This study compared the performance of the stable isotope composition of carbon (δ13C), oxygen (δ18O) and nitrogen (δ15N) by tracking plant response and genotypic variability of durum wheat to different salinity conditions. To that end, δ13C, δ18O and δ15N were analysed in dry matter (dm) and the water-soluble fraction (wsf) of leaves from plants exposed to salinity, either soon after plant emergence or at anthesis. The δ13C and δ18O of the wsf recorded the recent growing conditions, including changes in evaporative conditions. Regardless of the plant part (dm or wsf), δ13C and δ18O increased and δ15N decreased in response to stress. When the stress conditions were established just after emergence, δ15N and δ13C correlated positively with genotypic differences in biomass, whereas δ18O correlated negatively in the most severe treatment. When the stress conditions were imposed at anthesis, relationships between the three isotope signatures and biomass were only significant and positive within the most severe treatments. The results show that nitrogen metabolism, together with stomatal limitation, is involved in the genotypic response to salinity, with the relative importance of each factor depending on the severity and duration of the stress as well as the phenological stage that the stress occurs.