Synthesis Of Polylactide/Chitosan Copolymer And Application As Gene Delivery Vectors

Poly(D,L-lactide)-graft-chitosan(PLA-g-CS) copolymer were synthesized by the introduction of PL A side chains into chitosan molecules with D,L-lactide and chitosan and applied as gene vector for DNA delivery complex. Firstly, trimethylsilyl chloride (TMS-Cl) was used to alternately protect the-NH2 and-OH groups of chitosan molecules for the formation of modified chitosan. Then, by changing the molar ratios of D,L-LA and modified chitosan, the amphipathic PLA-g-CS were prepared with different PLA molecular chains contents and kept the capacity to combine with DNA molecules. FT-IR,DSC,H-NMR,TG and elemental analysis were applied to characterize the copolymer molecular structure and properties. By the complex coacervation process and ultrasonic wave, PLA-g-CS/pEGFP-N1 nanoparticles were achieved. Laser light scattering, Zeta potential measurements, atomic force microscope (AFM) and agrose gel electrophoresis were used to investigate the characterization and the stability of PLA-g-chitosan/DNA in vitro. The DNA release curves were determined in vitro to estimate the PLA-g-chitosan/DNA complex. The cytotoxicity of PLA-g-chitosan carriers for SCG and 3T3 cells was assessed by MTT assays respectively.The analytic results proved the formation of PLA-g-CS with defferent graft degrees of PLA molecular chains. Laser light scattering (LLS) analysis showed that PLA-g-CS/ pEGFP-N1 complex nanoparticles came into being, with diameters between 91.59nm to 234.56nm and performed with irregular spheres. Zeta potential measurements of PLA-g-CS carriers were between 8.45-11.20mV. The PLA-g-CS carrier which prepared with the molar ratios more than 3 for D,L-LA to chitosan monomers, combined with DNA under the N/P ratio also greater than 3, the PLA-g-CS and DNA complex particles could be protected well from the degradation of DNase. PLA-g-CS carrier had good sustained release for DNA molecules in vitro. The cells trials indicated that the PLA-g-chitosan could be low cytotoxicity. PLA-g-chitosan will be used as a noval non-virus DNA carrier.