Abstract
BackgroundWhole cells are usually employed for biocatalytic reduction reactions to ensure efficient coenzyme regeneration and to avoid problems with enzyme purification and stability. The efficiency of whole cell-catalyzed bioreduction is frequently restricted by pronounced toxicity of substrate and/or product to the microbial cells and in many instances the use of two-phase reaction systems can solve such problems. Therefore, we developed new, biphasic reaction systems with biocompatible water-immiscible ionic liquids (ILs) as alternatives to conventional organic solvents, in order to improve the asymmetric reduction of 4-(trimethylsilyl)-3-butyn-2-one (TMSB) to (S)-4-(trimethylsilyl)-3-butyn-2-ol {(S)-TMSBOL}, a key intermediate for synthesis of 5-lipoxygenase inhibitors, using immobilized Candida parapsilosis CCTCC M203011 cells as the biocatalyst.ResultsVarious ILs exerted significant but different effects on the bioreduction. Of all the tested water-immiscible ILs, the best results were observed with 1-butyl-3-methylimidazolium hexafluorophosphate (C4MIM·PF6), which exhibited not only good biocompatibility with the cells but also excellent solvent properties for the toxic substrate and product, thus markedly improving the efficiency of the bioreduction and the operational stability of the cells as compared to the IL-free aqueous system. 2-Propanol was shown to be the most suitable co-substrate for coenzyme regeneration, and it was found that the optimum volume ratio of buffer to C4MIM·PF6, substrate concentration, buffer pH, 2-propanol concentration and reaction temperature were 4/1 (v/v), 24 mM, 5.5, 130 mM and 30°C, respectively. Under these optimized conditions, the maximum yield and the product e.e. wer 97.7% and >99%, respectively, which are much higher than the corresponding values previously reported. The efficient whole-cell biocatalytic process was shown to be feasible on a 250-mL scale.ConclusionThe whole cell-catalyzed asymmetric reduction of TMSB to (S)-TMSBOL can be substantially improved by using a C4MIM·PF6/buffer biphasic system instead of a single-phase aqueous system and the resulting biocatalytic process appears to be effective and competitive on a preparative scale.
Highlights
Whole cells are usually employed for biocatalytic reduction reactions to ensure efficient coenzyme regeneration and to avoid problems with enzyme purification and stability
There have been a number of reports on biocatalytic reduction of ketones using microbial cells in various ionic liquids (ILs)-containing reaction systems, where the catalytic performances exhibited by the biocatalysts were closely related to the cation and anion types of ILs, and the effect of various ILs on the biocatalytic reactions has been found to vary widely [25,27,29,30,31]
TbFuhigfefuebrieobri1pehdauscictiosynsotefmTsMSB to (S)-TMSBOL with immobilized Candida parapsilosis CCTCC M203011 cells in water-immiscible IL/ The bioreduction of TMSB to (S)-TMSBOL with immobilized Candida parapsilosis CCTCC M203011 cells in water-immiscible IL/buffer biphasic systems
Summary
Whole cells are usually employed for biocatalytic reduction reactions to ensure efficient coenzyme regeneration and to avoid problems with enzyme purification and stability. As the silicon counterparts of chiral alcohols, enantiopure silicon-containing alcohols are becoming increasingly attractive, in that these silicon-containing compounds play an important role in asymmetric synthesis and functional materials, and in the preparation of silicon-containing drugs [3,4], such as Zifrosilone [5], Cisobitan [6] and TAC-101{4-[3,5-bis(trimethylsilyl)benzamido]benzoic acid} [7]. Such silicon-containing molecules generally have greater pharmaceutical activity, higher selectivity and lower toxicity than their carbon counterparts. Enzyme inactivation is frequently less of a problem when the enzyme is kept within the natural environments of living cells
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