CN102634043B - Supermolecular hydrogel and preparation method and application thereof - Google Patents
Supermolecular hydrogel and preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses supermolecular hydrogel and preparation method and application thereof. The preparation method of the supermolecular hydrogel comprises the following steps of: synthesizing a polyethylene glycol modified cation copolymer; mixing with a nucleic acid solution and standing to obtain a copolymer/nucleic acid composite solution; and finally, mixing with alpha-cyclodextrin solution, and standing to obtain the supermolecular hydrogel capable of acting on a gene vector material. In the forming process of the hydrogel disclosed by the invention, a chemical crosslinking reaction does not happen, and thus the influence of a byproduct of the chemical crosslinking reaction on the activity of nucleic acid is effectively avoided, an efficient gene transfer effect is realized, and the transfection efficiency is obviously improved over the conventional gel material; and moreover, the preparation method is convenient and quick, realizes formation at room temperature, imposes relatively low requirements on concentration and temperature, and is favorable for maintaining the biological activity of embedded gene and the biocompatibility of the material.
Description
Technical field
The present invention relates to a kind of biomedical materials field, be specifically related to a kind of supramolecular hydrogel that can be used as genophore and its preparation method and application.
Background technology
Gene therapy refer to the external source normal gene import target cell with correct or compensation because of genetic flaw and the disease that extremely causes to reach therapeutic purpose, be at present for a kind of effective ways of cancer and innate immune system disease treatment.The key of this technology implementation is to select suitable genophore and method of gene introduction, and then makes gene can obtain safe, efficient, controlled and stable expression in cell.Hydrogel is as the bio-medical material of a class useful as drug carrier; have load factor high, can protect macromolecular drug to avoid the degraded of organism or removing and to the characteristics such as sustainable release of drug molecule, studied load and controlled release for gene in recent years.For example, Kasper etc. take polyethyleneglycol modified fumaric acid esters as raw material, N, the N'-methylene-bisacrylamide is linking agent, prepare by radical polymerization and obtain a class chemical gel, this gellike is by swelling action load gene, the rate of release of DNA can regulate by the change of polyoxyethylene glycol weight-average molecular weight (Journal of Controlled Release 2005,104:521-539).Shea etc. pass through photopolymerization reaction, with DNA/PEI(polyvinyl imines) mixture is embedded in the hydrogel matrix that the polyoxyethylene glycol modified by two keys and hyaluronic acid form, and by the feed change proportioning regulate gel-strength and DNA rate of release (Journal of Controlled Release 2007,120:233-241).Hu etc. are at first with DNA and the compound nano-complex that obtains of PEI, and then with this nano-complex be carried on dextran hydrogel matrix based in.Due to the controlled-release function of gel matrix, this material has the effect of lasting transfection, and still kept in serum good gene delivery ability (Journal of Materials Chemistry 2009,19:3189-3197).Yet the formation of these hydrogel gene vectors be unable to do without the use of high temperature, chemical cross-linking agent or light trigger usually, is unfavorable for keeping the biocompatibility by the biological activity of embedding gene and material.
(patent No. is Chinese invention patent: ZL200910213939.5) disclose a kind of novel supramolecular hydrogel gene vector, synthetic linear polyethylene glycol-polylysine the block polymer that obtains take Pluronic F-68 as initiator, and then obtain the hydrogel solid support material with the alpha-cylodextrin assembling.But the efficiency gene transfection of this gelatinous material is very low, and its highest embodiment is no more than 20%, and actual application value is little; And the design of material is more loaded down with trivial details, and synthesis step is more.
In sum, how to build easily efficient hydrogel gene vector material under mild conditions, just become the important topic that current biomedical engineering field needs to be resolved hurrily.
Summary of the invention
The object of the invention is to overcome the defective of above-mentioned prior art, a kind of preparation method of supramolecular hydrogel is provided, it prepares take alpha-cylodextrin as main body, by Subjective and Objective assembling effect the supramolecular hydrogel that can be used as genophore, and preparation condition is gentle, transfection efficiency lower to temperature and concentration requirement, product is high; In addition, gel-strength can be regulated by polymer concentration or cyclodextrin concentration, and is easy to operate, has thixotropy, can be used as load and release that the novel injectable gel of a class is applied to macromolecular drug.
