Parameters for the Evaluation of Immobilized Enzymes Under Process Conditions.

The characterization of immobilized enzymes allows the evaluation of the immobilization process itself and also the projection of the immobilized enzyme performance under process operation conditions. Based on such characterization, strategies for support functionalization and enzyme immobilization into the activated support can be selected, determining the best conditions for conducting such steps in view of the intended use of the biocatalyst, establishing a linkage between biocatalyst production and biocatalyst use. The determination of the catalytic potential of the immobilized enzyme under operational conditions is a priceless parameter that takes into account both activity and stability, including the effect of both mass transfer limitations (diffusional restrictions) and intrinsic enzyme inactivation upon the immobilization process.

Diffusional restrictions in glyoxyl-agarose immobilized penicillin G acylase of different particle size and protein loading / Restricciones para la difusión de penicilina G acilasa inmovilizada en glicosil agarosa

Particle size and enzyme protein loading are design parameters of enzyme immobilization affecting biocatalyst performance that can be varied within broad margins. Their effect on mass transfer limitations at different bulk penicillin G concentrations has been studied with glyoxyl agarose immobilized penicillin G acylase biocatalysts of average particle size of 5·10-5m and 10·10-4m at protein loadings from 15 to 130 mg/g gel. Internal diffusional restrictions were evaluated for such biocatalysts: Thiele modulus varied from 1.17 for the small particles at the lower protein load to 5.84 for the large particles at the higher protein load. Effectiveness factors at different bulk substrate concentrations were determined for all biocatalysts, values ranging from 0.78 for small particle size at 25 mM penicillin G to 0.15 for large particle size at 2 mM penicillin G. Enzyme protein loading had a strong impact on the effectiveness factors of immobilized penicillin G acylase, being it more pronounced in the case of large particle size biocatalysts. At conditions in which 6-aminopenicillanic acid is industrially produced, all biocatalysts tested were mass-transfer limited, being this information valuable for reactor design and performance evaluation.