Ionic Liquid/Water Continuous-Flow System with Compartmentalized Spaces for Automatic Product Purification of Biotransformation with Enzyme Recycling

Abstract

In this work, we developed a biphasic designer solvent system for enzymatic biotransformation, to demonstrate automatic purification and enzyme reuse, in the frame of a new process concept reported recently (solvent-enabled factory, One-Flow project). "Automatic" refers to instantaneous operation by preferential solubility between two immiscible phases, which does not require (sophisticated) process control ("automated"). The reaction studied is lipase-catalyzed hydrolysis of 4-nitrophenyl acetate. As a designer solvent, the class of ionic liquids (ILs) has been chosen, and their usage is largely documented in the biocatalysis literature. Three ILs are chosen; all with the 1-butyl-3-methyl-imidazolium cation and differing in their anions, being tetrafluoroborate, bis(trifluoromethylsulfonyl)-imide, and hexafluoro-phosphate. The first IL forms a monophasic system and thus was left out of consideration. The other two form the desired biphasic system, when being contacted with water, respectively. The operation of that system is hampered by foaming, that is, the formation of an interfacial emulsion layer as the third phase, which makes phase separation difficult. Therefore, we investigated in detail the phase behavior of the batch and flow-processed fluid systems under various process conditions. Batch processing, which causes tremendous foaming, needs intense stirring because of the high IL viscosity. Continuous-flow reactors provide an alternative because they stir more softly by their shear-enforced convection in their liquid slugs. As a result, they do not show foaming, and therefore, separation in phases is facile. With that issue solved, we report here for the continuous-flow biocatalytic reaction that we achieved high-level product purification and (three times) recycling of the enzyme.

Applications of Ionic Liquids in Whole-Cell and Isolated Enzyme Biocatalysis

“…Biphasic- aqueous buffer and hydrophobic ionic liquid ([BMIM][NTf 2 ] and [BMIM][PF 6 ]) mixtures were developed for the continuous flow biocatalysis of Amano lipase from Pseudomonas fluorescens . In the hydrolysis of para-Nitrophenyl acetate the continuous system in buffer/[BMIM][NTf 2 ] achieved 1.3 times product formation in 0.5 h compared to the reaction in a batch reactor [ 121 ].…”

Life Cycle Assessment of an Enzymatic Ibuprofen Production Process with Automatic Recycling and Purification

“…Recently, an entirely new reactor and processing concept were proposed in the One-Flow EU project, to develop and explore in a bionic manner soft-matter compartmentalized processing systems . One of those concepts is the “Functional Solvent-enabled Factory,” representing a multiphase system which (by virtue of solvent modeling) can be so fine-tuned that ideally pure product result (automatically) and the reactant and catalyst are recycled in their own phases ().…”

“…We have recently reported the proof of concept through a de-esterification reaction using lipase as an enzyme and ionic liquids to uptake the low-polar reactant (ester) as a highly pure product and facilitating downstream processing while water taking up the polar products (i.e., acid, alcohol, etc.) for recycling . In light of the green chemistry approach, we are developing a new synthetic route for ibuprofen in which the toxic solvent, methanol, was replaced by a greener reagent which is water.…”

“…Recently, an entirely new reactor and processing concept were proposed in the One-Flow EU project, to develop and explore in a bionic manner soft-matter compartmentalized processing systems. 7 One of those concepts is the "Functional Solventenabled Factory," representing a multiphase system which (by virtue of solvent modeling) can be so fine-tuned that ideally pure product result (automatically) and the reactant and catalyst are recycled in their own phases (www.one-flow.org). We have recently reported the proof of concept through a deesterification reaction using lipase as an enzyme and ionic liquids to uptake the low-polar reactant (ester) as a highly pure product and facilitating downstream processing while water taking up the polar products (i.e., acid, alcohol, etc.)…”

“…for recycling. 7 In light of the green chemistry approach, we are developing a new synthetic route for ibuprofen in which the toxic solvent, methanol, was replaced by a greener reagent which is water. To this end, we propose to modify the third step of the synthetic route proposed by Bogdan et al, the saponification reaction, using an enzyme catalyst 8 in the presence of buffer and ionic liquid to enable the de-esterification reaction.…”