Hydroxynitrile lyase catalyzed cyanohydrin synthesis at high pH-values

Abstract

The application of unusual high pH-values within enzymatic cyanohydrin synthesis has been investigated. Usually enzymatic cyanohydrin synthesis in two-phase systems requires low pH-values within the aqueous phase to suppress the non-enzymatic side reaction. In contrast, we investigated the usage of pH-values above pH 6 by using the highly enantioselective (S)-selective hydroxynitrile lyase from Manihot esculenta. With these unusual reaction conditions also the unfavorable substrate 3-phenoxy-benzaldehyde can be converted by the wild type enzyme with excellent conversion and enantiomeric excess yielding pure (S)-3-phenoxy-benzaldehyde cyanohydrin with an enantiomeric excess of 97%. Although the variant MeHNL-W128A shows a higher activity with respect to this reaction, the enantioselectivity was reduced (85% e.e.(S)). Additionally, a new continuous spectroscopic cyanohydrin assay monitoring the formation of 3-phenoxy-benzaldehyde cyanohydrin was developed.

Uneven twins: Comparison of two enantiocomplementary hydroxynitrile lyases with α/β-hydrolase fold

“…In the case of m-phenoxy cyanohydrin synthesis the chemical side reaction is very slow and the product is stable also in the neutral pH range. This allows the enzymatic reaction to be carried out at neutral pH at the pH-and stability optimum of the enzyme which increases the half-life time of HNLs significantly (Von Langermann et al, 2008).…”

Hydroxynitrile Lyase‐Catalyzed Synthesis of Enantiopure Cyanohydrins in Biocatalytic Active Static Emulsions (BASE) with Suppression of the Non‐Enzymatic Side Reaction

“…Diisopropyl ether was used throughout this study as solvent since it exhibits only low water solubility and has proven its compatibility with the investigated hydroxynitrile lyase from Manihot esculenta. [27,36,37] Under the chosen conditions within 80 min ca. 5400 ppm water were observed in diisopropyl ether.…”

“…[26] Unfortunately these unphysiological low-pH conditions usually cause a significant loss of enzyme activity and (most importantly) enzyme stability. [24,27] A representative example is the synthetically interesting HNL from Arabidopsis thaliana (AtHNL), which is currently under investigation to increase low-pH stability. [28,29] Second, since chemical reactions are temperature-dependent, a reduction of reaction temperature also reduces to some extent the non-enzymatic reaction.…”

Biochemical engineering science

“…As a consequence, the first three contributions of this special issue address selected reaction engineering aspects of biocatalysis, followed by a contribution combining chemistry and biocatalysis. The paper of the Udo Kragl group (Institute of Chemistry at the University of Rostock, Germany) who was the first to head the enzyme group at the Institute of Christian Wandrey from 1989 to 1998, investigates the application of unusual high pH within enzymatic cyanohydrin synthesis [8]. Udo Kragl's successor in Jülich, Andreas Liese (Institute of Technical Biocatalysis, Technical University of Hamburg-Harburg, Germany) and co-authors report on the practical application of different enzymes immobilized in a highly porous methacrylic polymer matrix [4].…”