Sugar-oxidizing enzymes are widely detected in many fungal species. In this paper, a new cellooligosaccharide oxidase (COOX) was purified to homogeneity from wheat bran culture of a soil-isolated rice pathogen fungus strain, Sarocladium oryzae F137. This new sugar oxidase was composed of a single polypeptide chain with a molecular mass of 55 kDa and isoelectric point of 4.8-4.9. This enzyme contained 1 mol of FAD per mole of enzyme but no heme domain. This enzyme selectively bound to cellulose, but not to starch, chitin, or xylan. This enzyme oxidized oligosaccharides with reducing-end glucosyl residues linked by an beta-1,4 glucosidic bond, such as lactose, cellobiose, and cellooligosaccharids. No significant substrate inhibition was observed at high concentrations of lactose, cellobiose, and maltose. COOX was used to produce a natural antioxidant, lactobionic acid, in a batch type reaction. The enzyme showed a preference for the two-electron acceptor 2,6-dichlorophenol-indophenol over the one-electron acceptors cytochrome c or ferricyanide. COOX did not contain a heme domain and used only two-electron acceptor. This feature could distinguish between the extracellular hemoflavoenzyme, cellobiose dehydrogenase, which is produced by a number of wood-degrading and phytopathogenic fungi. (c) 2005 Elsevier Inc. All rights reserved.