CN111848855A

CN111848855A - Injectable hydrogel dressing with pH response and preparation method and application thereof

Info

Publication number: CN111848855A
Application number: CN202010632292.6A
Authority: CN
Inventors: 郭保林; 初大可; 何刚; 贺佳辉
Original assignee: Xian Jiaotong University
Current assignee: Xian Jiaotong University
Priority date: 2020-07-03
Filing date: 2020-07-03
Publication date: 2020-10-30

Abstract

The invention discloses an injectable hydrogel dressing with pH response and a preparation method and application thereof. The double-bonded amino acid or the double-bonded amino acid derivative is subjected to esterification reaction by-COOH of the double-bonded amino acid or the double-bonded amino acid derivative and-OH on NHS (N-hydroxysuccinimide) to prepare the tissue cross-linking agent monomer. The preparation method disclosed by the invention is simple to operate, the used raw materials are easy to obtain and low in price, and the prepared hydrogel dressing has good adhesiveness, pH responsiveness, injectability, self-healing performance and hemostatic performance, can effectively promote gastric hemostasis and wound healing after endoscopic treatment, and has a great application value in the fields of gastric hemostasis and wound healing.

Description

Injectable hydrogel dressing with pH response and preparation method and application thereof

Technical Field

The invention belongs to the technical field of degradable biomedical materials, and relates to an injectable hydrogel dressing with pH response, and a preparation method and application thereof.

Background

Gastrointestinal cancer is one of the most common malignancies worldwide. Although surgical treatment, chemotherapy and immunotherapy of gastrointestinal cancer have advanced in recent decades, gastrointestinal cancer still constitutes a major cause of death for related cancer patients due to its aggressiveness and metastasis (a. poursham, s.g.sephanlou, k.s.ikuta, c.bisignano, s.safiri, g.roshandel, m.sharf, m.khatibian, c.fitzmauure, m.r. Nixon, The lancet Gastroenterology & hepatogolology 2019,4, 934; a.etemaddi, s.safiri, s.g.sepalon, k.ikuta, c.bisignano, r.shakeri, m.amani, c.fitzmamnix, m.nitz, m.nixon, n.absient, The lancet & 5. Endoscopic submucosal resection (EMR) and Endoscopic Submucosal Dissection (ESD) are recognized therapies for gastrointestinal tumors, and these endoscopic techniques have many advantages such as being more invasive than surgical procedures, having a radical resection effect, and being able to preserve physiological gastrointestinal structures. Complications following EMR and ESD surgery, such as bleeding and perforation, will result in further complications. These complications may lead to prolonged hospital stays, additional treatments, high morbidity, and even life-threatening (m.banks, n.uedo, p.bhandari, t.gotoda, Gut2019, gutjnl; p.j.basford, r.george, e.nixon, t.chaudhuri, r.mead, p.bhandari, Surgical endipair 2014,28, 1594; y.zhu, j.x.xu, j.cheng, z.zhang, b.q.zhu, t.y.chen, x.xu, y.wang, m.y.cai, p.h.zhou, United European gasterlogue 2019,7, 782). In addition, following endoscopic treatment, infection and physicochemical irritation can also exacerbate chronic inflammation, slowing the extracellular matrix (ECM) degradation and wound healing process. Although many studies have focused on ideal wound dressing materials that accelerate coagulation, tissue formation, angiogenesis and re-epithelialization, to address the problem of rapid bleeding and promote wound healing after endoscopic treatment. But no endoscope sprayable material is available to effectively promote healing of gastric wounds after endoscopic treatment.

In situ injectable hydrogels have fluidity prior to gelation, can be injected into a wound site without causing gel fragmentation, undergo a transition or respond to changes in pH and temperature in a short time, form hydrogels in situ, and rapidly crosslink with tissue. And then integrated in the form of a whole gel to heal the wound.

Given the long-term use of hydrogels in gastric wound healing (e.g., bleeding following EMR/ESD surgery may develop after 2 weeks) and the long-term gastric motility (approximately 1000 times per day, 5-10kPa each time; gastric motility, dilation, contraction may damage hydrogels) (x.liu, c.steiger, s.lin, g.a.parada, j.liu, h.f.chan, h. Yuk, n.v.phan, j.collins, s.tamang, Nature communications 2019, 10.). Thus, the basic requirements of a hydrogel sprayed around a gastric wound are: first, the hydrogel is required to withstand prolonged mechanical forces from the stomach; second, the hydrogel is required to be self-healing after being subjected to external mechanical forces; third, the incidence of bleeding and perforation in the stomach will be increased due to the gastric acid environment and loss of mucus-bicarbonate and postoperative mucosal barrier. Therefore, the hydrogel is required to have stable pH response performance, and to provide a suitable microenvironment to accelerate the healing process of the wound after endoscopic treatment; fourth, hydrogels are required to have stable and long-term adhesive properties to prevent them from falling off the wound site, and thus provide better and longer protection of the wound from stomach acid. However, the existing hydrogel can not completely meet the above requirements, and therefore, there is a great need to develop a new hydrogel which can be used in the fields of gastric hemostasis and wound healing.

Disclosure of Invention

In order to overcome the defects of the prior art, the invention aims to provide an injectable hydrogel dressing with pH response, and a preparation method and application thereof, so as to solve the technical problems that the existing hydrogel has poor adhesion in an acidic environment, has insufficient hemostatic and healing promoting performances and is difficult to apply in the fields of gastric hemostasis and wound healing.

In order to achieve the purpose, the invention adopts the following technical scheme to realize the purpose:

a preparation method of an injectable hydrogel dressing with pH response comprises the steps of directly mixing a polymer monomer, a tissue cross-linking agent monomer and a cross-linking agent for reaction for 200-800 s to obtain the injectable hydrogel dressing with pH response; the tissue cross-linking agent monomer is obtained by reacting double-bonded amino acid or double-bonded amino acid derivative with NHS, and the cross-linking agent is N, N' -methylene bisacrylamide;

when the polymer monomer is any one of double-bonded polysaccharide, double-bonded polysaccharide derivative, double-bonded gelatin and double-bonded gelatin derivative, the mass ratio of the polymer monomer to the tissue cross-linking agent monomer to the cross-linking agent is (10-20): (0-20): (0.1 to 0.6); when the polymer monomer is double-bonded amino acid or double-bonded amino acid derivative, the mass ratio of the polymer monomer to the tissue cross-linking agent monomer to the cross-linking agent is (70-110): (0-20): (0.1-0.6).

