Photophysiological parameters provide useful insights into the effects of environmental forcings on phytoplankton physiology. We present data on the short-term photoacclimative responses of phytoplankton throughout the water column during a diel sampling (every 1.5–3 h for 33 h) at a coastal station in the Gulf of Naples (Italy) in November 1996. Liposoluble pigments (high performance liquid chromatography), variable fluorescence (Prim-Prod probe) and picoplankton cell counts and autofluorescence (flow cytometry) were investigated every 1.5–3 h over a period of 33 h. The phytoplankton was phased to the alternation of light and dark and also showed acclimation to the different light intensities. Photoprotective pigments were synthesized during the day at the surface (0–20 m) and were significantly correlated with light intensity changes, as well as with the effective quantum yield of fluorescence. At night, recovery of photosystems from excess light was observed as was redistribution of nutrients and algae due to vertical convective motions caused by thermal dissipation. Equations linking photobiological parameters and time or light evolution were inferred to obtain kinetic coefficients. These were then used as biological tracers of vertical mixing whose velocity in the surface layer was estimated to be <0.05 cm s−1.