Polysaccharide-based self-gel powder adhered to wet tissue and preparation method of polysaccharide-based self-gel powder

Abstract

The invention discloses tissue adhesion self-gel powder and a preparation method thereof, and belongs to the field of hydrogel. The preparation method comprises the following steps: firstly, modifying chitosan to obtain quaternized chitosan (QCS), and then grafting urushiol onto the QCS to obtain urushiol-gold quaternized chitosan (US-QCS); the preparation method comprises the following steps: reacting sodium hyaluronate (HA) with calcium salt to obtain calcium hyaluronate (Ca-HA); and mixing the US-QCS and the Ca-HA, so as to obtain US-QCS / Ca-HA self-gel powder. The tissue adhesion self-gel powder prepared by the invention can quickly absorb interface water and then adhere to tissues, various wounds can be blocked, wound healing is promoted, calcium ions in the powder can accelerate blood coagulation, and the powder has the potential ability of promoting bone repair.

Description

Technical Field

[0001] The invention relates to a polysaccharide-based self-gelling powder with wet tissue adhesion and a preparation method thereof, belonging to the field of medical materials. Background Art

[0002] Tissue-adhesive hydrogels have been a research hotspot in the field of biomedical materials in recent years. They have demonstrated significant advantages in wound closure, hemostasis, drug delivery, and tissue repair. However, interfacial water hinders effective contact and bonding between hydrogels and tissues. Research has developed adhesive materials that can absorb or remove interfacial water, thereby forming a close contact with tissues. Zhao Xuanhe's team prepared a polyacrylic acid / N-succinimidyl ester double-sided tape based on a "dry cross-linking" mechanism. The dry tape rapidly absorbs interfacial water, hydrating and swelling, rapidly forming covalent bonds and strong adhesion to the tissue surface (Hyunwoo Yuk, Claudia E. Varela, Christoph S. Nabzdyk, et al. Dry double-sided tape for adhesion of wet tissues and devices. Nature, 2019, 169–174). The powder obtained by drying and crushing the polyelectrolyte coacervate can quickly absorb water and expand, connecting with each other to form a blocky hydrogel. At the same time, it forms close contact with the substrate and diffuses into the substrate network, thereby enhancing wet adhesion (Xin Peng, Xianfeng Xia, Xiayi Xu, et al. Ultrafast self-gelling powder mediates robust wet adhesion to promote healing of gastrointestinal perforations. Science Advance, 2021, eabe8739).

[0003] However, previously reported self-gelling powders rely solely on absorbing interfacial water to enhance interfacial adhesion. When there is a large amount of moisture on the surface, the powders are unable to fully absorb the interfacial water or absorb too much water, resulting in decreased mechanical properties and interfacial adhesion of the resulting hydrogel. By introducing hydrophobic groups into the self-gelling powder, while the powder absorbs interfacial water and forms a hydrogel, the hydrophobic effect of the hydrophobic groups rapidly removes the interfacial water and inhibits the swelling of the hydrogel, effectively improving the mechanical properties and interfacial adhesion of the hydrogel.

[0004] Chitosan (CS) is a naturally occurring aminopolysaccharide polymer. Quaternization of chitosan effectively improves its water solubility and antimicrobial activity. The enhanced charge makes it more susceptible to reacting with oppositely charged polyanions (such as hyaluronic acid) to form polyelectrolyte aggregates. The present invention modifies quaternized chitosan (QCS) with urushiol to produce urushiol-quaternized chitosan (US-QCS), effectively enhancing its hydrophobic interaction and adhesion properties. Calcium hyaluronate (Ca-HA), a derivative of hyaluronic acid, is more stable than common sodium hyaluronate (HA). The presence of calcium ions accelerates coagulation and enhances osteogenic activity. US-QCS / Ca-HA self-gelling powder, which exhibits excellent wet tissue adhesion, can be used for rapid hemostasis, wound healing, and accelerated bone defect repair. Summary of the Invention

[0005] The method first modifies chitosan (CS) to obtain quaternized chitosan (QCS), which is then modified with urushiol to obtain urushiol-quaternized chitosan (US-QCS). Then, sodium hyaluronate (HA) is modified to obtain calcium hyaluronate (Ca-HA). US-QCS and Ca-HA are mixed in a specific ratio, precipitated, and freeze-dried to obtain a US-QCS / Ca-HA self-gelling powder.

