A multi-arm polyethylene glycol polymer hydrogel, a preparation method and application thereof

Abstract

The application discloses a multi-arm polyethylene glycol high-molecular hydrogel, a preparation method and application thereof, and belongs to the technical field of hydrogels. The multi-arm polyethylene glycol high-molecular hydrogel is obtained by cross-linking of a first component containing a first functionalized multi-arm polyethylene glycol precursor and a second component containing a second functionalized multi-arm polyethylene glycol precursor. The multi-arm polyethylene glycol high-molecular hydrogel has the advantages of fast solidification speed, cross-linking without addition of a cross-linking agent or an initiator, and no need for external condition stimulation, and does not swell in a 37 DEG C phosphate buffer solution simulating a body fluid, does not break and lose effectiveness after long-time use, can continuously and smoothly release drugs as a drug delivery carrier, and has a wide application prospect in the field of biomedical materials.

Description

Technical Field

[0001] This invention belongs to the field of hydrogel technology, specifically relating to a multi-arm polyethylene glycol polymer hydrogel, its preparation method, and its application. Background Technology

[0002] Due to their similarity to the mechanical properties of human tissues and organs and their good biocompatibility, polymer hydrogels have significant clinical applications, such as drug delivery, prevention of postoperative adhesions, wound healing dressings, and tissue adhesives. Polymer hydrogels are composed of cross-linked hydrophilic polymers, forming a three-dimensional polymer network structure that can rapidly absorb and retain most of the water while remaining insoluble in water. They are a type of flexible material that possesses both solid and liquid properties.

[0003] Based on the rich structure, functional regulation, and diversified design of polymer hydrogels, various high-performance hydrogels have been developed in the existing technology, such as self-healing hydrogels, high-strength hydrogels, and nanocomposite hydrogels. However, polymer hydrogels still have the following shortcomings in medical applications: (1) Polymer hydrogels have a slow curing speed, usually requiring several minutes, which can easily lead to additional risks such as infection in clinical applications; (2) Polymer cross-linking requires external stimuli, such as chemical cross-linking agents, ultraviolet light, extreme pH, and high temperature, which makes it difficult to implement in treatment or cause side effects; (3) The prepared polymer hydrogels are prone to swelling, which can lead to hydrogel breakage, loss of mechanical properties, and damage to the surrounding tissues at the application site.

[0004] Polyethylene glycol (PEG) is a high molecular weight polymer. Hydrogel products developed based on PEG have advantages such as good biocompatibility, non-toxicity, non-irritation, and biodegradability. For example, Chinese patent document CN113797385A discloses a chitosan / polyethylene glycol hydrogel and its preparation method and application. This hydrogel is obtained by reacting a four-armed polyethylene glycol active ester with a primary amine in the chitosan molecule to form amide bonds. The prepared hydrogel has a three-dimensional network porous structure, good water absorption and swelling properties in acidic environments (the pH of the wound is slightly acidic), and excellent mechanical strength, with a compressive strength of up to 440 kPa. However, four-armed polyethylene glycol hydrogels do not have an advantage in crosslinking speed, and the obtained hydrogel is prone to swelling.

[0005] Chinese patent document CN111218011A discloses a polyethylene glycol-based hydrogel, its preparation method, and its applications. The preparation method includes: synthesizing a four-armed PEGNB using four-armed polyethylene glycol amino and norbornene as raw materials, with the assistance of condensing agent HATU and organic base DIPEA; then initiating a click chemistry reaction between PEGNB, polyethylene glycol dithiol, and photoinitiator I2959 under ultraviolet light to form a gel; and finally modifying the PEG-based hydrogel with arginyl-glycyl-aspartic acid tripeptide. This invention has simple preparation steps, but the polymer crosslinking process requires external stimulation. Summary of the Invention

[0006] To address the issues of slow curing speed and the need for complex external stimuli for curing in polymer hydrogels, this invention provides a multi-arm polyethylene glycol polymer hydrogel. This multi-arm polyethylene glycol polymer hydrogel has a fast curing speed, crosslinking without the need for crosslinking agents or initiators, and no external stimuli. Furthermore, it does not swell at all in a 37°C phosphate buffer solution simulating body fluids and will not break down or fail after prolonged use. As a drug delivery carrier, it can continuously and stably release drugs.

