CN102924724B - Arborization macromolecule poly (acid amide-amine) grafting glucan and preparation method thereof - Google Patents
Arborization macromolecule poly (acid amide-amine) grafting glucan and preparation method thereof Download PDFInfo
- Publication number
- CN102924724B CN102924724B CN201210428081.6A CN201210428081A CN102924724B CN 102924724 B CN102924724 B CN 102924724B CN 201210428081 A CN201210428081 A CN 201210428081A CN 102924724 B CN102924724 B CN 102924724B
- Authority
- CN
- China
- Prior art keywords
- pamam
- azide
- dextran
- reaction
- grafting
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Landscapes
- Polysaccharides And Polysaccharide Derivatives (AREA)
Abstract
The invention provides an arborization macromolecule poly (acid amide-amine) grafting glucan. The glucan is represented as the formula I. The main chain is glucan of 1000-100000 of number average molecular weight, the total grafting ratio of Q1 and Q2 is in a range from 0.1% to 85%, the grafting ratio of the Q2 accounts for 0.1-75% of the total grafting ratio, the Q1 is a substitution group which is represented as the formula II, the Q2 is a substitution group which is represented as the formula III, and R is a substitution group formed by arborization macromolecule poly (acid amide-amine) of different number average molecular weight. The glucan has characteristics of the glucan and polyamidoamine (PAMAA), and cytotoxicity of the PAMAA is reduced. The invention also provides a preparation method of the grafting glucan.
Description
Technical field
The present invention relates to the synthetic preparation field of polymkeric substance, be specifically related to dendritic macromole poly-(acid amides-amine) of a kind of dextran grafting and preparation method thereof.
Background technology
PAMAM is that the first is synthesized and commercial branch-shape polymer.Along with the development of life science, people have carried out more deep research and application to PAMAM.Its intramolecular cavity can be used for packaging medicine molecule, and a plurality of surface functional groups can be used for carrying out various modifications, as connected targeted molecular, fluorescence molecule, drug molecule etc.Surface primary amine can be used in conjunction with DNA, as genophore.At present, PAMAM branch-shape polymer has been widely used in biomedicine field, as pharmaceutical carrier, and genophore, magnetic resonance imaging contrast etc.
Although the applied research of PAMAM branch-shape polymer makes great progress, a large amount of primary amine on PAMAM surface make it have certain cytotoxicity.Utilize low toxicity, it is to reduce toxicity that the immunologic inertia macromole of highly water-soluble is modified PAMAM, improves the important means of Biocompatibility.Forefathers have carried out a lot of relevant research work.Yet forefathers' research focuses mostly on and is utilizing PEG to modify PAMAM surface, as Bioco nj ugate Chemistry (volume: 19, phase: 11, page: 2239-2252) and Macromolecule s (volume: 35 phases: 9 pages: 3456-3462), all obtained good effect, but falling the hypotoxic while and also reduced the functional group densities of PAMAM modified surface, limited its application.Utilize dextran to modify and rarely have report PAMAM.
Dextran is one of bacillary polysaccharide, claims again dextran.It is good water solubility not only, and toxicity is low, and modified surface site is many, can also activating immune cell, stimulate the immunity system of body comprehensively.The ability that beta-glucan can make the lymphocyte of injured body produce cytokine (IL-1) recovers rapidly normal, effectively regulates human body immune function, can also promote the generation of IgM antibody in body, to improve the immunological competence of body fluid.The cell of dextran activation can excite the non-specific defense mechanism of host, therefore deeply attracting attention aspect tumour, infection disease and trauma care.Through specific step, extract and do not contain endotoxic β-1,3-dextran has assert it is a kind of safe material at U.S. FDA, is widely used in pharmaceutical industries.
Click chemistry(click chemistry) by the graduate Sharple s of U.S. Scripps s, proposed at first.The core of Click reaction is to open up a whole set of to take containing heteroatoms link unit C-X-C be basic combinatorial chemistry novel method, with simple and reliable on a small quantity, obtain molecular diversity widely with chemical transformation highly selective, started fast, effectively, or even 100% reliably, highly selective is manufactured the synthetic chemistry frontier of all kinds of new compounds.Cu(I wherein) trinitride-alkynes cycloaddition reaction of catalysis, embodies the most abundant to the target of click chemistry and thought.
In prior art, dendritic macromole poly-(acid amides-amine) be PAMAM owing to thering is a large amount of amino, amino with positive charge, cell is had to certain destruction.At present for reducing the cytotoxicity of cationic polymers, often at the polymer molecule of surface grafting good biocompatibility.Can reduce like this surface amino groups density of cationic polymers and still do like this activity that meeting reduces PAMAM, thereby when reducing its carrying capacity as carrier, also reduce the functional group densities of PAMAM modified surface, limit its application." at field of medicaments, PAMAM is often used to genophore as a kind of cationic polymers, if surface amino groups density declines, its carrying capacity for electronegative genomic medicine will significantly decline.
Summary of the invention
Dextran providing a kind of PAMAM grafting and preparation method thereof is provided the technical problem to be solved in the present invention, has both had the character of dextran and PAMAM concurrently, reduces again the cytotoxicity of PAMAM,
In order to solve above technical problem, the invention provides the dextran of poly-(acid amides-amine) grafting of a kind of dendritic macromole, there is structure shown in formula I:
Wherein, main chain is the dextran of number-average molecular weight 1000 ~ 100000; Q
1and Q
2total percentage of grafting be 0.1% ~ 85%; Q
2percentage of grafting account for 0.1% ~ 75% of total percentage of grafting; Q
1for substituting group, the Q shown in formula II
2for the substituting group shown in formula III;
R is the compound shown in formula II ~ formula VII;
Preferably, the number-average molecular weight of the compound shown in formula I is that the number-average molecular weight of the compound shown in formula I is 5328 ~ 1207253.
The present invention also provides the preparation method of the dextran of poly-(acid amides-amine) grafting of a kind of dendritic macromole, comprises the following steps:
A) provide 4-oxo-4-(propargyloxy) Succinic anhydried and dextran; The number-average molecular weight of described dextran is 1000 ~ 100000; By described 4-oxo-4-(propargyloxy) Succinic anhydried and dextran be blended in organic solvent, under the effect of catalyzer, ester condensation reaction occurs, and obtains 4-oxo-4-(propargyloxy) the Succinic anhydried dextran of modifying;
B) provide azide PAMAM, and the 4-oxo-4-(propargyloxy obtaining with step a)) dextran that Succinic anhydried is modified is blended in organic solvent, under the effect of copper sulfate and xitix, carry out click chemistry reaction, obtain the dextran of poly-(acid amides-amine) grafting of dendritic macromole; The number-average molecular weight of described azide PAMAM is 100 ~ 7182.
