Probing the biophysical interactions of chitosan derivatives with cell membrane models using Sum-Frequency Generation Spectroscopy

Chitosan, a biopolymer, has been used in many biological applications such as drug delivery, gene therapy, tissue engineering and antimicrobial systems. Although these applications have grown greatly in recent years, little is known about the modes of interaction at the molecular level of chitosan with biological membranes. Therefore, in this study we investigated the interactions of chitosan with lipids in Langmuir monolayers as model membranes using two chitosan samples: unmodified chitosan (CH) and positively charged chitosan functionalized with quaternary amino groups (CH-P40). The anionic dipalmitoyl phosphatidylglycerol (DPPG) and zwitterionic dipalmitoyl phosphatidylcholine (DPPC) lipids were used as membrane models. The interactions at the air-water interface were assessed by surface pressure-area isotherms, Brewster angle microscopy and sum-frequency generation (SFG) spectroscopy. It was found that both samples had an insignificant effect on DPPC monolayers formed on PBS buffer containing the chitosans, but it noticeably affected DPPG monolayers. The SFG spectra showed a well-packed DPPC monolayer, with few gauche defects in the acyl chains probably due to the insertion of the chitosans, pushing laterally the lipid molecules. Oppositely, an increase in gauche defects was observed for DPPG. This can be explained by the uniform expansion of the film molecules due to the interaction between chitosan and DPPG anionic groups occurring underneath monolayer. Morphological changes from the BAM imaging confirmed the association of both samples into the film. The results obtained may be helpful in identifying the interaction mode of chitosan with phospholipids.