Efficient phase separation and product recovery in organic‐aqueous bioprocessing using supercritical carbon dioxide

Abstract

Biphasic hydrocarbon functionalizations catalyzed by recombinant microorganisms have been shown to be one of the most promising approaches for replacing common chemical synthesis routes on an industrial scale. However, the formation of stable emulsions complicates downstream processing, especially phase separation. This fact has turned out to be a major hurdle for industrial implementation. To overcome this limitation, we used supercritical carbon dioxide (scCO(2)) for both phase separation and product purification. The stable emulsion, originating from a stereospecific epoxidation of styrene to (S)-styrene oxide, a reaction catalyzed by recombinant Escherichia coli, could be destabilized efficiently and irreversibly, enabling complete phase separation within minutes. By further use of scCO(2) as extraction agent, the product (S)-styrene oxide could be obtained with a purity of 81% (w/w) in one single extraction step. By combining phase separation and product purification using scCO(2), the number of necessary workup steps can be reduced to one. This efficient and easy to use technique is generally applicable for the workup of biphasic biocatalytic hydrocarbon functionalizations and enables a cost effective downstream processing even on a large scale.

The dynamic influence of cells on the formation of stable emulsions in organic–aqueous biotransformations

“…Emulsion formation during biotransformation is often advantageous, particularly when dealing with mass transfer limited systems, but stable emulsions are notoriously difficult to handle downstream [40,47]. Recently, we have reported methods for phase separation and product extraction based on supercritical carbon dioxide, which would be amenable to implementation on industrial scale [6,7].…”

Integrated organic–aqueous biocatalysis and product recovery for quinaldine hydroxylation catalyzed by living recombinant Pseudomonas putida

“…52 mL in each run) with supercritical carbon dioxide (scCO 2 ) in a high-pressure vessel (Büchi, Uster, Switzerland). The conditions applied during the scCO 2 treatment were as follows: pressure (p) = 250 bar, temperature (T) = 45°C, CO 2 mass fraction (w CO 2 ) = 0.75, and time (t) = 60 min [4]. After scCO 2 treatment, the organic phase (15 mL) was separated by centrifugation at 4,500 g for 40 min and placed into a pre-warmed separatory funnel.…”

“…The emulsion formed during biotransformation was very stable, and the separation of organic and aqueous phases was not possible by centrifugation, even at high centrifugal force (13,000 g) and after prolonged centrifugation time (1 h). As an alternative, we tested treatment with scCO 2 for phase separation [4] followed by product isolation from the organic phase by liquid-liquid extraction. The separation of the organic phase by the scCO 2 treatment allowed the recovery of 15 ml of the organic phase (out of an initial volume of approx.…”

“…scCO 2 has recently been shown to destabilize such emulsions efficiently and irreversibly [4]. This technique was successfully used for the separation of the stable 1-dodecanol-aqueous emulsion, which was presumably formed due to the emulsifying effect of microbial cells, as shown for E. coli in a 2LP system with (bis)-ethylhexyl phthalate as organic phase.…”

“…In 2LP processes, this includes phase separation and product isolation [34,43]. Phase separation in particular can be critical when stable emulsions are formed [4].…”