PREPARATION METHOD FOR AND USE OF REDOX-RESPONSIVE CHITOSAN-LIPOSOME

Abstract

The present invention provides a preparation method of a redox-responsive chitosan-liposome and use thereof, wherein the method uses dithiobis succinimidyl-substituted ester to synthesize a redox-responsive and disulphide bonded double fatty chain substituent phosphatidylethanolamine-s-s-chitosan. Using the synthesized double fatty chain substituent phosphatidylethanolamine chitosan, by a post-insertion and self-assembly method, to modify liposome, to construct a double fatty chain substituent phosphatidylethanolamine chitosan-liposome drug carrier having a redox-responsive chitosan brush on the surface thereof. The chitosan-liposome constructed in the present invention not only has the strong cell adhesion property and the antiserum property, but also has environmental response properties, being suitable for the intravenous injection. The present invention also provides the use of the chitosan-liposome encapsulating super-paramagnetic ferroferric oxide nanoparticles in drug delivery, which has high drug delivery efficiency and high biocompatibility, and has broad application prospects.

Claims

1. A redox-responsive chitosan, wherein the chitosan has a structure of formula (I): ##STR00002## wherein, L=CO(CH.sub.2).sub.aSS(CH.sub.2).sub.bCO, a=1-5, b=1-5; and R and R are identical or different C.sub.xH.sub.y, wherein x=11-17, y=21-35.

2. The redox-responsive chitosan according to claim 1, wherein L=CO(CH.sub.2).sub.2SS(CH.sub.2).sub.2CO, R and R are identical or different C.sub.11H.sub.23, C.sub.13H.sub.27, C.sub.17H.sub.35 or C.sub.17H.sub.33.

3. A method of preparing the redox-responsive chitosan according to claim 1, wherein firstly the chitosan is dissolved in water, sufficiently dissolved, under stirring, dropwise added to a DMSO solution of dithiobis succinimidyl-substituted ester, after reacting at 20-60 C. for 1-24 h, an ethanol solution of double fatty chain substituent phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 20-60 C. for 1-24 h, after rotary evaporation, the reaction solution is subjected to dialysis, lyophilize, to prepare the redox-responsive chitosan.

4. The method of preparing the redox-responsive chitosan according to claim 3, wherein a weight average molecular weight of the chitosan is 500-10000 Da, and a degree of deacetylation is 65-95%.

5. The method of preparing the redox-responsive chitosan according to claim 3, wherein the double fatty chain substituent phosphatidylethanolamine is one or more of 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine, 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine, 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine, 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine, and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine.

6. The method of preparing the redox-responsive chitosan according to claim 3, wherein the double fatty chain substituent phosphatidylethanolamine is used in an amount of 0.1-1 times a molar equivalent of a repeating unit of the chitosan, and a reaction condition is stirring and reacting at 20-50 C. for 2-48 h.

7. The method of preparing the redox-responsive chitosan according to claim 3, wherein the double fatty chain substituent phosphatidylethanolamine is used in an amount of 0.3-0.6 times a molar equivalent of a repeating unit of the chitosan, and a reaction condition is stirring and reacting at 30-50 C. for 4-12 h.

8. A method of preparing a redox-responsive chitosan-liposome, wherein using the double fatty chain substituent phosphatidylethanolamine according to claim 1 to modify a cationic liposome by a post-insertion and self-assembly method, to prepare the redox-responsive chitosan-liposome.

9. The method of preparing the redox-responsive chitosan-liposome according to claim 8, wherein the cationic liposome is any one of DOTAP, Lipofectin and Lipofectamin 2000, and a hydrophilic core of the cationic liposome encapsulating super-paramagnetic ferroferric oxide nanoparticles having a particle diameter of 1-30 nm.

10. Use of the redox-responsive chitosan-liposome according to claim 8 in drug delivery.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0025] FIG. 1 is a FTIR spectrum of redox-responsive chitosan prepared in example 1;

[0026] FIG. 2 is a .sup.1HNMR spectrum of redox-responsive chitosan prepared in example 1;

[0027] FIG. 3 is a TEM photograph of a redox-responsive chitosan-DOTAP liposome-SPIO composite carrier prepared in example 1;

[0028] FIG. 4 is a graph showing the retardation ability to DNA of the redox-responsive chitosan-liposome prepared in Example 1;

[0029] FIG. 5 shows the gene transfection efficiency of redox-responsive chitosan-liposome prepared in the present invention; and

[0030] FIG. 6 shows the cytotoxicity of redox-responsive chitosan-liposome prepared in the present invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0031] The present invention is described in details below with reference to the drawings and specific examples, but it is not intended to limit the scope of the invention. Unless otherwise stated, the experimental methods used in the present invention are all conventional methods, and the used experimental equipment, materials, reagents, and the like can be purchased from chemical companies.

Example 1

[0032] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 0.5 g an ethanol solution of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, after reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0033] 1 mg/mL the redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 2

[0034] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 0.5 g an ethanol solution of 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0035] 1 mg/mL the redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 3

[0036] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 0.5 g an ethanol solution of 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan.

[0037] 1 mg/mL the redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 4

[0038] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolved it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 0.5 g an ethanol solution of 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0039] 1 mg/mL the redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of Lipofectin cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 5

[0040] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 0.5 g an ethanol solution of 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0041] 1 mg/mL the redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of Lipofectin cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 6

[0042] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 1.0 g an ethanol solution of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0043] 1 mg/mL the redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of Lipofectamin 2000 cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 7

[0044] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 1.0 g an ethanol solution of 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0045] 1 mg/mL the redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 8

[0046] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 1.0 g an ethanol solution of 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan.

