Injectable amino acid temperature-sensitive hydrogel material as well as preparation method and application thereof

Abstract

The invention belongs to the field of high polymer materials, and discloses a material of injectable amino acid temperature-sensitive hydrogel as well as a preparation method and application of the injectable amino acid temperature-sensitive hydrogel, and the method comprises the following steps: dissolving DL-aspartic acid in heated deionized water to obtain a solution A, and then adding methacrylic anhydride to react to obtain a double-bond-containing DL-aspartic acid solution B; chitosan reacts with the solution B to obtain an MA-ASP-CS cross-linking agent, solid C is obtained after freeze drying, the solid C is taken out and reacts with N-isopropylacrylamide, and the hydrogel with the temperature-sensitive effect is obtained. The temperature-sensitive hydrogel disclosed by the invention has relatively good biodegradability and swelling property, and can be used as a drug carrier for targeted therapy of liver cancers.

Description

Technical Field

[0001] The invention belongs to the field of polymer materials, and carries out preparation of a thermosensitive hydrogel material and research on its performance, in particular to a material of injectable amino acid thermosensitive hydrogel and a preparation method and application thereof. Background Art

[0002] Hydrogel has very good hydrophilicity and is widely used in various fields such as medicine and diet. Hydrogel with a three-dimensional structure has the effect of efficient loading. The gaps in the body can be used as a box to wrap drugs. This feature makes it a good drug carrier. Thermosensitive hydrogel is a type of hydrogel. It mainly produces changes in its own structure through changes in the external temperature of the hydrogel. It is a solution when the temperature is low. When the temperature rises to the phase transition point, it will produce structural changes and become a gel. The drug is mixed with the hydrogel solution at low temperature. When injected for treatment, it is affected by the temperature change and becomes a gel to wrap the drug to achieve the effect of sustained drug release.

[0003] DL-aspartic acid is an amino dibasic acid, an acidic amino acid, and a non-essential amino acid in the human body. DL-aspartic acid is commonly found in biosynthesis, which can improve myocardial contractile function, reduce oxygen consumption, reduce the amount of nitrogen and carbon dioxide in the blood, enhance the function of the body, and eliminate fatigue. In medicine, it is generally used to treat heart disease, liver disease, and hypertension. It can be used with a variety of amino acids to prepare amino acid infusions, which are used as ammonia detoxifiers, liver function promoters, and fatigue recovery agents. As an amino acid, it has very good biocompatibility and is an excellent grafting object.

[0004] Chitosan is the only natural cationic polysaccharide with amino and hydroxyl groups in its structure. It has good hydrophilicity and biodegradability and can be used in medicine, agriculture, food, environmental protection and other fields. It is widely used. However, when it comes to drug sustained release, the mechanical properties and swelling properties of chitosan alone cannot meet its application, so it needs to be grafted and modified.

[0005] With the development of science and technology, more and more polymer materials are used in the medical field. Materials with good biocompatibility and biodegradability are commonly used in the medical field. However, the single performance of a single material cannot meet the requirements. Therefore, composite and grafting are widely used in this field. The modified polymer materials have multiple or unique characteristics, which make them very suitable for specific applications. The drug-loaded hydrogels, dressings, microspheres and other products currently used in the medical field are basically made by this method, so it is necessary to formulate specific materials according to application requirements and make specific adjustments to their performance. Summary of the invention

[0006] Compared with the existing technology, the present invention uses chitosan, DL-aspartic acid, and methacrylic anhydride to prepare a cross-linking agent containing unsaturated double bonds, which undergoes free radical polymerization with N-isopropylacrylamide to form a hydrogel. The hydrogel has good biocompatibility and biodegradability, and is temperature responsive.

[0007] Single N-isopropylacrylamide has poor degradation performance and will be harmful if it remains in the human body for a long time. In addition, its cross-linking agent NN-methylenebisacrylamide (BIS) is toxic. Therefore, the cross-linking agent prepared by chitosan, DL-aspartic acid and methacrylic anhydride is used to replace BIS to reduce toxicity and increase its biocompatibility and degradation performance.

[0008] In order to achieve the above object, the present invention provides a method for preparing an injectable amino acid thermosensitive hydrogel material, the method comprising the following steps:

[0009] (1) Dissolve DL-aspartic acid in deionized water at 75°C-90°C and stir evenly to obtain a 0.16 mol / L-0.5 mol / L solution A.

[0010] (2) Add 0.2 mol / L-0.6 mol / L methacrylic anhydride to solution A, heat in a water bath at 70°C-80°C or above, and protect from light for 2-4 hours to obtain solution B.

[0011] (3) 14.8 g / L-44.5 g / L chitosan was added to solution B, heated in a water bath at 40°C-60°C to obtain a solution, and freeze-dried to obtain solid C.

[0012] (4) 3.3 g / L-19.8 g / L of solid C was dissolved in deionized water and stirred, 0.29 mol / L-0.58 mol / L of N-isopropylacrylamide was added, and 1%-2% of the initiator was added, and the reaction was stirred for 6-8 hours to obtain DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel.