[0007] The specific technical solution adopted is as follows:

[0008] A functionalized multi-arm polyethylene glycol precursor, with the structural formula shown in formula (I):

[0009]

[0010] In equation (I), n is an integer from 4 to 300, m is an integer from 40 to 230, and R is selected from one of the structural formulas shown in equation (II). To replace the position;

[0011]

[0012] The present invention also provides a method for preparing the functionalized multi-arm polyethylene glycol precursor, comprising:

[0013] (1) Polymerization of norbornene-modified polyethylene glycol as monomer under the action of Grubb catalyst yields multi-arm polyethylene glycol precursor;

[0014] (2) Modify the end groups of the multi-arm polyethylene glycol precursor with crosslinkable functional groups, or deprotect the end groups of the multi-arm polyethylene glycol precursor and then modify them with crosslinkable functional groups to obtain the functionalized multi-arm polyethylene glycol precursor.

[0015] The structural formula of norbornene-modified polyethylene glycol is shown in formula (III):

[0016]

[0017] Where m is defined in the same way as in formula (I), and R1 is methyl, amino, hydroxyl, protected amino or protected hydroxyl.

[0018] The present invention also provides a multi-arm polyethylene glycol polymer hydrogel, which is obtained by crosslinking two of the functionalized multi-arm polyethylene glycol precursors as raw materials.

[0019] Specifically, the multi-arm polyethylene glycol precursor containing a first functionalized multi-arm polyethylene glycol precursor and the second component containing a second functionalized multi-arm polyethylene glycol precursor are crosslinked to obtain the multi-arm polyethylene glycol polymer hydrogel.

[0020] The structural formula of the first functionalized multi-arm polyethylene glycol precursor is any one of the following formulas:

[0021]

[0022] The structural formula of the second functionalized multi-arm polyethylene glycol precursor is any one of the following formulas:

[0023]

[0024] The definitions of n and m are the same as in equation (I).

[0025] The functionalized multi-arm polyethylene glycol precursor is a multi-arm polyethylene glycol polymer composed of a main chain and densely arranged side chains. Based on this structure, the functionalized multi-arm polyethylene glycol precursor can avoid complex intermolecular / intramolecular entanglement, exhibits multivalence, and has an extended morphology of side chains. The elimination of intermolecular / intramolecular entanglement improves the fluidity of high-concentration polymer solutions and increases the crosslinking efficiency of side chain ends. Multivalence increases the crosslinking speed of the polymer gel, and the extended side chain morphology can limit the swelling of the polymer gel. Therefore, the multi-arm polyethylene glycol polymer hydrogel prepared from the functionalized multi-arm polyethylene glycol precursor does not require the addition of crosslinking agents or initiators, nor does it require external stimulation. It also has a fast curing speed and is not prone to swelling.

[0026] Preferably, the first component and / or the second component contain a drug; the drug includes an antitumor drug, an antibacterial drug, an anti-inflammatory drug, or a biological macromolecule; the antitumor drug includes paclitaxel, doxorubicin, erlotinib, or gemcitabine; the antibacterial drug includes metronidazole, cefuroxime sodium, or rifampin; the anti-inflammatory drug includes dexamethasone or chloramphenicol; the biological macromolecule includes DNA, RNA, polypeptide, or protein.

[0027] Preferably, the drug is gemcitabine and / or glucagon-like peptide-1.

[0028] The present invention provides a method for preparing the multi-arm polyethylene glycol polymer hydrogel, comprising the following steps: dissolving a first component containing a first functionalized multi-arm polyethylene glycol precursor and a second component containing a second functionalized multi-arm polyethylene glycol precursor in water or a buffer solution to obtain a first solution and a second solution; mixing the first solution and the second solution to obtain the multi-arm polyethylene glycol polymer hydrogel.

[0029] Preferably, in the first solution, the mass fraction of the first functionalized multi-arm polyethylene glycol precursor is 0.5-80.0 wt%; and in the second solution, the mass fraction of the second functionalized multi-arm polyethylene glycol precursor is 0.5-80.0 wt%.

[0030] Specifically, the first solution is stored in syringe 1 of the dual-syringe syringe, and the second solution is stored in syringe 2 of the dual-syringe syringe. The first solution and the second solution are mixed to prepare the multi-arm polyethylene glycol polymer hydrogel.

