Nitrogenous nutrients—such as nitrate (NO3−), nitrite (NO2−), ammonium (NH4+), and urea—are key drivers of phytoplankton productivity and biogeochemical cycles in marine ecosystems, often entering coastal zones through estuarine discharge. Estuaries, particularly tropical ones, play a vital role in regulating the composition and export of nitrogen (N) species to downstream coastal waters. Despite their ecological significance, the factors influencing N utilization in tropical estuaries remain poorly understood. In this study, we used 15N isotope tracer incubation techniques to explore the seasonal variations in the uptake rates of NO3−, NO2−, NH4+, and urea in the Dongzhai Harbor estuary, a tropical system in China. We found that while NO3− was the most abundant reactive N species in both seasons, NH4+ was preferentially taken up by phytoplankton during the warm season. Conversely, urea uptake increased unexpectedly during the cold season, likely due to shifts in phytoplankton community composition favoring urea-preferred taxa like Peridiniopsis. Seasonal variations in N uptake rates were primarily governed by temperature and phytoplankton community structure. Enhanced inorganic N uptake during the warm season reduced the export of reactive inorganic N, whereas higher organic N uptake in the cold season facilitated the inorganic N exported to coastal waters. These findings highlight the interplay between environmental drivers and community dynamics in shaping nitrogen cycling in tropical estuaries. They underscore the importance of managing nutrient inputs to potentially safeguard sensitive downstream ecosystems, such as coral reefs, from eutrophication and degradation.