Reaction and diffusion of enzyme molecules in condensed polymer matrices

The objective of this work is to study the reaction and diffusion of enzyme molecules in condensed polymer matrices as well as the structure and interactions in polysaccharides aqueous solutions. Enzymatic degradation of guar galactomannan is investigated over a wide range of polymer concentrations using gel permeation chromatography (GPC) and steady shear viscometry. In dilute and intermediate polymer concentrations, the degradation kinetics is in accord with the Michaelis-Menton model. As the solution increases in concentration, enzyme diffusion through the concentrated polymer gel becomes a limiting factor for the reaction. A reaction-diffusion model is presented to express the competition between enzyme reaction and diffusion. The scaling theory and kinetic data are used to define the boundaries of the polymer concentration regimes between substrate (i.e. polymer strand) limited reactions, enzyme limited reactions, and hindered diffusion limited reactions. The influence of polymer derivatization on the degradation kinetics is also explored. The degradation rate is shown to be greatly affected by the type of substituent groups as well as the degree of substitution. Furthermore, a mechanism using oppositely charged polyelectrolyte as a reversible inhibitor for enzymatic reactions with polymeric substrates is found. This mechanism holds promise for controlled degradation of biopolymers in industry.; The diffusion of mesoscopic probe molecules in aqueous polymer solutions is measured using fluorescence recovery after photobleaching (FRAP). Two different FRAP techniques: fringe pattern bleaching and recovery (FPBR) and confocal scanning laser microscopy (CSLM) are used, and yield consistent diffusion results. The morphology of various probe particles are compared based on their fractal dimensions (d_f). Probe diffusion depends on the fractal dimension of the probe particle: the diffusion of rigid spheres is more hindered in semi-dilute and concentrated polymer solutions than random coil molecules with the same hydrodynamic size in free solution. The scaling equation D/D _o = exp [−β(R/ξ)δ] is used to fit experimental results. It is found that this equation is a good model for mesoscopic, rigid spherical probes. Furthermore, the effect of matrix polymer stiffness and polymer molecular weight is addressed. Polymers with higher chain rigidities tend to hinder more the probe diffusion than do flexible polymers.; In addition, the structure and molecular interactions between galactomannan polymer chains are studied by the osmotic stress method combined with X-ray scattering. As osmotic stress increases, there is a transition from liquid crystalline phase to crystalline phase for guar matrices, due to the propensity of guar to form intermolecular hydrogen bonds in solutions. Molecular interactions between polysaccharide chains can be controlled by altering the polymer chemistry. Adding hydroxypropyl (HP) substituent groups sterically decreases the hydrogen bonding attractions between guar chains. For moderately HP substituted guars, the phase transition occurs at much higher pressures compared to that of native guar, i.e. more work is needed to crystallize HPG than guar. For highly substituted guars, no phase transition has been found even under very high osmotic pressures, and the intermolecular interactions at close separations are dominated by the entropic fluctuations of polymer chains. The osmotic stress results are consistent with dilute solution viscometry measurements.