In this study, we prepared novel amphiphilic phenyl-polysucrose microspheres by a two-step pathway. Crosslinked polysucrose microspheres were synthesized with soluble polysucrose and epichlorohydrin by inversed suspension polymerization first. Then, phenyl-polysucrose microspheres were obtained through the reaction between the polysucrose microspheres and glycidyl phenyl ether. Fourier transform infrared spectrometry and X-ray photoelectron spectroscopy proved that the microspheres had both hydroxyl groups and phenyl ligands. The quantitative determination of the phenyl groups indicated the optimal conditions for synthesis of the phenyl-polysucrose microspheres. The properties of the phenyl-polysucrose beads showed that the dry density increased from 1.38 to 2.29 g/mL, the equilibrium water content decreased from 77.30 to 33.68%, and the hydroxyl content remained at about 41.70 mmol/g when the phenyl content was increased from 0.00 to 2.64 mmol/g. The results of protein adsorption showed that the saturated adsorption capacities of the phenyl-polysucrose microspheres increased with increasing ion strength compared with the polysucrose microspheres. (C) 2009 Wiley Periodicals, Inc. J Appl Polym Sci 114: 4042-4050, 2009