For achieving the above object, the present invention adopts following technical scheme:
A kind of preparation method of supramolecular hydrogel, it comprises the following steps:
1) polyethyleneglycol modified cationic copolymer is synthetic;
2) be that 8～20% polyethyleneglycol modified cationic copolymer solution mixes with the nucleic acid solution of 0.2～0.8 μ g/mL and mass percent concentration, standing under room temperature condition, obtain multipolymer/nucleic acid complexes solution;
3) to step 2) to add the equal-volume mass concentration in the multipolymer of gained/nucleic acid complexes solution be 10～20% alpha-cylodextrin solution, mixes, standing under room temperature condition, obtains supramolecular hydrogel gene vector material.
Preferably, the concentration of step 1) amplifying nucleic acid solution is 0.5 μ g/mL.
Preferably, in step 1), the synthetic method of polyethyleneglycol modified cationic copolymer is:
The activation of a, polyglycol polymer
The polyglycol polymer of drying is dissolved in the mixing solutions of methylene dichloride and pyridine, add p-methyl benzene sulfonic chloride, lucifuge reaction under room temperature condition, after reaction finishes, reaction solution extracts with isopyknic hydrogen chloride solution, organic layer neutralizes, filters with alkali, and rotary evaporation obtains polyoxyethylene glycol-p-toluenesulfonic esters after removing methylene dichloride;
Synthesizing of b, polylysine grafting ethylene glycol copolymer
Polyoxyethylene glycol-p-toluenesulfonic esters and polylysine are dissolved in deionized water, react under 40～50 ℃ of conditions, reaction is cooled to room temperature with mixture after finishing, dialysis is 3 days in pure water, and the polyethyleneglycol modified cationic copolymer that obtains is polylysine grafting ethylene glycol copolymer.
Preferably, in step 1), the synthetic method of polyethyleneglycol modified cationic copolymer is:
The activation of a, polyglycol polymer
The polyglycol polymer of drying is dissolved in the mixing solutions of methylene dichloride and pyridine, add p-methyl benzene sulfonic chloride, lucifuge reaction under room temperature condition, after reaction finishes, reaction solution extracts with isopyknic hydrogen chloride solution, organic layer neutralizes, filters with alkali, and rotary evaporation obtains polyoxyethylene glycol-p-toluenesulfonic esters after removing methylene dichloride;
Synthesizing of c, polymine grafting ethylene glycol copolymer
Polyoxyethylene glycol-p-toluenesulfonic esters and polymine are dissolved in deionized water, react under 40～50 ℃ of conditions, reaction is cooled to room temperature with mixture after finishing, dialysis is 3 days in pure water, and the polyethyleneglycol modified cationic copolymer that obtains is polymine branch ethylene glycol copolymer.
Preferably, described polyglycol polymer is polyoxyethylene glycol or poly glycol monomethyl ether, and the weight-average molecular weight of described polyoxyethylene glycol is 4000,6000,8000 or 10000, and the weight-average molecular weight of described polyoxyethylene glycol monoether is 1000,2000 or 5000.
Preferably, the volume ratio of methylene dichloride and pyridine described in step a is 2:1; The mol ratio of described polyglycol polymer and p-methyl benzene sulfonic chloride is 1:1～2; The volumetric molar concentration of described hydrogen chloride solution is 3mol/L; Described alkali is sodium bicarbonate, and the consumption of described alkali is to add 3～5 grams in every 100 milliliters of organic phases; The time of described lucifuge reaction is 24～48 hours; Temperature during described rotary evaporation operation is 40～60 ℃.
Preferably, in step b, the weight-average molecular weight of described polylysine is 30,000～70,000; The mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polylysine is 0.1～1:1; Contain 10～15g polylysine in every 100ml deionized water; Reaction times is 48～72h; The weight-average molecular weight of holding back of the dialysis tubing that uses in the dialysis operation is 5000～14000.
Preferably, in step c, the weight-average molecular weight of polymine is 10000 or 2500; The mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polymine is 0.1～1:1; Every 100ml deionized water contains 10～15g polymine; Reaction times is 48～72h; The weight-average molecular weight of holding back of the dialysis tubing that uses in the dialysis operation is 5000～14000.
