Development of an Enzymatic Process for the Synthesis of (S)-2-Chloro-1-(2,4-dichlorophenyl) Ethanol

Abstract

Chloro-1-(2,4-dichlorophenyl) ethanol ( 3) is a chiral intermediate in the synthesis of luliconazole ((R)-E-1). Here, we report a novel biopreparation of 3 by bioreduction of 2-chloro-1-(2,4-dichlorophenyl) ethanone (2) using recombinant Escherichia coli expressing LK08, a ketoreductase mutant from Lactobacillus kefiri, as a biocatalyst. The reaction conditions for the biotransformation including pH, temperature, and concentration of isopropanol and NADP + , as well as the amount of recombinant E. coli cells, were optimized to improve the process productivity. When the enzymatic process was carried out on a 300 g scale under the optimized conditions, the ketone 2 was fully converted to chiral alcohol 3 with a product ee value of >99%. Furthermore, 3 was isolated and used to chemically synthesize luliconazole with 38% yield and 99% ee. This study presents an efficient and cost-effective chemoenzymatic process for the production of (R)-E-1.

Development of a Novel Chemoenzymatic Process for (S)-1-(Pyridin-4-yl)-1,3-propanediol

“…However, the conversion was found to decrease sharply when the substrate concentration was increased to 150 g/L. In our previous research, we found that the pH and temperature are essential for improving the enzyme activity and the cosubstrate equivalent is a key factor for the biocatalytic process. , Thus, the process optimization for the reaction conditions, including the pH, temperature, cosubstrate loading, and catalyst loading, was subsequently performed (Figure ).…”

“…13,14 For example, Wei et al reported an enzymatic process to synthesize (S)-2-chloro-1-(2,4-dichlorophenyl) ethanol, the chiral intermediate of luliconazole, by a novel KRED mutant, LK08, from Lactobacillus kefiri. 15 In 2004, Berendes Frank reported the bioreduction of ketone 4 to 5 using Saccharomyces cerevisiae, with a substrate concentration of 20 mmol/L (3.9 g/L) and an of ee 81% (the methyl ester derivative had an ee of 96%). 16 Herein, we aimed to develop a novel chemoenzymatic process for (S)-1-(pyridin-4-yl)-1,3propanediol to overcome the shortcomings of existing routes.…”

“…The biotransformation of ketones to chiral alcohols is carried out in the presence of cofactor NAD­(P)­H . With the increasing use of KREDs, an increasing number of chiral intermediates are being synthesized by KREDs instead of chemical catalysts. , For example, Wei et al reported an enzymatic process to synthesize ( S )-2-chloro-1-(2,4-dichlorophenyl) ethanol, the chiral intermediate of luliconazole, by a novel KRED mutant, LK08, from Lactobacillus kefiri . In 2004, Berendes Frank reported the bioreduction of ketone 4 to 5 using Saccharomyces cerevisiae, with a substrate concentration of 20 mmol/L (3.9 g/L) and an of ee 81% (the methyl ester derivative had an ee of 96%) .…”

Application of Ketoreductase in Asymmetric Synthesis of Pharmaceuticals and Bioactive Molecules: An Update (2018–2020)

“…In the first report, Chen and co‐workers developed a chemo‐enzymatic synthesis route to luliconazole, with the aforementioned bioreduction being employed to set the required stereochemistry (Scheme 22). [36a] Specifically, by screening an in‐house preserved library of ketoreductases, the authors found a Lk CR variant with five mutations (E67D, A94T, F147L, L199H, A202L) displayed best enzyme activity and stereoselectivity. After optimizing reaction conditions such as pH, temperature, as well as the amounts of isopropanol, NADP + and biocatalyst, the biotransformation was upscaled to 300 g. In the presence of 300 g/L wet E. coli cells, 0.1 g/L NADP + , and 70 % (v/v) isopropanol, 300 g/L of 2‐chloro‐1‐(2,4‐dichlorophenyl) ethanone ( 44 ) was completely reduced within 30 h at pH 6.0 and 35 °C, providing enantiomerically pure ( S )‐2‐chloro‐1‐(2,4‐dichlorophenyl) ethanol (>99 % ee) in 94.3 % yield with a HPLC purity of 99 %.…”

Development of an Enzymatic Process for the Synthesis of the Key Intermediate of Telotristat Ethyl

“…Wei et al. reported an enzymatic process for the synthesis of the chiral intermediate of luliconazole by the Lk ADH‐catalyzed bioreduction [8] . Hu et al.…”