An object of the present invention is to provide a novel polysaccharide derivative used for a separating agent for optical isomers. The novel polysaccharide derivative contains a structure in which a hydrogen atom of a hydroxyl group or an amino group at the 2-position in a structural unit of the polysaccharide is substituted with a monovalent group represented by the following general formula (1), and a hydrogen atom of a hydroxyl group or an amino group at the 3-position in the structural unit is substituted with a monovalent group represented by the following general formula (2): R1—NH—CO—  (1) R2—NH—CO—  (2) where R1 and R2 represent substituted or unsubstituted aryl groups which are different from each other.