Interactions of Phospholipid Bilayer with Chitosan: Effect of Molecular Weight and pH

Chitosan has demonstrated its potentials as a gene carrier and a membrane perturbant for subsequent drug
delivery to cells. However, there is currently a lack of experimental correlation between the physiochemical
properties of chitosan and the resulting degree of lipid bilayer destabilization. In this study, the effect of pH
and chitosan molecular weight on the interaction between chitosan and dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) bilayer was examined with cross-polarization microscopy, differential scanning
calorimetry (DSC), and Fourier transform- (FT-) Raman spectroscopy. Cross-polarized images showed that
the direct hydration of the DPPC/chitosan mixture led to the formation of larger DPPC multilamellar vesicles
(MLV), and pure chitosan also induced fusions of individual MLV. Under the influence of chitosan, the
calorimetric enthalpy of DPPC was reduced in a concentration-dependent manner, and a new phase appeared
at 28 °C during sample cooling. Even the lowest chitosan mole fraction of 0.04% reduced the cooperative
unit of the DPPC bilayer by more than 70%. In addition, the electrostatic effect between chitosan and
DPPC tuned the degree of membrane bilayer perturbation. Reduction of pH increased the number of protonated
amines on the chitosan backbone and caused further disruption on the membrane organization. Mixing
DPPC with chitosan in an organic medium before hydration enhanced the hydrophobic interactions between
the two molecules and greatly reduced the cooperative unit among individual lipids during the main phase
transition. The increase of chitosan molecular weight also affected the cooperativity in the thermotropic
transition of DPPC bilayer. FT-Raman spectroscopy provided additional evidence that chitosan directly
perturbed the organizations of the hydrophobic inner core of the DPPC bilayer.