously affects the quality of life and survival time of patients after esophageal surgery. Excessive anastomotic tension is the main cause of anastomotic leakage, and currently, the only method to reduce anastomotic tension is to increase the number of sutures, which completely depends on the surgical experience of surgeons. However, this method can cause tissue damage and leakage, resulting in the spillage of digestive contents into the mediastinum or thoracic and abdominal cavities, and is prone to local or systemic infections. Therefore, there is an urgent need to develop adhesive materials with high efficiency, non-invasiveness, and excellent sealing properties.

[0003] Since the esophageal environment is humid and complex, it is necessary to develop a hydrogel material with high wet-state adhesiveness and low swelling properties, as well as a ready-to-use adhesive material with high biosecurity, portability, and adjustable mechanical strength. This material will provide significant practical application value for solving the problem of esophageal anastomotic leakage. Summary of the Invention

[0004] The purpose of the present invention is to provide a succinic acid polymer and its preparation method and derivatives, an adhesive hydrogel and its preparation method and applications.

[0005] To achieve the above-mentioned invention purpose, the technical solutions adopted by the present invention are as follows: In the first aspect, the present application provides a succinic acid polymer, and the structural formula of the succinic acid polymer is as follows:

[0006] n is an integer from 1 to 10; m is a positive integer.

[0007] Furthermore, the number-average molecular weight of the succinic acid polymer is 5000 - 8000 Da, and more preferably 6000 - 7000 Da.

[0008] Furthermore, the molecular weight distribution coefficient of the succinic acid polymer is 1.0 - 2.0, and more preferably 1.0 - 1.5.

[0009] Second aspect, the present application provides a preparation method of the succinic acid polymer described in the first aspect. The preparation method includes the following steps: using polyethylene glycol-1000, glycerol, and succinic acid as raw materials, N,N'-dicyclohexylcarbodiimide as a dehydrating agent, and 4-dimethylaminopyridine as a catalyst, adding them into a three-necked flask in sequence, using N,N'-dimethylformamide as a solvent, heating and reacting in a water bath at 60°C for 4 h; filtering to remove impurities, precipitating with methyl tert-butyl ether, standing overnight, filtering off the supernatant to obtain a white waxy solid, which is the succinic acid polymer.

[0010] Third aspect, the present application provides a derivative of the succinic acid polymer described in the first aspect, named succinic acid acrylate polymer. The chemical structural general formula of the succinic acid acrylate polymer is as follows:

[0011] n is an integer from 1 to 10; m is a positive integer.

[0012] Fourth aspect, the present application provides a preparation method of the succinic acid acrylate polymer described in the third aspect, using acryloyl chloride to modify the succinic acid polymer to prepare the succinic acid acrylate polymer.

[0013] Further, the preparation method includes the following steps: under the protection of an inert gas, using the succinic acid polymer and acryloyl chloride as raw materials, triethylamine as an acid-binding agent, and DMF as a solvent, reacting under ice bath conditions for 16 h; after the reaction, filtering to remove impurities, crystallizing, and filtering to obtain a filter cake, which is the succinic acid acrylate polymer.

[0014] Fifth aspect, the present application provides an adhesive hydrogel. The adhesive hydrogel is a crosslinked network prepared by using the succinic acid acrylate polymer described in the fourth aspect as a crosslinking agent, and acrylic acid, dimethylaminoethyl methacrylate, and N-hydroxysuccinimide acrylate as crosslinking monomers. The crosslinking method is chemical crosslinking, and the crosslinked structure is a three-dimensional network structure.

[0015] Preferably, the adhesive hydrogel formulation includes: the succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate, N-hydroxysuccinimide acrylate, and a photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphite.

[0016] Preferably, by mass percentage, the content of the succinic acid acrylate polymer is 1 - 15%; and / or; the content of acrylic acid is 30 - 50%; and / or; the content of dimethylaminoethyl methacrylate is 5 - 30%; and / or; the content of N-hydroxysuccinimide acrylate is 1 - 10%; and / or; the content of the photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphite is 0.1 - 1%.

[0017] In a sixth aspect, the present application provides a method for preparing the adhesive hydrogel described in the fifth aspect. Dissolve succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate, N-hydroxysuccinimide acrylate, and photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphite in ultrapure water, and mix well to obtain a uniform precursor solution. Further, cure the precursor solution under ultraviolet light irradiation for 20 to 600 s to obtain the adhesive hydrogel.