Preferably, the concentration of the double-bonded amino acid and the double-bonded amino acid derivative is 100-200 mg/mL; the concentrations of the double-bonded polysaccharide, the double-bonded polysaccharide derivative, the double-bonded gelatin and the double-bonded gelatin derivative are 10-20 mg/mL; the tissue cross-linking agent monomer is prepared by taking dimethyl sulfoxide as a solvent, and the concentration of the tissue cross-linking agent monomer is 0-400 mg/mL.

Preferably, the double-bonded amino acid and the double-bonded amino acid derivative are prepared by the following steps:

1) according to the feeding ratio of (15-20) g of amino acid, deionized water and sodium hydroxide: (100-140) mL: (4-12) preparing an amino acid solution;

according to the proportion of (15-20) g of amino acid derivatives, deionized water and sodium hydroxide: (100-140) mL: (4-12) g preparing an amino acid derivative solution;

2) according to the volume ratio of acryloyl chloride to tetrahydrofuran (8-24): (20-24) preparing an acryloyl chloride solution;

3) according to the molar ratio of acryloyl chloride to amino acid (1.0-1.4): (0.8-1.2), adding the acryloyl chloride solution into the amino acid solution, mixing and reacting for 18-32 h, and extracting, precipitating and drying to obtain the double-bonded amino acid derivative.

According to the molar ratio of acryloyl chloride to amino acid derivative (1.0-1.4): (0.8-1.2), adding the acryloyl chloride solution into the amino acid derivative solution, mixing and reacting for 18-32 h, and extracting, precipitating and drying to obtain the double-bonded amino acid derivative.

Further preferably, the amino acid is glycine;

further preferably, the amino acid derivative is 4-aminobutyric acid, 6-aminocaproic acid or 8-aminocaprylic acid;

further preferably, the extraction process described in step 3) is carried out by ethyl acetate solution; the volume ratio of the amino acid polymer precursor solution or the amino acid derivative polymer precursor solution to ethyl acetate is (20-50): (60-150).

Preferably, the double-bonded polysaccharide is obtained by double-bonded grafting reaction of any one of carboxymethyl chitosan, carboxyethyl chitosan, aminated hyaluronic acid and aminated sodium alginate; the double-bonded polysaccharide derivative is obtained by performing double-bonded grafting reaction on a chitosan quaternary ammonium salt derivative; the double-bonded gelatin derivative is obtained by double-bonded grafting reaction of quaternized gelatin or aminated gelatin.

Preferably, a catalyst is also added in the mixing reaction process; the catalyst is a mixture of APS and TEMED, and the volume ratio of N, N' -methylene bisacrylamide to the mixture of APS and TEMED is (150-250): (75-125): (75-125); the concentration of N, N' -methylene bisacrylamide is 1-4 mg/mL, the concentration of APS is 40-60 mg/mL, and the concentration of TEMED is 5-20 muL/mL.

Preferably, the preparation process of the tissue cross-linking agent monomer comprises:

1) using acetone as a solvent, preparing a double-bonded amino acid solution or a double-bonded amino acid derivative solution with the concentration of 100-150 mg/mL, and mixing the double-bonded amino acid solution or the double-bonded amino acid derivative solution with NHS according to the ratio of (100-150): (110-160), and stirring for 20-60 min to obtain a mixed solution;

2) the mass ratio of NHS to DCC is (110-160): (220-320), adding DCC into the mixed solution obtained in the step 1), and reacting for 18-36 hours to obtain a tissue cross-linking agent monomer precursor solution;

3) sequentially filtering, concentrating, filtering and drying the tissue cross-linking agent monomer precursor solution obtained in the step 2) to obtain the tissue cross-linking agent monomer.

Further preferably, the concentration in the step 3) is to concentrate the filtered tissue cross-linking agent monomer precursor solution to 30-60 mL; after concentration, adding 300-480 mL of petroleum ether into the concentrated solution, standing until the precipitation is complete, and then sequentially performing suction filtration and drying.

The injectable hydrogel dressing with the pH response function is of a porous net structure, and the pore diameter is 95-156 microns.

Use of an injectable hydrogel dressing with pH response in the field of gastric hemostasis and wound healing.

Compared with the prior art, the invention has the following beneficial effects:

the invention discloses a preparation method of an injectable hydrogel dressing with pH response. Any one of double-bonded polysaccharide, double-bonded polysaccharide derivative, double-bonded gelatin, double-bonded amino acid and double-bonded amino acid derivative is used as a polymer monomer, most of carboxyl at the tail end of a side chain can be protonated under an acidic environment, and then hydrogen bonds are formed between other tail carboxyl or amido on an interface; the double-bonded amino acid or the double-bonded amino acid derivative is subjected to esterification reaction between-COOH of the double-bonded amino acid or the double-bonded amino acid derivative and-OH on NHS (N-hydroxysuccinimide) to prepare a tissue cross-linking agent monomer, and the tissue cross-linking agent monomer can be used as a micro cross-linking agent and can be used for being capable of being in contact with-NH (nitrogen) of a cell membrane of a wound part₂The groups are subjected to covalent crosslinking to form amido bonds (-CO-NH-), thereby improving the adhesion of the hydrogel to gastric mucosa. Meanwhile, the tissue cross-linking agent monomer is a hydrophobic monomer, hydrophobic interaction exists, namely reversible non-covalent interaction, and the hydrophobic interaction can promote self-repairing of the dressing. The hydrogel dressing is placed in an environment with certain pH, and the equivalent series resistance value is changed in the process of changing the pH The change is small, and the hydrogel dressing prepared by the method is proved to have good pH response characteristics and can maintain the stability of the self structure in an acidic environment. Meanwhile, the hydrogel dressing has the same adhesive strength as a commercial fibrin glue adhesive, and has stable and long-term adhesive performance so as to prevent the hydrogel dressing from falling off from a wound part, thereby providing better and longer protection for the wound and avoiding the wound from being invaded by gastric acid. The preparation method is simple to operate, the used raw materials are easy to obtain and low in price, and the prepared hydrogel dressing has good adhesiveness, pH responsiveness, injectability, self-healing performance and hemostatic performance, and can effectively promote gastric hemostasis and wound healing after endoscopic treatment.