Preferably, step 2) add-on of described nucleic acid is 10～20 to add according to the N/P value with polyethyleneglycol modified cationic copolymer, described polyethyleneglycol modified cationic copolymer solution is that polyethyleneglycol modified cationic copolymer is dissolved in water or phosphate buffer soln, and time of repose is 15～30min; The described alpha-cylodextrin solution of step 3) is that alpha-cylodextrin is dissolved in water or phosphate buffer soln, and time of repose is 6～12h.
Another object of the present invention is to provide the supramolecular hydrogel of aforesaid method preparation.
Above-mentioned supramolecular hydrogel is applied to prepare in injectable drug carriers or injectable gene vector.
Principle of the present invention is:
Cationic polymers polymine (PEI), polylysine (PLL) have gene delivery effect preferably, polyoxyethylene glycol and cyclodextrin all have good biocompatibility, are one of minority synthetic compounds of being can be used as medicine and foodstuff additive by drugs approved by FDA; At first the present invention utilizes polyethyleneglycol modified cationic polymers, and this multipolymer forms stable nano-complex by the electrostatic interaction of polylysine or polymine segment and nucleic acid in solvent; And then assemble by the Subjective and Objective of cationic copolymer and alpha-cylodextrin the supramolecular structured hydrogel that interaction has further obtained the embedding in situ gene; The Main Function power that hydrogel forms is the crystallization after the cyclodextrin assembling; The introducing of polyoxyethylene glycol not only can reduce the cytotoxicity of cationic polymers, also can form gel with the cyclodextrin assembling, has dual function.
Compared with prior art, the invention has the beneficial effects as follows:
1, the preparation method is convenient and swift, can the room temperature compacted under, and lower to concentration and temperature requirement, be beneficial to and keep by the biocompatibility of the biological activity of embedding gene and material;
2, there is not chemical crosslink reaction in the forming process of hydrogel, has effectively avoided chemical crosslink reaction and byproduct of reaction on the impact of nucleic acid activity;
3, the intensity of hydrogel and gelation time can regulate and control by polymer concentration, cyclodextrin concentration;
4, polymkeric substance plays dual parts genophore and gel matrix in system, can simplify gel formula, saves cost;
5, hydrogel of the present invention is physical gel, and it can be loose to dissolving fully gradually along with liquid medium in organism constantly washes away corrosion, and is high to the release rate of nucleic acid;
6, hydrogel of the present invention has efficient gene delivery effect, and transfection efficiency is significantly improved than existing gelatinous material;
7, the composition of hydrogel of the present invention is simple, bio-compatibility good, the transfection successful, the characteristic that possesses good shear shinning, use convenient and swiftly, can be used as bio-medical engineering material, be widely used in preparing in injectable drug carriers or injectable gene vector.
Description of drawings
Fig. 1 is the nucleus magnetic hydrogen spectrum figure of polylysine grafting ethylene glycol copolymer;
Fig. 2 is the nucleus magnetic hydrogen spectrum figure of polymine grafting ethylene glycol copolymer;
Fig. 3 be supramolecular hydrogel F, G, H to the transfection efficiency data plot of rat fibroblast, control group is PEI-25k(N/P=10);
Fig. 4 is the fluorescence photo figure of the cell after supramolecular hydrogel G transfection rat fibroblast.
Embodiment
Below in conjunction with the drawings and specific embodiments, a kind of supramolecular hydrogel of the present invention and its preparation method and application is described in further detail.
Embodiment 1
1, the drying of polyoxyethylene glycol: with polyoxyethylene glycol vacuum-drying 24h under 80 ℃ of conditions.
2, the drying of methylene dichloride and pyridine: the hydrogenation calcium powder is joined (every 500ml solution adds 1-2 gram hydrolith) in methylene dichloride or pyridine solution, stir distillation after 24 hours under room temperature condition.
3, polyoxyethylene glycol-to the toluic acid ester synthesis:
The polyoxyethylene glycol (weight-average molecular weight 5000) of 15 gram dryings is dissolved in the mixing solutions of 100 milliliters of methylene dichloride and pyridine, and the volume ratio of methylene dichloride and pyridine is 2:1; Then adding mole number is the p-methyl benzene sulfonic chloride of 2 times of polyoxyethylene glycol, and under 30 degrees celsius, the lucifuge reaction is 48 hours; After reaction finished, reaction solution extracted with isopyknic 3mol/L hydrogen chloride solution, and organic layer neutralizes, filters with 3 gram sodium bicarbonates; Then obtain polyoxyethylene glycol-p-toluenesulfonic esters after under 60 degrees celsius, rotary evaporation is removed methylene dichloride.
