We investigated the carbon acquisition of three marine microalgae, Skeletonema costatum, Phaeocystis globosa, and Emiliania huxleyi in response to different light regimes. Rates of photosynthetic O 2 evolution and CO 2 and HCO uptake were measured by membrane inlet mass spectrometry in cells acclimated to cycles of 16 : 8 light :Ϫ 3 dark (LD; h : h) and 12 : 12 LD and were compared with those obtained under continuous light. In addition, cellular leakage was estimated for different photoperiods and ambient CO 2 concentrations during growth. Maximum rates of photosynthesis more or less doubled under LD cycles compared with continuous light. In S. costatum and E. huxleyi, a remarkably higher contribution of HCO to the overall carbon uptake was observed under LD cycles. Incontrast, P. globosa did not change its CO 2 : HCO uptake ratio in response to daylength. Half saturation concen-Ϫ 3 trations (K 1/2 ) for O 2 evolution and inorganic carbon (Ci) uptake were also influenced by the photoperiod. Under LD cycles K 1/2 values for photosynthesis in S. costatum and P. globosa were similar or higher compared with continuous light, whereas they were much lower in E. huxleyi. With the exception of CO 2 uptake in E. huxleyi and P. globosa, affinities for Ci decreased under the LD cycles. Cellular leakage was highest for E. huxleyi and lowest for S. costatum and generally decreased with increasing CO 2 concentration. Although this study confirms speciesspecific differences in the CO 2 -concentrating mechanisms (CCMs), the effect of daylength on CO 2 and HCO uptake Ϫ 3 has hitherto not been described. We put forward the idea that variations in light condition influence the cellular carbon demand, thereby imposing a stronger control on CCM regulation than the naturally occurring changes in CO 2 supply.