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Abstract
In the Cetus process crystalline D-fructose is produced from D-glucose via the intermediate 2-keto-D-glucose. Whereas the first step in the traditional process is catalyzed by the immobilized enzyme pyranose 2-oxidase, the ensuing reduction is performed by catalytic hydrogenation. In an entirely enzymatic variation of this process, soluble pyranose 2-oxidase from Trametes multicolor was employed. This biocatalyst could be efficiently stabilized under operational conditions by the addition of bovine serum albumin (BSA) together with catalase which decomposes hydrogen peroxide formed as a by-product. D-Glucose could be converted into 2-keto-D-glucose in yields above 98%. When the biocatalyst together with both stabilizing agents was separated from the product solution by ultrafiltration, it could be reutilized for several subsequent batch operation cycles. 2-Keto-D-glucose thus obtained was quantitatively reduced to D-fructose by NAD(P)-dependent aldose reductase from Candida tenuis. Two different enzymatic systems were successfully employed for the continuous regeneration of the coenzyme necessary in this reaction. In this way, D-fructose essentially free of D-glucose can be prepared by this simple and convenient method.