Anaerobic bacteria have been used to produce 2,4-dihydroxylamino-nitrotoluene (2,4DHANT), a reductive metabolite of 2,4,6-trinitrotoluene (TNT). Here, an aerobic TNT biodegrader Pseudomonas sp. strain TM15 produced 2,4DHANT as evidenced by the molecular ion with m/z of 199 identified from LC-TOFMS analyses. TNT biodegradation with a high cell concentration (10(9) cells/ml) led to a significant accumulation of 2,4DHANT in the culture medium, as well as hydroxylamino-dinitrotoluenes (HADNTs), although these products were not accumulated when a low cell concentration was used; also, the accumulation of diamino-nitrotoluene and of an unidentified metabolite were observed in the culture medium with the high cell concentration (10(10) cells/ml). 2,4DHANT overproduction was a function of the aeration speed since cultures with low aeration speeds (30 rpm) had a 19-fold higher DHANT productivity than those aerated with high speeds (180 rpm); this indicates that molecular oxygen was related to the formation of 2,4DHANT. The quantification of dissolved oxygen (DO) in the media demonstrated that the productivity of 2,4DHANT was increased at low DO values. Moreover, supplying oxygen to the culture media produced a remarkable decrease of 2,4DHANT accumulation; these results clearly indicate that high 2,4DHANT production was a consequence of the oxygen deficit in the culture medium. This finding is useful for understanding the TNT biodegradation (bioremediation technology) in an anoxic environment.