Synthesis of d-xylulose from d-arabitol by enzymatic conversion with immobilized mannitol dehydrogenase from Rhodobacter sphaeroides
Abstract d -Xylulose was produced by enzymatic conversion of d -arabitol using mannitol dehydrogenase from Rhodobacter sphaeroides. The purified enzyme was immobilized on CNBr-activated Sepharose® 4B resulting in a yield of 40.5% with a binding capacity of 20.7 U g−1. The Kmapp value of the immobilized enzyme for d -arabitol was determined to be 1.9 mM, the inhibitor constants for d -xylulose and NADH were 1 mM and 0.2 mM, respectively. Bioconversion of d -arabitol was carried out in a batch procedure, using a methylene blue/O2/diaphorase system for regeneration of NAD+. The yield of the enzymatic conversion could be improved from 42% to 80% by complexing the produced d -xylulose with borate and thereby preventing a product inhibition of mannitol dehydrogenase. d -Xylulose was purified from the reaction mixture by ligand exchange chromatography on Ca2+ loaded DOWEX 50W X8 with a yield of 77%.