Although microalgae exploitation is very promising, process sustainability is undermined by biomass production and harvesting. Among the various bottlenecks of the production process, particular attention should be paid to the water footprint. Indeed, a huge volume of water is required in microalgae production. Water reuse can support both the water footprint and medium cost reduction, saving water and unconverted substrates. The present study reports preliminary results regarding the utilization of a water reuse strategy for two Chlorophyta microalgae under batch conditions. Growth parameters and chlorophyll content are monitored and the optimal amount of reused medium is assessed. The results show that 70% of the medium can be reused with no loss of specific growth rate and chlorophyll fraction for Pseudococcomyxa simplex in three consecutive batch cultivations. By contrast, a significant decline in Chlorella vulgaris growth was observed after the first cultivation in reused medium, across all tested conditions.