The Bioproduction of L-phenylacetylcarbinol in solid-liquid two phase partitioning bioreactors
Khan, Tanya Razia
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Biphasic systems such as two-phase partitioning bioreactors (TPPBs) have been used to alleviate biological inhibition by sequestering inhibitory compounds within an immiscible phase. The use of solid polymer beads as this auxiliary phase provides a fully biocompatible alternative to commonly used yet potentially toxic organic solvents. This work focused on the application of solid-liquid TPPBs to the bioproduction of the pharmaceutical precursor L-phenylacetylcarbinol (PAC), a biotransformation which suffers from substrate (benzaldehyde), product (PAC), and by-product (benzyl alcohol) inhibition, and simple strategies to improve TPPB performance in general. A wide range of commercially available, biocompatible, and non-bioavailable polymers were screened for their affinity for benzaldehyde, PAC, and benzyl alcohol. Hytrel G3548L demonstrated the highest affinity for all three target compounds and was subsequently used in solid-liquid TPPBs for PAC production. Using 15% v/v polymer beads, PAC concentration was increased by 104% and benzyl alcohol concentration decreased by 38% over the single phase control. The delivery of benzaldehyde from polymer beads demonstrated only a 6-8% reduction in mass productivity with improved operational simplicity and reduced operator intervention. The final objective of this work was to independently investigate various aspects of the aqueous phase composition and determine how each factor affects the partition coefficient of benzaldehyde in Hytrel G3548L. Temperature and pH were observed to have no significant effect on partitioning. Salt and glucose additions increased the partition coefficient by 173% and 30% respectively compared to RO water, while ethanol was found to decrease the partition coefficient from 44 (±1.6) to 1 (±0.3). These findings may be applied to solid-liquid TPPBs to increase or decrease partitioning as required, leading to improved bioreactor performance. This work has successfully shown that with careful polymer selection, solid-liquid TPPBs can be used to increase the productivity of a biotransformation without the associated biocompatibility problems that have sometimes been observed with organic solvents. The delivery of inhibitory substrate from the polymer phase was successfully accomplished, which is a novel demonstration in the field of solid-liquid TPPBs for biocatalysis. Finally this work contributes a range of simple strategies to improve the partitioning behavior of solid-liquid TPPBs using the aqueous phase composition.