Abstract
Ni2+-functionalized porous ceramic/agarose composite beads (Ni-NTA Cerose) can be used as carrier materials to immobilize enzymes harboring a metal affinity tag. Here, a 6×His-tag fusion alcohol dehydrogenase Mu-S5 and glucose dehydrogenase from Bacillus megaterium (BmGDH) were co-immobilized on Ni-NTA Cerose to construct a packed bed reactor (PBR) for the continuous synthesis of the chiral intermediate (S)-(4-chlorophenyl)-(pyridin-2-yl) methanol ((S)-CPMA) NADPH recycling, and in situ product adsorption was achieved simultaneously by assembling a D101 macroporous resin column after the PBR. Using an optimum enzyme activity ratio of 2:1 (Mu-S5: BmGDH) and hydroxypropyl-β-cyclodextrin as co-solvent, a space-time yield of 1560 g/(L·d) could be achieved in the first three days at a flow rate of 5 mL/min and substrate concentration of 10 mM. With simplified selective adsorption and extraction procedures, (S)-CPMA was obtained in 84% isolated yield.
Highlights
In comparison to batch reactions, flow reactions are relatively new for biocatalytic preparation of value-added chemicals and active pharmaceutical ingredients, such as (S)-1-phenylethanol, (R)-flurbiprofen, fatty acid methyl esters etc. [1, 2]
For continuous synthesis of (S)-CPMA, a prototype combined device consisting of four main parts was designed (Fig. 1), including a bottle of flow reaction solution, a packed bed reactor (PBR) containing Ni-NTA Cerose with immobilized Mu-S5 and BmGDH, a macroporous resin column for adsorbing hydrophobic products, and a beaker for effluent recovery
When the reaction solution flowed through the PBR, the ketone substrate CPMK was reduced into the corresponding chiral alcohol (S)-CPMA
Summary
In comparison to batch reactions, flow reactions are relatively new for biocatalytic preparation of value-added chemicals and active pharmaceutical ingredients, such as (S)-1-phenylethanol, (R)-flurbiprofen, fatty acid methyl esters etc. [1, 2]. In comparison to batch reactions, flow reactions are relatively new for biocatalytic preparation of value-added chemicals and active pharmaceutical ingredients, such as (S)-1-phenylethanol, (R)-flurbiprofen, fatty acid methyl esters etc. Fluid containing substrates is pumped to the reactor to produce a product stream [3,4,5,6]. When a flow process is applied to biocatalysis, it can be performed by using immobilized cells or enzymes [7,8,9,10,11]. Flow reactions have advantages of improved productivity, safety, ease of modifying scale, and are especially useful for reactions with substrate/product inhibitory issues [12]. For coupling or multienzymatic reaction systems, co-immobilization is an ideal strategy
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