Results and Discussion
Optimization of the Enzymatic Imine Reductase
Reaction
Initial experiments targeted an increase of the substrate concentration
to facilitate higher productivities within the biocatalytic process
using the highly enantioselective imine reductase fromPaenibacillus elgii B69. Considering the chosen conditions, the
starting substrate concentration leads to a full conversion in the range
of 25 mmol·L‑1 to 150 mmol·L-1, but
a deactivation of the catalyst was found at even higher substrate
concentrations (see Figure 2A). This is solely caused by a substrate
inhibition and not a pH-shift due to the cofactor regeneration-based
formation of gluconic acid by glucose dehydrogenase. Noteworthy,
attempts using an excessive pH control with a
1 mol·L-1 phosphate buffer pH 7.5 were also not
successful due to a complete deactivation of the biocatalyst. A full
conversion was reached within 24 h under the chosen reaction conditions
(Figure 2B). The final optimized reaction conditions facilitated full
conversions at 30 °C, a substrate concentration of
150 mmol·L-1 and 500 mmol·L-1 of
d‑glucose. These experiments clearly indicate that further
improvements of the catalytic robustness towards higher substrate
loadings and higher specific activities are required to improve the
overall process productivity, which was not the main aim of this study.
Other noticeable examples of imine reductase-catalyzed reactions at
larger scale were recently reported for the biocatalytic reductive
amination reaction between cyclohexanone and cyclopropylamine with an
excellent volumetric productivity of
12.9 g·L-1·h-1 and a TON above 48000