[0013] In the above technical solution, further, in the step (1), the purity of DL-aspartic acid is not less than 98%, and it is a white powder.

[0014] In the above technical solution, further, the purity of methacrylic anhydride in step (2) is not less than 94%.

[0015] In the above technical solution, further, the chitosan in step (3) has a purity of not less than 95%, a deacetylation degree of 85-99%, and a molecular weight of 800,000-1,000,000.

[0016] In the above technical solution, further, the purity of N-isopropylacrylamide in step (4) is not less than 98%, and it is a light yellow crystalline powder.

[0017] In the above technical solution, further, in the step (4), the stirring rate is 300-600 r / min, the water bath heating temperature is 40° C.-60° C., and the stirring time is 6-8 h.

[0018] In the above technical solution, further, the initiator described in step (4) is one of (ammonium persulfate (APS), azobisisobutyronitrile (AIBN), dibenzoyl peroxide (BPO)).

[0019] The present invention replaces the toxic cross-linking agent with a self-made chitosan-DL-aspartic acid-methacrylic anhydride cross-linking agent with good biocompatibility, introduces amino acids such as DL-aspartic acid to enhance the biocompatibility and biodegradability of the hydrogel, and also improves its mechanical properties. Chitosan and DL-aspartic acid are connected through amide bonds, and then react with methacrylic anhydride to achieve the purpose of introducing double bonds, and finally react with N-isopropylacrylamide to form a thermosensitive hydrogel.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] Chitosan has good biocompatibility and adhesion, as well as good bactericidal properties. Using it as a hydrogel material can greatly increase the water swelling properties of the hydrogel. It has good adhesion when injected into the body and can stably adhere to the surface of the target to achieve the effect of stable drug release. DL-aspartic acid, as an amino acid, has excellent hydrophilicity. Its grafting introduction can increase the biocompatibility and degradation properties of the hydrogel, reduce the degradation time and the safety of the degradation products.

[0022] The preparation method of the present invention does not generate pollutants and harmful substances, and its production process is simple. The prepared polymer hydrogel material has good biocompatibility and degradability, and has temperature sensitivity. The intended use is the delivery and administration of cancer, which provides convenience in medical treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 This is the infrared spectrum of the DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel prepared in Example 1;

[0024] Figure 2 This is a sample picture of the DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel prepared in Example 2;

[0025] Figure 3Contact angle characterization diagram of DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel with different cross-linking degrees. DETAILED DESCRIPTION

[0026] The following examples are provided to facilitate a better understanding of the present invention, but are not intended to limit the present invention. In addition, in the following examples, unless otherwise specified, all materials and reagents used can be purchased from biological or chemical reagent companies.

[0027] Example 1

[0028] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0029] (2) Then, 1.77 mL of methacrylic anhydride solution was added to step 1 and reacted for 1-2 h at a stirring rate of 500 r / min.

[0030] (3) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 2, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0031] (4) Take 0.1 g of the freeze-dried product in step 3 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0032] After the experiment, the infrared absorption of the hydrogel was measured by potassium bromide tablet method. Figure 1 This is the infrared spectrum of the DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel prepared in Example 1. It can be seen from this that Figure 1 1690cm in (a) -1 、1700cm -1 It is the C=O stretching vibration absorption peak of aspartic acid, 2910cm -1 -NH 2 The absorption peak of Figure 1 2800-2900cm in (b) -1 The broad peak between local regions is the absorption peak of the OH bond of chitosan, 2330 cm -1 The medium-strong peak is the -NH 2 + The absorption peak of Figure 1 (c) 2980cm-1 It is the -NH 2 The absorption peak is 1460cm -1 Thermosensitive hydrogel -CH 3 The in-plane vibration peak of the bending -1 The strong peak is the absorption peak of the hydrogel C=CC=O structure, 1540cm -1 It is the absorption peak of O=C-NH, and it can be seen that the amidation reaction occurred, indicating the success of the hydrogel grafting modification.

[0033] Figure 2 This is a contact angle characterization diagram of the DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel with different crosslinking agents prepared in Example 1. It can be seen from the figure that the hydrogel exhibits good hydrophilic properties as a whole.

[0034] Figure 3 The DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel prepared in Example 1 was prepared from Figure 3 It can be seen that the hydrogel is endowed with temperature-sensitive properties, being in the state of a solution at low temperatures and in the state of a gel when heated.

