Agricultural industries produce vast amounts of liquid waste, which contains a significant concentration of nutrients. In the context of emphasizing the huge increase in population, climate changes, and pollution which results in depletion of fresh water resources, a sustainable solution for a greener future is needed. Wastewater treatment by the use of microalgae can mitigate a part of the problem by restoring water for irrigating agricultural crops. Little studies give insights on the physiological responses and ultrastructure of the Chlorophyta alga Desmodesmus sp. as it grows in cheese whey (CW). The algal strain was mixotrophically grown in a growth medium composed of CW only and CW supported with Bold’s basal medium (BBM) for 14 days. The potent response was observed with algal cultures fed by 15% CW enriched with 50% BBM. Fifteen percent CW in combination with 50% BBM significantly improved Desmodesmus sp. growth (303%), productivity (325%), and accumulation of cell metabolites, mainly lipids (3.89%), and carbohydrates (1.95%). On the contrary, protein and photosynthetic pigment (chlorophyll a, chlorophyll b, and carotenoids) contents were higher in BBM than in all treatments. Fatty acid composition demonstrated that the predominantly accumulated fatty acids were palmitic (25.86%), oleic (35.31%), and linoleic acid (13.22%). In conclusion, Desmodesmus sp. can be a good candidate for phycoremediation when cultivated on CW, whereas it can reduce the nutrition costs and water demand of algal cultivation by 50% and 15%, respectively. Therefore, it may be an effective strategy for algal mass production in sustainable agricultural systems.