4, polylysine grafting ethylene glycol copolymer is synthetic:
Polyoxyethylene glycol-p-toluenesulfonic esters obtained above and polylysine are dissolved in deionized water, and the mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polylysine is 1:1; This mixed solution reacted 48 hours under 40 ℃ of conditions; After reaction finishes, mixture being cooled to room temperature, is that 8000 dialysis tubing was dialysed product 3 days in pure water with holding back weight-average molecular weight, obtains polylysine grafting ethylene glycol copolymer.
5, the preparation of multipolymer/nucleic acid complexes solution:
Be 8%(w/w with the nucleic acid solution of every milliliter of 0.5 microgram with mass percent concentration) polylysine grafting ethylene glycol copolymer solution mix, the N/P value is 10; Mix, standing 15 minutes, obtain multipolymer/nucleic acid complexes solution.
6, the formation of supramolecular hydrogel:
Adding the equal-volume mass concentration in above-mentioned multipolymer/nucleic acid complexes solution is 16% alpha-cylodextrin solution, mixes, and standing 12 hours, obtains can be used as the supramolecular hydrogel F of gene vector material.
Embodiment 2
1, the drying of polyoxyethylene glycol: with polyoxyethylene glycol vacuum-drying 24h under 80 ℃ of conditions.
2, the drying of methylene dichloride and pyridine: the hydrogenation calcium powder is joined (every 500ml solution adds 1-2 gram hydrolith) in methylene dichloride or pyridine solution, stir distillation after 24 hours under room temperature condition.
3, polyoxyethylene glycol-p-toluenesulfonic esters is synthetic:
The polyoxyethylene glycol (weight-average molecular weight 2000) of 15g drying is dissolved in the mixing solutions of 100ml methylene dichloride and pyridine, and the volume ratio of methylene dichloride and pyridine is 2:1; Then adding mole number is the p-methyl benzene sulfonic chloride of 2 times of polyoxyethylene glycol, and under 25 ℃ of conditions, the lucifuge reaction is 36 hours; After reaction finished, reaction solution extracted with isopyknic 3mol/L hydrogen chloride solution, and organic layer neutralizes, filters with 4 gram sodium bicarbonates; Then obtain polyoxyethylene glycol-p-toluenesulfonic esters after under 50 ℃ of conditions, rotary evaporation is removed methylene dichloride.
4, polymine grafting ethylene glycol copolymer is synthetic:
Polyoxyethylene glycol-p-toluenesulfonic esters obtained above and polymine (weight-average molecular weight 25000) are dissolved in deionized water, and the mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polymine is 1:1; This mixed solution reacted 72 hours under 40 ℃ of conditions; After reaction finishes, mixture being cooled to room temperature, is that 14000 dialysis tubing was dialysed product 3 days in pure water with holding back weight-average molecular weight, obtains polymine grafting ethylene glycol copolymer.
5, the preparation of multipolymer/nucleic acid complexes solution:
Be 10%(w/w with the nucleic acid solution of every milliliter of 0.5 microgram with mass percent concentration) polymine grafting ethylene glycol copolymer solution mix, the N/P value is 10; Mix, standing 15 minutes, obtain multipolymer/nucleic acid complexes solution.
6, the formation of supramolecular hydrogel:
Adding the equal-volume mass concentration in above-mentioned multipolymer/nucleic acid complexes solution is 12% alpha-cylodextrin solution, mixes, and standing 8 hours, obtains can be used as the supramolecular hydrogel G of gene vector material.
Embodiment 3
1, the drying of polyoxyethylene glycol: with polyoxyethylene glycol vacuum-drying 24h under 80 ℃ of conditions.
2, the drying of methylene dichloride and pyridine: the hydrogenation calcium powder is joined (every 500ml solution adds 1-2 gram hydrolith) in methylene dichloride or pyridine solution, stir distillation after 24 hours under room temperature condition.