Further, in the preparation method of the present invention, based on 6-aminocaproic acid (a), acryloyl chloride (a) is grafted onto 6-aminocaproic acid through amidation reaction, and double-bonding grafting reaction is performed to obtain acryloyl 6-Aminocaproic Acid (AA) monomer, which can be used as a main raw material of a hydrogel dressing. On this basis, acryloyl-6-aminocaproic acid-g-N-hydroxysuccinimide (AA-NHS) monomer was prepared by esterification reaction. And carrying out crosslinking reaction on the obtained AA-NHS monomer and AA in a physiological environment, and taking Bis as a crosslinking agent to obtain the injectable hydrogel dressing (AA/AA-NHS) with pH response. 6-aminocaproic acid, a main component in the AA/AA-NHS hydrogel dressing, is used as an antifibrotic solvent and is proved to be used for promoting the healing of a wound surface;

Furthermore, APS/TEMED is used as a catalytic system, N, N' -methylene bisacrylamide is used as a cross-linking agent, and the content of the catalyst TEMED in the hydrogel dressing is adjusted to enable the hydrogel dressing to show proper gelling time, so that the hydrogel dressing is endowed with proper gelling time, has good injectability, can be administered under more convenient and simplified conditions, and is suitable for emergency treatment of severe gastrorrhagia in emergency situations.

The invention discloses an injectable hydrogel dressing with pH response, which is obtained based on the preparation method and has the following advantages:

(1) has effective and stable viscosityAdhesive properties because the hydrogel dressing contains NHS activated amino acids or amino acid derivatives as tissue cross-linker monomers with-NH of the cell membrane of the wound site₂The groups are covalently crosslinked to form amido bonds (-CO-NH-); hydrogel dressing adhesive strength can also be enhanced by the formation of hydrogen bonds between the hydrogel dressing and the tissue. Thus, these two actions together impart effective and stable adhesive properties to the hydrogel dressing.

(2) The hydrogel dressing has quick self-healing performance, because the side chain of the main component polymer monomer of the hydrogel dressing has proper length and flexibility, and the carboxyl at the tail end of the side chain can be mostly protonated under an acid environment, and then hydrogen bonds are formed between other tail carboxyl or amide on the interface; in addition, the tissue cross-linker monomer is a hydrophobic monomer, and the presence of hydrophobic interactions therein also promotes self-healing of the hydrogel dressing. Thus, the two actions jointly endow the hydrogel dressing with quick and strong self-healing performance; the rapid self-healing capability allows for self-healing of the hydrogel dressing due to the damage to the hydrogel dressing caused by the tension forces caused by gastric motility, expansion and contraction, and together with the tensile strength and strong adhesion of the hydrogel dressing, extends the useful life of the hydrogel dressing.

(3) The hydrogel dressing has pH response performance, because in a low pH value solution, carboxyl in the hydrogel dressing can form intramolecular hydrogen bonds, so that the swelling behavior of the hydrogel dressing is reduced, and the obvious pH response performance is further shown.

(4) Has good hemostatic performance because the hydrogel dressing has stable structure and excellent adhesive performance and can be bonded with-NH of cell membranes of wound parts₂The covalent crosslinking of the groups forms amido bond (-CO-NH-) to improve the adhesive strength of the gel at the wound part, and the gel forming time is shorter.

(5) With injectability, the precursor solution of the hydrogel dressing can be prepolymerized for about 3 minutes and then still injectable when transferred to a 23g endoscope.

Because the hydrogel dressing prepared by the invention has good performance of promoting the healing of the gastric mucosa wound, the hydrogel dressing can be firmly adhered to the wound partAct as a barrier to protect the wound from the external environment, protect the wound site from bacterial infection, maintain a moisture microenvironment, and allow the presence of oxygen, which may promote hydrogel dressings to effectively promote gastric wound healing by controlling inflammation. I.e. capable of providing a suitable microenvironment to accelerate the healing process of the wound following endoscopic treatment; the carboxyl at the tail end of the side chain can be protonated in an acidic environment, then other tail carboxyl can form hydrogen bonds with amide groups, and the hydrogel dressing contains a tissue cross-linking agent monomer which can be in contact with-NH of cell membranes of wound sites ₂The groups are covalently cross-linked. Namely, the injectable hydrogel dressing has high-efficiency and stable adhesive property, and has quick and good self-healing property in an acid environment. Therefore, the hydrogel dressing prepared by the invention can be sprayed to a stomach wound area through an endoscope without causing gel fragmentation to form the hydrogel dressing in situ, is quickly crosslinked with tissues and is stably and long-term adhered to the stomach wound area, and the hydrogel dressing has stable structure, excellent adhesion performance and short gel forming time, so the hydrogel dressing shows good hemostatic performance. In addition, the hydrogel dressing is able to withstand long-term mechanical forces from the stomach and self-heal after being subjected to external mechanical forces, which in turn may provide better, longer protection for the wound and provide a suitable microenvironment to accelerate the healing process of the wound after endoscopic treatment. Therefore, the injectable hydrogel dressing with pH response has great application value in the fields of stomach hemostasis and wound healing.

Drawings

FIG. 1 is AA¹H NMR chart;

FIG. 2 is of AA-NHS¹H NMR chart;

FIG. 3 is a graph of storage modulus versus time measured on a rheometer for a hydrogel dressing AA/AA-NHS made in accordance with the invention;

FIG. 4 is a self-healing performance of a hydrogel dressing AA/AA-NHS hydrogel dressing made in accordance with the present invention, wherein the rheometer measured storage modulus, loss modulus versus stress plot (a); storage modulus, loss modulus versus time plot (b);

FIG. 5-1 is a scanning electron microscope microscopic morphology image of the AA/AA-NHS hydrogel dressing prepared by the invention after freeze-drying after simulated artificial gastric juice (pH 2.0) swelling equilibrium, with a ruler: 150 μm, wherein (a) is AA/AA-NHS0, (b) is AA/AA-NHS5, (c) is AA/AA-NHS10, (d) is AA/AA-NHS15, (e) is AA/AA-NHS 20;

fig. 5-2 is a pore size statistical graph of the hydrogel dressing after swelling equilibrium (artificial gastric fluid, pH 2) of the AA/AA-NHS hydrogel dressing prepared according to the present invention;

fig. 5-3(a) is the equilibrium swelling ratio of AA/AA-NHS hydrogel dressing prepared according to the present invention at 37 ℃ (PBS, pH 7.4); FIGS. 5-3(b) are graphs showing the equilibrium swelling ratio of AA/AA-NHS hydrogel dressings prepared in accordance with the present invention at 37 deg.C (artificial gastric fluid, pH 2.0);

