CN111437438A

CN111437438A - Intelligent drug-loaded hydrogel responding to inflammatory microenvironment and preparation method and application thereof

Info

Publication number: CN111437438A
Application number: CN202010490459.XA
Authority: CN
Inventors: 王云兵; 胡成
Original assignee: Sichuan University
Current assignee: Sichuan University
Priority date: 2020-05-08
Filing date: 2020-06-02
Publication date: 2020-07-24
Also published as: WO2021223756A1; CN114585396B; CN114585396A

Abstract

The invention provides an intelligent medicine-carrying hydrogel responding to an inflammation microenvironment as well as a preparation method and application thereof, wherein the preparation method of the intelligent medicine-carrying hydrogel responding to the inflammation microenvironment comprises the following steps: reacting functional polymer containing ortho hydroxyl with phenylboronic acid containing amino or carboxyl to prepare phenylboronic acid polymer; preparing an amphiphilic drug carrier; dissolving the prepared amphiphilic drug carrier and hydrophobic drug by using a benign solvent, and then adding water into the amphiphilic drug carrier and the hydrophobic drug under the stirring state to prepare a drug-loaded micelle solution; dissolving hydrophilic drug and the prepared phenylboronic acid polymer in the prepared drug-loaded micelle solution, and then adjusting the pH value of the mixed solution to 8-9 to obtain the drug-loaded micelle solution. The hydrogel responsively releases the medicine through pH and active oxygen, and has the advantages of few preparation components and simple synthesis operation.

Description

Intelligent drug-loaded hydrogel responding to inflammatory microenvironment and preparation method and application thereof

Technical Field

The invention belongs to the technical field of medical materials, and particularly relates to an intelligent drug-loaded hydrogel responding to an inflammation microenvironment as well as a preparation method and application thereof.

Background

Hydrogels have been widely used in biomedical applications over the past decades, for example as tissue engineering scaffolds, cell culture matrices, and drug delivery vehicles. Traditional hydrogels, while functional for drug delivery, do not achieve controlled on-demand release of the drug in the disease microenvironment. Therefore, it has been a hot spot to develop a novel intelligent hydrogel whose shape can be adjusted according to environmental changes for intelligent controlled release of drugs. At present, intelligent hydrogel with stimulation response is widely concerned by people due to potential application in the biomedical field. These smart hydrogels release drugs in a rapid response to external stimuli such as enzymes, pH, glucose, and Reactive Oxygen Species (ROS). However, most of the work reported at present is single response, and as the research progresses, two to three components are often required to be used in the preparation process of the hydrogel in order to realize multiple responses of the hydrogel, which often causes other problems such as low solubility, multi-step synthesis, high cost and high labor consumption. More importantly, the complexity of the hydrogel components would be detrimental to clinical transformation. It is well known that increased metabolic activity of microorganisms in the environment of microbial infected local wound inflammation results in higher ROS levels and lower pH values than normal tissue. Thus, for inflammatory diseases, materials are designed to target inflammation, and pH and ROS are suitable trigger points. However, how to build multi-responsive smart hydrogels of single and specific polymer components using existing molecular units remains a challenge. Meanwhile, the hydrogel formula which realizes multiple responses, is simple in design and low in cost through the least synthesis steps and operation processes has important significance.

Disclosure of Invention

Aiming at the problems in the prior art, the invention provides an intelligent drug-loaded hydrogel responding to an inflammation microenvironment, and a preparation method and application thereof, and the drug-loaded hydrogel can effectively solve the problems that the existing hydrogel is single in response, and the existing hydrogel has a plurality of preparation components, so that the solubility is low, the synthesis is complex and the cost is high.

In order to achieve the purpose, the technical scheme adopted by the invention for solving the technical problems is as follows:

a preparation method of an intelligent drug-loaded hydrogel responding to an inflammation microenvironment comprises the following steps:

(1) reacting functional polymer containing ortho hydroxyl with phenylboronic acid containing amino or carboxyl to prepare phenylboronic acid polymer;

(2) preparing an amphiphilic drug carrier;

(3) dissolving the amphiphilic drug carrier and the hydrophobic drug prepared in the step (2) by using a benign solvent, and then adding water into the mixture under a stirring state to prepare a drug-loaded micelle solution;

(4) and (2) dissolving the hydrophilic drug and the phenylboronic acid polymer prepared in the step (1) into the drug-loaded micelle solution prepared in the step (3), and then adjusting the pH value of the mixed solution to 8-9 to obtain the drug-loaded micelle solution.

