**4.3 Improving transfection efficiency**

There have also some attempts to modify the chemical structure of chitosan to improve transfection efficiency, which have involved hydrophilic and hydrophobic modifications. The main purpose of hydrophilic modification of chitosan is to increase solubility and reduce sensitivity of chitosan-DNA complexes to pH, as well as reduce the chitosan-DNA complexes aggregation, which may improve transfection efficiency. The hydrophilic chitosan modification includes quaternised chitosan (Thanou et al., 2002), PEGylated (covalent attachment of polyethylene glycol polymer chains to another molecule) chitosan (Jiang et al., 2006) and low Mw soluble chitosan (Ercelen et al., 2006). Interestingly, Brannon-Peppas & Blanchette (2004) found that particles with more hydrophobic surfaces were also preferentially taken up by the liver, followed by the spleen and the lungs.

Hydrophobic modifications of chitosan have been performed in many studies. The main objectives of these modifications were increasing transfection efficiency by modulating complex interactions with cells, especially in the complexes' adsorption on the cell surfaces and cell uptake (Kurisawa et al., 2000). Some hydrophobic units also help in the dissociation between the chitosan DNA complexes to release DNA to enter the nucleus after cellular uptake, as well as protecting it from enzymatic degradation and facilitating intra cellular pDNA (plasmid DNA) association, which can enhance transfection efficiency. These hydrophobic modifications included deoxycholic chitosan, N-alkylated chitosan, thiolated chitosan and hybrid chitosan (Mao et al., 2010). The combination of hydrophilic and hydrophobic modification of chitosan structure has been another interesting area that looks highly promising for the development of high transfection efficiency in a non-viral vector, using chitosan as a core structure.
