Chitosan nanoparticles produced with the gradual temperature decrease technique for sustained gene delivery.

Different techniques have been employed to produce chitosan nanoparticles; thus, suitable alternatives and easy-handling production methods are highly desired. We used a tank reactor with baffles and mechanical stirring with Cowles impellers that can feasibly be up-scaled and allows for the production of homogenous chitosan nanoparticles. First, we explored the effects of temperature on the production of chitosan (CHI)/pentasodium tripolyphosphate (TPP) nanoparticles. We compared the effect of gradual temperature decrease between 40 to 55 °C with the effect of isothermal processes (25, 45 and 55 °C). CHI/TPP nanoparticles produced by the gradual temperature decrease technique had polydispersity indices (PDIs) that were significantly smaller than the nanoparticles produced by isothermal processes. To further control the production process, we employed an experimental design to determine the effects of CHI and TPP concentrations and CHI/TPP mass ratios in the gradual temperature decrease technique. Under all conditions, the sizes of the CHI/TPP nanoparticles were on the nanoscale. Gene delivery capacities of the nanoparticles were evaluated; the plasmid pEGFP-N1 was incorporated (10 and 40% of the chitosan weight) into the nanoparticles and incubated with 293A cells. The in vitro transfection efficiencies were evaluated over 120 h and indicated sustained gene delivery at 10% pDNA. The polyplexes demonstrated some cytotoxicity after 120 h. These findings will contribute to the development of processes for generating CHI/TPP nanoparticles for gene delivery applications. [ABSTRACT FROM AUTHOR]