Fe 3+ Preparation method and application of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel

Abstract

The present invention discloses a Fe 3+ Preparation method and application of catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel. The preparation method is specifically as follows: (1) preparing quaternary ammonium chitosan; (2) preparing catechol-modified quaternary ammonium chitosan; (3) preparing oxidized carboxymethyl cellulose; (4) preparing catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose composite hydrogel. The present invention also discloses Fe prepared by the method. 3+ Application of a catechol-modified quaternary ammonium chitosan-oxidized hydroxymethyl cellulose double-crosslinked hydrogel. The double-crosslinked hydrogel designed by the present invention does not require the addition of additional chemical reagents. The final product is only water and hydrogel, which does not require purification and is environmentally friendly. The prepared composite hydrogel has good viscoelasticity and strong adhesion, and has great application prospects in wound healing and wound treatment.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical and functional materials technology, and specifically relates to an Fe 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel. Background Art

[0002] Since trauma has a high incidence rate, promoting wound healing has become a key scientific issue that needs to be urgently addressed in the medical community today. Wound dressings can protect wounds from bacterial infection and play an important role in accelerating the wound healing process. In recent years, the hydrophilic polymer network structure of hydrogels has given them advantages such as high hydrophilicity, good biocompatibility, and transparency (facilitating observation of wound healing), making them ideal materials for wound dressings. Although hydrogels are considered to be ideal materials for wound dressings, the hydrogel materials currently under development have many problems, such as poor mechanical strength, poor solubility, and high cytotoxicity, which limit their scope of application.

[0003] Chitosan and carboxymethyl cellulose, as relatively abundant biomacromolecules in nature, are widely used in wound dressings due to their excellent biocompatibility, biodegradability, non-toxicity, and low cost. Carboxymethyl cellulose-based hydrogels, however, are often considered an ideal drug corrosion inhibitor due to their excellent biocompatibility and easy degradation. However, chitosan suffers from poor water solubility and low drug loading efficiency, while carboxymethyl cellulose-based hydrogels exhibit poor strength, brittleness, and high water absorption. Previous studies have shown that combining chitosan and carboxymethyl cellulose can produce hydrogels with improved performance. However, existing chitosan-carboxymethyl cellulose hydrogels exhibit large surface pores, leading to significant drug loss during loading. While small molecule crosslinkers such as glutaraldehyde and epichlorohydrin can increase drug loading efficiency, the crosslinkers themselves have certain biotoxicity. Furthermore, single-crosslinked hydrogels exhibit weak elasticity, long gelation times, and insufficient adhesion strength, making them unsuitable for dynamic wounds such as those at joints, significantly hindering their practical application.

[0004] The self-healing behavior of hydrogels is primarily achieved through reversible physical non-covalent interactions and chemical covalent bonds. Physical crosslinking forms weaker bonds, but it forms bonds and reestablishes dynamic equilibrium more quickly. Physically crosslinked hydrogels have weak mechanical strength and are not suitable for dynamic trauma. Chemical crosslinking can form stronger bonds, but it forms bonds and reestablishes dynamic equilibrium more slowly. Summary of the Invention

[0005] Purpose of the invention: In view of the problems existing in the prior art, the present invention provides a Fe 3+- Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel.

[0006] Another object of the present invention is to provide a Fe 3+ -Application of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel in wound dressing field.

[0007] Technical solution: In order to achieve the above purpose, the present invention provides a Fe 3+ A method for preparing a catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double-crosslinked hydrogel comprises the following steps:

[0008] (1) Preparation of quaternary ammonium chitosan: dissolving chitosan in deionized water and glacial acetic acid solution, slowly adding different amounts of modifier to obtain a crude quaternary ammonium chitosan solution, filtering, separating, purifying, and drying to obtain quaternary ammonium chitosan (QCS) with different degrees of substitution;

[0009] (2) Preparation of catechol-modified quaternary ammonium chitosan: quaternary ammonium chitosan was dissolved in deionized water, and catechol (CHA) was added to obtain QCS / CHA solution. Then, an ethanol solution containing a catalyst was added dropwise to the QCS / CHA solution. After dialysis and freeze-drying, catechol-modified quaternary ammonium chitosan (QCS-C) was obtained.