[0031] Preferably, the first component and / or the second component contain a drug; the drug concentration is 0.0001 to 0.5 wt% based on the mass of the multi-arm polyethylene glycol polymer hydrogel.

[0032] The present invention also provides the application of the aforementioned multi-arm polyethylene glycol polymer hydrogel in the field of biomedical materials.

[0033] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0034] (1) The present invention provides a functionalized multi-arm polyethylene glycol precursor, which has an easily adjustable structure and low aqueous solution viscosity;

[0035] (2) The present invention uses two-component functionalized multi-arm polyethylene glycol precursor as raw material to prepare multi-arm polyethylene glycol polymer hydrogel. The crosslinking process does not require the addition of crosslinking agent or initiator, nor does it require external stimulation. Crosslinking can occur at room temperature and the curing speed is fast (within a few seconds). The preparation process is simple and easy to operate.

[0036] (3) The multi-arm polyethylene glycol polymer hydrogel does not swell at all in simulated body fluid, will not break down and fail after long-term use, and when the multi-arm polyethylene glycol polymer hydrogel is loaded with drugs, the release of drugs has the advantage of being continuous and stable. Attached Figure Description

[0037] Figure 1 The norbornene active lipid in Example 1 1 H and 13 C NMR spectrum, where A is 1H NMR spectrum, B is 13 C NMR spectrum.

[0038] Figure 2 The molecular weight change of norbornene-modified polyethylene glycol before and after synthesis in Example 1 is shown.

[0039] Figure 3 The images show the gel permeation chromatogram of the multi-arm polyethylene glycol precursor in Example 1 and its nuclear magnetic resonance (NMR) spectra before and after deprotection, where A is the gel permeation chromatogram and B is the NMR spectra before and after deprotection.

[0040] Figure 4 The image shows the UV-Vis absorption spectrum of the dibenzocyclooctynyl modified multi-arm polyethylene glycol precursor in Example 1.

[0041] Figure 5 The image shows the infrared spectrum of the azide-modified multi-arm polyethylene glycol precursor in Example 1.

[0042] Figure 6 The image shows the 1H NMR spectrum of the dibenzocyclooctynyl modified multi-arm polyethylene glycol precursor in Example 1.

[0043] Figure 7 The images show the gel permeation chromatogram of the multi-arm polyethylene glycol precursor in Example 2 and its NMR spectra before and after deprotection, where A is the gel permeation chromatogram and B is the NMR spectra after deprotection.

[0044] Figure 8 The image shows the 1H NMR spectrum of the aldehyde-modified multi-arm polyethylene glycol precursor in Example 2.

[0045] Figure 9 The image shows the 1H NMR spectrum of the hydrazine-modified multi-arm polyethylene glycol precursor in Example 2.

[0046] Figure 10 The curing rate curve of the multi-arm polyethylene glycol polymer hydrogel is shown.

[0047] Figure 11 The results show the swelling process monitoring of the multi-arm polyethylene glycol polymer hydrogel.

[0048] Figure 12 The cytotoxicity analysis of the multi-arm polyethylene glycol polymer hydrogel prepared in Example 2 is shown in the figure. In the figure, A represents SKOV3 cells, B represents NCI-358 cells, and C represents HEK 293 cells.

[0049] Figure 13 The drug release kinetic curves are for the multi-arm polyethylene glycol polymer hydrogels prepared in Examples 3 and 4. Detailed Implementation

[0050] The present invention will be further illustrated below with reference to the embodiments and accompanying drawings. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Operating methods not specifically specified in the following embodiments are generally performed under conventional conditions or as recommended by the manufacturer.

[0051] Example 1

[0052] (1) The norbornene-modified polyethylene glycol tert-butoxycarbonyl was obtained by double-modifying polyethylene glycol with norbornene active ester and amino tert-butoxycarbonyl (Beijing Jiankai Technology Co., Ltd.).

