Increases in chloride concentration have been observed in water from numerous wells tapping the Floridan aquifer system in northeastern Florida. Although most increases have been in the eastern part of Duval County, Florida, no spatial pattern in elevated chloride concentrations is discernible. Possible sources of the mineralized water include modern seawater intrusion; unflushed Miocene-to-Pleistocene-age seawater or connate water in aquifer sediments; or mineralized water from deeper zones of the aquifer system or from formations beneath the Floridan aquifer system. The purpose of this study was to document the chemical and isotopic characteristics of water samples from various aquifer zones, and from geochemical and hydrogeologic data, to infer the source of the increased mineralization. Water samples were collected from 53 wells in northeastern Florida during 1997-1999. Wells tapped various zones of the aquifer including: the Fernandina permeable zone (FPZ), the upper zone of the Lower Floridan aquifer (UZLF), the Upper Floridan aquifer (UFA), and both the UFA and the UZLF. Water samples were analyzed for major ions and trace constituents and for isotopes of carbon, oxygen, hydrogen, sulfur, strontium, chlorine, and boron. Samples of rock from the aquifer were analyzed for isotopes of oxygen, carbon, and strontium. In general, water from various aquifer zones cannot be differentiated based on chemistry, except for water from FPZ wells. Major-ion concentrations vary as much within the upper zone of the Lower Floridan aquifer and the Upper Floridan aquifer as between these two zones. Simple models of mixing between fresh ground water and either modern seawater or water from the FPZ as a mineralized end member show that many water samples from the UZLF aquifer and the UFA are enriched in bicarbonate, calcium, magnesium, sulfate, fluoride, and silica and are depleted in sodium and potassium (as compared to concentrations predicted by simple mixing). Chemical mass-balance models of mixing and reactions between a hypothetical initial seawater and aquifer minerals cannot account for the observed water chemistry in a few wells, implying a source other than seawater, either ancient or modern, or the occurrence of other more complex rock-water reactions. Hydrogeologic and geochemical data from water and aquifer samples indicate that the most likely source of mineralized water in some wells yielding water with increasing chloride concentrations is water from the FPZ. In other wells, the flushing of Miocene-to-Pleistocene-age seawater can account for the observed chloride concentrations. The fact that most of the water samples collected are a mixture of less than one percent of mineralized water with more than 99 percent fresh or recharge water makes identifying the source of the mineralized water difficult. Differences in carbon-14 and sulfur-34 values probably reflect areal differences in aquifer mineralogy and distribution of organic carbon related to paleokarst features. Geochemical mass-balance models of seawater-rock interaction are unable to account for the chemical and isotopic composition of mineralized water from the FPZ, which implies another source of mineralized water, such as a brine, or the occurrence of more complex water-rock reactions.