Further, the specific reaction process in the step (1) is as follows: dissolving a functional polymer containing ortho hydroxyl to obtain a functional polymer solution containing the ortho hydroxyl, adding a condensing agent and phenylboronic acid containing amino or carboxyl into the solution, stirring the solution for 20 to 30 hours at the temperature of between 30 and 40 ℃, dialyzing the solution in deionized water, and then carrying out freeze drying to obtain a purified phenylboronic acid polymer, wherein the mass ratio of the functional polymer containing the ortho hydroxyl, the condensing agent and the phenylboronic acid containing the amino or carboxyl is 4-6:4-6: 1-2.

Further, the specific reaction process in the step (1) is as follows: dissolving a functional polymer containing ortho hydroxyl to obtain a functional polymer solution containing the ortho hydroxyl, adding a condensing agent and phenylboronic acid containing amino or carboxyl into the solution, stirring the solution at 37 ℃ for 24 hours, dialyzing the solution in deionized water for 3 days, and then carrying out freeze drying to obtain a purified phenylboronic acid polymer, wherein the mass ratio of the functional polymer containing the ortho hydroxyl, the condensing agent and the phenylboronic acid containing the amino or carboxyl is 5:4.8: 1.95.

Further, the condensing agent includes 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride and N-N-hydroxysuccinimide.

Further, the specific reaction process in the step (2) is as follows: completely dissolving a hydrophilic high molecular polymer and a hydrophobic molecule in dimethyl sulfoxide at 80 ℃, adding N, N '-dicyclohexylcarbodiimide and 4-dimethylaminopyridine, continuously stirring for 48 hours at 80 ℃, dialyzing the mixture in water for 2 days, removing excessive cholesterol, and finally freeze-drying for later use, wherein the mass ratio of the hydrophilic high molecular polymer to the hydrophobic molecule to the N, N' -dicyclohexylcarbodiimide to the 4-dimethylaminopyridine is 2:1.5:1: 0.5.

Further, the hydrophilic high molecular polymer includes hyaluronic acid, starch, cellulose, polyacrylic acid, polyacrylamide, polyvinyl alcohol, glycolic acid, and polylysine.

Further, hydrophobic molecules include cholesterol, polyolefins, polycarbonates, polyamides, polyacrylonitriles, polyesters, polylactic acids, and acrylates.

Further, the specific reaction process in the step (3) is that the amphiphilic drug carrier and the hydrophobic drug prepared in the step (2) are dissolved in a benign solvent, then the mixture is heated to 70-90 ℃, water is dropwise added into the mixture under the stirring condition, and the drug-loaded micelle solution with the concentration of 0.5-1.5mg/m L is prepared after dialysis.

Further, the specific reaction process in the step (3) is that 0.04 part of the amphiphilic drug carrier prepared in the step (2) and 0.008 part of hydrophobic drug are dissolved in a benign solvent, then the mixture is heated to 80 ℃, water is dropwise added into the mixture under the stirring condition, and the drug-loaded micelle solution with the concentration of 1mg/m L is prepared after dialysis.

Further, the benign solvent in step (3) includes DMSO, DMF, methanol, and acetone.

Further, the specific reaction process in the step (4) is as follows: dissolving a hydrophilic drug and the phenylboronic acid polymer prepared in the step (1) into the drug-loaded micelle solution prepared in the step (3) to enable the concentration of the phenylboronic acid polymer in the drug-loaded micelle solution to be 7-11% w/v, then adding an alkaline solution, and adjusting the pH value to be 8.5, wherein the mass ratio of the hydrophilic drug to the phenylboronic acid polymer is 1: 10.

Further, the concentration of the phenylboronic acid polymer in the micelle loading solution in step (4) is 9% w/v.

Further, the functional polymer containing ortho hydroxyl is a biological macromolecule containing amino and ortho hydroxyl or a biological macromolecule containing carboxyl and ortho hydroxyl.