[0018] In a seventh aspect, the present application provides the use of the succinic acid acrylate polymer or the succinic acid acrylate polymer prepared by the preparation method or the adhesive hydrogel in adhering biomaterials, glass, and metal. Any combination of biomaterials, glass, and metal can be adhered, for example, a biomaterial can be adhered to metal / glass, or a biomaterial can also be adhered to another biomaterial, etc.

[0019] Preferably, the adhesive hydrogel prepared from the succinic acid acrylate polymer is used in the treatment of esophageal anastomotic fistula, and can adhere the esophagus to the esophagus.

[0020] The present invention has the following beneficial effects: The present invention provides a new succinic acid acrylate polymer and provides a specific preparation method. The succinic acid acrylate polymer can be used to prepare an adhesive hydrogel. The adhesive hydrogel is a polymer material with excellent adhesion, mechanical properties, and biocompatibility, having strong adhesion to wet tissues, high toughness, and low swelling, and at the same time having the advantages of high efficiency, safety, convenience, and mildness. It is an ideal tissue adhesive material and can be used for the adhesive closure in the treatment of esophageal anastomotic fistula. Description of the Drawings

[0021] Figure 1 It is the chemical structural formula and nuclear magnetic resonance hydrogen spectrum of the succinic acid polymer.

[0022] Figure 2 It is the chemical structural formula and nuclear magnetic resonance hydrogen spectrum of the succinic acid acrylate polymer.

[0023] Figure 3 It is the Fourier transform infrared spectrum of the succinic acid polymer.

[0024] Figure 4 It is the Fourier transform infrared spectrum of the succinic acid acrylate polymer.

[0025] Figure 5 It is a schematic diagram of the biocompatibility effect of the adhesive hydrogel. Detailed Embodiments

[0026] The present invention provides a new succinic acid polymer, and the chemical structural formula of the succinic acid polymer is as follows:

[0027] n is an integer from 1 to 10; m is a positive integer.

[0028] Preferably, the number-average molecular weight of the succinic acid polymer is 5000 - 8000 Da, and more preferably, the number-average molecular weight of the succinic acid polymer is 6000 - 7000 Da.

[0029] Preferably, the molecular weight distribution coefficient of the succinic acid polymer is 1.0 - 2.0, and more preferably, the molecular weight distribution coefficient of the succinic acid polymer is 1.0 - 1.5.

[0030] The present invention also provides a preparation method of the succinic acid polymer, which specifically includes the following steps: Using polyethylene glycol-1000, glycerol and succinic acid as raw materials, N,N'-dicyclohexylcarbodiimide as a dehydrating agent, and 4-dimethylaminopyridine as a catalyst, adding them into a three-necked flask in sequence, using N,N'-dimethylformamide as a solvent, and reacting in a water bath at 60 °C for 4 h. Filter to remove impurities, precipitate with methyl tert-butyl ether, let stand overnight, filter off the supernatant, and obtain a white waxy solid, which is the succinic acid polymer.

[0031] The present invention provides a succinic acid polymer derivative: succinic acid acrylate polymer, which is obtained by modifying polyethylene glycol with acryloyl chloride.

[0032] The chemical structural formula of the succinic acid acrylate polymer is as follows:

[0033] n is an integer from 1 to 10; m is a positive integer.

[0034] The present invention also provides a preparation method of the succinic acid acrylate polymer, which specifically includes the following steps: Under the protection of an inert gas, using the succinic acid polymer and acryloyl chloride as raw materials, triethylamine as an acid-binding agent, and DMF as a solvent, reacting under ice bath conditions for 16 h. After the reaction, filter to remove impurities, crystallize, and filter to obtain a filter cake, which is the succinic acid acrylate polymer.

[0035] The present invention also provides the application of the succinic acid acrylate polymer in the preparation of an adhesive hydrogel. The succinic acid acrylate polymer can also maintain high adhesion strength in a humid and complex physiological environment and can be used for the treatment of esophageal fistula.

[0036] The present invention also provides an adhesive hydrogel prepared using the succinic acid acrylate polymer. The formulation of the adhesive hydrogel includes: succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate, N-hydroxysuccinimide acrylate, and a photoinitiator. Among them, by mass percentage, the content of the succinic acid acrylate polymer is 1-5%, preferably 1-5%; the content of acrylic acid is 30-50%, preferably 35-40%; the content of dimethylaminoethyl methacrylate is 5-30%, preferably 5-10%; the content of N-hydroxysuccinimide acrylate is 1-10%, preferably 2-5%; the content of the photoinitiator is 0.1-1%, preferably 0.1-0.3%.