Preferably, the oxo-4-(of 4-described in step a) propargyloxy) being prepared as follows of Succinic anhydried:
Propiolic alcohol, DMAP, Succinic anhydried are blended in organic solvent, carry out esterification, obtain succinic acid list propine alcohol ester;
By described succinic acid list propine alcohol ester and N, N'-dicyclohexylcarbodiimide is blended in organic solvent, carries out dehydrating condensation, obtains 4-oxo-4-(propargyloxy) Succinic anhydried.
Preferably, the mol ratio of described Succinic anhydried and propiolic alcohol, 1 ~ 2:1
Preferably, described 4-oxo-4-(propargyloxy) mol ratio of Succinic anhydried and described dextran is 1 ~ 525:1; Described azide PAMAM and described 4-oxo-4-(propargyloxy) mol ratio of the Succinic anhydried dextran of modifying is 1 ~ 394:1.
Preferably, being prepared as follows of azide PAMAM in step b):
By NaN
3, bromine propylamine hydrogen bromide salt is water-soluble, under alkaline condition, Michael reaction occurs, and through purification drying process, obtains G0 for azide PAMAM;
Or methyl acrylate and G0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G0.5 for azide PAMAM after purifying; Quadrol and described G0.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtain G 1.0 generation azide PAMAM;
Or methyl acrylate and described G1.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G1.5 for azide PAMAM after purifying; Quadrol and described G1.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G2.0 for azide PAMAM;
Or methyl acrylate and described G2.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G2.5 for azide PAMAM after purifying; Quadrol and described G2.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G3.0 for azide PAMAM;
Or methyl acrylate and described G3.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G3.5 for azide PAMAM after purifying; Quadrol and described G3.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G4.0 for azide PAMAM;
Or methyl acrylate and described G4.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G4.5 for azide PAMAM after purifying; Quadrol and described G4.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G5.0 for azide PAMAM.
Preferably, described azide PAMAM is selected from G0 generation, G1.0 generation, and G2.0 generation, G3.0 generation, G4.0 generation or G5.0 are for branch-shape polymer PAMAM.
Preferably, step b) is specially:
B 1) by 4-oxo-4-(propargyloxy) the Succinic anhydried dextran and the CuSO that modify
4.5H
2o, sodium ascorbate are dissolved in organic solvent, obtain the first organic solution;
B2) by azide PAMAM solvent in organic solvent, adjust pH=7, obtain the second organic solution;
B3) the first organic solution and the second organic solution are mixed, repeatedly carry out in order freezing, vacuumize, oxygen that melting operation is removed in reaction system for three times, oil bath lower seal stirs; The product obtaining is obtained after dialysis, freeze-drying to the dextran of poly-(acid amides-amine) grafting of dendritic macromole.
Preferably, described organic solvent is selected from a kind of in acetone, methyl alcohol, methylene dichloride, dimethyl sulfoxide (DMSO) or DMF.
The invention provides (PAMAM) dextran of grafting of a kind of dendritic macromole poly-(acid amides-amine), due to PAMAM molecule is grafted in dextran, so had the performance of dextran and PAMAM concurrently, forming compound, and while using, dextran can form amino being coated on PAMAM molecule, thereby reduced the cytotoxicity of PAMAM, compound shown in formula I after Shi Suoshu modification has a plurality of PAMAM side chains can be used as biology or pharmaceutical carrier, in addition, contain alkynyl side chain can also with other functional groups, the compound of preparing difference in functionality, expanded the application prospect of its this compound.
The present invention also provides (PAMAM) preparation method of the dextran of grafting of a kind of dendritic macromole poly-(acid amides-amine), first use 4-oxo-4-(propargyloxy) Succinic anhydried and dextran carry out esterification, by 4-oxo-4-(propargyloxy) Succinic anhydried is grafted on dextran main chain, then the dextran that 4-oxo-4-(propargyloxy just) Succinic anhydried is modified is carried out click chemistry and is reacted under the effect of copper catalyst with azide PAMAM, obtain being grafted with the dextran of PAMAM, the method is simple, mild condition, be swift in response, being applicable to sending out technical scaleization produces.
Accompanying drawing explanation
The alkynyl-modified dextran that Fig. 1 provides for the embodiment of the present invention 1 be take the hydrogen nuclear magnetic resonance spectrogram of DM S O during as solvent;
Fig. 2 for the embodiment of the present invention 6 provide with CDCl
3hydrogen nuclear magnetic resonance spectrogram during for solvent;
Fig. 3 for the embodiment of the present invention 7 provide with CDCl
3hydrogen nuclear magnetic resonance spectrogram during for solvent;
Fig. 4 for the embodiment of the present invention 16 provide with D
2hydrogen nuclear magnetic resonance spectrogram when O is solvent;
Fig. 5 for the embodiment of the present invention 17 provide with D
2hydrogen nuclear magnetic resonance spectrogram when O is solvent;
The biocompatibility correlation curve figure that Fig. 6 provides for embodiment 27;
Fig. 7 for embodiment 28 provide utilize embodiment 17 synthetic polymers and DNA mass ratio for 50:1 time transfection situation map.
Embodiment
In order further to understand the present invention, below in conjunction with embodiment, the preferred embodiments of the invention are described, but should be appreciated that these are described is the restriction for further illustrating the features and advantages of the present invention rather than patent of the present invention being required.
The dextran that the invention provides poly-(acid amides-amine) grafting of a kind of dendritic macromole, has structure shown in formula I:
Wherein, main chain is the dextran of number-average molecular weight 1000 ~ 100000; Q
1and Q
2total percentage of grafting be 0.1% ~ 85%; Q
2percentage of grafting account for 0.1% ~ 75% of total percentage of grafting; Q
1for substituting group, the Q shown in formula II
2for the substituting group shown in formula III;
R is the compound shown in formula II ~ formula VII;
According to the present invention, the substituting group shown in described IV to IX represents respectively the substituting group that the PAMAM of different algebraically forms, and is respectively G0 ~ G5.0 generation.The position being connected with main chain in described substituting group is the connected ethylidene of center nitrogen-atoms.
