CN108148867A - A kind of poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid and preparation method thereof - Google Patents
A kind of poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid and preparation method thereof Download PDFInfo
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- CN108148867A CN108148867A CN201711475288.8A CN201711475288A CN108148867A CN 108148867 A CN108148867 A CN 108148867A CN 201711475288 A CN201711475288 A CN 201711475288A CN 108148867 A CN108148867 A CN 108148867A
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- acetimide
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/87—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
- C08B37/0024—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid beta-D-Glucans; (beta-1,3)-D-Glucans, e.g. paramylon, coriolan, sclerotan, pachyman, callose, scleroglucan, schizophyllan, laminaran, lentinan or curdlan; (beta-1,6)-D-Glucans, e.g. pustulan; (beta-1,4)-D-Glucans; (beta-1,3)(beta-1,4)-D-Glucans, e.g. lichenan; Derivatives thereof
- C08B37/0027—2-Acetamido-2-deoxy-beta-glucans; Derivatives thereof
- C08B37/003—Chitin, i.e. 2-acetamido-2-deoxy-(beta-1,4)-D-glucan or N-acetyl-beta-1,4-D-glucosamine; Chitosan, i.e. deacetylated product of chitin or (beta-1,4)-D-glucosamine; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G81/00—Macromolecular compounds obtained by interreacting polymers in the absence of monomers, e.g. block polymers
Abstract
The invention discloses poly- acetimide nano-gene carriers of a kind of chitosan magnetic of nucleocapsid and preparation method thereof, including magnetic Nano kernel, the shell being coated on outside magnetic Nano kernel 1 and shell 2, magnetic Nano kernel and shell 1 are fitted close, and shell 1 and shell 2 are fitted close；The material of magnetic Nano kernel is Fe3O4, the material of shell 1 is that (molecular formula is (C to chitosan6H11NO4)m), the material of shell 2 is that (molecular formula is (CH to polyethyleneimine2CH2NH)n).The genophore of the present invention has magnetic response characteristic, enhance gene transfection can guidance quality, improve the transfection efficiency of gene, there is high-biocompatibility and high gene transfection efficiency, can recycle, the cost of gene transfection can be reduced.
Description
Technical field
The present invention relates to technical field of biological material, more particularly to a kind of poly- acetimide nanometer of the chitosan magnetic of nucleocapsid
Genophore and preparation method thereof.
Background technology
Genophore is a kind of tool by gene delivery into cell, and main function is that delivery target gene enters host
Cell simultaneously successfully can be replicated and be expressed, and be widely used in biological medicine, cell engineering, enzyme engineering, molecular biosciences
Etc. fields.But common genophore is not recyclable, improves the application cost of genetic engineering.Genophore is difficult to back
The main reason for receipts, is that genophore does not have magnetic responsiveness, can not be in situation existing for magnetic field after gene transfection is completed
It is lower by enriching and recovering.Therefore, there is an urgent need to research and develop a kind of genophore with magnetic responsiveness, the recyclable of genophore is improved
Performance reduces the cost of gene transfection.
Chitosan is a kind of natural polymer, has excellent biocompatibility, blood compatibility, safety, microorganism
Degradability, in the fields such as medicine, food, chemical industry, cosmetics, water process, METAL EXTRACTION and recycling, biochemistry and biomedical engineering
Application study achieve major progress.Polyethyleneimine (PEI) is applied to biological doctor as cationic gene delivery vector
Research is learned, but PEI also increases cytotoxicity, therefore its application in vivo while efficiency gene transfection is improved
It is restricted.How to realize the less toxic high efficiency gene transfection of PEI becomes a great problem.
Invention content
It is an object of the invention to overcome shortcoming in the prior art, a kind of chitosan magnetic of nucleocapsid is provided
Poly- acetimide nano-gene carrier makes genophore obtain magnetic responsiveness, passes through chitosan by the ferroso-ferric oxide of internal layer
Outer shell makes genophore have high-biocompatibility, material is made to obtain high gene transfection efficiency by polyethyleneimine.
Another object of the present invention is to provide the poly- acetimide nano gene of the chitosan magnetic of above-mentioned nucleocapsid to carry
The preparation method of body.
