Pleurotus eryngii was cultivated on four different sterilized agro wastes as wheat, rice, sugarcane and wood. Wheat substrate gave the maximum yield of P. eryngii fruiting bodies (63.4 and 11.3 gm ) for 100 gram of substrate for first flush and second flush respectively. Also, cultivation of tested mushroom on wheat substrate produced high quantity of polysaccharides (0.703 mg/ml) which was colormetrically evaluated by Phenol-Sulfuric acid method. The structure of endopolysaccharides was studied by Fourier Transform-Infrared spereectroscopy (FT-IR) noticed presence of O-H, C-H, C-O-C, CH2 and C-O bonds that indicated the appearance of c1-c5 by 1H & 13C NMR. Polysacchraides analysis with HPLC indicated that it is consisting of glucose, galactose, glucournic acid, ribose, rhamnose and mannose. Polysaccharides were obtained frome Pleurotus eryngii approved high antioxidant activity by Di phenyl Pichryl hydrocarbon (DPPH) radical scavenging activity which increased by increasing the concentration of polysaccharides. Polysaccharides had IC50 at 0.32 mg/ml.