The St. Johns River is Florida's longest river and a valued resource. The river is important for the region's ecology and socioeconomics. Human disturbances, flooding from hurricanes, and runoff from industrial and wastewater treatment facilities and agricultural fields in recent years has prompted the need for an increased frequency in monitoring of the water quality conditions in the St. Johns River. The objective of this study was to measure various water chemistry parameters and metal (cadmium, copper, lead, nickel, silver, and zinc) concentrations at 8 sites on the Lower St. Johns River associated with a variety of anthropogenic sources from 2017 to 2019. Water chemistry and metal concentrations varied to some degree across sites. Seasonal variation and variation due to episodic storm events were much more pronounced than the spatial variability. For example, salinity was significantly different on every sampling date. This parameter, among others, can directly influence aquatic life, as well as the bioavailability and toxicity of metals to aquatic organisms. All metals tested except zinc fluctuated at levels above EPA class III water quality criterion values, thus raising concerns about the future health of flora and fauna. This research provides reference data to improve the understanding of spatial and temporal variability of water quality parameters in the Lower St. Johns River.