Sardina pilchardus and Engraulis encrasicolus are of great economic importance in Tunisia. However, little is known about their stock structure. Otolith chemistry was used to assess the feasibility of using this natural tag to discriminate populations of sardine and anchovy in the open sea and the coastal area of the Gulf of Gabes. The chemical composition of whole otoliths was analyzed using solution-based inductively coupled plasma mass spectrometry (ICPMS). Linear discriminant analysis classified fish into open sea and inshore groups with close to 97% accuracy for the anchovy and to 94% for the sardine. Respectively, the discrimination was based on P, Fe, Zn, Sc and Sr for the anchovy and P, Sr and B for the sardine. In addition, when we pool together the two species data in each area (inshore/open sea), the discrimination was also accurate reaching 88% of well classification based on P, Fe, Li and Mg. The results reveal a clear discreteness of the open sea and inshore small pelagic groups. Such information will have major implications for fisheries management of sardine and anchovy in Tunisia.