FIG. 6 shows the adhesion strength of the AA/AA-NHS hydrogel dressing prepared by the present invention on the pig stomach matrix;

FIG. 7 is a test of the hemolytic rate of the AA/AA-NHS hydrogel dressing prepared by the present invention for mouse blood cells;

FIG. 8 is a cell viability test of the AA/AA-NHS hydrogel dressing prepared by the present invention for mouse fibroblast (L929);

FIG. 9-1 is a histological observation of AA/AA-NHS hydrogel dressing after 7 days of subcutaneous implantation in rats;

FIG. 9-2 is a toluidine blue stain observation of AA/AA-NHS hydrogel dressing 7 days after subcutaneous implantation in rats;

FIG. 10 shows statistical and quantitative data of thickness of inflammatory fibers in an implanted region 7 days after subcutaneous implantation of an AA/AA-NHS hydrogel dressing prepared according to the present invention in a rat;

FIG. 11(a) is the amount of bleeding of the AA/AA-NHS hydrogel dressing prepared by the present invention in a mouse liver wound model; FIG. 11(b) is the amount of bleeding of the AA/AA-NHS hydrogel dressing prepared by the present invention in a mouse liver incision model hemostasis test; FIG. 11(c) is the amount of bleeding of the AA/AA-NHS hydrogel dressing prepared by the present invention in the mouse tail-broken model hemostasis test;

FIG. 12 shows the application of the AA/AA-NHS hydrogel dressing prepared by the present invention to a pig gastrorrhagia model; wherein, fig. 12(a) is a pig gastrorrhagia model observed by an endoscope; FIG. 12(b) is a spray tube for gastroscopy to the bleeding site; FIG. 12(c) shows the hydrogel dressing adhered to the stomach wall in a pig; FIG. 12(d) shows hydrogel dressing adhered to the outer stomach wall of a pig;

fig. 13 is the delayed bleeding rate (n-4) for the different treatment groups;

FIG. 14 is a graph of wound healing performance in a porcine gastric ESD model on day 21 for an AA/AA-NHS hydrogel dressing injected, esomeprazole, and a blank group; (a) histological observation of wound regenerated tissue for day 21; (b) histological observations of wound type I collagen staining on day 21; (c) histological observations for alpha-smooth muscle actin staining (alpha-SMA) on day 21; (d) histological observations of CD34 staining for wound day 21;

FIG. 15 is a graph showing the results of AA/AA-NHS hydrogel injection, esomeprazole and blank group, (a) is a statistical result of the relative number of inflammatory cells in the wound-regenerating tissue on day 21; (b) semi-quantitative statistics for wound type I collagen on day 21; (c) semi-quantitative statistics for alpha-smooth muscle actin staining (alpha-SMA) at day 21; and (d) statistics of the revascularization status of wound on day 21.

Detailed Description

The invention is described in further detail below with reference to the accompanying drawings:

example 1

1) Preparation of doubly-bonded amino acid derivative, acryloyl 6-Aminocaproic Acid (AA):

19.8g of 6-aminocaproic acid were added to 120ml of deionized water, and 6.6g of sodium hydroxide was added to the solution. Vigorously stirred in an ice bath until 6-aminocaproic acid was dissolved. 13.35mL of acryloyl chloride was added to 22.5mL of tetrahydrofuran solution, followed by dropwise addition of the acryloyl chloride solution to the aminocaproic acid solution. Adjusting and maintaining the pH value at 7.5-7.8, and reacting for 24 hours. After the reaction was completed, the mixture was extracted 3 times with ethyl acetate (100 mL/time). The combined organic phases were collected. The aqueous phase was acidified to pH 3 and extracted again with ethyl acetate 3-5 times (100 mL/time). All the extracted organic phases were collected and combined, dried over sodium sulfate for 18 hours, and then filtered with suction using a buchner funnel. And performing rotary evaporation to concentrate the organic phase to 50mL, adding 400mL of petroleum ether into the concentrated organic phase, performing suction filtration and vacuum drying after complete precipitation to obtain acryloyl 6-Aminocaproic Acid (AA), and performing repeated precipitation to further purify to obtain the double-bonded amino acid derivative AA.

2) Preparation of tissue crosslinker monomer, acryloyl-6-aminocaproic acid-g-N-hydroxysuccinimide monomer (AA-NHS):

1.0g of AA (acryloyl 6-aminocaproic acid) monomer in step 1) was dissolved in 15mL of acetone, and the solution was pale yellow. Then, 1.24g of NHS (N-hydroxysuccinimide) was added thereto, and after stirring for 20 to 60 minutes, 2.7g of DCC (dicyclohexylcarbodiimide) was added to obtain a mixed solution, and the mixed solution was allowed to stand at room temperature for 24 hours. And after the reaction is finished, filtering the mixed solution, carrying out rotary evaporation and concentration on the obtained solution to 50mL, adding 400mL of petroleum ether into the concentrated solution, and carrying out suction filtration and vacuum drying after complete precipitation to obtain the AA-NHS monomer serving as the tissue cross-linking agent.

3) Preparation of injectable hydrogel dressing with pH response:

preparing 650 mu L of AA (acryloyl 6-aminocaproic acid) into 154mg/mL suspension by using deionized water, adding 33.26mg of sodium hydroxide into the suspension, vigorously stirring until the AA is completely dissolved, adding 50 mu L of dimethyl sulfoxide without AA-NHS, preparing 200 mu L of Bis (N, N' -methylene-bisacrylamide) into 2.5mg/mL solution by using deionized water, preparing 100 mu L of APS (ammonium persulfate) into 50mg/mL solution by using deionized water, preparing 100 mu L of TEMED (tetramethylethylenediamine) into 10 mu L/mL solution by using deionized water, and mutually crosslinking for 300-480 seconds at the temperature of 25 ℃ to obtain the injectable hydrogel dressing AA/AA-NHS0 with pH response.

Example 2

Diluting the AA concentration in the step 3 to 146mg/mL, and adding 50 mu L of 100mg/mL AA-NHS solution prepared by dimethyl sulfoxide. Otherwise, the AA/AA-NHS5 hydrogel dressing was obtained as in example 1.

Example 3

The AA/AA-NHS10 hydrogel dressing was obtained by diluting the AA concentration in step 3 to 138mg/mL and adding 50. mu.L of 200mg/mL AA-NHS solution thereto under the same conditions as in example 2.