[0047] 1 mg/mL the redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of Lipofectamin 2000 cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 9

[0048] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 2 h, 1.0 g an ethanol solution of 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0049] 1 mg/mL the redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 10

[0050] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to dissolve it sufficiently. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobis(succinimidyl propionate), after reacting at 30 C. for 2 h, 1.0 g an ethanol solution of 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 2 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0051] 1 mg/mL the redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 100 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 11

[0052] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobis(succinimidyl propionate), after reacting at 30 C. for 4 h, 1.0 g an ethanol solution of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0053] 1 mg/mL the redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 12

[0054] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobis(succinimidyl propionate), after reacting at 30 C. for 4 h, 1.0 g an ethanol solution of 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0055] 1 mg/mL the redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 13

[0056] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 4 h, 1.0 g an ethanol solution of 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan.

[0057] 1 mg/mL the redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 14

[0058] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 4 h, 1.0 g an ethanol solution of 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0059] 1 mg/mL the redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 15

[0060] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 30 C. for 4 h, 1.0 g an ethanol solution of 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 30 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0061] 1 mg/mL the redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 1 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 16

[0062] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 40 C. for 4 h, 1.0 g an ethanol solution of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 40 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0063] 1 mg/mL the redox-responsive 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 2 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 17

[0064] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 40 C. for 4 h, 1.0 g an ethanol solution of 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 40 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0065] 1 mg/mL the redox-responsive 1,2-distearoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 2 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 18

[0066] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 40 C. for 4 h, 1.0 g an ethanol solution of 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 40 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan.

[0067] 1 mg/mL the redox-responsive 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 2 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 19

[0068] 0.5 g chitosan (CSO) having a molecular weight of 1 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobis(succinimidyl propionate), after reacting at 40 C. for 4 h, 1.0 g an ethanol solution of 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine is continuously added dropwise to the reaction solution, reacting at 40 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan.

[0069] 1 mg/mL the redox-responsive 1,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 2 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

Example 20

[0070] 0.5 g chitosan (CSO) having a molecular weight of 5 kDa is dissolved in 100 mL water, and sonicated for 30 min to sufficiently dissolve it. Under stirring, the aqueous solution of chitosan is dropwise added to a DMSO solution of dithiobissuccinimidyl propionate, after reacting at 40 C. for 4 h, 1.0 g an ethanol solution of 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine is continuously added dropwise to the reaction solution, reacting at 40 C. for 4 h, thereafter the reaction solution is subjected to rotary evaporation, dialysis and lyophilize to prepare redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan.

[0071] 1 mg/mL the redox-responsive 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine chitosan aqueous solution is prepared, and take 500 L to mix with 1 mL of DOTAP cationic liposome containing SPIO by ultrasonic method, then left stand for 2 h, and modifying the liposome by means of post-insertion and self-assembly to prepare a liposome drug carrier having a redox-responsive chitosan brush on a surface thereof.

[0072] Gene Transfer Assay of Liposome Having a Redox-Responsive Chitosan Brush.

[0073] A pGL3 plasmid is used as a reporter gene to evaluate gene transfer properties of the liposome carrier having a redox-responsive chitosan brush, and the cells used are human non-small cell lung cancer A549 cell lines. The cultured cells are plated, cultured in an incubator until the cell fusion degree reaches 80%, and then subject to gene transfer. During transporting, the complete medium is aspirated, and washed twice with PBS, and transported under serum conditions, 400 L of medium containing 10% serum and different N/P ratios (mass ratio) of the redox-responsive polysome@SPIO (Example 3)DNA complexes are added, after culture for 18 h, the medium is aspirated, and after replacing with a fresh medium containing 10% serum and continually culture for 32 h. The photon intensity is measured by a BioTek Synergy2 multi-function microplate reader according to the instruction book provided by the luciferase kit, and the total protein concentration is detected by BCA, thereby normalizing the results to RLU/mg protein (relative photon number per mg of protein).

[0074] Cytotoxicity of Liposome Having a Redox-Responsive Chitosan Brush.

[0075] The cytotoxicity of the carrier is evaluated by MTT method. Cells are grown in a 96-well cell culture plate, 3 wells in parallel, 510.sup.4 cells were grown per well, and cultured at 37 C. in a 5% CO.sub.2 incubator until the cell fusion degree reaches 85% or more. The medium is removed, PBS is used to wash twice, a fresh medium and the carrier to be measured are added, after culturing for 24 h, 20 L 5 mg/mL of MTT solution is added to each well, continuously culturing at 37 C. for 4 h, the medium is removed, and the culture is terminated. Succinate dehydrogenase in mitochondria of living cells reduces MTT to generate formazan, which is dissolved by adding 150 L DMSO to each well, and continually incubating at 37 C. for 30 min. The absorbance value of each well at 570 nm was measured by a multi-functional microplate reader (Sunrise Tecan), the 96-well plate was automatically mixed for 600 s before detection, and a cell-free medium being used to zero the microplate reader. Cell viability is calculated according Formula 1.1:

cell viability (%)=A570.sub.SMP/A570.sub.CTL100(1.1)

[0076] Wherein A570.sub.SMP refers to an absorbance value of a cell plate to which the carrier or complex to be measured is added, and A570.sub.CTL refers to an absorbance value of a cell plate containing only the medium.

[0077] The above is merely the preferred examples of the present invention, and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, and the like that made within the spirit and principles of the present invention should be included in the protection of the present invention.