[0037] The present invention also provides a method for preparing the adhesive hydrogel, which specifically includes the following steps: Dissolve the succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate, N-hydroxysuccinimide acrylate, and the photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphinate in ultrapure water, and mix well to obtain a uniform precursor solution. Further, cure the precursor solution under ultraviolet light irradiation to obtain the adhesive hydrogel.

[0038] The present invention also provides a method for using the adhesive hydrogel, which specifically includes the following steps: Pre-clean the bonding site and fully expose it to the field of view for visual operation. Cut the adhesive hydrogel into a suitable size and shape, place it on the site to be bonded, apply a certain pressure (1-5 kPa) to the biological adhesive material for a certain period of time (5-20 s), ensure that the pressing object is clean and smooth, avoid metal utensils, and stop pressing after the biological adhesive material is in full contact with and bonded to the site to be bonded.

[0039] The adhesive hydrogel can still maintain excellent adhesive ability in a humid environment. Therefore, the adhesive hydrogel can be used for the treatment of esophageal anastomotic fistula.

[0040] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. If not specifically specified, the technical means used in the embodiments are conventional means well known to those skilled in the art. The obtained data are all averages obtained after at least 3 repetitions, and all repetitions obtained are valid data.

[0041] Example 1: Preparation of Succinic Acid Polymer Polyethylene glycol - 1000 (5 g, 0.005 mol), glycerol (0.46 g, 0.005 mol), N,N'-dicyclohexylcarbodiimide (2.06 g, 0.01 mol), 4-dimethylaminopyridine (0.122 g, 0.001 mol), and succinic acid (0.94 g, 0.008 mol) were successively added into a 100 mL three-necked flask, and 10 mL of N,N'-dimethylformamide was added. The mixture was heated in a water bath at 60 °C for 4 h. After the reaction, the by-product dicyclohexylurea was removed by suction filtration. 10 times the volume of methyl tert-butyl ether was added, and the mixture was allowed to stand overnight for precipitation. The supernatant was filtered off to obtain a white waxy solid with an esterification rate of 71.2%, which was the succinic acid polymer.

[0042] Deuterated dimethyl sulfoxide was used as the deuterated reagent to measure the nuclear magnetic resonance hydrogen spectrum of the product, and the KBr pellet method was used to measure the Fourier transform infrared spectrum of the product to determine the molecular structure of the succinic acid polymer. The chemical structural formula and nuclear magnetic resonance hydrogen spectrum of the succinic acid polymer are as Figure 1 shown. The peaks appearing at a (2.5 ppm) and b (3.5 ppm) in the figure are mainly attributed to the methylene functional group of succinic acid and the polyethylene glycol part. The Fourier transform infrared spectrum of the succinic acid polymer is as Figure 3 shown, and the peaks appearing at 1730 cm -1 and 1100 cm -1 are mainly attributed to the C=O stretching vibration and C-O-C stretching vibration of the succinic acid ester, indicating that the succinic acid polymer was successfully prepared.

[0043] Example 2: Preparation of succinic acid acrylate polymer 1. Preparation of succinic acid acrylate polymer Under the protection of an inert gas, succinic acid polymer (0.00006 mol) and triethylamine (0.0072 mol) were added to 10 mL of N,N'-dimethylformamide, and acryloyl chloride (0.0072 mol) was added dropwise under ice bath conditions. The reaction was carried out for 16 h. After the reaction, the by-product triethylamine salt was removed by suction filtration. 10 times the volume of methyl tert-butyl ether was added, and the mixture was allowed to stand overnight for precipitation. The supernatant was filtered off to obtain a white waxy solid with an esterification rate of 63.5%, which was the succinic acid acrylate polymer.

[0044] Deuterated dimethyl sulfoxide was used as the deuterated reagent to measure the nuclear magnetic resonance hydrogen spectrum of the product, and the KBr pellet method was used to measure the Fourier transform infrared spectrum of the product to determine the molecular structure of the succinic acid acrylate polymer. The products of the succinic acid acrylate polymer obtained by the two methods are the same, both proving that the succinic acid acrylate polymer was successfully obtained. The chemical structural formula and nuclear magnetic resonance hydrogen spectrum of the succinic acid acrylate polymer are as Figure 2As shown, new peaks c, d, and e appeared at 5.9, 6.2, and 6.3 ppm, mainly attributed to the hydrogen on the acryloyl functional group, proving that the acryloyl group was successfully modified on the succinic acid polymer. The Fourier transform infrared spectroscopy diagram of the succinic acid acrylate polymer is as Figure 4 shown. Compared with the succinic acid polymer, the succinic acid acrylate polymer showed obvious characteristic peaks at 1643 cm -1 , mainly attributed to the C=C stretching vibration of the acryloyl group, further indicating the successful preparation of the succinic acid acrylate polymer.