Example 4

The AA/AA-NHS15 hydrogel dressing was obtained by diluting the AA concentration in step 3 to 130mg/mL and adding 50. mu.L of 300mg/mL AA-NHS solution thereto under the same conditions as in example 2.

Example 5

The AA/AA-NHS20 hydrogel dressing was obtained by diluting the AA concentration in step 3 to 122mg/mL and adding 50. mu.L of 400mg/mL AA-NHS solution thereto under the same conditions as in example 2.

The raw material composition of the hydrogel dressings prepared in examples 1 to 5 is shown in table 1:

TABLE 1 addition amount of raw material components of hydrogel dressings of examples 1 to 5

The kinds of the substances selected and the amounts added in the production processes of the hydrogel dressings of examples 6 to 20 are shown in tables 2 and 3, and the rest is the same as in example 1.

TABLE 2 compositions of raw materials and amounts thereof added in tissue cross-linking agent monomers of examples 6-20

Remarking: a2 is glycine; a4 is aminobutyric acid; a6 is aminocaproic acid; a8 is aminocaprylic acid.

Table 3 addition amount of raw material components of hydrogel dressings of examples 6 to 20

Remarking: CSG is double-bond chitosan; QCSG is double-bond quaternary ammonium salt chitosan derivative; CMCG is double-bonded carboxymethyl chitosan; CECG is double-bond connected carboxyethyl chitosan; CSAG is double-bond-linked aminated chitosan, GTG is double-bond-linked gelatin; QGTG is double-bonded quaternized gelatin; GTAG is double-bond-linked aminated gelatin; HAA is double-bond aminated hyaluronic acid; SAA is double-bond aminated sodium alginate.

The structural formula of AA required by the invention is shown as the following formula

The structural formula of AA-NHS required by the invention is shown as the following formula

Nmr hydrogen spectra of injectable dressings with pH response prepared in example 1 and example 2: (¹H NMR), the results are shown in fig. 1 and fig. 2, and the analytical patterns can be obtained: in AA¹In H NMR, the characteristic peak at 12.00ppm was assigned to the AA carboxyl hydrogen atom, and the ratio of the number of protons (g-protons) in acryloyl chloride (a-proton) to 6-aminocaproic acid (g-proton) integrated intensity was 1: 2, indicating that each 6-aminocaproic acid monomer chain is linked to acryloyl chloride. In AA-NHS¹In H NMR, the 12.00ppm carboxyl proton peak disappeared, while the newly formed peak appeared at 2.81ppm, corresponding to the proton of NHS (j-proton). In addition, the integrated intensity ratio of protons to g protons for AA-NHS was 4: 2, the result shows that carboxyl in AA and hydroxyl in NHS are completely subjected to esterification reaction, and the successful synthesis of AA-NHS monomers is shown. Above that ¹H NMR results prove that AA, AA-NHS monomers are successfully synthesized.

The storage modulus of the hydrogel dressings AA/AA-NHS prepared in examples 1-5 was determined using a TA rheometer (DHR-2) with the experimental procedure that the storage modulus (G') of the AA/AA-NHS hydrogel was evaluated by time scanning at 37 ℃ with a constant strain of 1% and a constant frequency of 10 rad/s. 300 μ L of the hydrogel polymer solution was placed between 20mm parallel plates with a gap of 1000 μm. As shown in FIG. 3, it can be seen from FIG. 3 that the storage modulus of the hydrogel samples is continuously increased with the increase of the AA-NHS content in the gel, and the storage moduli of the hydrogel dressings AA/AA-NHS0, AA/AA-NHS5, AA/AA-NHS10, AA/AA-NHS15 and AA/AA-NHS20 prepared in examples 1-5 are 130Pa, 490Pa, 580Pa, 650 Pa and 760Pa, respectively.

The hydrogel dressing AA/AA-NHS10 prepared in example 3 was studied for storage modulus and loss modulus using the following experimental procedures: first a sample of AA/AA-NHS10 hydrogel with a diameter of 20mm and a thickness of 1mm was prepared, and then the values of the critical strain region were recorded by performing a strain amplitude scanning method (gamma from 1% to 2500%). Other hydrogel disks were then used to evaluate self-healing performance by performing an alternating stepped strain sweep test at a fixed angular frequency (10 rad/s). The amplitude oscillatory strain switches from a small strain (γ ═ 1.0%) to a subsequent large strain (γ ═ 2000%), each strain interval being 100 s. The hydrogel sample was found to reach the critical point of gel and solution state of the gel when 1900% of the external force was applied, as shown in fig. 4 (a). When 2000% strain force is applied to the hydrogel, the storage modulus of the hydrogel is lower than the loss modulus of the hydrogel, the hydrogel structure collapses, and when the strain force is reduced to 1%, the storage modulus of the hydrogel rapidly rises again, the hydrogel structure returns to normal, and after 5 cycles, the hydrogel still can show good self-healing performance, which indicates that the self-healing performance of the hydrogel is stable and excellent, as shown in fig. 4 (b).

The hydrogel dressings prepared in examples 1 to 5 were subjected to microscopic morphology characterization, and Scanning Electron Microscope (SEM) pictures thereof are shown in fig. 5-1, and it was observed that the AA/AA-NHS hydrogel dressings had a uniform and interconnected network structure. After swelling equilibration (artificial gastric fluid, pH 2), the pore size histogram of the hydrogel dressing is shown in fig. 5-2, with diameters of about 156 μm, 145 μm, 113 μm, and 95 μm for AA/AA-NHS5, AA/AA-NHS10, AA/AA-NHS15, and AA/AA-NHS20, respectively.

The hydrogel dressings prepared in examples 1 to 5 had an equilibrium swelling ratio in PBS (pH 7.4) and artificial gastric juice (pH 2.0) solutions at 37 ℃, and as a result, as shown in fig. 5-3(a) and 5-3(b), all of the hydrogel dressings were swollen sharply (ESR increased from 4.3 to 6.7) at pH 7.4 and ESR was increased from 0.3 to 0.9 at pH 2.0, respectively. The AA/AA-NHS hydrogel dressing has good water absorption capacity in a neutral environment; can maintain the stability of the structure under the acidic environment. Namely, the hydrogel dressing prepared by the invention has good pH corresponding performance.

