Diffusion and chemical reaction rates with nonuniform enzyme distribution: An experimental approach

Abstract

The study of the effects of nonuniform distributions of immobilized β‐galactosidase on the overall reaction rate of the hydrolysis of lactose are presented. Diffusion inside the particles has been characterized by measuring the diffusion rates of two β‐galactosidase substrates: lactose and ONPG in a commercial silica‐alumina support. Effective diffusivities have been determined by the chromatographic method under inert conditions. The results obtained for tortuosity can be explained assuming that the transport only takes place in the macropores. The distribution of the immobilized enzyme has been measured by means of confocal microscopy technique. The enzyme has been tagged with FITC and immobilized in particles of different diameters, the internal local concentrations of the enzyme have been determined with the aid of an image computer program. As expected, a more nonuniform internal profile of the enzyme was found when the particle diameter was bigger. Experiments under reaction conditions were carried out in batch reactors using lactose and ONPG as substrates and particles of the immobilized β‐galactosidase of different diameter (1 · 10−4 to 5 · 10−3 m) as catalyst, employing a temperature of 40°C for lactose and 25 and 40°C for ONPG, respectively. The mass balance inside the particle for the substrates has been solved for the internal profiles of the immobilized enzyme inside particles of different size and the enzymatic reactions considered. The calculated and the experimental effectiveness factor values were similar when particles under 2.75 · 10−3 m in diameter were employed. For the same Thiele modulus, a particle with nonuniform distribution of enzyme showed a higher effectiveness as a catalyst than particles with a more uniform distribution. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 72: 458–467, 2001.

Design of the Enzyme–Carrier Interface to Overcome the O2 and NADH Mass Transfer Limitations of an Immobilized Flavin Oxidase

“…In this scenario, internal diffusional restrictions may limit the apparent activity, especially that of the uniformly distributed enzymes, 32,39 as described for several hydrolases. 17,40 To assign the magnitude of the influence of NADH and O 2 limitations and design strategies for their further mitigation, we studied the kinetics of oxygen time consumption courses provided by different NOX immobilized preparations.…”

Hindered diffusion of proteins and polymethacrylates in controlled-pore glass: An experimental approach

“…The slope of the first plot is (1 + 0 ), this value can be used to calculated a K value (Ladero et al, 2001). The slope of the second plot, G, is used to obtain the effective diffusivity, knowing k f , with the simplified van Deemter equation, appropriate for diffusion in liquid phases (Boyer and Hsu, 1992).…”

Synthesis of Ibuprofen Monoglyceride in Solventless Medium with Novozym®435: Kinetic Analysis

“…Enzyme immobilization allows for its easy separation from the reaction medium [26] and facile reuse, or utilization in continuous-or flow-reactors [27,28]. In spite of this, the activity may be reduced by losing access to active centers and by the hindered diffusion of substrates and products [15,28,29]. In particular, a major limitation is the slow mass transport within the porous structure (internal mass transfer) or in the outer liquid layer (external mass transfer), in comparison to the reaction rate [26,30].…”