[0045] Example 3: Preparation and Effect Demonstration of Adhesive Hydrogel 1. Preparation of Adhesive Hydrogel The adhesive hydrogel was obtained by free radical polymerization under light irradiation initiated by a photoinitiator from succinic acid acrylate polymer, acrylic acid monomer, dimethylaminoethyl methacrylate, and N-hydroxysuccinimide acrylate monomer. The specific preparation method is as follows: Dissolve succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate, N-hydroxysuccinimide acrylate, and the photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphite in ultrapure water, and mix well to obtain a uniform precursor solution. Then, inject the precursor solution into a mold and cure it under ultraviolet light irradiation to obtain the adhesive hydrogel. The dosage values in Table 1 are mass percentages.

[0046] Table 1 Comparison Table of Preparation Parameters of Each Biological Adhesive Material

[0047] 2. Demonstration of Biosafety Effect The in vitro toxicity of the hydrogel was evaluated using HUVECs and L929 cell lines. The adhesive hydrogel was extracted by the immersion method, and the extraction ratio was film area: 10% serum medium = 6 cm 2 : 1 mL, and the extraction condition was extraction at 37 °C for 72 h. Digest the cells and inoculate them into a 96-well plate at a rate of 2×10 3 cells / well. After 24 h, treat them with the hydrogel extract, culture them in an incubator (5% CO2) at 37 °C for 24 h, and use a CCK-8 kit to measure the cell viability by measuring the absorbance at 450 nm. At the same time, set a blank group without adding biological adhesive material, and the other conditions are exactly the same. As Figure 5 shown, the results show that when the content of dimethylaminoethyl methacrylate is greater than 10%, the adhesive hydrogel has good cell compatibility.

[0048] 3. Demonstration of Tissue Adhesion Strength Effect Fresh porcine liver was used as a biological tissue sample, and the adhesion strength of the hydrogel was evaluated by lap shear tests. Briefly, the porcine liver was cut into rectangles of 25 mm × 60 mm and soaked in PBS before use. Subsequently, a certain area of the hydrogel was pasted onto the surface of the porcine liver, and another piece of porcine liver was placed on top of the hydrogel to achieve a contact area of 25 mm × 10 mm. Then, it was left standing at room temperature for 5 min, after which a lap shear test was performed using a material testing system equipped with a 100 N weighing unit and a shear rate of 50 mm / min to evaluate the adhesion performance. All these tests were repeated more than 3 times. The results are shown in Table 2.

[0049] Table 2 Comparison table of the adhesion performance of each adhesive hydrogel

[0050] 4. Demonstration of swelling performance Weigh 0.1 g of each adhesive hydrogel prepared in Step 1 and record it as the initial weight (W0). Place them separately in 15 mL of physiological saline. After a fixed period of time, take out the swollen hydrogels from the physiological saline, gently wipe off the excess moisture on the surface and record the mass of the gels, denoted as Wt, and then put the hydrogels back into the physiological saline. The swelling ratio (SR) of the hydrogel can be calculated by the formula: SR = (Wt - W0) / W0 × 100%. Place gel patches of fixed mass in a physiological saline solution and observe and record the mass of the gels at fixed time intervals. The results are shown in Table 3.

[0051] Table 3 Comparison table of the swelling performance of each bioadhesive material

[0052] 5. In vivo animal experiments Nine male New Zealand white rabbits (4 months old, body weight 2.5 ± 0.5 kg) were conventionally raised for 1 week and randomly divided into 3 groups. After the rabbits were anesthetized by intramuscular injection of Sumianxin II (0.2 ml / kg), their hair was shaved and disinfected. A longitudinal incision of about 3 cm was made along the left edge of the trachea starting from the level of the lower edge of the cricoid cartilage. The fascia and muscle tissues were separated, and the trachea was freed. The esophagus is located behind the right of the trachea, and the esophagus was separated and exposed.

[0053] Group A: A 2-mm slit was longitudinally cut in the esophagus with fine scissors. After the esophageal mucosa layer was visible, a 1*0.5 cm PG2.5D10 hydrogel was used to wrap around the esophagus for one week, and the muscle and the incision were closed layer by layer. (Direct wrapping) Group B: A 2-mm slit was longitudinally cut in the esophagus with fine scissors. After the esophageal mucosa layer was visible, the esophageal slit was closed with 5-0 prolene sutures, and the muscle and the incision were closed layer by layer. (Direct suture) Group C: A 2-mm longitudinal incision was made in the esophagus with fine scissors. After the esophageal mucosa was visible, the esophageal incision was closed with 5-0 prolene suture. A 1*0.5 cm PG2.5D10 hydrogel was wrapped around the esophagus for one week, and finally the muscle and incision were closed layer by layer. (Wrapped after suture) At 14 days after surgery, the animals were sacrificed by air embolism via the marginal ear vein, and the surgical segment of the esophagus (including 1 cm at both ends of the anastomosis) was removed intact.

