This study assesses the unique chemistry of the Spring River of Arkansas. The Spring River supports one of the most diverse fish fauna in the south-central region of the United States and is an economic base for this region of Arkansas. The mouth of the Spring River in northeast Arkansas is located at Mammoth Spring where 34 million L of water per hour flow out of the spring mouth at a constant temperature of about 15°C. This slightly alkaline, high-Ca water, “warm” end member mixes with cooler downstream waters derived primarily from surface runoff. The variation in temperature and activity of complexing ligands leads to speciation of metals. We examined the temporal and spatial variations in metal species abundances and found that the availability of toxic forms of metals such as Pb, V, Cd, and Cu is highest during base flow and decreases at times of high runoff. Bioavailability of toxic metal species, in the context of ecotoxicology, was also explored. Metal content in fish gills, liver, and muscle was analyzed by inductively coupled plasma mass spectrometry. These concentrations were compared to the chemistry of dissolved load, with particular attention to the relative abundance of metal species. This study establishes the link between metal accumulation in predatory fish with high abundances of bioavailable metals in regions of the river dominated by base flow.