Microalgae cultures have an excellent ability to capture CO2 and produce high, medium, and low valuable biocompounds such as proteins, carbohydrates, lipids, pigments, and polyhydroxyalkanoates; those compounds have shown excellent properties in the pharmaceutical, cosmetic, food, and medical industries. Recently, the supplementation of carbon dots (CDs) in microalgae cultures has been explored as a new strategy to increase light capture and improve photoluminescence, which in turn enhances biomass growth and biocompounds production. In this work, we synthesized CDs through a simple carbonization method using orange juice as a natural precursor. The green synthesized CDs were analyzed in detail through characterization techniques such as Fourier-transform infrared spectroscopy (FTIR), UV–visible, fluorescence spectroscopy, and ζ potential analysis. Moreover, CDs were added to Chlorella vulgaris to analyze the response under different photoperiod cycles and CDs dosages. The optimal results were obtained with the addition of 0.5 mg/L of CDs under a photoperiod cycle of 16h:8h (light:dark). In these conditions, a maximum biomass production of 2.12 g/L was observed, which represents an enhancement of 112% and 17% in comparison to the control samples under the photoperiod of 12h:12 h and 16h:8h (light/dark), respectively. Furthermore, the production of lipids, proteins, and carbohydrates was significantly increased to 249 mg/g, 285 mg/g, and 217 mg/g dry weight, respectively. These results suggest that the addition of CDs enhances cell growth and increases the production of lipids and proteins, being a strategy with great potential for the food and pharmaceutical industries.