Structural Hierarchy in Linear Polysaccharides - from the Nano- to Macroscale

The fundamental study of self-assembly pathway of biomacromolecules are of great importance to understand, design and fabricate new smart materials in many fields ranging from nanoelectronics and food science to biomedical engineering. Polysaccharides are abundant, affordable, biocompatible and self-assembling biopolymers making them ideal candidates for this purpose. In particular, charged polysaccharides offer a multitude of applications due their gelling ability. However, the formation and structure of the network building blocks on the nanoscale remain elusive in the semi-dilute regime. This thesis aims to shed light on the ion induced gelation pathway of linear charged polysaccharides on a single molecule level and its impact on the macroscopic properties. The self-assembly of κ- and ι-carrageenan, with one and two sulphate groups per ideal subunit, was investigated in varying environments of monovalent chloride salts. The ion-specificity of the disorder-order conformation change, the first step of gelation, was quantified and confirmed by analysing high-resolution atomic force microscopy (AFM) height images of single polymer chains. The formation of higher order structures, identified as secondary, tertiary and quaternary, was independent to the polysaccharide concentration, since the ion-induced higher order structures remained the same. In more detail, a two-step mechanism was observed with first a random coil to single helix conformation change, identified as primary structure and secondary structure, respectively. The conformation change was accompanied with a chain stiffening, subsequently followed by a coiled coiling in flexible chains or lateral aggregation in more rigid cases forming so-called tertiary and quaternary structures ultimately inducing the gelation of the system. Ordered domains formed by rigid quaternary structures induced heterogeneous macroscopic birefringence. The gel properties such as strength and deformability were directly linked to the flexibility of the first fundamental ordered state, the single helix. Based on the observed hierarchical nanostructures, a general nomenclature, inspired by the well-known one in protein science, was suggested and further supported by similar findings in Gellan gum, another linear charged polysaccharide. Motivated by the findings on the monovalent ion induced conformational change in Gellan gum, the influence of divalent ions was analysed in more detail. Again, a two-step mechanism was observed, in which first single helices as secondary structures were formed, which aggregate laterally into rigid fibrillar supramolecular bundles as quaternary structures. The molecular interaction of the two charged species was followed by Fourier-transformed infrared spectroscopy, revealing the coordination of the inorganic cation with one of the oxygen atoms in the carboxyl group on the polysaccharide backbone, inducing the stiffening of the polymer. Furthermore, the flexibility of the quaternary structures was extracted, reaching values of several micrometres for the persistence length, explaining the observed macroscopic properties such as high gel strength, low deformability and intense birefringence. Last, the impact of an organic charged species on linear polysaccharides was investigated by replacing the inorganic chloride salts with the anti-malarial drug chloroquine (CQ). The interaction between λ-carrageenan, the only carrageenan not known to undergo disorder-order transition, and divalent CQ resulted in complexes as revealed by distinct isosbestic points in UV- Vis spectra. The complexes were investigated by AFM imaging, revealing the formation of secondary structure followed by tertiary and quaternary structure formation, not yet reported in λ-carrageenan. The higher order structures disappeared by introducing physiologically relevant inorganic cations replacing the condensed CQ and polysaccharide chain in primary structure was recovered, suggesting ion-mediated drug release applications