Further, the biomacromolecule containing carboxyl and ortho-hydroxyl is sodium alginate, hyaluronic acid or a modified product thereof, and phenylboronic acid containing amino groups is grafted to a biomacromolecule side chain containing carboxyl groups and ortho-hydroxyl groups on the side chain through amidation reaction; the biomacromolecule containing amino and ortho-hydroxyl is chitosan or a modified product thereof, and phenylboronic acid with carboxyl groups is grafted to a biomacromolecule side chain containing amino groups and ortho-hydroxyl on the side chain through amidation reaction.

Further, the phenylboronic acid containing an amino group or a carboxyl group is ortho-aminophenylboronic acid, para-aminophenylboronic acid, ortho-carboxyphenylboronic acid or para-carboxyphenylboronic acid.

Further, the hydrophilic drug includes one of an antibiotic, a growth factor, a gene drug and a water-soluble protein drug.

Further, the hydrophobic drug includes one of an anti-inflammatory drug, an angiogenesis promoting drug, a cell proliferation promoting drug and a cell migration promoting drug.

Further, the anti-inflammatory agent comprises one of aspirin, paracetamol, amoxicillin and phenylbutazone.

The intelligent medicine-carrying hydrogel responding to the inflammation microenvironment is applied to preparation or application as wound dressing.

The beneficial effects produced by the invention are as follows:

1. the hydrogel is prepared from a single and definite polymer component, has simple synthesis steps, convenient operation and mild reaction conditions, still has good rheological property and complete structure after being loaded with two different medicaments, has multiple dynamic functions of self-healing, remodeling, injection and the like, has good cell compatibility, and has no adverse reaction in vivo.

2. The hydrogel has super-strong responsive drug release characteristics in an inflammation area, and as the pH value of a microorganism infected wound area is reduced compared with that of a normal tissue, the active oxygen level is increased, the hydrogel can rapidly release hydrophilic drugs and hydrophobic drugs by taking the pH value and the active oxygen as response mediators, and the two different ways accelerate the healing of the microorganism infected wound.

3. The phenylboronic acid ester bond in the intelligent drug-loaded hydrogel is rapidly broken under the conditions of high active oxygen and low pH value, and the hydrogel structure is damaged, so that the hydrophilic drug and the hydrophobic drug are rapidly released in a responding manner.

Drawings

FIG. 1 is a nuclear magnetic hydrogen spectrum of A L G, BA and A L G-BA according to the invention;

FIG. 2 is a nuclear magnetic hydrogen spectrum of CHO L, HA and HA-CHO L according to the present invention;

FIG. 3 is an IR spectrum of A L G, BA and A L G-BA according to the invention;

FIG. 4 is an IR spectrum of CHO L, HA and HA-CHO L according to the invention;

FIG. 5 is a particle size, potential and scanning electron microscope image of the drug-loaded micelle of the present invention;

FIG. 6 is a transmission electron microscope image of the drug-loaded micelle of the present invention;

FIG. 7 is a graph of particle size stability of drug-loaded micelles of the invention;

FIG. 8 is a preparation diagram of a drug-loaded hydrogel according to the present invention;

fig. 9 is a self-healing diagram of a drug-loaded hydrogel according to the present invention;

FIG. 10 is a scanning electron microscope image of the drug-loaded hydrogel of the present invention;

figure 11 is a graph of injectability and re-plasticity of drug-loaded hydrogels according to the present invention;

figure 12 is a frequency scan test chart of a drug-loaded hydrogel of the invention;

figure 13 is an amplitude scan test plot of a drug-loaded hydrogel of the invention;

fig. 14 is a self-healing performance graph of the intelligent drug-loaded hydrogel of the invention;

figure 15 is a dual response visualization of a drug-loaded hydrogel according to the invention;

FIG. 16 shows that the drug-loaded micelle of the invention has pH5.0 and H₂O₂Transmission electron microscopy images after treatment;

figure 17 is a drug release profile of a drug-loaded hydrogel according to the invention;

FIG. 18 is a graph and a quantitative graph of the healing efficacy of a drug-loaded hydrogel body of the invention for treating infected wounds in rats at various times;

FIG. 19 is a graph of HE staining and Masson staining of skin tissue from infected wounds of rats according to the invention;

FIG. 20 is a picture of TNF- α, I L-10 and CD31 in skin tissue from infected wounds of rats according to the present invention;

fig. 21 is a process diagram of the preparation of the intelligent drug-loaded hydrogel responding to the inflammatory microenvironment in the invention.