3, polyoxyethylene glycol-to the toluic acid ester synthesis:
The polyoxyethylene glycol (weight-average molecular weight 10000) of 15g drying is dissolved in the mixing solutions of 100ml methylene dichloride and pyridine, and the volume ratio of methylene dichloride and pyridine is 2:1; Then adding mole number is the p-methyl benzene sulfonic chloride of 2 times of polyoxyethylene glycol, and under 25 ℃ of conditions, the lucifuge reaction is 48 hours; After reaction finished, reaction solution extracted with isopyknic 3mol/L hydrogen chloride solution, and organic layer neutralizes, filters with the 3g sodium bicarbonate; Then obtain polyoxyethylene glycol-p-toluenesulfonic esters after under 60 ℃ of conditions, rotary evaporation is removed methylene dichloride.
4, polymine grafting ethylene glycol copolymer is synthetic:
Above-mentioned polyoxyethylene glycol-p-toluenesulfonic esters and polymine (weight-average molecular weight 25000) are dissolved in deionized water, and the mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polymine is 0.5:1; This mixed solution reacted 48 hours under 50 ℃ of conditions; After reaction finishes, mixture being cooled to room temperature, is that 8000 dialysis tubing was dialysed product 3 days in pure water with holding back weight-average molecular weight, obtains polymine grafting ethylene glycol copolymer.
5, the preparation of multipolymer/nucleic acid complexes solution:
Be 10%(w/w with the nucleic acid solution of every milliliter of 0.8 microgram with mass percent concentration) polymine grafting ethylene glycol copolymer solution mix, the N/P value is 20; Mix, standing 15 minutes, obtain multipolymer/nucleic acid complexes solution.
6, the formation of supramolecular hydrogel:
Adding the equal-volume mass concentration in above-mentioned multipolymer/nucleic acid complexes solution is 12% alpha-cylodextrin solution, mixes, and standing 8 hours, obtains can be used as the supramolecular hydrogel H of genetic material.
Embodiment 4
With embodiment 1 system to polylysine grafting ethylene glycol copolymer be dissolved in heavy water, carry out hydrogen spectrum nuclear-magnetism and characterize, result as shown in Figure 1, chemical shift appear at the peak at 4.22,2.92,1.63 and 1.36 places corresponding be proton peak on polylysine; 3.62 the proton peak of locating is the proton peak on the polyoxyethylene glycol segment.The nuclear-magnetism result of Fig. 1 confirms that polylysine grafting ethylene glycol copolymer is successfully synthesized.
Embodiment 5
The polymine grafting ethylene glycol copolymer that embodiment 2 is made is dissolved in heavy water, carrying out hydrogen spectrum nuclear-magnetism characterizes, result as shown in Figure 2, chemical shift appears at the proton peak at 3.65 places corresponding to the polyoxyethylene glycol segment, the proton peak at 2.5 places is corresponding to polymine.The nuclear-magnetism result of Fig. 2 confirms that polymine grafting ethylene glycol copolymer is successfully synthesized.
Embodiment 6
Supramolecular hydrogel F, G, H that embodiment 1,2,3 is obtained carry out extracorporeal releasing experiment under phosphate buffer soln/PBS environment, collect the releasing product of gelatinous material release 24 hours sections, the quantitatively rear transfection experiment that is used for rat fibroblast of the multipolymer that discharges/pEGFP mixture, the transfection efficiency result as shown in Figure 3.The fluorescence photo of the cell after the release liquid enforcement transfection of the supramolecular hydrogel G of embodiment 2 as shown in Figure 4.Positive controls PEI/pEGFP shows higher transfection efficiency, reaches approximately 86%; And the transfection efficiency of negative control naked DNA to be close to be 0%.The transfection efficiency of embodiment 1,2,3 supramolecular hydrogel F, G, multipolymer that H discharged at the 24th hour/pEGFP composite sample is respectively 24%, 46% and 40%, show good transfection effect, its transfection efficiency is significantly better than hydrogel gene vector material of the same type.
Embodiment 7
1, the drying of poly glycol monomethyl ether: with poly glycol monomethyl ether vacuum-drying 24h under 80 ℃ of conditions.
2, the drying of methylene dichloride and pyridine: the hydrogenation calcium powder is joined (every 500ml solution adds 1-2 gram hydrolith) in methylene dichloride or pyridine solution, stir distillation after 24 hours under room temperature condition.
