Abstract: To explore the comprehensive utilization value of polysaccharides from Haematococcus pluvialis residue, ethanol gradient precipitation (40%, 60% and 80%) was used to isolate polysaccharides from H. pluvialis residue in this study. Three fractions with crude polysaccharides (HP40, HP60 and HP80) were obtained by 40%, 60% and 80% ethanol precipitation successively. The monosaccharide composition, molecular weight, structure, particle size and Zeta potential of each fraction were analyzed by high performance liquid chromatography (HPLC), Fourier transform infrared spectroscopy (FT-IR), high pressure gel chromatography- multiple angle light scattering detector- differential refractive index detector (HPSEC-MALS-RI), and their antioxidant activity was also evaluated. The results showed that the physicochemical properties and antioxidant activity of polysaccharides precipitated from H. pluvialis residue were significantly affected by ethanol concentration. The yield and purity of the crude polysaccharides decreased with the increase of ethanol concentration. Furthermore, the lower molecular weight polysaccharides were precipitated by higher concentration of ethanol. High-purity crude polysaccharides were obtained in this study, in which, the total sugar content of HP40 was up to 99.46%±0.015%. All fractions contained 8 monosaccharides, including Glc, Man, Gal, Ara, Rha, Xyl, Fuc and Glc-UA, while the composition and proportion of monosaccharides were different. Compared with HP40, ribose was also involved in HP60 and HP80, which contained a total of 9 monosaccharides. HP80 possessed the smallest particle size (73.505±2.405 nm) and the largest Zeta potential absolute value (−11.6 mV). The results of antioxidant analysis showed that HP80 had the strongest antioxidant activity, and the scavenging rate of hydroxyl radicals was up to 96% (10.0 mg/mL). In summary, gradient ethanol precipitation method was effective to isolate polysaccharides from H. pluvialis residue preliminarily, and obtained polysaccharides with different monosaccharide composition, particle size, molecular weight and antioxidant activity. In which, the highest yield and sugar content was gained in fraction precipitated by 40 % ethanol (HP40), while the smallest molecular weight and the strongest antioxidant capacity were gained in fraction precipitated by 80 % ethanol (HP80).