Mechanical stability of immobilized biocatalysts (CLECs) in dilute agitated suspensions

Cross-linked enzyme crystals (CLECs) are a novel form of immobilized biocatalyst designed for application in industrial biotransformation processes. In this work we have investigated the mechanical stability of agitated CLEC suspensions in relation to the design and scale-up of bioconversions carried out in stirred-tank reactors. By careful control of the crystallization conditions yeast alcohol dehydrogenase I (YADHI) microcrystals of different size were first prepared having either an hexagonal (approximately 12 microm) or rod-shaped (approximately 4.6 microm) morphology. These were then cross-linked with glutaraldehyde to form CLECs. The rate of breakage of the CLEC suspensions was subsequently measured in a rotating disk shear device (total volume, 11 mL) by monitoring the change in crystal size distribution with time. This device is designed to mimic the shear and energy dissipation rates found in a range of process scale equipment and may be used to study the mechanical stability of any immobilized biocatalyst preparation. Experiments were performed as a function of the speed and duration of disk rotation, CLEC concentration (0.26-2.5 mg.mL(-1)) and energy dissipation rate (2.2 x 10(3) to 6.8 x 10(5) W.kg(-1)). No breakage of the rod-shaped CLECs was observed over the entire range of experimental conditions investigated. Breakage of the larger hexagonal-shaped CLECs did occur, however, at energy dissipation rates, epsilon(max), above 1.0 x 10(5) W.kg(-1), where the calculated length scale of turbulence was around 2.0 microm. Based on visual observation of the sheared CLEC suspensions and models of crystal breakage, it was concluded that breakage of the hexagonal-shaped CLECs occurred due to shear induced attrition. Measurement of the catalytic activity of both the hexagonal and rod-shaped CLECs showed no significant change in activity before and after shearing.