The purpose of the present invention is achieved through the following technical solutions：
A kind of poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid, including magnetic Nano kernel, cladding
Shell 1 and shell 2 outside magnetic Nano kernel, magnetic Nano kernel and shell 1 are fitted close, and shell 1 and shell 2 are closely matched
It closes；The material of magnetic Nano kernel is Fe3O4, the material of shell 1 is that (molecular formula is (C to chitosan6H11NO4)m), the material of shell 2
Expect that (molecular formula is (CH for polyethyleneimine2CH2NH)n)。
In the poly- acetimide nano-gene carrier of chitosan magnetic of nucleocapsid, poly- acetimide surface carries strong positive electricity
Lotus (zeta current potentials>20mV), magnetic Fe3O4Kernel carries strong negative electrical charge (zeta current potentials<-20mV).
The preparation method of the poly- acetimide nano-gene carrier of chitosan magnetic of above-mentioned nucleocapsid, including following steps
Suddenly：
(1) soluble divalent iron salt and trivalent iron salt, chitosan are added in into reaction vessel, are fully dissolved using solvent,
Then it adds in reagent A to be reacted, obtains the magnetic Fe containing chitosan shell 13O4；
(2) by the magnetic Fe containing chitosan shell 13O4, polyethyleneimine (molecular weight 25kDa-750kDa) and penta 2
Aldehyde is added in reaction vessel, is then added in reagent B, is stirred to react, and the poly- acetimide of chitosan magnetic that nucleocapsid is made is received
Rice genophore.
In step (1), the molal weight ratio 1 of divalent iron salt, trivalent iron salt and chitosan:(1.5~2.5):(1~2),
Middle divalent iron salt, trivalent iron salt are based on the molal weight of ferro element, and chitosan is based on the molal weight of aminoglucose saccharide residue；Two
Valency molysite is preferably ferrous sulfate, ferrous nitrate, frerrous chloride；Trivalent iron salt is preferably ferric sulfate, ferric nitrate, iron chloride.
Step (1), the solvent are 1% acetic acid solutions.
In step (1), the reagent A is sodium tripolyphosphate, water and ammonia spirit.
In step (1), the temperature of reaction is 60~90 DEG C, and the time of reaction is 0.5~3 hour.
In step (2), reagent B is 1- ethyl-3-methylimidazole hydrochlorides.
In step (2), the magnetic Fe containing chitosan shell 13O4, polyethyleneimine mass ratio be 1:(1~2)；Poly- second
The mass ratio of alkene imines and glutaraldehyde is 1:(1~2)；1- ethyl-3-methylimidazoles hydrochloride and three kinds of substances (contain chitosan
The magnetic Fe of shell 13O4, polyethyleneimine, glutaraldehyde) quality sum ratio be (5~100):1.
In step (2), the temperature being stirred to react is 30~90 DEG C, and the time is preferably 0.1~24 hour.
The present invention has the following advantages that compared with prior art and effect：
(1) genophore of the invention has magnetic response characteristic, enhance gene transfection can guidance quality, improve gene
Transfection efficiency.
(2) genophore of the invention has high-biocompatibility and high gene transfection efficiency.
(3) genophore of the invention can recycle, and can reduce the cost of gene transfection.
Description of the drawings
Fig. 1 is the structure diagram of the poly- acetimide nano-gene carrier of chitosan magnetic of nucleocapsid.
Specific embodiment
Further detailed description is done to the present invention, but the implementation of the present invention is not limited to this with reference to embodiment.
Embodiment 1
As shown in Figure 1, a kind of poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid, including magnetic Nano
Kernel, the shell 1 being coated on outside magnetic Nano kernel and shell 2, magnetic Nano kernel and shell 1 are fitted close, shell 1 and outer
Shell 2 is fitted close；The material of magnetic Nano kernel is Fe3O4, the material of shell 1 is that (molecular formula is (C to chitosan6H11NO4)m),
The material of shell 2 is that (molecular formula is (CH to polyethyleneimine2CH2NH)n)。
Preparation method is：By soluble frerrous chloride and iron chloride and chitosan (being calculated by aminoglucose saccharide residue)
According to molal weight than 1:1.5:1 adds in reaction vessel, after then adding in the dissolving of 1% acetic acid solution, addition sodium tripolyphosphate,
Water, ammonium hydroxide react 1h at 60 DEG C, and the magnetic Fe containing chitosan shell is made3O4；Then 10g is contained into chitosan shell
Magnetic Fe3O4, 10g polyethyleneimines (molecular weight 25kDa) and 15g glutaraldehydes add in reaction vessel, then add in 350g1-
Ethyl-3-methylimidazole hydrochloride is stirred to react 0.1 hour at 30 DEG C, and the poly- acetyl of chitosan magnetic of nucleocapsid is made
Imine nanometer genophore.
