Mercury is an element with potential risk to fish and those who consume it. Thus, this study aimed to determine the levels of total mercury (THg), carry out a health risk assessment related to the consumption of the freshwater stingrays Potamotrygon motoro, and determine the physical and chemical properties of the water where stingrays occur. Stingrays of the species P. motoro were obtained from the Amazon River, and samples of the animals’ musculature were collected to determine THg levels. Risk assessment was conducted using pre-established formulas of estimated monthly intake (EMI), maximum monthly intake rate (IRmm), and hazard quotient (HQ). Three population scenarios were evaluated, considering both sexes and differences between rural and urban areas. There was no relationship between weight and THg concentration nor between total length and THg concentration. Higher EMI values were observed in rural children; for the IRmm, male children had the lowest consumption levels. For the hazard quotient, there was a similarity between the three age groups when comparing the male and female sexes. In addition, the representatives of the rural area always had lower values than the urban area. Freshwater stingrays, like other elasmobranchs, can be crucial animal species because they act as sentinels in studies that assess harmful chemicals like mercury.