Euryhaline and coastal elasmobranchs are high-order predators that play important roles in ecosystem function and assemblage structure. Unfortunately, many of these species have declined in recent decades and are of increasing conversation concern. Understanding the trophic ecology of elasmobranchs in estuaries and coastal waters is important to develop our understanding of the ecological implications of these species and to inform conservation policy and management. Analyses of fatty acids (FA) and stable isotopes (SI) of δ13C and δ15N in body tissues are increasingly utilized, non-fatal approaches for obtaining integrated information on the dietary composition of animals (e.g., by collecting muscle biopsies or fin clips). In this study, we examined the dietary composition of four species of elasmobranchs (Carcharhinus leucas, Glyphis garricki, G. glyphis and Rhizoprionodon taylori), and compared them with putative prey species via analysis of muscle biopsies. By correlating muscle FA and SI within prey and sharks, we found significant seasonal differences in FA and SI profiles of prey species, but that such a pattern did not occur in elasmobranches. Both Glyphis spp. had similar FA and SI profiles that indicate the use of fresh and estuarine prey. In contrast, local prey species made only a minor contribution to the diet of C. leucas, suggesting that this species relies on alternative food sources. There was little evidence of intra-specific dietary specialization in these species, which may be due to the changes in sites and seasons of prey species. Our study shows that these sharks feed across multiple ecosystems, congregating in productive sections of the river.