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1.
PLoS One ; 14(10): e0224112, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31639172

RESUMO

Photolysis is one of the main transformation pathways for 2,4,6-trinitrotoluene (TNT) released into the environment. Upon exposure to sunlight, TNT is known to undergo both oxidation and reduction reactions with release of nitrite, nitrate, and ammonium ions, followed by condensation reactions of the oxidation and reduction products. In this study, compound classes of transformation products from the aqueous and solid phase photodegradation of 2,4,6-trinitrotoluene (TNT) have been identified by liquid and solid state 13C and 15N NMR. Aqueous phase experiments were performed on saturated solutions of T15NT in deionized water, natural pond water (pH = 8.3, DOC = 3.0 mg/L), pH 8.0 buffer solution, and in the presence of Suwannee River Natural Organic Matter (SRNOM; pH = 3.7), using a Pyrex-filtered medium pressure mercury lamp. Natural sunlight irradiations were performed on TNT in the solid phase and dissolved in the pond water. In deionized water, carboxylic acid, aldehyde, aromatic amine, primary amide, azoxy, nitrosophenol, and azo compounds were formed. 15N NMR spectra exhibited major peaks centered at 128 to 138 ppm, which are in the range of phenylhydroxylamine and secondary amide nitrogens. The secondary amides are proposed to represent benzanilides, which would arise from photochemical rearrangement of nitrones formed from the condensation of benzaldehyde and phenylhydroxylamine derivatives of TNT. The same compound classes were formed from sunlight irradiation of TNT in the solid phase. Whereas carboxylic acids, aldehydes, aromatic amines, phenylhydroxylamines, and amides were also formed from irradiation of TNT in pond water and in pH 8 buffer solution, azoxy and azo compound formation was inhibited. Solid state 15N NMR spectra of photolysates from the lamp irradiation of unlabeled 2,6-dinitrotoluene in deionized water also demonstrated the formation of aromatic amine, phenylhydroxylamine/ 2° amide, azoxy, and azo nitrogens.


Assuntos
Isótopos de Carbono/análise , Espectroscopia de Ressonância Magnética/métodos , Isótopos de Nitrogênio/análise , Fotólise/efeitos da radiação , Rios/química , Trinitrotolueno/metabolismo , Água/análise , Luz Solar , Trinitrotolueno/química , Trinitrotolueno/efeitos da radiação
2.
Org Biomol Chem ; 17(23): 5666-5670, 2019 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-30973549

RESUMO

Selective and sensitive detection of desired targets is very critical in sensor design. Here, we report a genetically engineered M13 bacteriophage-based sensor system evaluated by quantum mechanics (QM) calculations. Phage display is a facile way to develop the desired peptide sequences, but the resulting sequences can be imperfect peptides for binding of target molecules. A TNT binding peptide (WHW) carrying phage was self-assembled to fabricate thin films and tested for the sensitive and selective surface plasmon resonance-based detection of TNT molecules at the 500 femtomole level. SPR studies performed with the WHW peptide and control peptides (WAW, WHA, AHW) were well-matched with those of the QM calculations. Our combined method between phage engineering and QM calculation will significantly enhance our ability to design selective and sensitive sensors.


Assuntos
Bacteriófago M13/genética , Engenharia Genética , Trinitrotolueno/química , Regulação Viral da Expressão Gênica , Conformação Proteica , Teoria Quântica , Trinitrotolueno/metabolismo , Proteínas Virais
3.
Ecotoxicol Environ Saf ; 173: 452-460, 2019 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-30798189

RESUMO

Although laccase is involved in the biotransformation of 2,4,6-trinitrotoluene (TNT), little is known regarding the effect of E. coli laccase on TNT biotransformation. In this study, E. coli K12 served as the parental strain to construct a laccase deletion strain and two laccase-overexpressing strains. These E. coli strains were used to investigate the effect of laccase together with copper ions on the efficiency of TNT biotransformation, the variety of TNT biotransformation products generated and the toxicity of the TNT metabolites. The results showed that the laccase level was not relevant to TNT biotransformation in the soluble fraction of the culture medium. Conversely, TNT metabolites varied in the insoluble fraction analyzed by thin-layer chromatography (TLC). The insoluble fraction from the laccase-null strain showed fewer and relatively fainter spots than those detected in the wild-type and laccase-overexpressing strains, indicating that laccase expression levels were interrelated determinants of the varieties and amounts of TNT metabolites produced. In addition, the aquatic invertebrate Tigriopus japonicus was used to assess the toxicity of the TNT metabolites. The toxicity of the TNT metabolite mixture increased when the intracellular laccase level in strains increased or when purified E. coli recombinant Laccase (rLaccase) was added to the culture medium. Thus, our results suggest that laccase activity must be considered when performing microbial TNT remediation.


