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Transformation and detoxification of 2,4,6-trinitrotoluene by a Pseudomonas aeruginosa strain
Abstract
Although TNT (2,4,6-trinitrotoluene) can be degraded by numerous soil microorganisms, complete and effective bioremediation requires optimizing transformation pathways that maximize degradation and minimize toxic metabolite production. Under aerobic conditions, Pseudomonas aeruginosa strain MX, isolated from munitions-contaminated soil, transformed 100 mg TNT L-1 to 2-hydroxylamino-4,6-dinitrotoluene (2HADNT), 2-amino-4,6-dinitrotoluene (2ADNT), and 2,2′-azoxytoluene (2,2′AZT) in culture medium within 10 h. The 2HADNT and 2,2′AZT were relatively less toxic than TNT, while exposure to TNT and the aminodinitrotoluenes (ADNTs) resulted in condensation of DNA at the cell centers. Yeast extract, which provides a source of carbon and nitrogen, was the most effective energy source for TNT transformation by the isolate. Metabolite production and accumulation were affected by carbon source, NADPH, and pH. While TNT transformation was optimum at pH 7 to 8, ADNT and HADNT accumulated under acidic conditions and more AZT was produced at higher pH. A simple and rapid colorimetric assay based on TNT absorbance at 447 nm (pH 12.2) was developed and used to quantify nitroreductase-mediated TNT transformation to colorless metabolites. TNT-nitroreductase from crude extracts obtained from P. aeruginosa strain MX was purified by sequential phenyl-sepharose, hydroxyapatite, and anion exchange chromatography, and its molecular weight was approximately 54 kDa. This purified TNT-nitroreductase reduced TNT to HADNT and ADNT. TNT reduction increased with increasing NADH, NADPH, and protein concentrations. TNT reduction to HADNTs in the presence of P. aeruginosa strain MX was enzymatic, while AZT production from HADNTs occurred abiotically under aerobic conditions. Detection of 2,4,6-trihydroxytoluene and cyclohexanone indicated the potential for TNT metabolism by the strain to less toxic and more biodegradable products. The transient appearance of 2,6-diamino-4-nitrotoluene (2,6DANT) and 2,4-diamino-6-nitrotoluene (2,4DANT); triaminotoluene (TAT); dinitro- and nitrotoluene; and 3,5-dinitroaniline indicated that various metabolic routes occurred in microbial metabolism and detoxification of TNT. Results from this research indicate the importance of environmental conditions on TNT degradation and provide guidance in developing strategies for remediating contaminated soil.
Subject Area
Microbiology|Environmental engineering|Environmental science
Recommended Citation
Oh, Byung-Taek, "Transformation and detoxification of 2,4,6-trinitrotoluene by a Pseudomonas aeruginosa strain" (1999). ETD collection for University of Nebraska-Lincoln. AAI9952689.
https://digitalcommons.unl.edu/dissertations/AAI9952689