Abstract: Biodegradation of MTBE under various multi-substrate conditions by Pseudomonas aeruginosa was investigated in this research. The addition of BTEX in various combinations significantly inhibited MTBE biodegradation. This result was mainly due to the non-competitive inhibition between MTBE and BTEX compounds. The rate of MTBE biodegradation decreased with the increasing substrate number for multi-substrate conditions. Additionally, the kinetic models developed in this research successfully simulate the degradation of MTBE under various multi-substrate conditions. However, the accumulation of TBA during MTBE biodegradation revealed that P. aeruginosa was unable to degrade TBA during the period of time tested.