The extensive use of nanoparticles (NPs) in diverse applications causes their localization to aquatic habitats, affecting the metabolic products of primary producers in aquatic ecosystems, such as algae. Synthesized calcium oxide nanoparticles (CaO NPs) are of the scarcely studied NPs. Thus, the current work proposed that the exposure to CaO NPs may instigate metabolic pathway to be higher than that of normally growing algae, and positively stimulate algal biomass. In this respect, this research was undertaken to study the exposure effect of CaO NPs (0, 20, 40, 60, 80, and 100 µg mL −1 ) on the growth, photosynthesis, respiration, oxidative stress, antioxidants, and lipid production of the microalga Coccomyxa chodatii SAG 216-2. The results showed that the algal growth concomitant with chlorophyll content, photosynthesis, and calcium content increased in response to CaO NPs. The contents of biomolecules such as proteins, amino acids, and carbohydrates were also promoted by CaO NPs with variant degrees. Furthermore, lipid production was enhanced by the applied nanoparticles. CaO NPs induced the accumulation of hydrogen peroxide, while lipid peroxidation was reduced, revealing no oxidative behavior of the applied nanoparticles on alga. Also, CaO NPs have a triggering effect on the antioxidant enzymes such as superoxide dismutase, catalase, ascorbate peroxidase, and guaiacol peroxidase. The results recommended the importance of the level of 60 µg mL −1 CaO NPs on lipid production (with increasing percentage of 65% compared to control) and the highest dry matter acquisition of C. chodatii. This study recommended the feasibility of an integrated treatment strategy of CaO NPs in augmenting biomass, metabolic up-regulations, and lipid accumulation in C. chodatii.