Abstract: Off-line analysis of liquid phase samples have traditionally been used for the determination of organics and nitrogen load for process control or optimization in full-scale wastewater treatment plants. However, these sampling and analysis methods are labor-intensive and time-consuming. Besides, the results are not generally available on real-time to the operators who struggle on daily basis to meet the constant discharge license conditions of the effluent while the influent conditions change diurnally with varying loads. Combined respirometric titrimetric sensor is an on-line method that has been developed recently, indicating the oxygen demand exerted by organic and nitrogen loadings. This sensor monitors both the oxygen uptake as well as the proton production/consumption resulting from the biological reactions in activated sludge system. The basic-data can then be interpreted to find out the short-term BOD which can be used effectively for process control and optimization. Data interpretation and model development are still in progress as researchers keep on unraveling the mystery of how the substrates are biodegraded by activated sludge. Hence, in this study, it is aimed to develop an in-depth understanding about the aerobic biodegradation process using real-time biosensors through addition of different substrates such as acetate, surfactant (SDS) and ammonium to batch cultures of activated sludge by maintaining the pH of 7.8. Basic data interpretation was carried out to calculate Oxygen Uptake Rate (OUR) as well as proton production/consumption rate from the raw data. Besides, the improved bio-kinetic model calibration was presented in this study explaining the oxidation of different substrates in activated sludge system.