Chlorella vulgaris and Scenedesmus sp. are commonly used in wastewater treatment due to their fast growth rates and ability to tolerate a range of environmental conditions. This study explored the cultivation of Chlorella vulgaris and Scenedesmus sp. using wastewater from the food industry, particularly from Italian basil pesto production tanks. The experiment involved different carbon dioxide concentrations and light conditions with a dilution rate of basil pesto wastewater at 1:2. Both microalgae strains were able to grow on pesto wastewater, and biomass characterization highlighted the influence of CO2 supply and light irradiation. The highest lipid storage was 79.3 ± 11.4 mg gdry biomass−1 and 75.5 ± 13.3 mg gdry biomass−1 for C. vulgaris and S. obliquus under red light (5% CO2 supply) and white light (0.04% CO2 supply), respectively. Protein storage was detected at 20.3 ± 1.0% and 24.8 ± 1.3% in C. vulgaris and S. obliquus biomasses under white light with a 5% CO2 and 0.04% CO2 supply, respectively. The removal of P, N, chemical oxygen demand, and biological oxygen demand resulted in 80–100%, 75–100%, 26–35%, and 0–20%, respectively.