Please use this identifier to cite or link to this item: http://hdl.handle.net/1946/25466
Permeation enhancers have been studied to increase the bioavailability of peptide and protein based drugs through mucosal membranes. N-quaternary chitosan derivatives are water soluble, independent of the pH and have been investigated as such permeation enhancers. The precise mechanism explaining this effect is unknown but an interaction with tight junction (TJ) proteins has been suggested. Before initiating the PhD project, it was hypothesized that by increasing the N-alkyl length of N-quaternary chitosan derivatives, more degree of amphiphilicity could be achieved, augmenting further the permeation enhancement effect. This approach would require specific synthetic strategies for the production of highly N-quaternized derivatives, enabling detailed structure determination. In order to test the hypothesis, it is also important to use a differentiated epithelium with established TJ structure and function for the investigation.
Therefore, the aim of this PhD project was to develop strategies for the synthesis of highly N-quaternized chitosan derivatives with well defined structures and determine the relationship between their N-alkyl chain length and their permeation enhancement in a bronchial epithelial cell model.
The synthesis was done with the aid of di-tert-butyldimethylsilyl hydroxyl protected chitosan, which resulted in highly quaternized N-methyl-, N-propyl-, N-butyl- and N-hexyl-N,N-dimethyl chitosan derivatives (TMC, QuatPropyl, QuatButyl and QuatHexyl respectively), without O-methylation. A fluorophore was regioselectively conjugated to the reducing end of TMC by oxime formation and visualized in bronchial epithelial cells. This provided an important proof of concept for further reducing-end oxime modifications of chitosan derivatives with other functional groups. It was shown that the VA10 bronchial epithelial cell line was able to model the native bronchial epithelial integrity, permeability and morphology and can, therefore, be considered suitable to elucidate the relationship between the structure of N-quaternary chitosan derivatives and their activity as permeation enhancers. Those results demonstrated that increased amphiphilicity of the N-quaternary chitosan derivatives caused increased permeation, TJ disassembly and decreased viability in the order of QuatHexyl ≈ QuatButyl > QuatPropyl > TMC at pH 7.4, showing that the permeation is not only affected by the degree of quaternization but also by the degree of N-alkylation. The TMC homopolymer possesses promising permeation enhancer properties, being effective in low concentrations without causing prolonged adverse effects and should therefore be considered as a promising candidate for the development of future respiratory dosage forms.
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