[0006] The first object of the present invention is to provide a method for preparing a polysaccharide-based self-gelling powder having wet tissue adhesion, comprising the following steps: S1: Preparation of QCS. CS was dissolved in deionized water containing glacial acetic acid, reacted with 2,3-epoxypropyltrimethylammonium chloride (GTMAC), precipitated with acetone, dialyzed to remove small molecule impurities, and then freeze-dried to obtain the product QCS. S2: Preparation of US-QCS. Dissolve QCS in deionized water, add ammonium persulfate (APS), and react under nitrogen. Dissolve urushiol in ethanol and add to the QCS solution. Heat under nitrogen. Wash away any unreacted urushiol with ethanol, then dry under vacuum to yield US-QCS. S3: Preparation of Ca-HA. Dissolve the calcium salt in ethanol, adjust the pH with aqueous acetic acid, and then add HA to react. Wash with ethanol and perform gradient dehydration. Finally, vacuum dry to obtain the product, Ca-HA. S4: Preparation of self-gelling powder: US-QCS and Ca-HA were mixed to obtain US-QCS / Ca-HA self-gelling powder.

[0007] As a preferred embodiment of the present invention, the molecular weight of CS in step S1 is 100-1000 kDa and the degree of deacetylation is ≥90%. As a preferred embodiment of the present invention, the molecular weight of HA in step S3 is 200-2000 kDa.

[0008] As a preferred embodiment of the present invention, the calcium salt in step S3 is one or more of calcium chloride, calcium citrate, and calcium nitrate.

[0009] As a preferred embodiment of the present invention, the mass ratio of US-QCS to Ca-HA in step S4 is 1:5 to 5:1.

[0010] The present invention has the following beneficial effects: (1) The self-gelling powder prepared by the present invention is rapidly self-gelled by the electrostatic action of quaternary ammonium salt and carboxylate.

[0011] (2) The self-gelling powder prepared by the present invention has good biocompatibility and no obvious toxic side effects.

[0012] (3) The self-gelling powder prepared by the present invention can simultaneously absorb and repel interfacial water and has good wet tissue adhesion.

[0013] (4) The self-gelling powder prepared by the present invention can seal various wounds.

[0014] (5) The self-gelling powder prepared by the present invention can accelerate blood coagulation, promote wound healing, and has the application potential of promoting bone defect repair. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a photo of the polysaccharide-based self-gel powder prepared in Example 1 of the present invention adhered to pig skin after hydration. The hydrogel formed after hydration can still adhere to the pig skin after being twisted or even washed with water. DETAILED DESCRIPTION

[0016] The technical solutions of the present invention will be described in detail below with reference to specific embodiments. It should be noted that the following embodiments are only preferred implementation examples of the present invention, which are intended to help understand the core ideas and implementation methods of the present invention and do not constitute a limitation on the scope of protection. Based on the basic principles of the present invention, all technical solutions obtained by those skilled in the art through routine experimental means or reasonable technical extensions on the basis of a full understanding of the contents of this disclosure are within the scope of protection of the claims of the present invention.

[0017] Example 1 (1) Preparation of QCS: 3 g of CS was dissolved in 160 ml of deionized water containing 0.5% v / v glacial acetic acid, and 5.65 g of GTMAC was dissolved in 5 ml of deionized water. The mixture was then added to the CS solution and stirred at 55 °C for 18 h. The mixture was precipitated with acetone, dialyzed, and freeze-dried to obtain the product QCS. (2) Preparation of US-QCS: Dissolve 1 g of QCS in 100 mL of deionized water, then add 0.3 g of sodium dodecyl sulfate (SDS). Then add 0.25 g of APS and react under nitrogen for 15 min. Dissolve 5 g of urushiol in 25 ml of ethanol and then add it to the QCS solution. Under nitrogen, react at 60 °C for 6 h. Wash away the unreacted urushiol with ethanol, then dry in vacuum to obtain the product US-QCS. (3) Preparation of Ca-HA: 10 g of calcium chloride was dissolved in 240 g of 80% ethanol and the pH was adjusted to 5.6 with acetic acid. 2 g of HA was then added and stirred for 12 h, followed by a 2-h pause and an additional 12 h of stirring. After the reaction, the mixture was washed twice with 80% ethanol (pH 5.6), then dehydrated three times with 99% ethanol, and finally dried under vacuum to obtain the product Ca-HA.

[0018] (4) Preparation of self-gelling powder: US-QCS and Ca-HA were mixed in a mass ratio of 1:1 and ground to obtain US-QCS / Ca-HA self-gelling powder.

