Applied catastrophic phase inversion: a continuous non-centrifugal phase separation step in biphasic whole-cell biocatalysis

Biphasic whole-cell biotransformations are known to be efficient alternatives to common chemical synthesis routes, especially for the production of, e.g. apolar enantiopure organic compounds. They provide high stereoselectivity combined with high product concentrations owing to the presence of an organic phase serving as substrate reservoir and product sink. Industrial implementation suffers from the formation of stable Pickering emulsions caused by the presence of cells. State-of-the-art downstream processing includes inefficient strategies such as excessive centrifugation, use of de-emulsifiers or thermal stress. In contrast, using the catastrophic phase inversion (CPI) phenomenon (sudden switch of emulsion type caused by addition of dispersed phase), Pickering-type emulsions can be destabilized efficiently. Within this work a model system using bis(2-ethylhexyl) phthalate (BEHP) as organic phase in combination with E. coli, JM101 was successfully separated using a continuous mixer settler setup. Compared to the state-of-the-art centrifugal separations, this process allows complete phase separation with no detectable water content or cells in the organic phase with no utilities/additives required. Furthermore, the concentration of the product is not affected by the separation. It is therefore a simple applicable method that can be used for separation of stable Pickering-type emulsions based on the knowledge of the point of inversion.