Light controls the developmental, physiological, morphological, and metabolic responses of many fungi. Most fungi respond primarily to blue, red, and green light through their respective photoreceptors. In this study, a screening of different light wavelengths’ effects on submerged Pleurotus ostreatus cultivation in baffled flasks was conducted. P. ostreatus growth was not inhibited in all tested conditions, while an equal or higher protein content was observed in comparison with dark conditions. Red and green light favored exopolysaccharide (EPS) production while red and blue light favored intracellular polysaccharide (IPS) production. To focus on EPS production, the effect of red and green light wavelengths on the production of the polysaccharide via submerged cultivation of P. ostreatus LGAM 1123 was tested. Submerged cultivation using red light in baffled flasks resulted in EPS production of 4.1 ± 0.4 g/L and IPS content of 23.1 ± 1.4% of dry weight (dw), while green light resulted in EPS production of 4.1 ± 0.2 g/L and 44.8 ± 5.2% dw IPS content. Similar production levels were achieved in a 3.5 L bioreactor using red light. The EPS produced using red light revealed a polysaccharide with a higher antioxidant activity compared to the polysaccharides produced by green light. In addition, the analysis of the crude polysaccharides has shown differences in biochemical composition. The structural differences and β glucan’s existence in the crude polysaccharides were confirmed by FT-IR analysis. Overall, these polysaccharides could be used in the food industry as they can enhance the functional health-promoting, physicochemical, and sensory properties of food products.