Careful nitrogen (N) management will be needed to nourish the growing human population while minimizing adverse environmental impacts. Aquaponic systems (AS) have a great potential to become a sustainable technology making further use of N-rich aquaculture wastewater. In the present study, we observed the N retention and losses in a running prototype of decoupled AS with Nile tilapia (Oreochromis niloticus) and tomatoes (Solanum lycopersicum) over 24 days. N losses amounted to 32.5% of feed N input and were observed in the recirculating aquaculture system (RAS) of the AS. Fish retained 21.1% of N input while 25.2% of N input accumulated in the RAS water. About 14.1% of the loss of N was caused probably by anaerobic denitrification processes in the lamellar settler (LS). In addition, 18.4% of N input was discharged during the three cleanings of LS. In the hydroponic unit of the AS that has been due to space limitations much smaller than an optimized AS could be (only about 20% of the optimal size relative to fish biomass), the tomato plants, including fruits, leaves, and stems, recovered 3.1% of N input with water uptake of 1700 L. The fish culture management, system design, and environmental management in the greenhouse affect the N recovery in the decoupled AS.