[0053] Norbornene active ester was obtained by condensation of exo-bicyclo[2.2.1]heptane-5-en-2-carboxylic acid and N-hydroxysuccinimide. 2.0 g of exo-bicyclo[2.2.1]heptane-5-en-2-carboxylic acid, 2.3 g of N-hydroxysuccinimide, and 3.9 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride were dissolved in 20 mL of dichloromethane. The reaction was allowed to proceed overnight, and the norbornene active ester was obtained by purification using a silica gel column (n-hexane / ethyl acetate = 2:1). The reaction route is shown in the following formula; the norbornene active ester... 1 H and 13 The C NMR spectra are as follows: Figure 1 As shown in A and B in the diagram.

[0054]

[0055] 3.51 g of norbornene active lipid and 30 g of amino-tert-butyloxycarbonyl-modified polyethylene glycol (PEG with a degree of polymerization of approximately 80) were mixed in 1 L of dichloromethane, and 100 mL of triethylamine was added and the mixture was reacted overnight. After concentration, norbornene-modified PEG tert-butyloxycarbonyl was obtained by precipitation with diethyl ether. The molecular weight change of PEG was characterized by matrix-assisted laser desorption / ionization time-of-flight mass spectrometry to confirm the acquisition of norbornene-modified PEG tert-butyloxycarbonyl. The results are as follows: Figure 2 As shown, the increase in the molecular weight of polyethylene glycol is due to norbornene. The reaction route is shown in the following equation;

[0056]

[0057] (2) 2g of norbornene-modified polyethylene glycol was dissolved in 10mL of dichloromethane, followed by the addition of 1mL of a dichloromethane solution containing 0.0118g / mL Grubb catalyst. After stirring for three hours, 2mL of vinyl ether was added to remove the Grubb catalyst. After stirring for another three hours, the polymer was precipitated in 100mL of cold ether. Impurities were removed by centrifugation at 3500r / min. The precipitate was further redissolved in dichloromethane and precipitated repeatedly three times. 2g of the polymer precipitate was dissolved in trifluoroacetic acid and stirred overnight to remove the tert-butyloxycarbonyl protecting group. Small molecule impurities were then removed by dialysis in water using a 6000-8000 dialysis membrane. The product was then lyophilized to obtain a multi-arm polyethylene glycol precursor. The structural formula of the multi-arm polyethylene glycol precursor is shown below, and the gel permeation chromatogram is shown below. Figure 3 As shown in A, multi-arm polyethylene glycol has good dispersibility. Nuclear magnetic resonance spectra before and after deprotection, as shown in the figure Figure 3 As shown in Figure B, the disappearance of the tert-butyl carbonyl peak proves that the amino protecting group has been completely removed.

[0058]

[0059] (3) Dissolve 1g of the multi-arm polyethylene glycol precursor obtained in step (2) in 0.1mol / L sodium carbonate solution. Add 0.16g of diphenylcyclooctyne-active ester and 0.11g of azidobutyric acid-active ester (Xi'an Dianhua Biotechnology Co., Ltd.) to the multi-arm polyethylene glycol precursor solution to prepare dibenzocyclooctyne-modified multi-arm polyethylene glycol precursor and azido-modified multi-arm polyethylene glycol precursor. Stir overnight, remove small molecule impurities by dialysis in water using a 6000-8000 dialysis membrane, and freeze-dry to obtain dibenzocyclooctyne-modified multi-arm polyethylene glycol precursor and azido-modified multi-arm polyethylene glycol precursor. The structural formulas of the azido-modified multi-arm polyethylene glycol precursor and the dibenzocyclooctyne-modified multi-arm polyethylene glycol precursor are shown in the following formulas:

[0060]

[0061] The UV-Vis absorption spectrum and 1H NMR spectrum of the dibenzocyclooctynyl-modified multi-arm polyethylene glycol precursor are shown below. Figure 4 and Figure 6 As shown, the characteristic UV absorption of dibenzocyclooctyne is at ~310 nm, and the characteristic NMR peak of dibenzocyclooctyne is at 7.0-8.0 ppm. The infrared spectrum of the azido-modified multi-arm polyethylene glycol precursor is shown below. Figure 5 As shown, 2100cm -1 This is the characteristic infrared absorption spectrum of the azide functional group.

