A cationic gene delivery vector, guanidinylated disulfide-containing poly(amido amine)(CARCBA), was synthesized by Michael addition reaction between N,N′-cystaminebisacrylamide(CBA) and guanidine hydrochloride(CAR). ...A cationic gene delivery vector, guanidinylated disulfide-containing poly(amido amine)(CARCBA), was synthesized by Michael addition reaction between N,N′-cystaminebisacrylamide(CBA) and guanidine hydrochloride(CAR). Gel permeation chromatography(GPC) was used to evaluate the molecular weight of synthesized CAR-CBA. Polyethyleneimine(PEI) with molecular weight of 25 kDa was adopted as a reference, and polyethylene glycols(PEG) with different molecular weights were used to establish a standard curve for determining the molecular weight of CAR-CBA. The effects of two critical factors, namely columns and eluents,on the molecular weight measurement of CAR-CBA were investigated to optimize the GPC quantitative method. The results showed that Ultrahydrogel columns(120, 250) and HAc–NaAc(0.5 M, pH 4.5) buffer solution were the optimal column and GPC eluent, respectively.The molecular weight of the synthesized CAR-CBA was analyzed by the optimized GPC method and determined to be 24.66 kDa.展开更多
In this paper, a novel series of bis [(aminoethyl)]-amine cationic lipid derivatives have been synthesized and identified to purity by NMR and Elemental analysis. B16-F0 cells were transfected with cationic lipid/pEGF...In this paper, a novel series of bis [(aminoethyl)]-amine cationic lipid derivatives have been synthesized and identified to purity by NMR and Elemental analysis. B16-F0 cells were transfected with cationic lipid/pEGFP-N1 and cationic lipid/ß-gal lipoplexes complexed at +/- charge ratios of 1:1, 2:1, and 4:1. Dimyristoyl derivative showed highest activity at charge ratio 2:1 and both dimyristoyl and dioleoyl derivatives showed similar ß-gal activity at charge ratios 4:1. In 40 mM tris buffer pH 7.2 the dioleoyl derivative was able to fully complex with and retard pDNA at charge ratios above 2:1. None of the other lipid derivatives, dilauroyl, dimyristoyl, dipalmitoyl and distearoyl were able to fully neutralize the plasmid DNA at charge ratios similar to those used in the transfection experiment. The gel-to-liquid phase transition temperatures for dimyristoyl, dipalmitoyl and distearoyl were determined by a fluorescence anisotropy method to be 27.5°C, 32.5°C and 39°C, respectively. A gel-to-liquid crystalline phase transition temperature below 37°C, appears to be the crucial property that cationic lipids have to possess in order to mediate high levels of in vitro transfection activity in the absence of other helper lipids.展开更多
基金the National Natural Science Foundation of China for financial support(No.81373335)
文摘A cationic gene delivery vector, guanidinylated disulfide-containing poly(amido amine)(CARCBA), was synthesized by Michael addition reaction between N,N′-cystaminebisacrylamide(CBA) and guanidine hydrochloride(CAR). Gel permeation chromatography(GPC) was used to evaluate the molecular weight of synthesized CAR-CBA. Polyethyleneimine(PEI) with molecular weight of 25 kDa was adopted as a reference, and polyethylene glycols(PEG) with different molecular weights were used to establish a standard curve for determining the molecular weight of CAR-CBA. The effects of two critical factors, namely columns and eluents,on the molecular weight measurement of CAR-CBA were investigated to optimize the GPC quantitative method. The results showed that Ultrahydrogel columns(120, 250) and HAc–NaAc(0.5 M, pH 4.5) buffer solution were the optimal column and GPC eluent, respectively.The molecular weight of the synthesized CAR-CBA was analyzed by the optimized GPC method and determined to be 24.66 kDa.
文摘In this paper, a novel series of bis [(aminoethyl)]-amine cationic lipid derivatives have been synthesized and identified to purity by NMR and Elemental analysis. B16-F0 cells were transfected with cationic lipid/pEGFP-N1 and cationic lipid/ß-gal lipoplexes complexed at +/- charge ratios of 1:1, 2:1, and 4:1. Dimyristoyl derivative showed highest activity at charge ratio 2:1 and both dimyristoyl and dioleoyl derivatives showed similar ß-gal activity at charge ratios 4:1. In 40 mM tris buffer pH 7.2 the dioleoyl derivative was able to fully complex with and retard pDNA at charge ratios above 2:1. None of the other lipid derivatives, dilauroyl, dimyristoyl, dipalmitoyl and distearoyl were able to fully neutralize the plasmid DNA at charge ratios similar to those used in the transfection experiment. The gel-to-liquid phase transition temperatures for dimyristoyl, dipalmitoyl and distearoyl were determined by a fluorescence anisotropy method to be 27.5°C, 32.5°C and 39°C, respectively. A gel-to-liquid crystalline phase transition temperature below 37°C, appears to be the crucial property that cationic lipids have to possess in order to mediate high levels of in vitro transfection activity in the absence of other helper lipids.
