Simulation analysis based on a mathematical model is one of the powerful tools for determining the operational conditions for a full scale biological nutrient removal plant. The model that included the Activated Sludge Model No.2 was developed for simulating the performance of the plant in Sapporo City. The investigated plant has the biological reaction basin which consists of the four zones, anaerobic-aerobic-anoxic-aerobic phases with the step feed of the primary effluent to the anaerobic and anoxic zones. We performed three experiments to calibrate and verify our model: (i) Characterization of organic matters in the influent of the reaction basin with the OUR measurement method; (ii) Measurement of COD and nutrient concentration at the plant; (iii) Lab-scale batch experiments with the anaerobic-aerobic-anoxic-aerobic phases. The calibrating process of the model showed that no modification of parameter values was required to evaluate the performance of the plant and population of the activated sludge. Simulated results showed that the choice of the sewage step feed ratio did not affect the nitrogen removal and that the denitrification rate in the anoxic zone was controlled by the hydrolysis rate of the slowly biodegradable organic matters. The results of the lab-scale experiment and simulation results showed that the addition of the readily biodegradable organic matters like fermentation products of the primary settler sludge was effective to improve the performance of nitrogen removal.