Detailed Description

The following detailed description of embodiments of the invention refers to the accompanying drawings.

In the following examples, the chemical agents other than the matrix are chemically pure unless otherwise stated.

Example 1

A preparation method of intelligent drug-loaded hydrogel responding to inflammatory microenvironment comprises the following steps:

(1) synthesis of Phenylboronic acid-modified alginate (A L G-BA)

Sodium alginate (A L G, 5.00G) was precisely weighed, dissolved in 500m L MES buffer (0.1mol, pH5.0), to which was added 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC. HCl, 4.80G, 25.0mmol) and 3-aminophenylboronic acid (BA, 1.95G, 12.5mmol), followed by stirring at 37 ℃ for 24h, finally dialyzed in deionized water (pH7.4) for 3 days, after 3 days, freeze-dried with a freeze dryer to obtain purified A L G-BA;

(2) synthesis of Cholesterol-modified hyaluronic acid (HA-CHO L)

Completely dissolving HA (2.00g) and cholesterol (CHO L, 1.50g) in dimethyl sulfoxide (DMSO, 30m L) at 80 deg.C, adding N, N' -dicyclohexylcarbodiimide (DCC, 1.00g) and 4-dimethylaminopyridine (DMAP,0.50g), stirring at 80 deg.C for 48 hr, dialyzing the mixture in water for 2 days, and centrifuging to remove excessive cholesterol;

(3) preparation of drug loaded Micelle (MIC) solution

HA-CHO L (40.0mg) and naproxen (Nap, 8.00mg) were dissolved inIn DMSO (10m L), heated to 80 ℃ and then stirred slowly with 10m L H₂O is added into the mixed solution drop by drop, and finally, the mixture is dialyzed in water for 2 days to prepare a drug-loaded micelle solution;

(4) preparation of hydrogels

Respectively dissolving A L G-BA (1.00G) and amikacin (AM, 100mg) prepared in the step (1) in the drug-loaded micelle solution (1mg/m L) prepared in the step (3) to enable the final concentration of A L G-BA in the drug-loaded micelle solution to be 9% w/v, and adjusting the pH of the drug-loaded micelle solution to 8.5 by using an alkaline solution to rapidly gelatinize the drug-loaded micelle solution to form hydrogel.

Example 2

(1) synthesis of phenylboronic acid modified hyaluronic acid (HA-BA)

Precisely weighing 5g hyaluronic acid, dissolving in 500m L MES buffer (0.1mol, pH5.0), adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC. HCl, 4.80g, 25.0mmol) and one of 2-aminophenylboronic acid or 3-aminophenylboronic acid (BA, 1.95g, 12.5mmol), stirring at 37 deg.C for 24h, dialyzing in deionized water (pH7.4) for 3 days, and freeze-drying with a freeze dryer to obtain purified HA-BA;

(2) amphiphilic drug carrier

Completely dissolving sodium alginate (2.00g) and cholesterol (CHO L, 1.50g) in a mixed solution of dimethyl sulfoxide and water (DMSO, 30m L), adding N, N' -dicyclohexylcarbodiimide (DCC, 1.00g) and 4-dimethylaminopyridine (DMAP,0.50g), continuously stirring for 48h, dialyzing the mixture in water for 2 days, removing excessive cholesterol by a centrifugal separation method, and freeze-drying the finished product for later use;

(3) preparation of drug-loaded micelles

Dissolving 10mg of the amphiphilic drug carrier in step (2) and 1mg of the anti-inflammatory drug (piroxicam) in DMSO (1m L), then adding it dropwise to 10ml of water with slow stirring, and finally, dialyzing the mixture in water for 2 days;

(4) preparation of hydrogels

When the drug-loaded hydrogel is prepared, 1g of HA-BA polymer and 100mg of antibiotic (ciprofloxacin hydrochloride) are respectively dissolved in a drug-loaded micelle solution (1mg/m L), and a proper amount of sodium hydroxide solution is added to adjust the pH of the solution to 8.5, so that the solution can be rapidly gelled.

