D-Arabinose isomerase from Klebsiella pneumoniae 40bXX was purified 12-fold with a 62.5% yield indicated by its electrophoretic homogeneity. The purified enzyme showed the highest activities toward D-arabinose and L-fucose as substrates at optimum conditions (50 mM glycine-NaOH, pH 9.0, 40 degrees C). The enzyme had a broad range of substrate specificities toward various D/L-aldoses, i.e., D-arabinose, L-fucose, D/L-xylose, D-mannose, D/L-lyxose, L-glucose, D-altrose and D/L-galactose. The equilibrium ratios between D-arabinose and D-ribulose, L-fucose and L-fuculose, D-altrose and D-psicose, and L-galactose and L-tagatose were 90:10, 90: 10, 13:87 and 25:75, respectively. Using a combination of the immobilized D-tagatose 3-epimerase and D-arabinose isomerase, we achieved the production of D-altrose from D-fructose in a batch reactor. We successfully produced approximately 12 g of D-altrose from 200 g of D-fructose in a reaction series with an overall yield of 6%. The product obtained was confirmed to be D-altrose by HPLC and C-13- NMR. To the best of our knowledge, this is the first report on the production of D-altrose from a cheap sugar, D-fructose, using an enzymatic method.