Enzymatic production of pure D-mannitol at high productivity

An enzymatic, NAD(H)-dependent process for the efficient production of D-mannitol from D-fructose as one single product is described and optimized with respect to productivity at high substrate conversion. Stereospecific reduction of D-fructose is catalyzed by recombinant mannitol dehydrogenase from Pseudomonas fluorescens DSM 50106, overexpressed. in Escherichia coli. Regeneration of NADH is accomplished by formate dehydrogenase-mediated oxidation of formate into CO?, thus avoiding byproduct formation and yielding total turnover numbers for the coenzyme of approximately 1000 for a single round of D-fructose conversion. In optimized batchwise reduction of D-fructose, a D-mannitol productivity of 2.25 g/(Lh) was obtained for a final product concentration of 72 g/L and a D-fructose conversion of 80%. D-Mannitol was crystallized From the ultrafiltered product solution in 97% purity and 85% recovery, thus also allowing reuse of enzymes for repeated batchwise production of D-mannitol.