[0019] Example 2 The amount of urushiol in step (2) of Example 1 was adjusted to 10 g, and the rest remained the same as in Example 1.

[0020] Example 3 The mass ratio of US-QCS to Ca-HA in step (4) of Example 1 was adjusted to 2:1, and the rest remained the same as in Example 1.

[0021] Example 4 The mass ratio of US-QCS to Ca-HA in step (4) of Example 1 was adjusted to 1:2, and the rest remained the same as in Example 1.

[0022] Comparative Example 1 CS and HA were mixed in a mass ratio of 1:1 to obtain CS / HA self-gelling powder.

[0023] Comparative Example 2 QCS and HA were mixed in a mass ratio of 1:1 to obtain QCS / HA self-gelling powder.

[0024] Comparative Example 3 QCS and Ca-HA were mixed in a mass ratio of 1:1 to obtain QCS / Ca-HA self-gelling powder.

[0025] Performance Testing The self-gelling powders obtained in the examples and comparative examples were subjected to performance tests, and the test results are as follows: Self-gelling time: The self-gelling powder prepared in the present invention is mixed with water, and the self-gelling time is measured by an inversion method.

[0026] Table 1 shows the self-gelling time. It can be seen from Table 1 that the self-gelling time of Examples 1-3 and the comparative example is extremely short, which meets the requirements of clinical application.

[0027] Table 1 example Example 1 Example 2 Example 3 Example 4 Comparative Example 1 Comparative Example 2 Comparative Example 3 Self-gel time (s) 6±2 5±2 5±3 20±4 5±2 4±2 4±1 Adhesion performance: The self-gel powder was hydrated and applied to pig skin. A lap shear test was performed using a universal testing machine at a tensile speed of 10 mm / min.

[0028] Table 2 shows the adhesion performance test results. It can be seen from Table 2 that the adhesion strength of the samples of Examples 1-4 exceeds that of the comparative example.

[0029] Table 2 example Example 1 Example 2 Example 3 Example 4 Comparative Example 1 Comparative Example 2 Comparative Example 3 Adhesion strength (kPa) 38±5 40±4 36±4 28±3 20±4 22±3 21±4 Cytotoxicity: Mouse fibroblast cell line L929 was used as a biocompatibility evaluation model. The extracts of the self-gelling powders of the examples and comparative examples were co-cultured with the cells for different time periods, and CCK-8 was used to detect cell activity.

[0030] The samples of Examples 1-4 and Comparative Examples 1-3 all have good cell compatibility.

[0031] It should be noted that the preferred embodiments disclosed in this specification are only illustrative examples of the technical solutions of the present invention and do not constitute a limitation on the scope of protection of the patent right. Any person skilled in the art, on the premise of fully understanding the technical concept of the present invention, may make equivalent substitutions, reorganize technical features or adjust process parameters for the above-mentioned implementation schemes. Such technical variations, if they do not deviate from the substantial technical features stated in the claims, shall be deemed to be within the legal protection scope of the patent right of the present invention. The legal protection boundary of the present invention shall be based on the content of the claims authorized by the State Intellectual Property Office. Any non-substantial changes based on the technical solution of the present invention shall not be able to oppose the statutory protection effect of the patent right.

Claims

1. A method for preparing tissue-adhesive self-gelling powder, characterized in that: The steps include: (1) Chitosan (CS) was modified with 2,3-epoxypropyltrimethylammonium chloride to obtain quaternized chitosan (QCS), and quaternized chitosan was modified with urushiol to obtain urushiol-quaternized chitosan (US-QCS); (2) reacting sodium hyaluronate (HA) with calcium salt to obtain calcium hyaluronate (Ca-HA); (3) US-QCS and Ca-HA were mixed and ground to obtain US-QCS / Ca-HA self-gel powder.

2. The method according to claim 1, characterized in that The molecular weight of the water CS in step (1) is 100-1000 kDa, and the degree of deacetylation is ≥90%.

3. The method according to claim 1, characterized in that The molecular weight of HA in step (2) is 200-2000 kDa.

4. The method according to claim 1, wherein The calcium salt in step (2) is one or more of calcium chloride, calcium citrate and calcium nitrate.

5. The method according to claim 1, characterized in that The mass ratio of US-QCS to Ca-HA in step (3) is 1:5 to 5:

1.

6. A method according to any one of claims 1 to 5, wherein the self-gelling powder is prepared.

7. The self-gelling powder according to claim 6 can be used in medical fields such as tissue adhesion, hemostasis, drug delivery, and tissue repair.

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