Industrial Biotransformations

Dehydrogenase
Candida sorbophila
N
1) Reaction conditions
[1]: 0.2 M, 60 g · L –1 [299.28 g · mol –1 ]
pH: 5.6
T: 34 °C
medium: two-phase: aqueous, solid
reaction type: reduction of keto group
catalyst: suspended whole cells
enzyme: dehydrogenase
strain: Candida sorbophila
2) Remarks
O
H
N
1
O
NO2
● Here the biotransformation is preferred over the chemical reduction with commercially available
asymmetric catalysts (boron-based or noble-metal-based), since with the latter ones the
desired elevated enantiomeric excess is not achievable.
● To prevent foaming during the biotransformation 2 mL · L –1 defoamer is added.
E
EC 1.1.X.X
● Since the ketone has only a very low solubility in the aqueous phase, it is added as an ethanol
slurry to the bioreactor. By this method no formal sterilization is possible. Heat sterilization is
also not applicable because of the ketone’s instability at elevated temperature. In the large scale
production, 1 kg ketone is added as solution in 4 L 0.9 M H 2SO 4 to the bioreactor. This solution
is sterilized prior to addition by pumping through a 0.22 µm hydrophobic Durapore microfiltration
membrane (Millipore Opticap cartridge, Bedford, USA).
● After intensive investigation and optimization a glucose feed of 1.5 g · L –1 was found to support
the highest initial bioreduction rate.
● The bioreduction is essentially carried out in a two-phase system, consisting of the aqueous
phase and small beads 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 (pK a = 4.66) with NaOH.
N
OH
H
N
(R)-2
1 = 2-(4-nitro-phenyl)-N-(2-oxo-2-pyridin-3-ethyl)-acetamide
2 = (R)-N-(2-hydroxy-2-pyridin-3-yl-ethyl)-2-(4-nitro-phenyl)-acetamide Merck & Co, Inc.
Fig. 1.1.X.X – 1
O
NO 2
191