Example 3

(1) synthesis of phenylboronic acid modified chitosan

5g of chitosan was finely weighed, dissolved in 500m L MES buffer (0.1mol, pH5.0), added with EDC. HCl (4.80g, 25.0mmol) and one of 2-carboxyphenylboronic acid or 3-carboxyphenylboronic acid (BA, 1.95g, 12.5mmol), then stirred at 37 ℃ for 24h, finally dialyzed in deionized water (pH7.4) for 3 days, and freeze-dried with a lyophilizer.

(2) Amphiphilic drug carrier

The amphiphilic drug carrier is directly selected from polylactic acid-glycolic acid copolymer (P L GA) purchased from the company, which is polymerized by lactic acid and glycolic acid according to the proportion of 50: 50.

(3) Preparation of drug-loaded micelles

10mg of P L GA and an anti-inflammatory drug (ibuprofen, 1mg) were dissolved in DMSO (10m L), and then added dropwise to 10m L water with slow stirring, and finally, the mixture was dialyzed in water for 2 days to obtain;

(4) preparation of hydrogels

Respectively dissolving the phenylboronic acid modified chitosan polymer (1g) and 100mg of antibiotic (vancomycin) in the step (1) into a drug-loaded micelle solution (1mg/m L), and then adding a proper amount of sodium hydroxide solution to adjust the pH of the solution to 8.5, so that the solution can be rapidly gelatinized.

Example 4

(1) synthesis of phenylboronic acid modified hyaluronic acid (HA-BA)

(2) amphiphilic drug carrier

(3) preparation of drug-loaded micelles

(4) preparation of hydrogels

When the drug-loaded hydrogel is prepared, 1g of HA-BA polymer and 100mg of Epidermal Growth Factor (EGF) are respectively dissolved in a drug-loaded micelle solution (1mg/m L), and a proper amount of sodium hydroxide solution is added to adjust the pH value of the solution to 8.5, so that the solution can be rapidly gelled.

Example 5

(1) synthesis of phenylboronic acid modified chitosan

(2) Amphiphilic drug carrier

The amphiphilic carrier is directly selected from polylactic acid-glycolic acid copolymer (P L GA) purchased from the company.

(3) Preparation of drug-loaded micelles

(4) preparation of hydrogels

Respectively dissolving the phenylboronic acid modified chitosan polymer (1g) and the angiogenesis promoting drug (deferoxamine mesylate) of 100mg in the step (1) into a drug-loaded micelle solution (1mg/m L), and then adding a proper amount of sodium hydroxide solution to adjust the pH of the solution to 8.5, so that the solution can be rapidly gelatinized.

Test examples

Taking the substance prepared in the example 1 as an example, the detection is carried out, and the specific operation process and the result are as follows:

firstly, detecting the A L G-BA prepared in the step (1), wherein the specific result is shown in figure 1 and figure 3, as shown in figure 1, the successful preparation of the A L G-BA polymer is proved, the grafting degree of the BA obtained by detecting the ultraviolet absorption of 295nm is 10 +/-1.5 percent, as shown in the infrared spectrogram of figure 3, the successful preparation of the A L G-BA polymer is also proved;

secondly, detecting the HA-CHO L prepared in the step (2), specifically referring to fig. 2 and fig. 4, as shown in fig. 2, proving that HA-CHO L is successfully prepared, obtaining the grafting degree of CHO L to be 13.5% through nuclear magnetism quantification, and as shown in fig. 4, also proving that HA-CHO L polymer is successfully prepared;

detecting the drug-loaded micelle solution prepared in the step (3), specifically shown in fig. 5-7, as shown in an electron microscope image of fig. 5, wherein the particle size of the drug-loaded micelle is 189.7nm, and the potential is-22.6 mV; as shown in the transmission electron microscope image of fig. 6, the shape of the drug-loaded micelle is proved to be spherical; as shown in the stability chart of fig. 7, the micelle demonstrated better stability in PBS;

