Alginate-based microcapsules for mammalian cell culture and other biotechnological applications

Since life began, nature has been using the envelopment of systems for their protection or for providing a particular reaction space, the enveloping wall displaying additional specific membrane functions. The abundance of examples is quite overwhelming and extends from arboreal fruits and plant seeds to plant seed spores, from the hen egg and its shell to the cell and its membrane/wall. The technique of microencapsulation aims at immobilizing gases, liquids or solids in an envelope. The result is a core contained in a capsule ranging from the nanometer to the millimeter scale. Nowadays, microencapsulation is being studied and developed in many different backgrounds throughout the chemical and life sciences, biotechnology, medicine and related industries. The aims of this thesis were to study the potentials and limitations of alginate-based microcapsules, for biotechnological applications, especially mammalian cell culture. It was shown that the limitations of alginate poly-L-lysine (PLL) microcapsules, commonly used for mammalian cell entrapment, arises from the poor mechanical stability of the polyelectrolytic complex constituting the membrane, and the intracapsular alginate concentration preventing optimal cell growth and core colonization. These limitations may be overcome by finding new techniques preventing the accumulation of alginate in the core, and by reinforcing the membrane through the formation of covalent bonds. To remove alginate from the core of the capsule, the use of an alginate degrading enzyme might be a possibility. However, alginate lyase degrades both gelled, dissolved and complexed forms of alginate. Therefore, this system should be more appropriated for cell release from microcapsules. Another alternative for the formation of alginate-free core microcapsules may be the hydrolysis of an organic-core capsule by a lipase. Aqueous-core capsules surrounded by an alginate matrix were successfully obtained, without accumulation of alginate in the core. However, this system is not suitable for mammalian cell encapsulation due to the toxicity of the process, especially the degradation product resulting from the enzymatic hydrolysis. The third solution to avoid the presence of alginate in the core was the use of the alginate/casein aqueous two-phase system. The combination of these compounds was suitable for the formation of aqueous-core microcapsules, using the jet break-up technique with a concentric nozzle. Casein was not toxic to cells, however precipitated in the presence of Ca2+ (dissolved in the buffer to induce alginate gelation), and hindered further cell growth. Membrane stability can be reinforced by the formation of covalent bonds. Acrylamide monomers polymerization and PLL / propylene-glycol-alginate / BSA transacylation reaction were the two systems studied. The formation of covalent bonds was successful for both systems, and produced reinforced capsules. However, acrylamide monomer, as well as the polymerization proce