[0010] (3) Preparation of oxidized carboxymethyl cellulose: dissolve carboxymethyl cellulose in deionized water, add sodium periodate, react under acidic conditions in the dark, add ethylene glycol, and then dialyze. Place the solution in an ultra-low temperature freezer to freeze, thereby obtaining oxidized carboxymethyl cellulose.

[0011] (4)Fe 3+ Preparation of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel: Ferric chloride solution was added to oxidized carboxymethyl cellulose solution, and then mixed with catechol-modified quaternary ammonium salt chitosan solution to obtain Fe 3+ -Catechol modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel.

[0012] Furthermore, in the step (1), the modifier is trimethyl ammonium chloride, and the mass ratio of the modifier to chitosan is 1 to 3:1, so as to synthesize quaternary ammonium salt chitosan with low, medium and high degree of substitution.

[0013] Furthermore, the degree of substitution of the quaternary ammonium salt chitosan prepared in step (1) is determined by titration.

[0014] Furthermore, the catalyst in step (2) is 1-(3-dimethylaminopropyl)-3-ethyldiimide (EDC).

[0015] Furthermore, the dialysis in step (2) is performed in a HCl solution with a pH of 4.5 to 5.0 for 3 to 5 days, and then in deionized water for 1 to 3 days.

[0016] Furthermore, in step (3), the mass ratio of sodium periodate to carboxymethyl cellulose is 1 to 3:1.

[0017] Furthermore, in step (4), the mass ratio of catechol-modified quaternary ammonium salt chitosan to oxidized carboxymethyl cellulose is 1-3:3-6.

[0018] The present invention provides a preparation method for preparing Fe 3+ -Catechol modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel.

[0019] The present invention also provides a Fe 3+ -Application of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel in biomedical field and wound dressing.

[0020] As a preference, the Fe prepared by the present invention 3+ -Catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose double-crosslinked hydrogel for dynamic burn wound healing.

[0021] Fe 3+ The formation mechanism of the catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose double-crosslinked hydrogel is as follows: Glycidyl trimethylammonium chloride (GTMAC) reacts with chitosan, and the quaternary ammonium groups replace the H atoms on the chitosan amino groups, forming quaternary ammonium chitosan. Using the catalyst 1-(3-dimethylaminopropyl)-3-ethyldiimine (EDC), the quaternary ammonium chitosan reacts with catechol, where the catechol groups further replace the H atoms on the quaternary ammonium chitosan amino groups, ultimately forming catechol-modified quaternary ammonium chitosan. Carboxymethyl cellulose undergoes an oxidation reaction with sodium periodate, converting the hydroxyl groups on the carboxymethyl cellulose into aldehyde groups. The nucleophilic addition reaction is carried out by oxidized carboxymethyl cellulose containing aldehyde groups and quaternary ammonium chitosan modified with catechol containing amino groups. The nucleophilic reagent is the quaternary ammonium chitosan modified with catechol. The nitrogen atom with a lone electron pair in the compound structure attacks the carbon atom with a positive charge on the carbonyl group to complete the nucleophilic addition reaction and form an intermediate α-hydroxyamine compound, which is then further dehydrated to form a Schiff base. When the Schiff base is formed, ferric chloride solution is added to react with the catechol modified quaternary ammonium chitosan to form Fe 3+ -Catechol modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel.

[0022] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0023] 1. The preparation method of the present invention greatly improves Fe by optimizing and controlling parameters. 3+ -The mechanical properties of catechol-modified quaternary ammonium chitosan-oxidized hydroxymethyl cellulose double cross-linked hydrogel. The prepared double cross-linked hydrogel has stronger compression performance and better elasticity, and can reach 90% without damage.

[0024] 2. C=N is a dynamic covalent bond, and the reversible equilibrium reaction between imine bonds does not require any external stimulation. Hydrogels containing catechol groups have excellent tissue adhesion properties. Hydrogels with good interfacial adhesion can establish stable connections with dynamic wounds, thereby resisting damage and destruction.

