The antioxidant activity of the polysaccharide from Ramulus mori and its derivatives (sulfated, phosphorylated, acetylated and benzoylated polysaccharides) were determined, including scavenging activity against superoxide, hydroxyl and 1,1-diphenyl-2-picrylhydrazyl radicals, reducing power and chelating ability. Obvious differences in antioxidant activity between the natural polysaccharide and its derivatives were observed, the antioxidant activity of the sulfated, phosphorylated and acetylated derivatives was stronger than that of the natural polysaccharide. In addition, the effect of temperature (20, 40, 60, 80 and 90 °C) and pH (3, 5, 7, 9 and 11) on the antioxidant activity of the natural polysaccharide and its derivatives was also investigated. The antioxidant activity of all samples in the linoleate emulsion improved with increasing from pH 3 to 11, while it decreased with ascending storage temperature from 20 to 90 °C.