According to the present invention, described when different number-average molecular weight dextran are during as main chain, m+n+p is just different, and the number-average molecular weight of the dextran that the present invention selects is preferably 1000 ~ 100000, and more preferably 5000 ~ 50000.But for the dextran of any one number-average molecular weight, all can carry out similar modification.The dextran that the molecular weight of take is 40000 is example, and each repeating unit molecular weight is 162, so m+n+p=40000/162=247, and each chain link all can carry out alkynyl modification, and percentage of grafting is 0% to 85%, so the scope of n+p is 247*0%=0 to 247*85%=210.Take percentage of grafting as 85% being example, and when carrying out click chemistry reaction, percentage of grafting is 0% to 25%, and the number of p is 210*0=0 to 210*25%=53.So according to different situations, total percentage of grafting of side chain Q 1 and Q2 is preferably 20% ~ 85%, and the percentage of grafting of Q2 is total percentage of grafting 0.1% ~ 75%, more preferably 0.1% ~ 25%.Because R can be the substituting group of different molecular weight, so the number-average molecular weight of the dextran after grafting provided by the invention is preferably 5328 ~ 1207253, more preferably 9696 ~ 46850.
Graftomer prepared by the present invention does not reduce surface amino groups density, but when it carries drug molecule or gene as solid support material bag, can form assembly cationic moiety is wrapped up, shell is the good dextran molecule of wetting ability, thereby plays the hypotoxic effect of falling.
In order to obtain the compound shown in formula I, the invention provides a kind of preparation method, comprising:
A) provide 4-oxo-4-(propargyloxy) Succinic anhydried and dextran; The number-average molecular weight of described dextran is 1000 ~ 100000; By described 4-oxo-4-(propargyloxy) Succinic anhydried and dextran be blended in organic solvent, under the effect of catalyzer, ester condensation reaction occurs, and obtains 4-oxo-4-(propargyloxy) the Succinic anhydried dextran of modifying;
B) provide azide PAMAM, and the 4-oxo-4-(propargyloxy obtaining with step a)) dextran that Succinic anhydried is modified is blended in organic solvent, under the effect of copper sulfate and xitix, carry out click chemistry reaction, obtain the dextran of poly-(acid amides-amine) grafting of dendritic macromole; The number-average molecular weight of described azide PAMAM is preferably 100 ~ 7182.
Reaction principle of the present invention is to be reacted PAMAM macromole is grafted on dextran main chain by click chemistry, due to the functional group's hydroxyl containing in dextran and PAMAM molecule and amino reaction difficulty, so need to modify described dextran, in described dextran, form alkynyl side chain, and then utilize the click chemistry reaction of alkynes and azido group that PAMAM is connected with the side chain of dextran, obtain being grafted with the dextran of PAMAM.
According to the present invention, the propargyloxy of 4-oxo-4-(described in described step step a)) being prepared as follows of Succinic anhydried:
Propiolic alcohol, DMAP, Succinic anhydried are blended in organic solvent, carry out esterification, obtain succinic acid list propine alcohol ester;
By described succinic acid list propine alcohol ester and N, N'-dicyclohexylcarbodiimide is blended in organic solvent, carries out dehydrating condensation, obtains 4-oxo-4-(propargyloxy) Succinic anhydried.The mol ratio of described Succinic anhydried and propiolic alcohol is preferably 1 ~ 2:1.
Preparation method is more specifically:
By propiolic alcohol, DMAP (DMAP), Succinic anhydried is dissolved in methylene dichloride, and room temperature reaction spends the night.Add water one time, utilize NaHSO
4(10wt%) repeatedly extract three times, organic phase is utilized MgSO
4dry, then filter, room temperature vacuum-drying 24h obtains product succinic acid list propine alcohol ester.The volume ratio of a described water and described methylene dichloride is preferably 70 ~ 80:100.
Then, succinic acid list propine alcohol ester is dissolved in to methylene dichloride and obtains the first solution; DCC is dissolved in to methylene dichloride and obtains the second solution.Under ice-water bath and nitrogen atmosphere, the second solution is slowly dropped in the first solution.Dropwise, room temperature continues reaction 12h, filters to obtain white solid, and room temperature vacuum-drying 24h, obtains product 4-oxo-4-(propargyloxy) Succinic anhydried.
Finally will in reaction flask, add 4-oxo-4-(propargyloxy) Succinic anhydried and dextran, take pyridine as catalyzer, dimethyl sulfoxide (DMSO) (DMSO) is solvent, in 30 ℃ of water-baths, react 48h, utilize 3500 dialysis tubing to dialyse two days in a water, freeze-drying obtains 4-oxo-4-(propargyloxy) the Succinic anhydried dextran of modifying.According to the present invention, described 4-oxo-4-(propargyloxy) mol ratio of Succinic anhydried and described dextran is preferably 1 ~ 525:1.
According to the present invention, being prepared as follows of azide PAMAM in step b):
By NaN
3, bromine propylamine hydrogen bromide salt is water-soluble, under alkaline condition, Michael reaction occurs, and through purification drying process, obtains G0 for azide PAMAM;
Or methyl acrylate and G0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G0.5 for azide PAMAM after purifying; Quadrol and described G0.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtain G 1.0 generation azide PAMAM;
Or methyl acrylate and described G 1.0 generation azide PAMAM are dissolved in organic solvent, carry out Michael reaction, reaction product obtains G 1.5 generation azide PAMAM after purifying; Quadrol and described G 1.5 generation azide PAMAM are dissolved in organic solvent, carry out aminolysis reaction, reaction product is through purifying, obtaining G2.0 for azide PAMAM;
Or methyl acrylate and described G2.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G2.5 for azide PAMAM after purifying; Quadrol and described G2.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G3.0 for azide PAMAM;
Or methyl acrylate and described G3.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G3.5 for azide PAMAM after purifying; Quadrol and described G3.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G4.0 for azide PAMAM;
Or methyl acrylate and described G4.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G4.5 for azide PAMAM after purifying; Quadrol and described G4.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G5.0 for azide PAMAM.
More specifically, the preparation method of described azide PAMAM is:
By NaN3, bromine propylamine hydrogen bromide salt is dissolved in a water, then joins in reaction flask 90 ℃ of reaction 24h, standing to room temperature, in reaction flask, add KOH to regulate pH to alkalescence, with anhydrous diethyl ether extraction, organic phase is utilized to anhydrous magnesium sulfate drying 4h at twice, then solid filtering is fallen, be spin-dried for and obtain colourless transparent liquid, be the product azide PAMAM in the 0th generation, referred to as G0.
