Biofilm-based systems have recently attracted increasing attention as an alternative to the conventional cultivation in suspension. In Rotating Algal Biofilms (RAB), cells are periodically exposed to phases of light and darkness. In light/dark cycles, the exposition of the cells to darkness allows mitigating intense sunlight, thus protecting microalgae from photoinhibition. In order to better understand how light variations affect photosynthesis, a mechanistic model was developed based on Han’s model. It includes respiration dynamics and covers a wide range of cycle times and light fractions (Light/Dark fractions) and is calibrated with experimental data. Our model predicts accurately the biofilm behavior in various conditions of light intensity and duty cycles. Respiration variation in intermittent light regimes is for the first time considered to better predict biofilm growth under various cycle times and duty cycles. Data show that, for the same average light intensity, increasing the light frequency enhances growth thanks to reduced inhibition. Increasing the light fraction improves the growth rate as both the peak light intensity and the dark period decrease, which is beneficial at high and low frequencies. Model analysis provides clues to understand the biofilm response and suggests strategies for productivity enhancement, guiding reactor design and advanced process control.