Biomicrogels for biological and medicinal applications

This work describes the synthesis and self-assembly of biopolymer microgels for biological and medicinal applications that include drug delivery and cell encapsulation. We focused on the preparation of chitosan nanoparticles of a controlled size (below 100 nm) and narrow size distribution through the process of ionic gelation between chitosan and sodium tripolyphosphate (TPP). By combining post-deacetylation and fractionation of commercial chitosan, we achieved control over particle size and size distribution. Properties of chitosan nanoparticles were tested at various temperatures, pH, and ratios of chitosan to TPP. A model protein, bovine serum albumin, was encapsulated into the chitosan nanoparticles, and its in vitro release profiles were examined in physiologically relevant media at 37°C. Next, we rationally designed chitosan-based microgels for targeted (site-specific) drug delivery. A water-soluble chitosan derivative N-[(a-hydroxy-3-trimethylammonium)-propyl] chitosan chloride (HTCC) was synthesized by grafting an epoxy compound, glycidyltrimethylammonium chloride (GTMAC), to the polymer chain. The degree of quaternization was controlled by changing the weight ratio of chitosan to GTMAC. HTCC microgels were ionically cross-linked by TPP to form 180 nm sized microgels that appeared to be pH responsive. These microgels were incorporated with methotrexate disodium (MTX), a cytotoxic drug for cancer treatment, and conjugated to the targeting biomolecule apotransferrin. Uptake of the bioconjugated microgels occurred via receptor-mediated endocytosis. In the cell compartments, exposure of the microgels to low pH environment caused the chitosan nanoparticles to swell and release MTX, thus killing the cells. This rational drug delivery design may be useful in enhancing cancer therapy and reducing side effects. In the second part of this work, we developed a novel approach to generating capsules of biopolymer microgels in microfluidic devices. Biopolymer droplets of an aqueous solution were emulsified in an organic phase comprising a cross-linking agent. Polymer gelation was achieved in situ (on a microfluidic chip) by diffusion-controlled ionic cross-linking of the biopolymer, following the transfer of the cross-linking agent from the continuous phase to the droplets. Gelation was quenched by collecting the particles in a large pool of cross-linking agent-free liquid. The structure of microgels from capsules to gradient microgels and then to particles with a uniform structure was controlled by varying either the concentration of the cross-linking agent in the continuous phase, or the time of residence of droplets on the microfluidic chip. We also demonstrated the encapsulation of a controlled number of polystyrene beads in the microgel capsules. The described approach was applied to the preparation of capsules of several polysaccharides such as alginate, kappa-carrageenan, and carboxymethylcellulose. Finally, we investigated the microfluidic encapsulation of mouse stem cells in alginate and agarose microgels by using the external gelation or thermally induced gelation, respectively. Preliminary results showed that thermal gelation of agarose provided a promising approach to the encapsulation of stem cells.