Get methyl acrylate and be dissolved in methyl alcohol, add in reaction flask, ice bath stirs.Again G0 is utilized to dissolve with methanol, be slowly added drop-wise in reaction flask, drip off about half an hour.To after reaction flask sealing, move to stirring reaction 48h in 30 ℃ of oil baths, products therefrom utilizes silicagel column 1:1 normal hexane: ethyl acetate is that developping agent is separated, and product solution is collected together, and vacuumizes after 8h, obtains colorless oil transparent liquid, is product.The azide PAMAM in the 0.5th generation is referred to as G0.5.
After quadrol is dissolved in to methyl alcohol, add reaction flask, ice bath stirs.After G1.0 being utilized to dissolve with methanol again, be added drop-wise in reaction flask, dropwise about half an hour.To after reaction flask sealing, move in 35 ℃ of oil baths stirring reaction three days, after infrared tracking finds that ester bond disappears, stopped reaction, vacuumizes 2h by reaction flask.In reaction flask, add toluene that dope is dissolved, vacuum is drained, three times repeatedly, add methyl alcohol that dope is dissolved, vacuum is drained, repeatedly after three times yellow dope be product, the azide PAMAM of 1st generation is referred to as G1.0.
After the same method, recycle methyl acrylate and carry out Michael reaction, utilize quadrol aminolysis, can obtain for the 1.5th generation, the 2.0th generation, the 2.5th generation, the 3.0th generation, the 4.0th generation, the 5.0th generation azide PAMAM.
Prepared 4-oxo-4-(propargyloxy) after the Succinic anhydried dextran and azide PAMAM of modifying, both are carried out to click chemistry reaction, concrete steps are:
B 1) by 4-oxo-4-(propargyloxy) the Succinic anhydried dextran and the CuSO that modify
4.5H
2o, sodium ascorbate are dissolved in organic solvent, obtain the first organic solution;
B2) by azide PAMAM solvent in organic solvent, adjust pH=7, obtain the second organic solution;
B 3) the first organic solution and the second organic solution are mixed, repeatedly carry out in order freezing, vacuumize, oxygen that melting operation is removed in reaction system for three times, oil bath lower seal stirs.The product obtaining is obtained after dialysis, freeze-drying to the dextran of poly-(acid amides-amine) grafting of dendritic macromole.
According to the present invention, described azide PAMAM and 4-oxo-4-(propargyloxy) mol ratio of the Succinic anhydried dextran of modifying is 1 ~ 394:1.
According to the present invention, described Salzburg vitriol is as the catalyst click chemistry reaction of click chemistry reaction, and the click chemistry reaction of take G.0 for PAMAM is example, and step is specially:
First by CuSO
4.5H
2o, sodium ascorbate, the alkynyl that DMSO dissolves, dextran adds in reaction flask, then gets G 1.0 generation azide branch-shape polymer PAMAM, and after DMSO dissolves, with MHCl, pH test paper is reference, and dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain the dextran that brown solid is PAMAM grafting.
According to the present invention, described organic solvent is selected from a kind of in acetone, methyl alcohol, methylene dichloride, dimethyl sulfoxide (DMSO) or DMF.
The present invention also provides (PAMAM) preparation method of the dextran of grafting of a kind of dendritic macromole poly-(acid amides-amine), and the method is simple, and mild condition is swift in response, and is applicable to sending out technical scaleization and produces.
In order further to set forth technical solution of the present invention, be below the specific embodiment of the invention, it should be noted that main raw material of the present invention source.Different molecular weight dextran is purchased from Sigma-A De Ritchie company, propiolic alcohol, Succinic anhydried, quadrol, methyl acrylate is purchased from Sigma reagent company, 1-(3-dimethylamino-propyl)-3-ethyl-carbodiimide hydrochloride (EDC), dicyclohexylcarbodiimide (DC C), DMAP (DMAP) is purchased from Shanghai gill reagent company.All the other various medicines are all purchased from Beijing Chemical Plant.
In following embodiment, reaction raw materials is to be buied from the market or makes according to ordinary method, product quality * 100% that reaction yield=the actual product quality/theory obtaining obtains.
Embodiment 1:
Preparation 4-oxo-4-(propargyloxy) dextran (number-average molecular weight of dextran is 5000) of Succinic anhydried modification reaction:
Take respectively the dextran that 3 parts of 0.5g number-average molecular weights are 5000, put into respectively 3 reaction flasks, add respectively 9.376g, 7.500g, 4-oxo-4-(propargyloxy of 4.688g) Succinic anhydried, 2.4g pyridine, 40mLDM S O.Then solution is continued under stirring at room to reaction 48h, after reaction finishes, 400mL cold ethanol sedimentation for reaction system, centrifugal, with cold ethanol/cold diethyl ether, wash after three times, vacuum-drying 24h at 25 ℃, obtains the alkynyl-modified dextran of 3 kinds of different grafting densities.Products therefrom is in Table one.
The preparation of the different alkynyl grafting density of table one dextran
In upper table, A/I is 4-oxo-4-(propargyloxy) molar feed ratio of Succinic anhydried and dextran; The graft ratio of percentage of grafting (actual %) for calculating by nuclear-magnetism.
Embodiment 2:
Preparation 4-oxo-4-(propargyloxy) dextran (dextran number-average molecular weight is 40000) of Succinic anhydried modification reaction:
Take respectively the dextran that 3 parts of 1g number-average molecular weights are 40000, put into respectively 3 reaction flasks, add respectively 2.344g, 1.875g, 4-oxo-4-(propargyloxy of 1.172g) Succinic anhydried, 0.6g pyridine, 10mLDM S O.Then solution is continued under stirring at room to reaction 48h, after reaction finishes, 100mL cold ethanol sedimentation for reaction system, centrifugal, with cold ethanol/cold diethyl ether, wash after three times, vacuum-drying 24h at 25 ℃, obtains the alkynyl-modified dextran of 3 kinds of different grafting densities.Products therefrom is in Table one.
The preparation of the different alkynyl grafting density of table two dextran
In upper table, A/I is 4-oxo-4-(propargyloxy) molar feed ratio of Succinic anhydried and dextran; The graft ratio of percentage of grafting (actual %) for calculating by nuclear-magnetism.