3, polyoxyethylene glycol-p-toluenesulfonic esters is synthetic:
The poly glycol monomethyl ether (weight-average molecular weight 1000) of 10g drying is dissolved in the mixing solutions of 100 milliliters of methylene dichloride and pyridine, and the volume ratio of methylene dichloride and pyridine is 2:1; Then adding mole number is the p-methyl benzene sulfonic chloride of 2 times of polyoxyethylene glycol, and under 15 ℃ of conditions, the lucifuge reaction is 24 hours; After reaction finished, reaction solution extracted with isopyknic 3mol/L hydrogen chloride solution, and organic layer neutralizes, filters with the 5g sodium bicarbonate; Then obtain polyoxyethylene glycol-p-toluenesulfonic esters after under 40 ℃ of conditions, rotary evaporation is removed methylene dichloride.
4, polymine grafting ethylene glycol copolymer is synthetic:
Above-mentioned polyoxyethylene glycol-p-toluenesulfonic esters and polymine (weight-average molecular weight 10000) are dissolved in deionized water, and the mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polymine is 0.1:1; This mixed solution reacted 72 hours under 50 ℃ of conditions; After reaction finishes, mixture being cooled to room temperature, is that 5000 dialysis tubing was dialysed product 3 days in pure water with holding back weight-average molecular weight, obtains polymine grafting ethylene glycol copolymer.
5, the preparation of multipolymer/nucleic acid complexes solution:
Be 20%(w/w with the nucleic acid solution of every milliliter of 0.2 microgram with mass percent concentration) polymine grafting ethylene glycol copolymer solution mix, the N/P value is 10; Mix, standing 15 minutes, obtain multipolymer/nucleic acid complexes solution.
6, the formation of supramolecular hydrogel:
Adding the equal-volume mass concentration in multipolymer obtained above/nucleic acid complexes solution is 20% alpha-cylodextrin solution, mixes, and standing 6 hours, obtains can be used as the supramolecular hydrogel of gene vector material.
Above-described embodiment is only the preferred embodiment of the present invention, can not limit protection scope of the present invention with this, and the variation of any unsubstantiality that those skilled in the art does on basis of the present invention and replacement all belong to protection scope of the present invention.
Claims (8)
1. the preparation method of a supramolecular hydrogel is characterized in that comprising the following steps:
1) polyethyleneglycol modified cationic copolymer is synthetic;
2) be that 8～20% polyethyleneglycol modified cationic copolymer solution mixes with the nucleic acid solution of 0.2～0.8 μ g/mL and mass percent concentration, standing under room temperature condition, obtain multipolymer/nucleic acid complexes solution;
3) to step 2) to add the equal-volume mass concentration in the multipolymer of gained/nucleic acid complexes solution be 10～20% alpha-cylodextrin solution, mixes, standing under room temperature condition, obtains supramolecular hydrogel gene vector material;
The polyethyleneglycol modified positively charged ion of step 1) synthesizes by circuit one or circuit two, and the concrete steps of circuit one are as follows:
The activation of a, polyglycol polymer
The polyglycol polymer of drying is dissolved in the mixing solutions of methylene dichloride and pyridine, add p-methyl benzene sulfonic chloride, lucifuge reaction under room temperature condition, after reaction finishes, reaction solution extracts with isopyknic hydrogen chloride solution, organic layer neutralizes, filters with alkali, and rotary evaporation obtains polyoxyethylene glycol-p-toluenesulfonic esters after removing methylene dichloride;
Synthesizing of b, polylysine grafting ethylene glycol copolymer
Polyoxyethylene glycol-p-toluenesulfonic esters and polylysine are dissolved in deionized water, react under 40～50 ℃ of conditions, reaction is cooled to room temperature with mixture after finishing, dialysis is 3 days in pure water, and the polyethyleneglycol modified cationic copolymer that obtains is polylysine grafting ethylene glycol copolymer;
The concrete steps of circuit two are as follows:
The activation of a, polyglycol polymer
The polyglycol polymer of drying is dissolved in the mixing solutions of methylene dichloride and pyridine, add p-methyl benzene sulfonic chloride, lucifuge reaction under room temperature condition, after reaction finishes, reaction solution extracts with isopyknic hydrogen chloride solution, organic layer neutralizes, filters with alkali, and rotary evaporation obtains polyoxyethylene glycol-p-toluenesulfonic esters after removing methylene dichloride;
Synthesizing of c, polymine grafting ethylene glycol copolymer
Polyoxyethylene glycol-p-toluenesulfonic esters and polymine are dissolved in deionized water, react under 40～50 ℃ of conditions, reaction is cooled to room temperature with mixture after finishing, dialysis is 3 days in pure water, and the polyethyleneglycol modified cationic copolymer that obtains is polymine grafting ethylene glycol copolymer.