[0062] (4) The dibenzocyclooctyne-modified multi-arm polyethylene glycol precursor and the azide-modified multi-arm polyethylene glycol precursor were dissolved in water to obtain a first solution and a second solution, respectively. In the first solution, the mass concentration of the dibenzocyclooctyne-modified multi-arm polyethylene glycol precursor was 10%, and in the second solution, the mass concentration of the azide-modified multi-arm polyethylene glycol precursor was 10%. The multi-arm polyethylene glycol hydrogel could then be obtained by using a double-barrel syringe or by simply mixing the two solutions.

[0063] Example 2

[0064] (1) Norbornene-modified polyethylene glycol hydroxyl groups were obtained by reacting the norbornene active ester prepared in Example 1 with amino-hydroxyl-modified polyethylene glycol (Beijing Jiankai Technology Co., Ltd.). The specific steps were as follows: 3.51 g of norbornene active ester and 30 g of amino-hydroxyl-modified polyethylene glycol (polyethylene glycol degree of polymerization approximately 80) were mixed in 1 L of dichloromethane, and 100 mL of triethylamine was added and reacted overnight. After concentration, the norbornene-modified polyethylene glycol hydroxyl groups were obtained by precipitation with diethyl ether. The reaction route is shown in the following formula.

[0065]

[0066] (2) 2g of norbornene-modified polyethylene glycol was dissolved in 10mL of dichloromethane, followed by the addition of 1mL of a dichloromethane solution containing 0.0118g / mL Grubb catalyst. After stirring for three hours, 2mL of vinyl ether was added, and stirring was continued for another three hours. The polymer was then precipitated in 100mL of cold ether. Impurities were removed by centrifugation at 3500r / min, and the product was further redissolved in dichloromethane. After repeated precipitation three times, the multi-arm polyethylene glycol precursor was obtained. The structural formula of the multi-arm polyethylene glycol precursor is shown below, and the gel permeation chromatogram is shown below. Figure 7 As shown in A, the polymer polymerizes well, but the dispersion is low. The characteristic NMR peak of the hydroxyl group (~4.6 ppm) is as follows... Figure 7 As shown in B in the diagram.

[0067]

[0068] (3) Dissolve 2g of the multi-arm polyethylene glycol precursor obtained in step (2) in dichloromethane, add 0.3g of p-methylphenylsulfonyl chloride, 0.2g of 4-dimethylaminopyridine and 200μL of triethylamine, stir overnight, dialyze in water using a 6000-8000 dialysis membrane to remove small molecule impurities, and freeze dry to obtain multi-arm polyethylene glycol p-methylbenzenesulfonate; dissolve 1g of multi-arm polyethylene glycol p-methylbenzenesulfonate in 10mL of dimethylformamide, add 1g of potassium carbonate and 200mg of 4-hydroxybenzaldehyde respectively, stir overnight, dialyze in water using a 6000-8000 dialysis membrane to remove small molecule impurities, and freeze dry to finally obtain aldehyde-modified multi-arm polyethylene glycol precursor;

[0069] 1 g of multi-arm polyethylene glycol p-toluenesulfonate, 200 mg of methyl mercaptoacetate, and 200 μL of triethylamine were dissolved in 10 mL of dimethylformamide. After heating at 80 °C for 48 h, small molecule impurities were removed by dialyzing in water using a 6000-8000 dialysis membrane. The mixture was then lyophilized to obtain a methyl mercaptoacetate-modified multi-arm polyethylene glycol precursor. Subsequently, the methyl mercaptoacetate-modified multi-arm polyethylene glycol precursor was dissolved in 2 mL of hydrazine hydrate in a mixture of water and tetrahydrofuran (volume ratio 3:7) and refluxed for 24 h. Small molecule impurities were removed by dialyzing in water using a 6000-8000 dialysis membrane. The mixture was then lyophilized to obtain a hydrazine-modified multi-arm polyethylene glycol precursor.

[0070] The preparation routes for aldehyde-modified and hydrazine-modified multi-arm polyethylene glycol precursors are shown in the following formulas, and their proton NMR spectra are shown in the following figures. Figure 8 and Figure 9 As shown, the characteristic peaks of acylhydrazine (~9.1 ppm) and aldehyde (~9.8 ppm) are clearly visible.