[0025] 3. The present invention is a process of Schiff base reaction and catechol-Fe 3+ The physicochemical double-crosslinked hydrogel prepared by chelating bonds combines the advantages of non-covalent bonds and covalent bonds, which can make the pores smaller, more conducive to drug loading and controlled release, with fast gelation speed, enhanced mechanical properties, and strong tissue adhesion. The hydrogel is wear-resistant, has enhanced mechanical properties, and strong tissue adhesion, and is suitable for dynamic trauma treatment.

[0026] 4. The preparation method of the present invention prepares a composite hydrogel containing catechol-quaternary ammonium salt chitosan / oxidized carboxymethyl cellulose with different degrees of substitution and oxidation through Schiff base reaction. When the oxidation degree of oxidized carboxymethyl cellulose is 9.3 mmol / g and the substitution degree of quaternary ammonium salt chitosan is 34.14%, the prepared hydrogel has the best performance, and the Fe 3+ The coordination with catechol forms a double-cross-linked hydrogel, which does not require the addition of additional chemical reagents. The final product is only water and hydrogel, which does not require purification and is environmentally friendly. The prepared composite hydrogel has good viscoelasticity and strong adhesion, and has good application prospects in wound healing and wound treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 This is the H NMR spectrum of quaternary ammonium chitosan.

[0028] Figure 2 This is the H NMR spectrum of catechol-modified quaternary ammonium chitosan.

[0029] Figure 3 The infrared spectra of chitosan and quaternary ammonium salt chitosan.

[0030] Figure 4 The infrared spectra of carboxymethyl cellulose and oxidized carboxymethyl cellulose.

[0031] Figure 5 This is the UV spectrum of catechol quaternary ammonium salt chitosan.

[0032] Figure 6 Fe 3+-Infrared spectrum of catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose double-crosslinked hydrogel.

[0033] Figure 7 Comparison of the mechanical properties of single-crosslinked and double-crosslinked hydrogels. DETAILED DESCRIPTION

[0034] The present invention will be further described below with reference to the accompanying drawings and examples.

[0035] The experimental methods described in the examples are conventional methods unless otherwise specified.

[0036] Example 1

[0037] Preparation of quaternary ammonium salt chitosan

[0038] Weigh 1.0g of chitosan, dissolve it in 36mL of deionized water and 180μL of glacial acetic acid solution, stir for 30min, and then slowly drop different amounts of triglycerol trimethylammonium chloride (GTMAC, Adamas) into the mixture of chitosan and glacial acetic acid. The mass ratio of GTMAC to chitosan is 1:1, 2:1 and 3:1, respectively. Subsequently, the reaction is carried out at 55°C for 18 hours to obtain aqueous solutions of crude quaternary ammonium chitosan products with different degrees of substitution. After filtering through a Buchner funnel, the products are separated and purified by ethanol precipitation, dehydrated with acetone three times, and then dried in a vacuum oven at 45°C for 3 days to finally obtain quaternary ammonium chitosan (QCS) pure products with low, medium and high degrees of substitution; Figure 1 The following is the nuclear magnetic hydrogen spectrum of quaternary ammonium salt chitosan. The trimethylammonium and -NH-CH2- protons of quaternary ammonium salt chitosan have two characteristic peaks at 3.1ppm and 4.3ppm. Figure 3 The infrared spectra of chitosan and quaternary ammonium salt chitosan are shown, 1480 cm -1 The deformation vibration absorption peak of -CH3 of the quaternary ammonium group is located at , indicating that the preparation is successful.

[0039] Example 2

[0040] Determination of Substitution Degree of Quaternary Ammonium Salt Chitosan

[0041] The degree of substitution of quaternary ammonium salt chitosan was determined by titration. 100 mg of quaternary ammonium salt chitosan with different degrees of substitution was accurately weighed and dissolved in 25 mL of deionized water. The pH of the solution was adjusted with NaOH (controlled between 8.2 and 9.6). 1 mL of 8% potassium chromate was used as an indicator and added to each quaternary ammonium salt chitosan solution with different degrees of substitution. The solution was then slowly titrated with 0.05 mol / L silver nitrate. The titration endpoint was when the solution turned brick red. The degree of substitution DS was calculated using the following formula.