2. the preparation method of supramolecular hydrogel as claimed in claim 1, it is characterized in that: described polyglycol polymer is polyoxyethylene glycol or poly glycol monomethyl ether, the weight-average molecular weight of described polyoxyethylene glycol is 4000,6000,8000 or 10000, and the weight-average molecular weight of described poly glycol monomethyl ether is 1000,2000 or 5000.
3. the preparation method of supramolecular hydrogel as claimed in claim 1, it is characterized in that: the volume ratio of methylene dichloride described in step a and pyridine is 2:1; The mol ratio of described polyglycol polymer and p-methyl benzene sulfonic chloride is 1:1～2; The volumetric molar concentration of described hydrogen chloride solution is 3mol/L; Described alkali is sodium bicarbonate, and the consumption of described alkali is to add 3～5 grams in every 100 milliliters of organic phases; The time of described lucifuge reaction is 24～48 hours; Temperature during described rotary evaporation operation is 40～60 ℃.
4. the preparation method of supramolecular hydrogel as claimed in claim 1, it is characterized in that: in step b, the weight-average molecular weight of described polylysine is 30,000～70,000; The mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polylysine is 0.1～1:1; Contain 10～15g polylysine in every 100ml deionized water; Reaction times is 48～72h; The weight-average molecular weight of holding back of the dialysis tubing that uses in the dialysis operation is 5000～14000.
5. the preparation method of supramolecular hydrogel as claimed in claim 1, it is characterized in that: in step c, the weight-average molecular weight of polymine is 10000 or 2500; The mass ratio of polyoxyethylene glycol-p-toluenesulfonic esters and polymine is 0.1～1:1; Every 100ml deionized water contains 10～15g polymine; Reaction times is 48～72h; The weight-average molecular weight of holding back of the dialysis tubing that uses in the dialysis operation is 5000～14000.
6. the preparation method of supramolecular hydrogel as claimed in claim 1, it is characterized in that: step 2) add-on of described nucleic acid is 10～20 to add according to the N/P value with polyethyleneglycol modified cationic copolymer, described polyethyleneglycol modified cationic copolymer solution is that polyethyleneglycol modified cationic copolymer is dissolved in water or phosphate buffer soln, and time of repose is 15～30min; The described alpha-cylodextrin solution of step 3) is that alpha-cylodextrin is dissolved in water or phosphate buffer soln, and time of repose is 6～12h.
7. the supramolecular hydrogel of the described method preparation of a claim 1～6.
8. the application of supramolecular hydrogel claimed in claim 7 in preparation injectable drug carriers or injectable gene vector.
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| CN1757431A (en) * | 2005-09-16 | 2006-04-12 | 东南大学 | Visualized aquogel, and its preparing method |
| CN101288782A (en) * | 2008-06-18 | 2008-10-22 | 武汉科技学院 | Preparation method of high intensity biodegradable supramolecule hydrogel |
| CN101716346A (en) * | 2009-12-16 | 2010-06-02 | 中山大学 | Supramolecular hydrogel gene vector material, and preparation method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1757431A (en) * | 2005-09-16 | 2006-04-12 | 东南大学 | Visualized aquogel, and its preparing method |
| CN101288782A (en) * | 2008-06-18 | 2008-10-22 | 武汉科技学院 | Preparation method of high intensity biodegradable supramolecule hydrogel |
| CN101716346A (en) * | 2009-12-16 | 2010-06-02 | 中山大学 | Supramolecular hydrogel gene vector material, and preparation method and application thereof |
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| CN104069507A (en) * | 2014-06-11 | 2014-10-01 | 广州吉家庄生物科技有限公司 | High-strength supramolecular hydrogel, as well as preparation method and application thereof |
| CN104069507B (en) * | 2014-06-11 | 2017-04-12 | 广州吉家庄生物科技有限公司 | High-strength supramolecular hydrogel, as well as preparation method and application thereof |
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