**4.2.2 pH and degree of deacetylation (DDA)**

The study of Lavertu et al. (2006) also found that the low Mw chitosan, which had a numeric average Mw (Mn) of about 10 and 80% DDA at N/P ratio 10:1, gave higher transfection efficiency at the same level as lipofectamineTM at pH 6.5. However, the very low Mw (1.9-7.7 kDa) chitosans with high DDA were found to form aggregates easily, even at very low charge ratios (Morris et al., 2008), and this might lower the transfection. However, the depolymerised LW chitosan in this study had only 54% DDA, which may reduce the problem of particles aggregation and, after cells uptake the chitosan-DNA nanoparticles, the DNA may be released from the nanoparticles more easily, as DNA binding efficacy was reduced as DDA was decreased (Kiang et al., 2004). Hence, many factors may have to be considered for improving transfection efficiency of chitosan, not only the ligand binding, but also the method of binding or conjugation, the size and morphology of the particles, the aggregation of the complexes, and especially the chitosan itself, as Mw, DDA and charge ratio, which may have to be adjusted.
