Chlorination is a widely used method to prevent biofouling in power station cooling water systems in coastal and estuarine environments. This study evaluated the impact of chlorination together with temperature increase to simulate primary entrainment of a phytoplankton community. Biomass, diversity, and photosynthetic activity were monitored over 72 hours to establish impacts on the phytoplankton community. Biomass was significantly reduced after treatment. The mean cell size of the population significantly increased immediately after treatment highlighting an impact on the smaller cell size species of the community (picophytoplankton). Changes in accessory pigments composition suggest an effect on groups such as Prasinophyceae, Cyanobacteria and Chlorophycea. Species composition, dominated by diatoms, was also affected with Skeletonema marinoi and Asterionellopsis glacialis amongst the most sensitive species. Photosynthetic activity was affected in the short term but recovered after 48 hours. This study shows that by using a combination of measurements (e.g biomass, diversity, and physiology) the effects of entrainment in power station cooling water systems, that may be of longer-term significance for specific functional groups of phytoplankton communities, can be discerned. These changes would not necessarily be seen using individual techniques alone such as cell number counts or biomass assessment which may indicate apparent community recovery.