The hydrogel dressings prepared in examples 1 to 5 were tested for adhesive strength, and the experimental procedure was to select a fresh pig stomach as the adherend. Fresh pig stomach was made into a rectangular block of 1x 3cm in size, 100 μ L of AA/AA-NHS hydrogel polymer solution was injected onto the surface of the pig stomach, and then another pig stomach was placed on the hydrogel solution. The bonding area was 10X 10 mm. Lap shear testing was performed using a lap shear test on an Instron material testing system (mtscririon 43, MTS Criterion) equipped with a 50N weighing sensor at a speed of 5 mm/min.

As shown in FIG. 6, the adhesive strengths of the hydrogels AA/AA-NHS0, AA/AA-NHS5, AA/AA-NHS10, AA/AA-NHS15 and AA/AA-NHS20 were about 2.19kPa, 2.33kPa, 6.63kPa, 7.70kPa and 5.16kPa, respectively. The adhesive strength of the hydrogel was gradually increased by increasing the content of AA-NHS in the hydrogel. AA/AA-NHS15 showed the highest adhesive strength. The hydrogel prepared in examples 3-5 showed ideal adhesive strength with commercial fibrin glue adhesive

(about 5kPa) maintained a comparable strength.

The blood compatibility of the hydrogels prepared in examples 1-5 was evaluated by obtaining erythrocytes by centrifuging mouse blood (1000rpm) for 10 minutes, diluting to a final concentration of 5% (v/v) for use, and mixing 500. mu.L of erythrocyte suspension with 500. mu.L of AA/AA-NHS hydrogel in a 24-well plate. After standing at 37 ℃ for 1h with an oscillation speed of 100rpm, 500. mu.L of fresh PBS was added to each well of the hydrogel-filled microplate. All microplate contents were centrifuged at 1000rpm for 10 minutes to remove non-hemolytic erythrocytes. 100 μ L of the supernatant was carefully transferred to a new 96-well transparent plate. The absorbance of the solution at 540nm was read using a microplate reader (Molecular Devices). 0.1% Triton X-100 was used as a positive control and DPBS was used as a negative control.

As shown in FIG. 7, the hemolysis rates of the hydrogels AA/AA-NHS were 1.87%, 3.47%, 3.49%, 7.48%, and 12.17%, respectively. The results show that the hydrogels prepared in examples 1 to 3 have good blood compatibility, and the hemolysis rate is lower than 4%.

The cellular compatibility of the hydrogel was assessed by co-culturing fibroblasts (L929) with AA/AA-NHS hydrogel extracts (5 mg/mL). The experimental procedure was to soak the lyophilized AA/AA-NHS hydrogel samples in cell culture medium at 37 ℃ for 48h to prepare a series of sterile AA/AA-NHS hydrogel extracts at a fixed concentration of 5 mg/mL. 3000L 929 cells suspended in 100. mu.L of complete growth medium were seeded into each well of a 96-well plate. After 24 hours of incubation, the medium was changed to 100. mu.L of hydrogel extract from the sample. After 24 hours of co-incubation, the medium was removed and 10. mu.L of 100. mu.L of complete growth medium was added

Reagents were added to each well in the plate. The fluorescence of each well was read using a microplate reader to assess cell viability.

The statistics of the cell data of AA/AA-NHS in examples 1-5 are shown in FIG. 8, where it can be seen that the cell number measured by Alamar-Blue showed a tendency to increase after 72 hours of incubation of L929 with AA/AA-NHS hydrogel extracts for 24 hours, indicating good cell growth throughout the experiment. After 24h of co-culture, there was no significant difference in quantitative data of cell viability between the hydrogel group and the control group. After 72 hours of co-incubation, all hydrogel groups showed lower cell proliferation compared to the control group, but there was no difference in cell viability between the control and hydrogel groups (survival rates were greater than 85%), indicating that the hydrogel extracts were free of cytotoxic extracts. Shows good in vitro cell compatibility and can be used as a good biomedical material.

Evaluation experiments of biocompatibility of the hydrogel dressing prepared in examples 1 to 5 in vivo after subcutaneous implantation in rats. The experimental procedure was that AA/AA-NHS hydrogel was injected subcutaneously into SD rats on the back after sterilization and removed on day 7. The hydrogel was removed from the vicinity of the tissue, fixed with 4% paraformaldehyde for 24 hours, embedded in paraffin, and sectioned into thin sections for further histological examination. All species were stained with hematoxylin-eosin (H & E) and Toluidine Blue (TB) staining to assess the inflammatory response of the hydrogel. The stained sections of each sample were examined by microscopy (Olympus, japan) to assess tissue inflammatory responses.

The results are shown in FIG. 9, and FIG. 9-1 shows that all hydrogel groups showed mild acute inflammatory reactions 7 days after the hydrogel was implanted subcutaneously in rats. FIG. 9-2 shows that 7 days after the hydrogel is implanted into the subcutaneous tissues of rats, the number of mast cells in the peripheral tissues of the implanted hydrogel is close, and the hydrogel prepared in examples 1-5 can cause acute inflammatory reaction of the adjacent cell tissues.

When the hydrogel dressings prepared in examples 1 to 5 were subcutaneously implanted in rats for 7 days, quantitative data on the thickness of inflammatory fibers in the implanted region were counted, and the results are shown in fig. 10, in which the thickness of the fibrous inflammatory regions around the hydrogel groups of AA/AA-NHS0, AA/AA-NHS5, and AA/AA-NHS10 were similar. The thickness of the inflammation fibers in the implantation area of the hydrogel AA/AA-NHS0, AA/AA-NHS5, AA/AA-NHS10, AA/AA-NHS15 and AA/AA-NHS20 is respectively about 252.04 μm, 247.55 μm, 263.97 μm, 321.82 μm and 374.74 μm. The result shows that the hydrogel loaded with the tissue cross-linking agent monomer has good biocompatibility in the use process.

The hemostatic properties of the hydrogels prepared in examples 1 and 3 were evaluated and tested by the following experimental procedures:

(1) mouse liver hole-pricking model

Mice were anesthetized and then fixed on surgical cork plates. The liver of the mice was exposed by abdominal incision and the serous fluid around the liver was carefully cleared. A pre-weighed parafilm filter paper was placed under the liver. Liver bleeding was induced using a 20G needle, the cork plate was tilted about 30 °, and 300 μ L of AA/AA-NHS hydrogel solution was immediately applied to the bleeding site using a syringe. After 15 minutes, the weight of the filter paper having absorbed blood was measured and compared with the control group.

