Metals and metalloids can represent a significant global issue, due to the potential for high bioaccumulation and biomagnification along aquatic food webs. Elasmobranchs, a taxonomic group that includes rays and sharks, are highly vulnerable to fishing activities and chemical pollutants. Few studies have investigated the presence and impacts of metals and metalloids in these animals, and studies with non-lethal sampling methods are still scarce. This study evaluated the potential of bile as a biomarker of recent contamination in three threatened species of elasmobranchs artisanally captured in the city of Rio de Janeiro, Gymnura altavela, Dasyatis hypostigma, and Pseudobatos horkelii. Biliary metal and metalloid concentrations were determined by Inductively Coupled Plasma Mass Spectrometry (ICP-MS), metallothioneins by UV-Vis spectrometry, and associations between intracellular metals and metallothioneins were investigated by Size Exclusion-High-Performance Liquid Chromatography coupled to ICP-MS (SEC-HPLC-ICP-MS). Overall, Fe presented the highest concentrations (0.7 more or less 0.1 mg L-1) and Sn the lowest (max. 0.004 mg L-1), quantified in only five individuals of the total studied (107). Arsenic and copper showed higher averages (10.9 and 5.2 mg L-1, respectively) in bile samples from Brazilian guitarfish, the only species caught in Copacabana. Statistical correlations among metallothionein concentrations, biometric indices, and elemental concentrations in bile fractions suggest environmental contamination and potential sublethal effects on the development and reproduction of these animals. Results obtained from SEC-HPLC-ICP-MS indicate metabolic differences in detoxification pathways among the various species. In conclusion, the utilization of elasmobranch bile is an effective alternative that allows non-lethal sampling for monitoring recent environmental contamination by metals and metalloids.