Mangrove ecosystems are located in the critical land-sea interface in the tropics and subtropics and play a very important role in global primary productivity as well as in the coastal “blue carbon” sink. Biological nitrogen (N) fixation introduces new N to mangrove ecosystems to support part of the primary productivity and also has potential implications for biodiversity and the ecology of phytoplankton in the water column. Currently, most studies on nitrogen fixation in mangrove ecosystems have focused on soils, sediments, algal mats, and decomposing wood, while little is known about the N fixation in the water column of the mangrove system. In this study, we collected water samples from 12 sites in the mangrove ecosystem of Dongzhai Harbor, Haikou, and conducted incubation experiments on diel rhythm. The carbon and nitrogen fixation rates and the ability to utilize nutrients in mangrove water bodies were analyzed by combining ¹⁵N and ¹³C isotope tracing techniques with changes in nutrients during the incubation time. The results showed that there was obvious nitrogen fixation in the mangrove water body of Dongzhai Port, and it was mainly daytime nitrogen fixation. On the whole, the nitrogen fixation rate of water in the mangrove was higher than that in the outside, and the nitrogen fixation rate was 0.05μmol N L^-1 d^-1ï¿?.87 μmol N L^-1 d^-1, and there is a good positive correlation between nitrogen fixation rate and carbon fixation rate. It is suggested that nitrogen fixation in mangrove water is of great significance to the analysis of nutrient salt source and sink process in this system, and the carbon and nitrogen cycle process of mangrove ecosystem is more perfect.