Daily light–dark cycles drive the circadian rhythm of many ocean processes including photosynthesis, gene expression, and zooplankton diel vertical migration (DVM). In phosphate deplete surface ocean regions, microbes produce metalloenzymes, such as alkaline phosphatases (AP), to access dissolved organic phosphorus. Here, we provide novel evidence of diurnal variation in AP activity (APA) in the subtropical North Atlantic using two independent datasets, with APA being two‐ to three‐fold higher at night. We demonstrate that zooplankton are a source of AP and postulate that zooplankton DVM is a source of enhanced AP in the surface waters at night, with reduction or degradation of AP during the day. Our results challenge the current assumption that APA is linear over a 24‐h period. While future ocean scenarios predict intensification and expansion of oceanic phosphate limitation, our findings indicate a role for zooplankton in regenerating phosphate that is currently missing in conceptual and numerical models.