Embodiment 3:
Preparation 4-oxo-4-(propargyloxy) dextran (dextran number-average molecular weight is 100000) of Succinic anhydried modification reaction:
Take respectively the dextran that 3 parts of 2.5g number-average molecular weights are 100000, put into respectively 3 reaction flasks, add respectively 2.344g, 1.875g, 4-oxo-4-(propargyloxy of 1.172g) Succinic anhydried, 0.6g pyridine, 10mLDM S O.Then solution is continued under stirring at room to reaction 48h, after reaction finishes, 100mL cold ethanol sedimentation for reaction system, centrifugal, with cold ethanol/cold diethyl ether, wash after three times, vacuum-drying 24h at 25 ℃, obtains the alkynyl-modified dextran of 3 kinds of different grafting densities.Products therefrom is in Table one.
The preparation of the different alkynyl grafting density of table three dextran
In upper table, A/I is 4-oxo-4-(propargyloxy) molar feed ratio of Succinic anhydried and dextran; The graft ratio of percentage of grafting (actual %) for calculating by nuclear-magnetism.
Embodiment 4:
Azide G0 is for the preparation of PAMAM:
Take NaN
310.0g, bromine propylamine hydrogen bromide salt 10.0g, is dissolved in respectively in water of 30mL, then joins in reaction flask, 90 ℃ of reaction 24h, standing to room temperature, in reaction flask, add 10gKOH, at twice with the extraction of 90mL anhydrous diethyl ether, organic phase is utilized to anhydrous magnesium sulfate drying 4h, then solid filtering is fallen, be spin-dried for to obtain colourless transparent liquid, be product.The azide PAMAM in the 0th generation is referred to as G0.Productive rate is 58.5%.
Embodiment 5:
The preparation of azide G 1.0 generation PAMAM:
Get 2.0g methyl acrylate and be dissolved in 20mL methyl alcohol, add in reaction flask, ice bath stirs.Take 1.0gG0 again, utilize 10mL dissolve with methanol, be slowly added drop-wise in reaction flask, drip off about half an hour.To after reaction flask sealing, move to stirring reaction 48h in 30 ℃ of oil baths, products therefrom utilizes silicagel column 1:1 normal hexane: ethyl acetate is that developping agent is separated, and product solution is collected together, and vacuumizes after 8h, obtains colorless oil transparent liquid, is product.The azide PAMAM in the 0.5th generation is referred to as G0.5.Productive rate is 85.2%.
Get 10.0mL quadrol and be dissolved in 20mL methyl alcohol, add in reaction flask, ice bath stirs.Take 1.8gG 1.0 again, utilize after 10mL dissolve with methanol, be added drop-wise in reaction flask, dropwise about half an hour.To after reaction flask sealing, move in 35 ℃ of oil baths stirring reaction three days, after infrared tracking finds that ester bond disappears, stopped reaction, vacuumizes 2h by reaction flask.In reaction flask, add 10mL toluene, vacuum is drained, three times repeatedly, add 10mL methyl alcohol, vacuum is drained, repeatedly after three times yellow dope be product, the azide PAMAM of 1st generation is referred to as G 1.0.Productive rate is 99.0%.
Embodiment 6:
Azide G2.0 is for the preparation of PAMAM:
Get 5.0g methyl acrylate and be dissolved in 20mL methyl alcohol, add in reaction flask, ice bath stirs.Take 2.0gG 1.0 again, utilize 10mL dissolve with methanol, be slowly added drop-wise in reaction flask, drip off about half an hour.To after reaction flask sealing, move to stirring reaction 48h in 30 ℃ of oil baths, products therefrom utilizes silicagel column 10:1 normal hexane: ethyl acetate is that developping agent is separated, and product solution is collected together, and vacuumizes after 8h, obtains faint yellow dope, is product.The azide PAMAM in the 1.5th generation is referred to as G 1.5.Productive rate is 89.5%.
Get 15.0mL quadrol and be dissolved in 20mL methyl alcohol, add in reaction flask, ice bath stirs.Take 2.0gG 1.5 again, utilize after 10mL dissolve with methanol, be added drop-wise in reaction flask, dropwise about half an hour.To after reaction flask sealing, move in 35 ℃ of oil baths stirring reaction three days, after infrared tracking finds that ester bond disappears, stopped reaction, vacuumizes 2h by reaction flask.In reaction flask, add 10mL toluene, vacuum is drained, three times repeatedly, add 10mL methyl alcohol, vacuum is drained, repeatedly after three times deep yellow dope be product, the azide PAMAM of 2nd generation is referred to as G2.0.Productive rate is 99.2%
Embodiment 7:
Azide G3.0 is for the preparation of PAMAM:
Get 5.0g methyl acrylate and be dissolved in 20mL methyl alcohol, add in reaction flask, ice bath stirs.Take 2.0gG2.0 again, utilize 10mL dissolve with methanol, be slowly added drop-wise in reaction flask, drip off about half an hour.To after reaction flask sealing, move to stirring reaction 48h in 30 ℃ of oil baths, products therefrom utilizes silicagel column separated, first uses 10:1 normal hexane: ethyl acetate is rushed post, by after impurity wash-out, uses methyl alcohol instead and rushes post, obtains product.Product solution is collected together, vacuumizes after 8h, obtain yellow dope, be product.The azide PAMAM in the 2.5th generation is referred to as G2.5.Productive rate is 72.2%.
Get 15.0mL quadrol and be dissolved in 20mL methyl alcohol, add in reaction flask, ice bath stirs.Take 2.0gG2.5 again, utilize after 10mL dissolve with methanol, be added drop-wise in reaction flask, dropwise about half an hour.To after reaction flask sealing, move in 35 ℃ of oil baths stirring reaction three days, after infrared tracking finds that ester bond disappears, stopped reaction, vacuumizes 2h by reaction flask.In reaction flask, add 10mL toluene, vacuum is drained, three times repeatedly, add 10mL methyl alcohol, vacuum is drained, repeatedly after three times deep yellow dope be product, the azide PAMAM in the 3rd generation is referred to as G3.0.Productive rate is 99.3%.
Embodiment 8:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.250g) that the alkynyl molecular weight that the alkynyl substituted degree that DM S O (2mL) dissolves is 82.3% is 5000 adds in reaction flask, then get the 0th generation azide branch-shape polymer PAMAM0.081g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.Repeatedly freezing, vacuumize, melting circulation three times is except the oxygen in desolventizing, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 85.3%.Percentage of grafting is 25.3%.