[0035] Example 2

[0036] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0037] (2) Then, 1.77 mL of methacrylic anhydride solution was added to step 1 and reacted for 1-2 h at a stirring rate of 500 r / min;

[0038] (3) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 2, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0039] (4) Take 0.1 g of the freeze-dried product in step 3 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0040] Example 3

[0041] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0042] (2) Then, 1.77 mL of methacrylic anhydride solution was added to step 1 and reacted for 1-2 h at a stirring rate of 500 r / min;

[0043] (3) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 2, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0044] (4) Take 0.1 g of the freeze-dried product in step 3 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0045] Example 4

[0046] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0047] (2) Then, 1.77 mL of methacrylic anhydride solution was added to step 1 and reacted for 1-2 h at a stirring rate of 500 r / min;

[0048] (3) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 2, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0049] (4) Take 0.1 g of the freeze-dried product in step 3 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0050] Example 5

[0051] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0052] (2) Then, 1.77 mL of methacrylic anhydride solution was added to step 1 and reacted for 1-2 h at a stirring rate of 500 r / min;

[0053] (3) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 2, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0054] (4) Take 0.1 g of the freeze-dried product in step 3 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0055] Example 6

[0056] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0057] (2) Then, 1.77 mL of methacrylic anhydride solution was added to step 1 and reacted for 1-2 h at a stirring rate of 500 r / min;

[0058] (3) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 2, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0059] (4) Take 0.1 g of the freeze-dried product in step 3 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0060] Example 7

[0061] (1) Pipette 1.77 mL of methacrylic anhydride solution into 60 mL of deionized water.

[0062] (2) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 1, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0063] (3) Take 0.1 g of the freeze-dried product in step 2 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The mixture was reacted for 6-8 hours under the atmosphere, and after cross-linking, methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel was obtained.

[0064] Example 8

[0065] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0066] (2) Pipette 1.77 mL of methacrylic anhydride solution into the solution in step 1, react at 40°C for 7-8 hours, then take out and freeze-dry in a watch glass.

[0067] (3) Take 0.1 g of the freeze-dried product in step 2 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The reaction was carried out under atmosphere for 6-8 hours without gel formation.

[0068] Example 9

[0069] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0070] (2) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 1, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0071] (3) Take 0.1 g of the freeze-dried product in step 2 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 2 g of N-isopropylacrylamide and 0.021 g of initiator ammonium persulfate. 2 The reaction was carried out under atmosphere for 6-8 hours without gel formation.

[0072] Example 10

[0073] (1) Weigh 1.33 g of 98% pure DL-aspartic acid, dissolve it in 60 mL of 90°C deionized water, stir for 30 min, and then cool the temperature to 70°C;

[0074] (2) Weigh 0.89 g of chitosan with a degree of deacetylation of ≥95% and a molecular weight of 1 million, add it to the solution in step 1, react at 40° C. for 7-8 hours, then take it out and freeze-dry it in a watch glass.

[0075] (3) Take 0.1 g of the freeze-dried product in step 2 and add it to a three-necked flask with 30 mL of deionized water. After ultrasonic dissolution, add 0.021 g of initiator ammonium persulfate and heat at 40-60 ° C and N 2 The reaction was carried out under atmosphere for 6-8 hours without gel formation.

[0076] The above-described embodiments are only preferred embodiments of the present invention, but not all feasible embodiments of the present invention. For those skilled in the art, any obvious changes made thereto without departing from the principles and spirit of the present invention should be considered to be included in the scope of protection of the claims of the present invention.

Claims

1. A method for preparing an injectable amino acid thermosensitive hydrogel material, characterized in that: The method comprises the following steps: firstly dissolving DL-aspartic acid in heated deionized water to obtain solution A, then adding methacrylic anhydride to react to obtain DL-aspartic acid solution B containing double bonds; reacting chitosan with solution B to obtain MA-ASP-CS crosslinking agent, which is freeze-dried to obtain solid C, and taking out solid C to react with N-isopropylacrylamide to obtain a hydrogel with a temperature-sensitive effect.

2. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 1, characterized in that: It includes the following steps: (1) dissolving DL-aspartic acid in deionized water and stirring uniformly to obtain a 0.16 mol / L-0.5 mol / L solution A; (2) adding 0.2 mol / L-0.6 mol / L methacrylic anhydride to solution A and reacting for 2-4 hours in the dark to obtain solution B; (3) adding 14.8 g / L-44.5 g / L chitosan to solution B, dissolving and reacting for 6-8 hours, and freeze-drying to obtain solid C; (4) 3.3 g / L-19.8 g / L of solid C was dissolved in deionized water and stirred, 0.29 mol / L-0.58 mol / L of N-isopropylacrylamide was added, and 1%-2% of the total mass fraction of the initiator was added, and the reaction was stirred to obtain DL-aspartic acid / methacrylic anhydride / chitosan / N-isopropylacrylamide thermosensitive hydrogel.

3. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 1, characterized in that: The temperature of the deionized water is 75°C-90°C.

4. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 2, characterized in that: The reaction in step (2) is maintained in a water bath heated at 70°C-80°C or above.

5. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 1, characterized in that: During the dissolution in step (3), the mixture is heated in a water bath at 40°C-60°C.

6. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 2, characterized in that: The stirring rate in step (4) is 300-600 r / min.

7. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 2, characterized in that: In step (4), the water bath heating temperature is 40°C-60°C.

8. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 2, characterized in that: The reaction time in step (4) is 6-10h.

9. The method for preparing the injectable amino acid thermosensitive hydrogel material according to claim 2, characterized in that: The initiator in step (4) is one of ammonium persulfate, azobisisobutyronitrile and dibenzoyl peroxide.

10. An application of an injectable amino acid thermosensitive hydrogel material, characterized in that: As a drug carrier for targeted therapy of liver cancer.

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