Degradation of 2,4,6-trinitrotoluene (TNT), a nitroaromatic explosive found in the soil and ground water, was investigated using Pseudomonas aeruginosa in in vitroexperiments. Biodegradable abilitiy of this bacteria was performed with 50 and 75 mg L−1 TNT concentrations in a defined liquid medium for 96 h time period. Treatment of TNT in supernatant samples taken at 0, 6, 12, 24, 48, 72 and 96 h from agitated vessels was followed by reverse-phase high-performance liquid chromatography (HPLC). In cultures supplemented with 50 and 75 mgL−1 TNT, after 96 h of incubation 46% and 59% reduction were detected respectively. Two metabolites as degradation intermediates with nitrite release into the medium, 2,4-dinitrotoluene (2,4-DNT) and 4-aminodinitrotoluene (4-ADNT), were elucidated by thin layer chromatography (TLC) and gas chromatography-mass spectrometry (GC-MS). These findings clearly indicate that Pseudomonas aeruginosa can be used in bioremediation of TNT contaminated sites.