Aqueous systems, bioreduction

Here the biotransformation (Fig. 19-6) is preferred over the chemical reduction with commercially available asymmetric catalysts (BH3- or noble-metal-based), since with the chemocatalysts the desired high enantiomeric excess (ee > 98%, 99.8% after purification) is not achievable. Since the ketone has only a very low solubility in the aqueous phase, 1 kg ketone is added as solution in 4 L 0.9 M H2SO4 to the bioreactor. The bioreduction is essentially carried out in a two-phase system, consisting of the aqueous phase and small droplets made up of substrate and product. The downstream processing consists of multiple extraction steps with methyl ethyl ketone and precipitation induced by pH titration of the pyridine functional group (pfCa = 4.66) with NaOH. The (R)-amino alcohol is an important intermediate for the synthesis of (1-3-agonists that can be used for obesity therapy and to decrease the level of associated type II diabetes, coronary artery disease and hypertension. 

Figure 6.1 Schematic illustration of asymmetric hydrogen-transfer bioreduction with E. coli biocatalysts in IPA-aqueous solution system.

In this chapter, latest advancements in solvent engineering in bioreductions and greener needs for bioreaction media have been discussed in depth with recent examples. Solvents for bioreductions may be categorized as (i) aqueous (ii) water/water-miscible (monophasic aqueous-organic system) (iii) water/ water-immiscible (biphasic aqueous-organic system) (iv) nonaqueous (mono-phasic organic system, including solvent-free system) and (v) nonconventional media (e.g., ionic liquids, supercritical fluids, gas-phase media, and reverse micelles). 

In many cases, the substrates for reaction are poorly water-soluble, meaning it is necessary to consider organic solvent or two-liquid-phase systems to ensure the concentration of substrate (and hence product) in the reactor is sufficient. Two-liquid-phase systems may also be used for product removal as discussed later. There are relatively few reports of solvent effects on the various bioreduction enzymes, but a recent publication found ene-reductases to be robust in several aqueous-organic two-phase systems [9]. 

Using traditional organic cosolvents, the bioreduction of para-bromo-2,2,2-trifluoroacetophenone 83 (Scheme 6.34) was limited to a maximum product concentration of 10 g/L due to substrate-induced deactivation of the R. eryth-ropolis (ADH RE) biocatalyst. However, by employing 10% (v/v) [BMP][NTf2], a water immiscible ionic liquid, more than 5 times the ketone was converted to the chiral alcohol in less than 24 h. The ionic liquid improved the initial reaction rate by more than four times in the presence of ionic liquid compared to an aqueous-only reaction. Moreover, the ionic liquid improved the stability of both the ADH RE and the GDH coenzyme compared to reactions with either organic cosolvents or aqueous buffer systems [59]. 

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