Microalgal biofilms of Chlorella sorokiniana were cultivated under simulated day-night cycles at high productivity and high photosynthetic efficiency. Comparing day-night to continuous illumination did not demonstrate differences in the light utilization efficiency. This indicates that biomass consumed overnight represents sugar consumption for synthesis of new functional biomass and maintenance related respiration. Modelling microalgal biofilm growth was employed to calculate maximum productivities and photosynthetic efficiencies. A light limited microalgal biofilm growth model in which both diurnal carbon-partitioning and maintenance under prolonged dark conditions were taken into account was developed, calibrated, and validated experimentally. Extended periods of darkness resulted in reduced maintenance related respiration. Based on simulations with the validated biofilm growth model, it could be determined that the photosynthetic efficiency of biofilm growth can be higher than that of suspension growth. This is related to the fact that the maintenance rate in the dark zones of the biofilm is lower compared to that in the dark zones of suspension cultures which are continuously mixed with the photic zone.