[0042]

[0043] The calculated low, medium and high substitution degrees of quaternary ammonium chitosan are 15.23%, 34.14% and 50.74% respectively.

[0044] Example 3

[0045] Catechol-modified quaternary ammonium chitosan

[0046] 0.5 g of the quaternary ammonium chitosan (QCS) prepared in Example 1 with a degree of substitution of 15.23%, 34.14% and 50.74% was weighed and dissolved in 75 mL of deionized water. The pH was adjusted to 5.0 with 2 mol / L HCl to dissolve the chitosan completely. Next, 0.43 g of catechol (CHA) was added to the quaternary ammonium chitosan solution to obtain a QCS / CHA solution. A 50 mL ethanol solution containing 0.36 g of 1-(3-dimethylaminopropyl)-3-ethyldiimine (EDC) was added dropwise to the QCS / CHA solution. The reaction was allowed to react at room temperature for 12 h with a pH maintained at 4.5. After the reaction was complete, the mixture was dialyzed in a HCl solution at a pH of 4.5 for 3 days and then dialyzed in deionized water for 1 day. The mixture was freeze-dried to obtain catechol-modified quaternary ammonium chitosan (QCS-C). Figure 2 As shown in the H NMR spectrum of catechol-modified quaternary ammonium salt chitosan, there are characteristic peaks between 6.5-7.0 ppm, indicating that catechol chitosan was successfully prepared. Figure 5 As shown in FIG. 1 , the UV spectrum of catechol quaternary ammonium salt chitosan is shown. The catechol content is calculated to be 0.009 mg / mL according to the UV fitting relationship y=6.51638E-4+85.59733x.

[0047] Example 4

[0048] Preparation of oxidized carboxymethyl cellulose

[0049] Weigh 1.5g of carboxymethyl cellulose, dissolve it in 150mL of water, and add 1.5g, 3g, and 4g of sodium periodate respectively. After reacting in the dark for 3 hours at 40℃ and pH 3.0, add ethylene glycol (0.39mL, 0.78mL, 1.05mL) in an amount equimolar to the initial sodium periodate and react at 40℃ for 1 hour to terminate the reaction. Then, dialyze for 3 days using a dialysis bag with a barrier relative molecular weight of 3500, and freeze the solution in an ultra-low temperature freezer for 1 day to obtain oxidized carboxymethyl cellulose with different degrees of substitution. Figure 4 The infrared spectra of carboxymethyl cellulose and oxidized carboxymethyl cellulose are shown in Figure 1. -1 and 885cm -1 The absorption peaks of aldehyde carbonyl and hemiacetal are shown in Figure 2. Compared with the infrared spectrum of carboxymethyl cellulose, the absorption peak of oxidized carboxymethyl at 1650 cm -1The carbonyl absorption peak becomes stronger, indicating that the hydroxyl groups in the carboxymethyl cellulose molecules have been oxidized to aldehyde groups.

[0050] Example 5

[0051] Preparation for determination of oxidation degree in oxidized carboxymethyl cellulose

[0052] The degree of oxidation of oxidized carboxymethyl cellulose is determined based on the Schiff base reaction of aldehyde groups with hydroxylamine hydrochloride. 0.1 g of oxidized carboxymethyl cellulose with different degrees of substitution is fully dispersed in 50 mL of hydroxylamine hydrochloride in acetic acid buffer (0.2 mol / L, pH = 4.5). After reacting for 24 hours at room temperature, the mixture is titrated with 0.1 mol / L standard NaOH solution, using carboxymethyl cellulose as a blank control. The degree of oxidation (DO) of oxidized carboxymethyl cellulose is calculated according to the following formula:

[0053]

[0054] The calculated oxidation degrees of oxidized carboxymethyl cellulose (CMC) were 6.7 mmol / g, 9.3 mmol / g and 10.4 mmol / g, respectively.

