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The transition between phosphorus limitation of primary production in freshwater and nitrogen limitation in seawater was examined along an estuarine gradient simulated in 4 large 13 m3 enclosures connected in a series and containing pelagic and benthic subsystems. Nominal salinities of 0, 5, 10 and 25 ppt were maintained by exchanging appropriate volumes of water between enclosures. River water, which served as a freshwater endmember, was naturally high in N relative to P, while the oceanic endmember (water from Narragansett Bay, RI, USA) was low in N relative to P. Production in the water column was supported by external inputs and recycled nutrients. Bioassays, inorganic nutrient concentrations and N:P ratios of the seston and inorganic nutrients indicated that phosphorus was limiting at 0, 5 and 10 ppt, while nitrogen was limiting at 25 ppt. Coincident with this shift in limiting nutrient was a shift in the N:P ratio of nutrient supply from greater than the Redfield ratio of 16 to less than 16. External inputs established relative rates of supply in each enclosure. The relative proportion of N and P in external inputs was largely a function of the hydrodynamic mixing of fresh (high N, low P) and salt water (low N, high P) endmembers. At the scale of the estuarine segment or enclosure, neither recycled inputs from the benthos and water column, nitrogen fixation nor internal losses of N and P to sedimentation and/or denitrification materially altered relative supply rates, despite a hydrodynamic residence time of 27 d.