fourthly, detecting the hydrogel prepared in the step (4), and particularly referring to fig. 8-11, wherein fig. 8 is a preparation diagram of the hydrogel; FIG. 9 is a self-healing chart of hydrogel, demonstrating that the prepared hydrogel has strong self-healing properties; FIG. 10 is a scanning electron micrograph of a hydrogel, which is a uniform cellular structure, demonstrating the success of hydrogel preparation in FIG. 10; FIG. 11 is a graph of injectability and remodeling of a hydrogel demonstrating that the hydrogel has injectability and remodeling;

and fifthly, detecting the rheological property of the prepared hydrogel, wherein the specific detection process is as follows:

performing rheological property detection on the hydrogel sample by using an MCR302 rheometer, wherein frequency scanning detection adopts 0.5% strain, oscillation frequency is 0.1-100 rad/s, strain scanning detection frequency is 1Hz, and strain value is 0.01-100%; in the self-healing test, a step strain test was performed by repeating a large strain (500%, 90s) and a small strain (0.1%, 90 s); the results are shown in fig. 12 and 13, which are respectively a frequency scanning graph and an amplitude scanning graph of the hydrogel, and as shown in fig. 12 and 13, the formation of the hydrogel is shown, and the mechanical properties of the hydrogel are well maintained after the antibiotics and the drug-loaded micelles are added. FIG. 14 is a self-healing performance graph of a hydrogel, and the results of FIG. 14 show that after the hydrogel structure is destroyed, 98.9% of the modulus of the hydrogel is recovered, which proves that the hydrogel has strong self-healing performance;

sixth, detection of hydrogel response to pH and ROS

After formation of the A L G-BA hydrogel, an appropriate volume of hydrochloric acid was added to lower the pH to 5, and an appropriate volume of hydrogen peroxide (H) was added₂O₂) To raise the ROS level, after four hours, the state of the hydrogel is recorded by imaging, specifically as shown in fig. 15, a two-way response visualization graph, as shown in fig. 15, after the hydrogel is gelled, the hydrogel changes from a gel state to a sol state after the pH is lowered or hydrogen peroxide is added;

seventhly, the release experiment process of the medicine is as follows:

soaking a certain mass of hydrogel in PBS (pH7.4 or pH5.0), with or without the addition of H₂O₂And collecting external PBS buffer solution at predetermined time, quantitatively determining drug release amount by high performance liquid chromatography and ultraviolet-visible spectrophotometer, detecting amikacin with potassium dihydrogen phosphate solution (pH 6.5) and methanol (30:70) as mobile phase, and wavelength of 340 nm. Naproxen measurement at 332nm with UV spectrophotometerThe content, the specific result is shown in a perspective electron microscope image of figure 16 and a drug release image of figure 17;

as shown in fig. 16, the drug-loaded micelle disintegrates and changes the particle size in the presence of pH and hydrogen peroxide; FIG. 17 demonstrates the drug release results that the drugs amikacin and naproxen respond to release at pH5.0 and 10mM H2O2, with both drugs releasing more than 80% at 24 hours, while under normal conditions, the drug release of amikacin is only about 30% and that of naproxen is less than 20%;

eighthly, the in vivo healing test process is as follows: in the following experimental illustrations, the hydrogel groups 1 to 4 represent the following combinations, respectively, unless otherwise specified: hydrogel group 1: blank hydrogel; hydrogel group 2: an antibiotic loaded hydrogel; hydrogel group 3: loading hydrogel of the drug-loaded micelle; hydrogel group 4: a hydrogel loaded with antibiotics and drug-loaded micelles;

anesthetizing rats with 10% chloral hydrate (0.3m L/100 g), preparing 4 full-thickness wounds (1cm × 1cm) on the back of each rat by using medical scissors, dividing the rats into different groups, dripping 100 mu L of pseudomonas aeruginosa (1 × 108CFU m L-1) on the wounds of each rat to initiate infection, smearing different hydrogel samples on the infected wounds for treatment after one day, performing euthanasia on the rats on the 14 th day of treatment, measuring the sizes of the wounds by drawing wound boundaries on transparent drawing paper, fixing regenerated skin at the wound parts with 4% glutaraldehyde, performing histopathological examination, and specifically, detecting results are shown in figures 18-20;