Embodiment 9:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.250g) that 82.3% the alkynyl molecular weight that DMSO (2mL) dissolves is 5000 adds in reaction flask, then get G1.0 for azide branch-shape polymer PAMAM0.262g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 81.3%.Percentage of grafting is 26.3%.
Embodiment 10:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuS O
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 82.3% the alkynyl molecular weight that DMSO (2mL) dissolves is 5000 adds in reaction flask, then get G2.0 for azide branch-shape polymer PAMAM0.070g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate 82.1%.Percentage of grafting is 17.2%.
Embodiment 11:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 82.3% the alkynyl molecular weight that DMSO (2mL) dissolves is 5000 adds in reaction flask, then get G3.0 for azide branch-shape polymer PAMAM0.152g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 3500 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 44.3%.Percentage of grafting is 22.1%.
Embodiment 12:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 82.3% the alkynyl molecular weight that DMS O (2mL) dissolves is 5000 adds in reaction flask, then get G4.0 for azide branch-shape polymer PAMAM0.304g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 3500 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 53.1%.Percentage of grafting is 12.2%.
Embodiment 13:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 82.3% the alkynyl molecular weight that DMSO (2mL) dissolves is 5000 adds in reaction flask, then get G5.0 for azide branch-shape polymer PAMAM0.608g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 7000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 20.3%.Percentage of grafting is 7.1%.
Embodiment 14:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.250g) that the alkynyl molecular weight that the alkynyl substituted degree that DMSO (2mL) dissolves is 84.0% is 40000 adds in reaction flask, then get G0 for azide branch-shape polymer PAMAM0.081g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 85.3%.Percentage of grafting is 25.3%.
Embodiment 15:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.250g) that 84.0% the alkynyl molecular weight that DMS O (2mL) dissolves is 40000 adds in reaction flask, then get G1.0 for azide branch-shape polymer PAMAM0.262g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 83.4%.Percentage of grafting is 33.2%.
Embodiment 16:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 84.0% the alkynyl molecular weight that DMSO (2mL) dissolves is 40000 adds in reaction flask, then get G2.0 for azide branch-shape polymer PAMAM0.070g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate 87.1%.Percentage of grafting is 15.2%.
Embodiment 17:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 84.0% the alkynyl molecular weight that DMSO (2mL) dissolves is 40000 adds in reaction flask, then get G3.0 for azide branch-shape polymer PAMAM0.152g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 3500 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 40.6%.Percentage of grafting is 24.5%.
Embodiment 18:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 84.0% the alkynyl molecular weight that DMSO (2mL) dissolves is 40000 adds in reaction flask, then get G4.0 for azide branch-shape polymer PAMAM0.304g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 3500 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 54.8%.Percentage of grafting is 15.8%.
Embodiment 19:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 84.0% the alkynyl molecular weight that DMSO (2mL) dissolves is 40000 adds in reaction flask, then get G5.0 for azide branch-shape polymer PAMAM0.608g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 7000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 13.4%.Percentage of grafting is 6.5%.
Embodiment 20:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.010g) that the alkynyl molecular weight that the alkynyl substituted degree that DMSO (2mL) dissolves is 81.6% is 100000 adds in reaction flask, then get G0 for azide branch-shape polymer PAMAM0.032g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 89.7%.Percentage of grafting is 46.1%.
Embodiment 21:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.250g) that 81.6% the alkynyl molecular weight that DMSO (2mL) dissolves is 100000 adds in reaction flask, then get G 1.0 generation azide branch-shape polymer PAMAM0.262g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 80.4%.Percentage of grafting is 33.1%.
Embodiment 22:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 81.6% the alkynyl molecular weight that DMSO (2mL) dissolves is 100000 adds in reaction flask, then get 2nd generation azide branch-shape polymer PAMAM0.071g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 1000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate 90.1%.Percentage of grafting is 17.2%.
Embodiment 23:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 81.6% the alkynyl molecular weight that DMSO (2mL) dissolves is 100000 adds in reaction flask, then get G3.0 for azide branch-shape polymer PAMAM0.152g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 3500 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 35.3%.Percentage of grafting is 26.1%.
Embodiment 24:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 81.6% the alkynyl molecular weight that DMSO (2mL) dissolves is 100000 adds in reaction flask, then get G4.0 for azide branch-shape polymer PAMAM0.304g, after 2mLDMSO dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 3500 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 52.3%.Percentage of grafting is 14.2%.
Embodiment 25:
Synthesizing of branch-shape polymer PAMAM grafting dextran:
First by CuSO
4.5H
2o(15mg), sodium ascorbate (62.5mg), the dextran (0.025g) that 81.6% the alkynyl molecular weight that DMSO (2mL) dissolves is 100000 adds in reaction flask, then get G5.0 for azide branch-shape polymer PAMAM0.608g, after 2mLDM S O dissolves, with 0.1MHCl, pH test paper is reference, dissolving is adjusted to after pH=7, adds in reaction flask.The oxygen that repeatedly freezing---vacuumizing---melting circulation removes in desolventizing for three times, 60 ℃ of condition lower seals of oil bath stir three days.To in water of the product utilization obtaining 7000 dialysis tubing, dialyse three days, after freeze-drying, obtain brown solid and be reaction product.Productive rate is 12.6%.Percentage of grafting is 6.1%.
Embodiment 26:
Above dendritic macromole of a kind of dextran grafting provided by the invention poly-(acid amides-amine) and preparation method thereof is described in detail, having applied specific case herein sets forth principle of the present invention and embodiment, the explanation of above embodiment is just for helping to understand method of the present invention and core concept thereof, should be understood that, for those skilled in the art, under the premise without departing from the principles of the invention, can also carry out some improvement and modification to the present invention, these improvement and modification also fall in the protection domain of the claims in the present invention.
Embodiment 27:
Cell suppresses experiment:
In order to investigate the biocompatibility of the dendritic macromole poly-(acid amides-amine) of dextran grafting of the present invention.Take HEK293 cell as model, adopt cell toxicity test, test material to act on the survival rate situation of cell after cell.
Concrete operation step is as follows:
1), collect logarithmic phase HEK293 cell, adjust cell concn, inoculate into (every hole 10 in 96 orifice plates
4individual, 100 μ l);
2), by poly-(acid amides-amine) concentration of dendritic macromole of the synthetic dextran grafting of substratum dilution embodiment 16 and embodiment 17, make a series of, the solution sample of totally 5 ~ 8 concentration gradients, every hole adds 100 μ L, 6 multiple holes of every kind of concentration;
3), 37 ℃, saturated humidity, 5%CO
2in cell culture incubator, cultivate 24 hours;
4), after 24h, every hole adds 20 μ L MTT solution (5mg/mL), continues to cultivate 4 hours;
5), stop to cultivate, suck nutrient solution in hole, every hole adds 150 μ L DMSO, low-speed oscillation 10 minutes detects each hole in the absorption value at 492nm place by microplate reader.
