Quantum yields of photosystcm II (PSII) charge separation (ΦP and oxygen production () were determined by simultaneous measurements of oxygen production and variable fluorescence in four different aquatic microalgae representing three different taxonomic groups: the freshwater alga Scenedesmus protuberans (Chlorophyceae) and the marine algae Phaeocystis globosa (Prymnesiophyceae), Emiliania huxleyi (Prymnesiophyceae), and Phaeodactylum tricornutum (Bacillariophyceae). In S. protuberans, P. tricornutum, and E. huxleyi, light‐dependent variability was observed in the ratio of to ΦP i.e. in the number of oxygen molecules produced per electron generated by PSII. The ratio :ΦP was highly variable at low light intensities (E < 0.5 Ek), and at higher light intensities (E > 0.5 EK) :ΦP showed a nonlinear decrease with increasing light intensity. In contrast, in P. globosa a trend in :ΦP could not be distinguished, and this species showed a decrease in :ΦP during the day, indicating a dependency of :ΦP on light history. Additionally, considerable interspecific quantitative differences in :ΦP were observed. Two possible interpretations to explain the variability in :ΦP are discussed. Assuming that ΦP is a reliable measure of the quantum yield for charge separation at PSII, one interpretation is that net oxygen production is influenced by processes that consume oxygen or affect linear electron transport (e.g. cyclic electron transport around PSII, pseudocyclic electron transport in the Mehler reaction, Rubisco oxygenase activity, and lightdependent mitochondrial respiration). A second interpretation, however, suggests that at saturating light, changes in photosynthesis turnover time occur, such that ΦP does not predict the steady‐state O2 yield.