[0055] Example 6

[0056] Fe 3+ Preparation of catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel

[0057] 1.2 g of oxidized carboxymethyl cellulose with oxidation degrees of 6.7 mmol / g, 9.3 mmol / g and 10.4 mmol / g were dissolved in 12 mL of deionized water, and the pH of the solution was adjusted to 10 by adding NaOH (2 mol / L), and then FeCl3 solution (2 mol / L) was added. 0.4 g of catechol-modified quaternary ammonium salt chitosan prepared in Example 3 was dissolved in 4 mL of deionized water. The above two solutions were mixed to prepare FeCl3 solution. 3+ -Catechol modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel. Figure 6 As shown, Fe 3+ -In the infrared spectrum of catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, the peak at 1656 cm -1 There is an imine bond at the site, proving that the double-cross-linked hydrogel was successfully synthesized.

[0058] Example 7

[0059] Mechanical comparison of double cross-linked hydrogel and catechol-modified quaternary ammonium chitosan-oxidized carboxymethyl cellulose single cross-linked hydrogel

[0060] The mechanical properties of the hydrogels were evaluated through compression testing using a universal materials testing machine (WDW3020, China). Oxidized carboxymethyl cellulose (OCMC) with an oxidation degree of 9.3 mmol / g and quaternary ammonium chitosan with a degree of substitution of 34.14% exhibited the best performance, so these OCMC and quaternary ammonium chitosan were selected for hydrogel preparation. The hydrogel samples were prepared in a cylindrical shape with a base diameter of 22 mm and a height of 20 mm. The compression test speed was 6 mm / min, and compressive stress-strain curves were obtained at strains ranging from 10% to 90%. Figure 7 As shown, the double-crosslinked hydrogel has stronger compression performance and better elasticity, and can achieve 90% without damage, while the single-crosslinked hydrogel is damaged at 78%. This shows that the use of double-crosslinking technology can make the pores smaller, which is more conducive to drug loading and controlled release.

Claims

1. A kind of Fe 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: The steps include: (1) Preparation of quaternary ammonium chitosan: dissolving chitosan in deionized water and glacial acetic acid solution, slowly adding different amounts of modifier to obtain a crude quaternary ammonium chitosan solution, filtering, separating, purifying, and drying to obtain quaternary ammonium chitosan (QCS) with different degrees of substitution; (2) Preparation of catechol-modified quaternary ammonium chitosan: quaternary ammonium chitosan was dissolved in deionized water, and catechol (CHA) was added to obtain QCS / CHA solution. An ethanol solution containing a catalyst was then added dropwise to the QCS / CHA solution. After dialysis and freeze-drying, catechol-modified quaternary ammonium chitosan (QCS-C) was obtained. (3) Preparation of oxidized carboxymethyl cellulose: dissolve carboxymethyl cellulose in deionized water, add sodium periodate, react under acidic conditions in the dark, add ethylene glycol, and then dialyze. Freeze the solution in an ultra-low temperature freezer to obtain oxidized carboxymethyl cellulose; (4) Fe 3+ Preparation of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel: Ferric chloride solution was added to oxidized carboxymethyl cellulose solution, and then mixed with catechol-modified quaternary ammonium salt chitosan solution to obtain Fe 3+ -Catechol modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel.

2. Fe according to claim 1 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: In the step (1), the modifier is trimethyl ammonium chloride, and the mass ratio of the modifier to chitosan is 1-3:1, so as to synthesize quaternary ammonium salt chitosan with low, medium and high degree of substitution.

3. Fe according to claim 1 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: The degree of substitution of the quaternary ammonium salt chitosan prepared in the step (1) is determined by titration.

4. Fe according to claim 1 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: The catalyst in step (2) is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC).

5. Fe according to claim 1 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: The dialysis in step (2) is performed in a HCl solution with a pH of 4.5 to 5.0 for 3 to 5 days, and then in deionized water for 1 to 3 days.

6. Fe according to claim 1 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: In the step (3), the mass ratio of sodium periodate to carboxymethyl cellulose is 1-3:

1.

7. Fe according to claim 1 3+ - Preparation method of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel, characterized in that: In the step (4), the mass ratio of catechol-modified quaternary ammonium salt chitosan to oxidized carboxymethyl cellulose is 1-3:3-6.

8. Fe prepared by the preparation method according to claim 1 3+ -Catechol modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel.

9. Fe according to claim 8 3+ -Application of catechol-modified quaternary ammonium salt chitosan-oxidized carboxymethyl cellulose double cross-linked hydrogel in the preparation of wound dressing.

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