(2) Mouse liver incision model

Briefly, mice were anesthetized and then fixed on surgical cork plates. The liver of the mice was exposed through an abdominal incision and the serum around the liver was carefully cleared. A pre-weighed parafilm filter paper was placed under the liver. Then, liver bleeding was induced by creating a wound (5 mm long, 2mm deep) with a scalpel, and immediately applying 300 μ L of AA/AA-NHS hydrogel solution to the bleeding site using a syringe. After 15 minutes, the weight of the filter paper having absorbed blood was measured and compared with the control group.

(3) Model for amputation of mouse tail

Mice were anesthetized and then fixed on surgical cork plates. Fifty percent of the tail length was cut with surgical scissors. After cutting, the tail of the rat was left in the air for 15 seconds to ensure normal bleeding. Then, 300 μ L of AA/AA-NHS hydrogel solution was immediately applied to the bleeding site using a syringe. After 15 minutes, the weight of the filter paper with the absorbed blood and hydrogel sample was measured and compared to the control group (no treatment after puncturing the liver).

As shown in FIG. 11, the results in FIG. 11(a) show that in the mouse liver puncture model, the blood volume released by the liver of the control group mouse is close to 370.7 + -25.9 mg, the bleeding volume of the AA/AA-NHS0 hydrogel group mouse is 237.9 + -35.9 mg, and the bleeding volume of the AA/AA-NHS10 hydrogel group mouse is 151.8 + -14.9 mg, wherein the AA/AA-NHS10 is very different from the control group (P <0.005), and the AA/AA-NHS10 is very different from the AA/AA-NHS0 (P < 0.01). In addition, the mouse liver notching model of fig. 11(b) and the mouse tail-cutting model of fig. 11(c) both confirm that the hydrogel has good in vivo hemostatic properties.

The hydrogel dressing prepared in example 3 was used in a pig gastrorrhagia model.

The experimental procedure was that prior to the day of surgery, pigs were fasted but allowed complete access to drinking water for gastrointestinal tract preparation within 48 hours. A gastric bleeding model was established endoscopically. An endoscope with a water jet function was used for all the steps. After general anesthesia after tracheal intubation of the experimental pig, an endoscope was inserted into the stomach of the pig, and then a hook knife was inserted into the endoscope to perform electrical cutting. The mucosa and submucosa of the posterior wall of the stomach were then dissected with the electrosurgical generator set up and dissected with a slow pulse of 40W until the arterial network was severed and a venous bleed initiated. In the experimental group, AA/AA-NHS10 hydrogel was immediately sprayed to the bleeding area by gastroscope using a spray tube, and in the control group, 40 mg of esomeprazole was intravenously injected into the gastrorrhagia pig every day for 7 days. In the negative control group, pigs with artificial ulcers were not treated.

After resection of the posterior gastric wall and medial arterial network, continuous bleeding was observed, as shown in fig. 12 (a). The AA/AA-NHS10 hydrogel was then immediately sprayed onto the bleeding area by gastroscope using a spray tube, as shown in fig. 12 (b). The hydrogel was found to adhere firmly to the stomach wall by forming a hydrogel film by gastroscopy. Thereafter, the AA/AA-NHS10 hydrogel stopped hemostasis immediately within a few seconds by gelling at the bleeding point, thereby stopping hemostasis immediately, and the wound did not bleed further, as shown in fig. 12 (c). The results of the in vitro adhesion experiment in FIG. 12(d) show that the AA/AA-NHS hydrogel prepared by the present invention adheres to the stomach wall in vitro in pigs. By injecting AA/AA-NHS10 directly onto the surface of the stomach wall, a firm attachment of the hydrogel to the stomach wall was observed, mainly due to hydrogen bonding between the hydrogel and the tissue, and the synergistic enhancement of the adhesion strength of the hydrogel by the-CO-NH-bond formed between the hydrogel and the tissue mediated by the tissue cross-linker monomer.

Hemostatic use of the hydrogel prepared in example 3 to prevent delayed bleeding in a porcine gastrorrhagia model.

The experimental procedure was to collect a small stool sample from the experimental pig. The samples were then smeared in test tubes and immediately sent to a clinical laboratory. The measurement was carried out according to the product specification of colloidal gold method (Aibo occult blood test reagent; Aibo biomedicine, Hangzhou, China).

Hydrogels prepared in example 3, esomeprazole (PPI), and blanks were injected and subjected to fecal routine testing. The results are shown in Table 4.

TABLE 4 stool convention test results

Remarking: "+" indicates that the pig manure contains blood, and "-" indicates that the manure does not contain blood.

The results show that the positive rate of pigs sprayed in AA/AA-NHS10 was significantly reduced after 3 days of treatment compared to esomeprazole and control groups.

The results of the delayed bleeding rates of the hydrogel dressing, esomeprazole and the blank prepared in injection example 3 are shown in fig. 13, and the results show that the delayed bleeding rate of the hydrogel group was significantly reduced after 3 days of treatment and no significant bleeding was found 5 days after treatment. All these results show that the AA/AA-NHS10 hydrogel treated pigs were provided with haemostatic assurance after spray application and no secondary treatment was required.

The AA/AA-NHS hydrogel prepared by the invention, the esomeprazole (PPI) treatment group and the blank group are placed in a pig stomach ESD model for 21-day wound healing performance test. The experimental procedure was to insert an endoscope into the tip of an experimental pig, after general anesthesia and gastrointestinal tract preparation, with the tip fitted with a disposable distal transparent cap attachment, and to inject physiological saline solution into the posterior submucosal layer to enhance the mucosal layer. A double knife with a tip length of 2 mm was used to cut the mucosa to form mucosal defect wounds and artificial ulcers. For the experimental group, 500mg of AA/AA-NHS hydrogel was sprayed directly onto the ulcer via an endoscopic spray tube. In the PPI treatment group, 40mg of esomeprazole (amezeneca DEL, USA) was intravenously injected daily for 14 days. In the negative control group, pigs with artificial ulcers were not treated. On

days

7, 14 and 28, euthanasia was performed by intravenous injection of sodium pentobarbital in the ear vein and the stomach was excised.

