MASS-TRANSFER STUDIES ON IMMOBILIZED ALPHA-CHYMOTRYPSIN BIOCATALYSTS PREPARED BY DEPOSITION FOR USE IN ORGANIC MEDIUM

Mass transfer limitations were studied in enzyme preparations of alpha -chymotrypsin made by deposition on different porous support materials such as controlled pore glasses, Celite, and polyamides of different particle sizes, it is the onset of mass transfer limitations that dete rmines the position of the activity optimum with respect to enzyme loa ding on each support. The evidence of various experiments indicates th at internal diffusional limitations are the important mechanism for th e observed mass transfer limitations. External diffusion was not found to play an important role under the conditions used, and it was also found that when immobilizing multilayers of enzyme the buried enzyme m olecules are active to a large extent. An extreme situation is observe d on Celite at very high loadings. Under these conditions, this suppor t is expected to have its pores completely filled with packed enzyme m olecules, and then it is the diffusion within the enzyme layer that de termines the observed rate. As the enzyme loading increases, the area of contact between the deposited enzyme layers and the liquid solution inside the pores diminishes, causing a decrease on the observed rate of an intrinsically fast reaction which apparently is incongruous with the presence of more enzyme in the system. This work shows that mass transfer limitations can be an important factor when working with immo bilized enzymes in organic media, and its study should be carried out in order to avoid undesired reduced enzyme activities and specificitie s. (C) 1998 John Wiley & Sons, Inc.