Background Auricularia polytricha is known to be a highly nutritious foodstuff. We report here the purification, structure characterization and antimutagenic activity in vivo of a 0.9% NaCl solution-soluble polysaccharide ( SSP) from the mycelia of A. polytricha. Results Analysis by high-performance liquid chromatography coupled to a TSK-G5000PWXL column and gel filtration chromatography on Sephacryl S-400 HR indicated that SSP is homogeneous with an average molecular weight of about 9.30 × 105 Da. The structure of SSP was revealed by chemical methods, Fourier transform infrared spectroscopy and nuclear magnetic resonance spectroscopy. Results indicated that SSP is a glucan consisting of a1,3-β-glucan, 1,6-α-glucan, 1,4-α-glucan and 1,3-α-glucan backbone with a single 1,6-α- d-glucopyranosyl side-branching unit on every nine residues, on average, along the main chain. Atomic force microscopy indicates the presence of macromolecular species in morphology and shows a clear association of prolate particle. Meanwhile, SSP was found to significantly preventing micronuclei in polychromatic erythrocytes and reticulocytes of mice ( P < 0.01). Conclusion These results indicate that polysaccharide SSP from A. polytricha exhibits antimutagenic activity against the in vivo DNA-damaging effect of the indirectly acting alkylating agent cyclophosphamide.© 2013 Society of Chemical Industry [ABSTRACT FROM AUTHOR]