This study aims to validate the stable carbon isotopic composition (δ13C) of phytoplankton as a tool for detecting submarine leakages of anthropogenic CO2(g), since it is characterised by δ13C values significantly lower than the natural CO2 dissolved in oceans. Three culture experiments were carried out to investigate the changes in δ13C of the diatom Thalassiosira rotula during growth in an artificially modified medium (ASW). Three different dissolved inorganic carbon (DIC) concentrations were tested to verify if carbon availability affects phytoplankton δ13C. Simultaneously, at each experiment, T. rotula was cultured under natural DIC isotopic composition (δ13CDIC) and carbonate system conditions. The available DIC pool for diatoms grown in ASW was characterised by δ13CDIC values (−44.2 ± 0.9‰) significantly lower than the typical marine range. Through photosynthetic DIC uptake, microalgae δ13C rapidly changed, reaching significantly low values (until −43.4‰). Moreover, the different DIC concentrations did not affect the diatom δ13C, exhibiting the same trend in δ13C values in the three ASW experiments. The experiments prove that phytoplankton isotopic composition quickly responds to changes in the δ13C of the medium, making this approach a promising and low-impact tool for detecting CO2(g) submarine leakages from CO2(g) deposits.