As shown in Figure 6, with the positive contrast of polymine (25K), simple material has good biocompatibility.
Embodiment 28:
Green fluorescent protein plasmid DNA transfection experiment:
In order to investigate the ability of lipoid plastid transfection DNA of the present invention, take HEK293 cell as model, detect the expression of green fluorescent protein.
Concrete operation step is as follows:
1), collect logarithmic phase HEK293 cell, adjust cell concn, inoculate into (every hole 10 in 96 orifice plates
4individual, 100 μ l);
2), with the dendritic macromole of the synthetic dextran grafting of substratum dilution embodiment 17, gather (acid amides-amine) concentration compound with green fluorescent protein plasmid DNA, make a series of, the solution sample of totally 5 ~ 8 concentration gradients, room temperature is after compound half an hour, every hole adds 100 μ L, 6 multiple holes of every kind of concentration;
3), 37 ℃, saturated humidity, 5%CO
2in cell culture incubator, cultivate 48 hours;
4), stop to cultivate, under inverted microscope, utilize 475nm blue-light excited observation transfection situation and take pictures.Transfection situation while as shown in Figure 7, being 50:1 for embodiment 17 synthetic polymers and DNA mass ratio.
Claims (9)
1. dendritic macromole gathers the dextran of (acid amides-amine) grafting, has structure shown in formula I:
Wherein, main chain is the dextran of number-average molecular weight 1000～100000; Q
1and Q
2total percentage of grafting be 0.1%～85%; Q
2percentage of grafting account for 0.1%～75% of total percentage of grafting; Q
1for substituting group, the Q shown in formula II
2for the substituting group shown in formula III;
R is the substituting group shown in formula II～formula VII;
2. the dextran of poly-(acid amides-amine) grafting of dendritic macromole according to claim 1, is characterized in that, the number-average molecular weight of the compound shown in formula I is 5328～1207253.
3. a preparation method for the dextran of poly-(acid amides-amine) grafting of dendritic macromole, is characterized in that, comprises the following steps:
A) provide 4-oxo-4-(propargyloxy) Succinic anhydried and dextran; The number-average molecular weight of described dextran is 1000～100000; Described 4-oxo-4-(propargyloxy) Succinic anhydried and dextran are blended in organic solvent, under the effect of catalyzer, ester condensation reaction occur, obtain the dextran that 4-oxo-4-(propargyloxy) Succinic anhydried is modified;
B) provide azide PAMAM, and be blended in organic solvent with the dextran that 4-oxo-4-(propargyloxy) Succinic anhydried that step a) obtains is modified, under the effect of copper sulfate and xitix, carry out click chemistry reaction, obtain the dextran of poly-(acid amides-amine) grafting of dendritic macromole; The molecular weight of described azide PAMAM is 100～7182;
Described 4-oxo-4-(propargyloxy) Succinic anhydried is prepared as follows:
Propiolic alcohol, DMAP, Succinic anhydried are blended in organic solvent, carry out esterification, obtain succinic acid list propine alcohol ester;
By described succinic acid list propine alcohol ester and N, N'-dicyclohexylcarbodiimide is blended in organic solvent, carries out dehydrating condensation, obtains 4-oxo-4-(propargyloxy) Succinic anhydried.
4. preparation method according to claim 3, is characterized in that, the mol ratio 1～2:1 of described Succinic anhydried and propiolic alcohol.
5. preparation method according to claim 3, is characterized in that, the mol ratio of described 4-oxo-4-(propargyloxy) Succinic anhydried and described dextran is 1～525:1; The mol ratio of the dextran that described azide PAMAM and described 4-oxo-4-(propargyloxy) Succinic anhydried are modified is 1～394:1.
6. preparation method according to claim 3, is characterized in that, step b) in being prepared as follows of azide PAMAM:
By NaN
3, bromine propylamine hydrogen bromide salt is water-soluble, under alkaline condition, Michael reaction occurs, and through purification drying process, obtains G0 for azide PAMAM;
Or methyl acrylate and G0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G0.5 for azide PAMAM after purifying; Quadrol and described G0.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G1.0 for azide PAMAM;
Or methyl acrylate and described G1.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G1.5 for azide PAMAM after purifying; Quadrol and described G1.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G2.0 for azide PAMAM;
Or methyl acrylate and described G2.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G2.5 for azide PAMAM after purifying; Quadrol and described G2.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G3.0 for azide PAMAM;
Or methyl acrylate and described G3.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G3.5 for azide PAMAM after purifying; Quadrol and described G3.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G4.0 for azide PAMAM;
Or methyl acrylate and described G4.0 are dissolved in organic solvent for azide PAMAM, carry out Michael reaction, reaction product obtains G4.5 for azide PAMAM after purifying; Quadrol and described G4.5 are dissolved in organic solvent for azide PAMAM, carry out aminolysis reaction, reaction product is through purifying, obtaining G5.0 for azide PAMAM.
7. preparation method according to claim 5, is characterized in that, described azide PAMAM is selected from G0 generation, G1.0 generation, and G2.0 generation, G3.0 generation, G4.0 generation or G5.0 are for branch-shape polymer PAMAM.
8. preparation method according to claim 5, is characterized in that, step b) be specially:
B1) dextran and the CuSO that 4-oxo-4-(propargyloxy) Succinic anhydried are modified
4.5H
2o, sodium ascorbate are dissolved in organic solvent, obtain the first organic solution;
B2) by azide PAMAM solvent in organic solvent, adjust pH=7, obtain the second organic solution;
B3) the first organic solution and the second organic solution are mixed, repeatedly carry out in order freezing, vacuumize, oxygen that melting operation is removed in reaction system for three times, oil bath lower seal stirs; The product obtaining is obtained after dialysis, freeze-drying to the dextran of poly-(acid amides-amine) grafting of dendritic macromole.