Embodiment 2
By soluble ferrous nitrate and ferric nitrate and chitosan (being calculated by aminoglucose saccharide residue) according to 1：2.5:1 rubs
You add in reaction vessel by mass ratio, after then adding in the dissolving of 1% acetic acid solution, add in sodium tripolyphosphate, water, are reacted at 90 DEG C
The magnetic Fe containing chitosan shell is made in 3h3O4.By the magnetic Fe containing chitosan shell obtained by 10g3O4, the poly- second of 15g
Alkene imines (molecular weight 25kDa) and 30g glutaraldehydes are added in reaction vessel, then add in 2750g1- ethyl-3-methylimidazoles
Hydrochloride is stirred to react 18 hours at 60 DEG C, and the poly- acetimide nano-gene carrier of chitosan magnetic of nucleocapsid is made.
Embodiment 3
By soluble ferrous sulfate and ferric nitrate and chitosan (being calculated by aminoglucose saccharide residue) according to 1：2.5:1 rubs
You add in reaction vessel by mass ratio, after then adding in the dissolving of 1% acetic acid solution, add in sodium tripolyphosphate, water, are reacted at 75 DEG C
The magnetic Fe containing chitosan shell is made in 1.5h3O4.By the magnetic Fe containing chitosan shell obtained by 10g3O4, 10g gathers
Aziridine (molecular weight 750kDa) and 20g glutaraldehydes are added in reaction vessel, then add in 4000g1- ethyl -3- methyl
Imidazole hydrochloride is stirred to react 24 hours at 90 DEG C, and the poly- acetimide nano gene of chitosan magnetic of nucleocapsid is made
Carrier.
Embodiment 4
By soluble frerrous chloride and ferric sulfate and chitosan (being calculated by aminoglucose saccharide residue) according to 1：1.5:
.1.5 molal weight is than adding in reaction vessel, after then adding in the dissolving of 1% acetic acid solution, adds in addition sodium tripolyphosphate, water,
3h is reacted at 60 DEG C, the magnetic Fe containing chitosan shell is made3O4.The 10g of gained is contained to the magnetism of chitosan shell
Fe3O4, 10g polyethyleneimines (molecular weight 200kDa) and 10g glutaraldehydes add in reaction vessel, then add in 300g1- second
Base -3- methylimidazolium chlorides are stirred to react 6 hours at 30 DEG C, and the poly- acetimide of chitosan magnetic of nucleocapsid is made
Nano-gene carrier.
Embodiment 5
By soluble ferrous nitrate and ferric sulfate and chitosan (being calculated by aminoglucose saccharide residue) according to 1：2:1.5 it rubs
You add in reaction vessel by mass ratio, after then adding in the dissolving of 1% acetic acid solution, sodium tripolyphosphate, water are added in, at 90 DEG C
1h is reacted, obtained core contains the magnetic Fe of chitosan shell3O4.The 10g of gained is contained to the magnetic Fe of chitosan shell3O4、
20g polyethyleneimines (molecular weight 25kDa) and 20g glutaraldehydes are added in reaction vessel, then add in 250g1- ethyl -3- first
Base imidazole hydrochloride is stirred to react 18 hours at 80 DEG C, and the poly- acetimide nanometer base of chitosan magnetic of nucleocapsid is made
Because of carrier.
Claims (10)
1. a kind of poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid, it is characterised in that：Including magnetic Nano
Kernel, the shell 1 being coated on outside magnetic Nano kernel and shell 2, magnetic Nano kernel and shell 1 are fitted close, shell 1 and outer
Shell 2 is fitted close；The material of magnetic Nano kernel is Fe3O4, the material of shell 1 is that (molecular formula is (C to chitosan6H11NO4)m),
The material of shell 2 is that (molecular formula is (CH to polyethyleneimine2CH2NH)n)。
2. the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 1, feature exist
In：Poly- acetimide surface carries strong positive charge (zeta current potentials>20mV), magnetic Fe3O4Kernel carries strong negative electrical charge (zeta
Current potential<-20mV).