The staining results are shown in fig. 14(a) and 15(a), which show that the number of gastric wound invasive inflammatory cells was significantly reduced in the AA/AA-NHS 10-treated pigs compared to the PPI-treated and control groups. The results of fig. 14(b), 14(c), 15(b), and 15(c) show that the AA/AA-NHS hydrogel significantly inhibited the expression of α -SMA and type I collagen in the gastric wound surface, indicating that the hydrogel dressing prepared by the present invention can inhibit fibrosis during ECM remodeling. The staining results of FIGS. 14(d) and 15(d) show that the number of blood vessels in the gastric wound was significantly increased compared to the control group. These results indicate that AA/AA-NHS hydrogels can promote healing of gastric wounds following endoscopic treatment by controlling inflammation, promoting ECM remodeling and angiogenesis. And it is even more effective than PPI (first-line therapy of endoscopic resection therapy) and shows a wide application prospect in clinic.

The above experimental results show that: the injectable hydrogel dressing with pH response prepared by the method is used in a pig stomach wound bleeding model and a stomach wound healing model after simulated endoscopic treatment: in a pig gastric bleeding model, wound bleeding, excrement routine tests, delayed bleeding and the like are directly monitored and evaluated through an endoscope, and the hydrogel dressing is proved to show a better treatment effect than intravenous esomeprazole (proton pump inhibitor, PPI) in the gastric bleeding model; in a pig stomach wound healing model, the wound healing condition, the number of inflammatory cells, the type I collagen deposition, the alpha-SMA expression and the blood vessel formation are directly monitored and evaluated through an endoscope, and the hydrogel dressing is proved to show the similar effect to that of intravenous injection esomeprazole in a stomach bleeding model and even have more effective treatment effect.

In conclusion, the hydrogel dressing prepared by the invention has effective and stable adhesion performance, has quick and strong self-healing performance, injectability, good hemostatic performance and wound healing promotion performance, stable rheological property, proper gelling time and good pH response performance, can be firmly adhered to a wound part, can be used as a barrier to protect the wound from the external environment, and can provide a proper microenvironment to accelerate the healing process of the wound after endoscopic treatment. Therefore, the injectable hydrogel dressing with pH response prepared by the invention has great application value in the aspects of stomach hemostasis and wound healing after endoscopic treatment.

The above-mentioned contents are only for illustrating the technical idea of the present invention, and the protection scope of the present invention is not limited thereby, and any modification made on the basis of the technical solution according to the technical idea proposed by the present invention falls within the protection scope of the claims of the present invention.

Claims

1. The preparation method of the injectable hydrogel dressing with pH response is characterized in that a polymer monomer, a tissue cross-linking agent monomer and a cross-linking agent are directly mixed and react for 200-800 s to obtain the injectable hydrogel dressing with pH response; the tissue cross-linking agent monomer is obtained by reacting double-bonded amino acid or double-bonded amino acid derivative with NHS, and the cross-linking agent is N, N' -methylene bisacrylamide;

2. The method for preparing an injectable hydrogel dressing having pH response according to claim 1, wherein the concentration of the doubly-bonded amino acid and the doubly-bonded amino acid derivative is 100 to 200 mg/mL; the concentration of the double-bonded polysaccharide, the double-bonded polysaccharide derivative, the double-bonded gelatin and the double-bonded gelatin derivative is 10-20 mg/mL; the tissue cross-linking agent monomer is prepared by taking dimethyl sulfoxide as a solvent, and the concentration of the tissue cross-linking agent monomer is 0-400 mg/mL.

3. The method for preparing an injectable hydrogel dressing having pH response according to claim 1, wherein the double-bonded amino acid and the double-bonded amino acid derivative are prepared by:

1) according to the feeding ratio of (15-20) g of amino acid, deionized water and sodium hydroxide: (100-140) mL: (4-12) preparing an amino acid solution; the amino acid is glycine;

according to the proportion of (15-20) g of amino acid derivatives, deionized water and sodium hydroxide: (100-140) mL: (4-12) g preparing an amino acid derivative solution; the amino acid derivative is 4-aminobutyric acid, 6-aminocaproic acid or 8-aminocaprylic acid;

3) According to the molar ratio of acryloyl chloride to amino acid (1.0-1.4): (0.8-1.2), adding the acryloyl chloride solution into the amino acid solution, mixing and reacting for 18-32 hours, and extracting, precipitating and drying to obtain a double-bonded amino acid derivative;

4. The method for preparing an injectable hydrogel dressing having pH response according to claim 3, wherein the extraction process in the step 3) is performed by ethyl acetate solution; the volume ratio of the amino acid polymer precursor solution or the amino acid derivative polymer precursor solution to the ethyl acetate is (20-50): (60-150).

5. The method for preparing an injectable hydrogel dressing having pH response according to claim 1, wherein the double-bonded polysaccharide is obtained by double-bonded graft reaction of any one of carboxymethyl chitosan, carboxyethyl chitosan, aminated hyaluronic acid and aminated sodium alginate; the double-bonded polysaccharide derivative is obtained by performing double-bonded grafting reaction on a chitosan quaternary ammonium salt derivative; the double-bonded gelatin derivative is obtained by double-bonded grafting reaction of quaternized gelatin or aminated gelatin.

6. The method for preparing an injectable hydrogel dressing having pH response according to claim 1, wherein a catalyst is further added during the mixing process; the catalyst is a mixture of APS and TEMED, and the volume ratio of N, N' -methylene bisacrylamide to the mixture of APS and TEMED is (150-250): (75-125): (75-125); the concentration of N, N' -methylene bisacrylamide is 1-4 mg/mL, the concentration of APS is 40-60 mg/mL, and the concentration of TEMED is 5-20 muL/mL.

7. The method of preparing an injectable hydrogel dressing having pH response according to claim 1, wherein the tissue cross-linker monomer is prepared by a process comprising:

1) using acetone as a solvent, preparing a double-bonded amino acid solution or double-bonded amino acid derivative solution with the concentration of 100-150 mg/mL, and mixing the double-bonded amino acid solution or double-bonded amino acid derivative solution with NHS according to the ratio of (100-150): (110-160), and stirring for 20-60 min to obtain a mixed solution;

8. The method for preparing the injectable hydrogel dressing with pH response according to claim 7, wherein the concentration in step 3) is to concentrate the filtered tissue cross-linking agent monomer precursor solution to 30-60 mL; after concentration, adding 300-480 mL of petroleum ether into the concentrated solution, standing until the precipitation is complete, and then sequentially performing suction filtration and drying.

9. The injectable hydrogel dressing with pH response obtained by the preparation method according to any one of claims 1 to 8, wherein the injectable hydrogel dressing with pH response is a porous net structure, and the pore diameter is 95-156 μm.

10. Use of the injectable hydrogel dressing having pH response of claim 9 in the field of gastric hemostasis and wound healing.