To contribute towards effective exploitation and utilization of natural antioxidants, response surface methodology (RSM) was employed to optimize the medium composition for the production of exopolysaccharides from the medicinal mushroom Ganoderma lingzhi (GLEPS). An optimal medium for GLEPS production was gave through Plackett-Burman design, path of steepest ascent, and Box-Behnken design as follows: glucose (59.62 g/L), yeast extract (10. 03 g/L), CaCO (0.2 g/L), thiamine (45.13 mg/L), KHPO (1.0 g/L), peptone (1.5 g/L), Tween 80 (10.26 mL/L), ZnSO (0.3 g/L), mannitol (1.5 g/L), MgSO (0.5 g/L), and aspartate (8.86 g/L). The GLEPS yield obtained was 3.57 ± 0.21 g/L-3.16-fold higher than that produced in basal medium alone. The resulting GLEPS rich in uronic acid, d-mannose, l-rhamnose, and d-glucose, was a heteropolysaccharide with high-molecular weights (475,000 kDa and 21.6 kDa, 87.97%). It was demonstrated that the GLEPS with higher carbohydrate and uronic acid contents exhibited strong in vitro antioxidant activities via radical scavenging, reductive capacity, and chelation of transition metal catalysis. These findings indicated that RSM is an efficient tool to predict the composition of culture medium required for maximizing GLEPS yield, and GLEPS had potent antioxidant activities and could be explored as a novel natural antioxidant in functional food or medicine.