The water-soluble crude polysaccharides were obtained from the fruiting bodies of cultured Cordyceps militaris by hot water extraction followed by ethanol precipitation. The polysaccharides were successively purified by chromatography on DEAE-cellulose-52 and Sephacryl S-100 HR columns, giving main three polysaccharide fractions termed P50-1, P70-1, and P70-2. Structural features of P70-1 were investigated by a combination of chemical and instrumental analysis, such as partial acid hydrolysis, methylation analysis, periodate oxidation - Smith degradation, GC-MS, C-13 NMR, HPAEC-PAD. and FT-IR. The results indicated that P70-1 has a backbone of (1 -> 6)-linked beta-D-mannopyranosyl residues, which occasionally branches at O-3. The branches were mainly composed of (1 -> 4)-linked alpha-D-glucopyranosyl and (1 -> 6)-linked beta-D-galactopyranosyl residues, and terminated with beta-D-galactopyranosyl residues and alpha-D-glucopyranosyl residues. In the in vitro antioxidant assay, P70-1 was found to possess hydroxyl radical-scavenging activity with an IC50 value of 0.548 mg/ml. (C) 2007 Elsevier Ltd. All rights reserved.