[0071]

[0072] (4) Dissolve the hydrazine-modified multi-arm polyethylene glycol precursor and the aldehyde-modified multi-arm polyethylene glycol precursor in water to obtain a first solution and a second solution. In the first solution, the mass concentration of the hydrazine-modified multi-arm polyethylene glycol precursor is 10%, and in the second solution, the mass concentration of the aldehyde-modified multi-arm polyethylene glycol precursor is 10%. Then, the multi-arm polyethylene glycol hydrogel can be obtained by using a double-barrel syringe or by direct simple mixing.

[0073] Example 3

[0074] A gemcitabine-loaded multi-arm polyethylene glycol (PEG) hydrogel was prepared. First, 5 mL of gemcitabine aqueous solution was prepared; the concentration could be selected according to the target requirements. In this example, the gemcitabine aqueous solution was designed to be 2 mg / mL. The dibenzocyclooctylenyl-modified and azide-modified PEG precursors were dissolved in the gemcitabine aqueous solution to obtain a first solution and a second solution, respectively. In the first solution, the mass concentration of the dibenzocyclooctylenyl-modified PEG precursor was 10%, and in the second solution, the mass concentration of the azide-modified PEG precursor was 10%. The PEG hydrogel was then obtained by using a double-barrel syringe or by simple direct mixing.

[0075] In this embodiment, the mass concentration of the drug in the multi-arm polyethylene glycol polymer hydrogel is 0.18 wt% based on the mass of the multi-arm polyethylene glycol polymer hydrogel.

[0076] Example 4

[0077] A multi-arm polyethylene glycol hydrogel loaded with glucagon-like peptide-1 was prepared. First, 5 mL of glucagon-like peptide-1 aqueous solution was prepared; the concentration could be selected according to the target requirements. In this embodiment, the glucagon-like peptide-1 aqueous solution was set to 1 mg / mL. Hydrazine-modified and aldehyde-modified multi-arm polyethylene glycol precursors were dissolved in the glucagon-like peptide-1 aqueous solution to obtain a first solution and a second solution, respectively. In the first solution, the mass concentration of the hydrazine-modified multi-arm polyethylene glycol precursor was 10%, and in the second solution, the mass concentration of the aldehyde-modified multi-arm polyethylene glycol precursor was 10%. The multi-arm polyethylene glycol hydrogel was then obtained by using a double-barrel syringe or by simple direct mixing.

[0078] In this embodiment, the mass concentration of the drug in the multi-arm polyethylene glycol polymer hydrogel is 0.09 wt%, based on the mass of the multi-arm polyethylene glycol polymer hydrogel.

[0079] Sample Analysis

[0080] Characterization tests were performed on the curing rate, swelling process, and cytotoxicity of the multi-arm polyethylene glycol polymer hydrogel.

[0081] (1) Curing rate study

[0082] The curing rate was measured by immediately measuring the storage modulus G′ and loss modulus G″ of the multi-arm polyethylene glycol (PEG) polymer solution mixture placed on the rheometer sample stage. Taking aldehyde-modified and hydrazine-modified PEG precursors as examples, they were dissolved in water or phosphate buffer solution to prepare 10% solutions. After simple mixing, the mixture was placed on the rheometer sample stage and oscillated at 25°C (τ = 10 Pa, ω = 10 rad / s) to monitor the changes in G′ and G″. The polymer sample was sealed with a layer of mineral oil during the measurement to prevent dehydration. The curing time of the polymer gel was defined as when G′ was higher than G″; the test results are shown below. Figure 10 As shown, G′ was already significantly higher than G″ at the start of the measurement, indicating that the multi-arm polyethylene glycol hydrogel could be cured.

[0083] (2) Swelling performance test

[0084] In the swelling experiment, multi-arm polyethylene glycol hydrogels with polymer concentrations of 10 wt%, 5 wt%, and 3 wt% were prepared in microcentrifuge tubes (preparation method similar to Example 2). After gel solidification, 1 mL of phosphate buffer was added above the hydrogel, and the centrifuge tubes were then incubated at 37°C and 300 rpm on a constant-temperature stirrer. At predetermined time points, the phosphate buffer was removed, and the weights of the hydrogel and centrifuge tubes were measured. The change in hydrogel mass was used to quantitatively evaluate the swelling behavior of the hydrogel; the test results are shown below. Figure 11As shown, the volume of the multi-arm polyethylene glycol hydrogel did not increase at all over time, indicating that the multi-arm polyethylene glycol hydrogel has no swelling properties.