[0054] The experimental results showed that the animals in Group B (simple suture) died due to severe infection on the 3rd day after surgery, and the failure reason could be clearly attributed to the formation of anastomotic leakage. The animals in Group A and Group B were in good growth condition and were sacrificed 2 weeks later. After the neck incision was opened, no obvious inflammatory changes were seen, and the gel was still in place. After the gel was removed, the original injury incision was not visible to the naked eye, which was attributed to the high adhesiveness of the hydrogel, which tightly adhered to the esophageal surface in the wet environment. Although esophageal peristalsis could still maintain the integrity of the physical barrier and block the leakage of digestive fluid. The equivalence between Group A and Group C further indicated that the hydrogel could bypass the traditional suture steps and directly achieve functional closure through biological interface interaction. This not only simplified the surgical operation and reduced the technical dependence, but also avoided suture-related complications (such as foreign body reaction and suture breakage).

[0055] The embodiments described above are only descriptions of the preferred embodiments of the present invention, and do not limit the scope of the present invention. Without departing from the design spirit of the present invention, various deformations, variations, modifications, and substitutions made by those of ordinary skill in the art to the technical solutions of the present invention shall fall within the protection scope determined by the claims of the present invention.

Claims

1. A succinic acid polymer, characterized in that: The general chemical structure formula of the succinic acid polymer is as follows: n is an integer from 1 to 10; m is a positive integer.

2. The succinic acid polymer according to claim 1, wherein: The number-average molecular weight of the succinic acid polymer is 5000 - 8000 Da.

3. The succinic acid polymer according to claim 1, wherein: The molecular weight distribution coefficient of the succinic acid polymer is 1.0 - 2.

0.

4. The preparation method of the succinic acid polymer according to any one of claims 1 to 3, characterized in that: The preparation method includes the following steps: Using polyethylene glycol-1000, glycerol, and succinic acid as raw materials, N,N'-dicyclohexylcarbodiimide as a dehydrating agent, and 4-dimethylaminopyridine as a catalyst, add them into a three-necked flask in sequence, using N,N'-dimethylformamide as a solvent, heat and react in a water bath at 60°C for 4 h; filter to remove impurities, precipitate with methyl tert-butyl ether, let stand overnight, filter off the supernatant to obtain a white waxy solid, which is the succinic acid polymer.

5. A derivative of the succinic acid polymer according to any one of claims 1 to 3, characterized in that: Named succinic acid acrylate polymer, the general chemical structure formula of the succinic acid acrylate polymer is as follows: n is an integer from 1 to 10; m is a positive integer.

6. The preparation method of the succinic acid acrylate polymer according to claim 5, characterized in that: Modify the succinic acid polymer with acryloyl chloride to prepare the succinic acid acrylate polymer.

7. The preparation method according to claim 6, characterized in that: The preparation method includes the following steps: Under the protection of an inert gas, using the succinic acid polymer and acryloyl chloride as raw materials, triethylamine as an acid-binding agent, and DMF as a solvent, react under ice bath conditions for 16 h; after the reaction, filter to remove impurities, crystallize, and filter to obtain a filter cake, which is the succinic acid acrylate polymer.

8. A bonding hydrogel, characterized in that: The adhesive hydrogel is a crosslinked network prepared by using the succinic acid acrylate polymer described in claim 5 as a crosslinking agent, and acrylic acid, dimethylaminoethyl methacrylate, and N-hydroxysuccinimide acrylate as crosslinking monomers. The crosslinking method is chemical crosslinking, and the crosslinked structure is a three-dimensional network structure.

9. The preparation method of the adhesive hydrogel according to claim 8, characterized in that: Dissolve the succinic acid acrylate polymer, acrylic acid, dimethylaminoethyl methacrylate, N-hydroxysuccinimide acrylate, and the photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphite in ultrapure water, mix well to obtain a uniform precursor solution, and further cure the precursor solution under ultraviolet light irradiation for 20 - 600 s to obtain the adhesive hydrogel.

10. Use of the adhesive hydrogel according to claim 8 in adhering biomaterials, glass, and metals, characterized in that, The application in the adhesive biological material includes the use in the preparation of materials for treating esophageal fistula.

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