9. according to the preparation method described in claim 3,6 or 8, it is characterized in that, described organic solvent is selected from a kind of in acetone, methyl alcohol, methylene dichloride, dimethyl sulfoxide (DMSO) or DMF.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210428081.6A CN102924724B (en) | 2012-10-31 | 2012-10-31 | Arborization macromolecule poly (acid amide-amine) grafting glucan and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210428081.6A CN102924724B (en) | 2012-10-31 | 2012-10-31 | Arborization macromolecule poly (acid amide-amine) grafting glucan and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102924724A CN102924724A (en) | 2013-02-13 |
| CN102924724B true CN102924724B (en) | 2014-10-22 |
Family
ID=47639666
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201210428081.6A Active CN102924724B (en) | 2012-10-31 | 2012-10-31 | Arborization macromolecule poly (acid amide-amine) grafting glucan and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102924724B (en) |
Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106046382B (en) * | 2016-05-25 | 2018-10-09 | 暨南大学 | It is a kind of to load nitric oxide production cationic polymer and its preparation method and application |
| CN106589391B (en) * | 2016-12-16 | 2019-05-17 | 天津商业大学 | Chitosan/polylysine dendrimer core-shell nano grain and preparation method thereof |
| CN107459583B (en) * | 2017-08-08 | 2020-04-28 | 中山大学 | Highly hyperbranched cationic polysaccharide derivative containing dendritic polyamide-amine group and preparation method thereof |
| CN108003354B (en) * | 2017-12-28 | 2020-10-16 | 浙江大学 | Polymer responding to intracellular acidity and redox environment and preparation and application thereof |
| CN108926714B (en) * | 2018-06-28 | 2021-11-26 | 哈尔滨医科大学 | High-molecular gel for delivering pharmacological active substances to bladder cancer in bladder perfusion targeted manner and preparation method of high-molecular gel |
| CN110669223A (en) * | 2019-09-19 | 2020-01-10 | 暨南大学 | Glucan grafted dendritic polyamide-amine polymer and preparation method and application thereof |
| CN111693619B (en) * | 2020-05-18 | 2021-09-17 | 中国石油大学(北京) | P-mercaptobenzoic acid modified magnetic PAMAM dendritic polymer material |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7985401B2 (en) * | 2003-10-31 | 2011-07-26 | The Regents Of The University Of California | Peptides whose uptake by cells is controllable |
| CN101343359B (en) * | 2008-09-04 | 2011-01-19 | 上海交通大学 | Preparation method for aminophenol modified daiamid type tree shaped numerator |
| CN102002117B (en) * | 2010-10-21 | 2012-10-17 | 中山大学 | Branched chitosan derivative and preparation method thereof |
| CN102241790B (en) * | 2011-06-13 | 2013-03-06 | 中山大学 | Amphipathic chitosan derivative and preparation method and application thereof |
| CN102604114B (en) * | 2012-01-10 | 2014-06-25 | 中山大学 | Star-shaped cationic polymer containing dendriform polylysine element and preparation method thereof |
| CN102604115B (en) * | 2012-02-22 | 2013-07-10 | 天津大学 | Carboxymethyl chitosan quaternary ammonium salt/PAMAM(Polyamidoamine) core-shell nanoparticles and preparation method |
-
2012
- 2012-10-31 CN CN201210428081.6A patent/CN102924724B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN102924724A (en) | 2013-02-13 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102924724B (en) | Arborization macromolecule poly (acid amide-amine) grafting glucan and preparation method thereof | |
| CN102911368B (en) | Amphiphilic triblock copolymer with acid sensitivity and preparation method and application thereof | |
| CN102002117B (en) | Branched chitosan derivative and preparation method thereof | |
| CN103333301B (en) | Amphiphilic pH-responsive 4/6 heteroarm star-shaped copolymer and preparation method thereof | |
| CN103834002B (en) | Based on sensitivity to acid Zorubicin prodrug and preparation method thereof and the application of polyoxyethylene glycol | |
| CN107141323B (en) | Reduction/pH dual responsiveness adriamycin prodrug and the preparation method and application thereof | |
| CN103396554B (en) | Hydrogel, preparation method thereof and applications | |
| CN108017783A (en) | Polymer with high potency drugs load performance and preparation method and application | |
| CN103204998B (en) | Amphipathic polysaccharide/polypeptide block polymer containing azobenzene group and preparation method and application of block polymer | |
| CN102936337A (en) | Poly (gamma-propargyl-L-glutamate)-polyamino acid segmented copolymer, functional segmented copolymer and preparation method | |
| CN103804528A (en) | New method for preparing cyclodextrin (meth)acrylate | |
| CN102936338A (en) | Cationic lipoid plastid and preparation method thereof | |
| CN102167818B (en) | Poly(gamma-propynyl-L-glutamate) block copolymer and preparation method and hydrogel thereof | |
| CN104877092B (en) | The amphipathic copolymer of dual-target containing acetal bonds and its preparation and the application as antineoplastic drug carrier | |
| CN103467746B (en) | A kind of amphipathic nature polyalcohol and preparation method thereof | |
| CN103289082B (en) | A kind of segmented copolymer and preparation method thereof | |
| CN103304804B (en) | A kind of polyethylene glycol-amide-amine-polyamino acid is linear-dendritic block polymkeric substance and preparation method thereof | |
| CN105254867A (en) | Amphipathic polymer with main chain containing double anticancer drugs, as well as preparation method and nano-micelle of amphipathic polymer | |
| CN104974353A (en) | Poly(beta-aminoester)-based pH-responsive triblock linear polymer and micelle system | |
| CN110305301B (en) | Amphiphilic dendritic sugar-containing copolymer and synthesis method thereof | |
| CN101230139B (en) | Polycaprolactone/amylose amphiphilic block polymer as well as preparation method and use thereof | |
| CN111848685B (en) | Preparation method of amphiphilic PN = PS type phosphorus-containing tree crown macromolecule nano micelle and application of drug carrier of amphiphilic PN = PS type phosphorus-containing tree crown macromolecule nano micelle | |
| CN102977378A (en) | Amphipathic heparin-based block polymer and preparation method and application thereof | |
| CN102977358B (en) | Module compound with hydrogen bond sequence specificity combination and preparation method thereof | |
| CN102766222B (en) | P-containing modified chitosan lyotropic liquid crystal and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C41 | Transfer of patent application or patent right or utility model | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20161228 Address after: Changzhou City, Jiangsu province Hehai road 213000 No. 9 Patentee after: Changzhou Institute of Energy Storage Materials & Devices Address before: 130022 Changchun people's street, Jilin, No. 5625 Patentee before: Changchun Applied Chemistry Inst., Chinese Academy of Sciences |