[0085] (3) Cytotoxicity studies

[0086] The multi-arm polyethylene glycol (PEG) hydrogel from Example 2 was cut into cubic blocks (1 mm × 1 mm × 1 mm) and placed in 96-well plates containing cultured cells. The control group received phosphate solution. Cells involved included cancer cells and somatic cells, ovarian cancer cells (SKOV3), non-small cell lung cancer cells (NCI-358), and human embryonic kidney cells (HEK 293). Ovarian cancer cells and human embryonic kidney cells were grown in DMEM medium containing 10% heat-inactivated fetal bovine serum, 1% antibiotics, and 1% L-glutamine. Non-small cell lung cancer cells were grown in RPMI-1640 medium containing 10% heat-inactivated fetal bovine serum, 1% antibiotics, and 1% L-glutamine. All cells were incubated at 37°C in a humidified environment containing 5% carbon dioxide. The cytotoxicity of the multi-arm PEG hydrogel was assessed using the MTT assay after 3 days of incubation. 3-[4,5-dimethylthiazolyl-2-yl]-3,5-diphenyltetrazol bromide (MTT dye, final concentration 0.5 mg / mL) was added. Cells were incubated for 4 hours, and then the absorbance at 570 nm was measured using a microplate reader to determine the cytotoxicity of the multi-arm polyethylene glycol gel; the results are as follows. Figure 12 As shown in AC, the multi-arm polyethylene glycol hydrogel has little or no cytotoxicity.

[0087] The multi-arm polyethylene glycol hydrogels from Examples 3 and 4 were cut into three identical blocks (length: 7 mm; width: 5 mm; height: 2 mm). The gel blocks were then incubated with phosphate buffer at 300 rpm on a constant-temperature stirrer at 3°C. By collecting and replacing the phosphate buffer at predetermined time intervals, the absorbance at 280 nm was calculated using a UV-Vis spectrophotometer to obtain drug release curves for the small molecule drug and peptide. Figure 13 Release curves of multi-arm polyethylene glycol hydrogels loaded with gemcitabine and glucagon-like peptide-1, respectively.

[0088] The embodiments described above provide a detailed explanation of the technical solutions of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Any modifications, additions, or similar substitutions made within the scope of the principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A multi-armed polyethylene glycol macromer hydrogel, characterized in that, crosslinking the first component comprising the first functionalized multi-arm polyethylene glycol precursor and the second component comprising the second functionalized multi-arm polyethylene glycol precursor to obtain the multi-arm polyethylene glycol macromolecular hydrogel; the first functionalized multi-arm polyethylene glycol precursor has a structural formula as shown in the following formula: ； the second functionalized multi-arm polyethylene glycol precursor has a structural formula as shown in the following formula: ； wherein n is an integer of 4-300 and m is an integer of 40-230; respectively dissolving the first component comprising the first functionalized multi-arm polyethylene glycol precursor and the second component comprising the second functionalized multi-arm polyethylene glycol precursor in water or buffer to obtain a first solution and a second solution, mixing the first solution and the second solution to obtain the multi-arm polyethylene glycol macromolecular hydrogel; in the first solution, the mass fraction of the first functionalized multi-arm polyethylene glycol precursor is 10-80.0 wt%; in the second solution, the mass fraction of the second functionalized multi-arm polyethylene glycol precursor is 10-80.0 wt%; a drug is added to the first component and / or the second component; the mass concentration of the drug is 0.0001-0.5 wt% based on the mass of the multi-arm polyethylene glycol macromolecular hydrogel.

2. The multi-armed polyethylene glycol macromer hydrogel of claim 1, wherein, the drug comprises an anti-tumor drug, an antibacterial drug, an anti-inflammatory drug or a biological macromolecule; the anti-tumor drug comprises paclitaxel, doxorubicin, erlotinib or gemcitabine; the antibacterial drug comprises benznidazole, cefuroxime sodium or rifampicin; the anti-inflammatory drug comprises dexamethasone or chloramphenicol; the biological macromolecule comprises DNA, RNA, polypeptide or protein.

3. Use of the multi-arm polyethylene glycol macromolecular hydrogel according to claim 1 or 2 in the field of preparing biomedical materials.

Images