3. a kind of preparation of the poly- acetimide nano-gene carrier of the chitosan magnetic of the nucleocapsid described in claims 1 or 2
Method, it is characterised in that include the following steps：
(1) soluble divalent iron salt and trivalent iron salt, chitosan are added in into reaction vessel, is fully dissolved using solvent, then
It adds in reagent A to be reacted, obtains the magnetic Fe containing chitosan shell 13O4；
(2) by the magnetic Fe containing chitosan shell 13O4, polyethyleneimine (molecular weight 25kDa-750kDa) and glutaraldehyde add
Enter in reaction vessel, then add in reagent B, be stirred to react, the poly- acetimide nanometer base of chitosan magnetic of nucleocapsid is made
Because of carrier.
4. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：In step (1), the molal weight ratio 1 of divalent iron salt, trivalent iron salt and chitosan:(1.5~2.5):(1
~2), for wherein divalent iron salt, trivalent iron salt based on the molal weight of ferro element, chitosan presses mole matter of aminoglucose saccharide residue
Gauge；Divalent iron salt is ferrous sulfate, ferrous nitrate, frerrous chloride；Trivalent iron salt is ferric sulfate, ferric nitrate, iron chloride.
5. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：Step (1), the solvent are 1% acetic acid solutions.
6. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：In step (1), the reagent A is sodium tripolyphosphate, water and ammonia spirit.
7. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：In step (1), the temperature of reaction is 60~90 DEG C, and the time of reaction is 0.5~3 hour.
8. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：In step (2), reagent B is 1- ethyl-3-methylimidazole hydrochlorides.
9. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：In step (2), the magnetic Fe containing chitosan shell 13O4, polyethyleneimine mass ratio be 1:(1~
2)；The mass ratio of polyethyleneimine and glutaraldehyde is 1:(1~2)；1- ethyl-3-methylimidazoles hydrochloride (contains with three kinds of substances
There is the magnetic Fe of chitosan shell 13O4, polyethyleneimine, glutaraldehyde) quality sum ratio be (5~100):1.
10. the preparation side of the poly- acetimide nano-gene carrier of the chitosan magnetic of nucleocapsid according to claim 3
Method, it is characterised in that：In step (2), the temperature being stirred to react is 30~90 DEG C, and the time is 0.1~24 hour.
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Citations (4)
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|---|---|---|---|---|
| CN101130086A (en) * | 2007-08-01 | 2008-02-27 | 浙江大学 | Method for preparing drug administration carrier of gene with polyethylene imine beautify chitosan |
| CN103788380A (en) * | 2014-01-29 | 2014-05-14 | 大连民族学院 | Method for cleanly preparing chitosan grafted polyethylenimine non-viral gene vector |
| CN105195111A (en) * | 2015-10-26 | 2015-12-30 | 福建农林大学 | Preparation method of polyethylenimine modified chitosan-coated magnetic nanocellulose adsorbent and product thereof |
| CN105560190A (en) * | 2016-01-25 | 2016-05-11 | 河南工业大学 | Novel double-targeting gene conveying system based on magnetic nanoparticles and preparing method thereof |
-
2017
- 2017-12-29 CN CN201711475288.8A patent/CN108148867A/en not_active Withdrawn
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101130086A (en) * | 2007-08-01 | 2008-02-27 | 浙江大学 | Method for preparing drug administration carrier of gene with polyethylene imine beautify chitosan |
| CN103788380A (en) * | 2014-01-29 | 2014-05-14 | 大连民族学院 | Method for cleanly preparing chitosan grafted polyethylenimine non-viral gene vector |
| CN105195111A (en) * | 2015-10-26 | 2015-12-30 | 福建农林大学 | Preparation method of polyethylenimine modified chitosan-coated magnetic nanocellulose adsorbent and product thereof |
| CN105560190A (en) * | 2016-01-25 | 2016-05-11 | 河南工业大学 | Novel double-targeting gene conveying system based on magnetic nanoparticles and preparing method thereof |
Non-Patent Citations (1)
| Title |
|---|
| CHILI ZHAO等: ""A facile one-step method for preparation of Fe3O4/CS/INH nanoparticles as a targeted drug delivery for tuberculosis"", 《MATERIALS SCIENCE AND ENGINEERING C》 * |
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