Síntese, caracterização e estudo físico-químico das nanopartículas formadas pela interação de amino derivados de quitosana com DNA
Carregando...
Data
2011-08-11
Autores
Oliveira, Franciele de Paula Pansani [UNESP]
Título da Revista
ISSN da Revista
Título de Volume
Editor
Universidade Estadual Paulista (Unesp)
Resumo
O presente trabalho apresenta a síntese e caracterizaçao de derivados de quitosana obtidos pela reação com cloreto de 2-cloro dietilaminoetilamônio (DEAE Cl). Os derivados obtidos foram utilizados na síntese de nanopartículas para liberação controlada de genes. Inicialmente a quitosana comercial foi desacetilada e as sínteses foram realizadas em meio aquoso alcalino pela reação de substituição nucleofílica dos grupos amino da quitosana. A caracterização dos derivados foi realizada utilizando-se a espectroscopia de ressonância magnética nuclear de prótons (RMN-1 H), titulações potenciométricas e condutimétricas. Os derivados obtidos apresentaram proporções crescentes de DEAE (15%, 24%, 80% e 117%) e diferentes capacidades de tamponamento. A massa molecular viscosimétrica foi determinada por medidas de viscosidade e variou de 17 a 21 kDa. As titulações potenciométricas mostraram que a modificação de quitosana com o grupo DEAE permite aumentar a capacidade tamponante com respeito à quitosana e pode contribuir para aumentar a liberação de DNA e a eficiência do processo de transfecção gênica. O estudo dos derivados com DNA foi realizado utilizando-se as técnicas de fluorescência, eletroforese, espalhamento de luz dinâmico (tamanho das nanopartículas) e potencial zeta. O ensaio com brometo de etídio mostrou que a força de interação de DNA com os derivados depende tanto do grau de substituição por DEAE quanto da razão de cargas (N/P). Em altos valores de pH (7,4) todos os derivados interagem fortemente com DNA devido a presença dos grupos amino terciários e quaternários ligados a cadeia de quitosana. Os resultados de eletroforese mostram que a força de interação aumenta com o grau de substituição de DEAE e os resultados iniciais de espalhamento de luz mostram que nanopartículas estáveis podem ser obtidas em pH fisiólogico...
This work presents the synthesis and characterization of chitosan derivatives obtained by reaction of chitosan with diethylaminoethyl chloride - Cl DEAE. The obtained derivatives were employed in the synthesis of nanoparticles for controlled delivery of genes. Commercial chitosan was deacetylated and the reactions were carried out in aqueous solution at pH 8.0 by the nucleophilic subsitituion of the chitosan amino groups. The characterization was performed using 1H NMR spectroscopy, potentiometric and conductimetric titrations. The obtained derivatives contain increasing proportions of DEAE (15%, 24%, 80% and 117%) and different buffering capacities. Viscosity measurements were performed to determine the viscoimetric molecular weight (Mv), which varied from 17 to 21 kDa. The potentiometric titrations showed that DEAE groups increase the buffering capacity of chitosan and this may contribute to increase the DNA delivery from nanoparticles and the efficiency of gene transfection. The study of interaction between the derivatives and DNA was performed using fluorescence, gel electrophoresis, dynamic light scattering and zeta potential. The ethidium bromide (EtBr) assay showed that the strenght of interaction between the derivatives and DNA depends on both, the degree of substitution (DS) and N/P charge ratio. At physiological pH all derivatives interacted with DNA due to the presence of tertiary quaternary amino groups attached to the chitosan main chain. The gel electrophoresis results showed that the strength of interaction increases with DS and the initial results employing light scattering techniques show that stable nanoparticles can be obtained in phyisological pH, which is clearly an important property in comparison to deacetylated chitosan. Transfection studies carried out with these derivatives confirmed that the presence of DEAE groups improves the transfection efficiency... (Complete abstract click electronic access below)
This work presents the synthesis and characterization of chitosan derivatives obtained by reaction of chitosan with diethylaminoethyl chloride - Cl DEAE. The obtained derivatives were employed in the synthesis of nanoparticles for controlled delivery of genes. Commercial chitosan was deacetylated and the reactions were carried out in aqueous solution at pH 8.0 by the nucleophilic subsitituion of the chitosan amino groups. The characterization was performed using 1H NMR spectroscopy, potentiometric and conductimetric titrations. The obtained derivatives contain increasing proportions of DEAE (15%, 24%, 80% and 117%) and different buffering capacities. Viscosity measurements were performed to determine the viscoimetric molecular weight (Mv), which varied from 17 to 21 kDa. The potentiometric titrations showed that DEAE groups increase the buffering capacity of chitosan and this may contribute to increase the DNA delivery from nanoparticles and the efficiency of gene transfection. The study of interaction between the derivatives and DNA was performed using fluorescence, gel electrophoresis, dynamic light scattering and zeta potential. The ethidium bromide (EtBr) assay showed that the strenght of interaction between the derivatives and DNA depends on both, the degree of substitution (DS) and N/P charge ratio. At physiological pH all derivatives interacted with DNA due to the presence of tertiary quaternary amino groups attached to the chitosan main chain. The gel electrophoresis results showed that the strength of interaction increases with DS and the initial results employing light scattering techniques show that stable nanoparticles can be obtained in phyisological pH, which is clearly an important property in comparison to deacetylated chitosan. Transfection studies carried out with these derivatives confirmed that the presence of DEAE groups improves the transfection efficiency... (Complete abstract click electronic access below)
Descrição
Palavras-chave
Biofísica, Biologia molecular, Físico-química, Quitosana, Nanopartículas, Biofísica molecular, Chitosan, Gene therapy, Derivates
Como citar
OLIVEIRA, Franciele de Paula Pansani. Síntese, caracterização e estudo físico-químico das nanopartículas formadas pela interação de amino derivados de quitosana com DNA. 2011. 56 f. Dissertação (mestrado) - Universidade Estadual Paulista, Instituto de Biociências, Letras e Ciências Exatas, 2011.