as shown in fig. 18, the drug-loaded hydrogel treatment group can promote the healing of the wound of the rat more quickly, the wound healing rate of the drug-loaded hydrogel treatment group reaches one hundred percent in 14 days, while the wound healing rate of the blank group is less than 70%, as shown in fig. 19, HE results show that the wound skin tissue structure of the rat in the drug-loaded hydrogel treatment group is more regular, Masson staining results show that more collagen is generated, as shown in fig. 20, the drug-loaded hydrogel treatment group can reduce the inflammation of the wound of the rat, which is shown as the reduction of TNF- α positive points (red), the increase of I L-10 positive points (red), the TNF- α positive cell rate of the drug-loaded hydrogel treatment group (hydrogel group 4) is about 10%, while the positive hydrogel rate of the control group exceeds 80%, the I L-10 positive cell rate of the drug-loaded hydrogel treatment group (hydrogel group 4) exceeds 40%, while the positive cell rate of the control group is about 5%.

Claims

1. The preparation method of the intelligent drug-loaded hydrogel responding to the inflammation microenvironment is characterized by comprising the following steps:

(2) preparing an amphiphilic drug carrier;

2. The preparation method of the intelligent drug-loaded hydrogel responding to inflammatory microenvironment of claim 1, wherein the specific reaction process in the step (1) is as follows: dissolving a functional polymer containing ortho hydroxyl to obtain a functional polymer solution containing the ortho hydroxyl, adding a condensing agent and phenylboronic acid containing amino or carboxyl into the solution, stirring the solution for 20 to 30 hours at the temperature of between 30 and 40 ℃, dialyzing the solution in deionized water, and then carrying out freeze drying to obtain a purified phenylboronic acid polymer, wherein the mass ratio of the functional polymer containing the ortho hydroxyl, the condensing agent and the phenylboronic acid containing the amino or carboxyl is 4-6:4-6: 1-2.

3. The preparation method of the intelligent drug-loaded hydrogel responding to inflammatory microenvironment according to claim 1, wherein the specific reaction process in the step (3) is that the amphiphilic drug carrier and the hydrophobic drug prepared in the step (2) are dissolved in a benign solvent, then the benign solvent is heated to 70-90 ℃, water is dropwise added into the benign solvent under stirring, and the drug-loaded micelle solution with the concentration of 0.5-1.5mg/m L is prepared after dialysis.

4. The preparation method of the intelligent drug-loaded hydrogel responding to inflammatory microenvironment of claim 1, wherein the specific reaction process in the step (4) is as follows: dissolving a hydrophilic drug and the phenylboronic acid polymer prepared in the step (1) into the drug-loaded micelle solution prepared in the step (3) to enable the concentration of the phenylboronic acid polymer in the drug-loaded micelle solution to be 7-11% w/v, then adding an alkaline solution, and adjusting the pH value to be 8.5, wherein the mass ratio of the hydrophilic drug to the phenylboronic acid polymer is 1: 10.

5. The method for preparing the intelligent drug-loaded hydrogel responding to inflammatory microenvironment, wherein the functional polymer containing the ortho-hydroxyl is a biological macromolecule containing amino and ortho-hydroxyl or a biological macromolecule containing carboxyl and ortho-hydroxyl.

6. The method for preparing the intelligent drug-loaded hydrogel responding to inflammatory microenvironment, wherein the amino-or carboxyl-containing phenylboronic acid is ortho-aminophenylboronic acid, para-aminophenylboronic acid, ortho-carboxyphenylboronic acid or para-carboxyphenylboronic acid.

7. The method for preparing an intelligent drug-loaded hydrogel responding to inflammatory microenvironment of claim 1, wherein the hydrophilic drug comprises one of antibiotics, growth factors, gene drugs and water-soluble protein drugs.

8. The method of preparing an inflammatory microenvironment responsive smart drug-loaded hydrogel of claim 1, wherein the hydrophobic drug comprises one of an anti-inflammatory drug, a pro-angiogenic drug, a pro-cell proliferative drug, and a pro-cell migratory drug.

9. The intelligent drug-loaded hydrogel responding to the inflammatory microenvironment is prepared according to the preparation method of any one of claims 1 to 8.

10. Use of the intelligent drug-loaded hydrogel responsive to inflammatory microenvironments of claim 9 in the preparation or as a wound dressing.