Assuntos
Proteínas de Bactérias/metabolismo , Copépodes/efeitos dos fármacos , Cobre/farmacologia , Escherichia coli/metabolismo , Lacase/metabolismo , Trinitrotolueno/toxicidade , Animais , Proteínas de Bactérias/genética , Biotransformação , Cromatografia em Camada Delgada , Escherichia coli/genética , Trinitrotolueno/metabolismo
4.
Phys Chem Chem Phys ; 21(22): 11589-11598, 2019 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-30801593

RESUMO

Biodegradation is a cost-effective and environmentally friendly alternative to removing 2,4,6-trinitrotoluene (TNT) pollution. However, mechanisms of TNT biodegradation have been elusive. To enhance the understanding of TNT biotransformation by the Old Yellow Enzyme (OYE) family, we investigated the crucial first-step hydrogen-transfer reaction by molecular dynamics simulations, docking technologies and empirical valence bond calculations. We revealed the significance of the π-π stacking conformation between the substrate TNT and the reduced flavin mononucleotide (FMNH2) cofactor, which is a prerequisite for the aromatic ring reduction of TNT. Under the π-π stacking conformation, the barrier of the hydrogen-transfer reaction in the aromatic ring reduction is about 16 kcal mol-1 lower than that of nitro group reduction. Then, we confirmed the mechanism of controlling the π-π stacking, that is, the π-π interaction competition mechanism. It indicates that the π-π stacking of TNT and FMNH2 occurs only when the π-π interaction between FMNH2 and TNT is stronger than that between TNT and several key residues with aromatic rings. Finally, based on the competition mechanism, the formation of π-π stacking of TNT and FMNH2 can be successfully enabled by removing the aromatic ring of those key residues in enzymes that originally only transform TNT through the nitro group reduction. This testified the validity of the π-π interaction competition mechanism. This work theoretically clarifies the molecular mechanism of the first-step hydrogen-transfer reaction for the biotransformation of TNT by the OYE family. It is helpful to obtain the enzymes that can biodegrade TNT through the aromatic ring reduction.


Assuntos
Flavoproteínas/metabolismo , NADPH Desidrogenase/metabolismo , Trinitrotolueno/metabolismo , Animais , Bactérias/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Biotransformação , Domínio Catalítico , Mononucleotídeo de Flavina/química , Flavoproteínas/química , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Himenópteros/enzimologia , Proteínas de Insetos/química , Proteínas de Insetos/metabolismo , Modelos Químicos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , NADPH Desidrogenase/química , Oxirredução , Ligação Proteica , Saccharomyces/enzimologia , Eletricidade Estática , Trinitrotolueno/química
5.
Planta ; 249(4): 1007-1015, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30488285

RESUMO

MAIN CONCLUSION: Transgenic western wheatgrass degrades the explosive RDX and detoxifies TNT. Contamination, from the explosives, hexahydro-1, 3, 5-trinitro-1, 3, 5-triazine (RDX), and 2, 4, 6-trinitrotoluene (TNT), especially on live-fire training ranges, threatens environmental and human health. Phytoremediation is an approach that could be used to clean-up explosive pollution, but it is hindered by inherently low in planta RDX degradation rates, and the high phytotoxicity of TNT. The bacterial genes, xplA and xplB, confer the ability to degrade RDX in plants, and a bacterial nitroreductase gene nfsI enhances the capacity of plants to withstand and detoxify TNT. While the previous studies have used model plant species to demonstrate the efficacy of this technology, trials using plant species able to thrive in the challenging environments found on military training ranges are now urgently needed. Perennial western wheatgrass (Pascopyrum smithii) is a United States native species that is broadly distributed across North America, well-suited for phytoremediation, and used by the US military to re-vegetate military ranges. Here, we present the first report of the genetic transformation of western wheatgrass. Plant lines transformed with xplA, xplB, and nfsI removed significantly more RDX from hydroponic solutions and retained much lower, or undetectable, levels of RDX in their leaf tissues when compared to wild-type plants. Furthermore, these plants were also more resistant to TNT toxicity, and detoxified more TNT than wild-type plants. This is the first study to engineer a field-applicable grass species capable of both RDX degradation and TNT detoxification. Together, these findings present a promising biotechnological approach to sustainably contain, remove RDX and TNT from training range soil and prevent groundwater contamination.


Assuntos
Substâncias Explosivas/metabolismo , Poaceae/genética , Poluentes do Solo/metabolismo , Triazinas/metabolismo , Trinitrotolueno/metabolismo , Biodegradação Ambiental , Engenharia Genética/métodos , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Poaceae/metabolismo , Reação em Cadeia da Polimerase em Tempo Real
6.
Mar Pollut Bull ; 135: 1072-1078, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30301003

RESUMO

Bioaccumulation of 2,4,6-trinitrotoluene (TNT) and its main metabolites 2-amino-4,6-dinitrotoluene (2-ADNT) and 4-amino-2,6-dinitrotoluene (4-ADNT) leaking from corroded munitions at a munitions dumping site (Kolberger Heide, Germany) was evaluated in transplanted blue mussels (Mytilus edulis). Six moorings with mussel bags were placed east and west at varying positions near the mine mound. In order to monitor any differences resulting from changing seasons, three exposure times were chosen. First exposure period: April-July 2016 (106 days); second exposure period: July-December 2016 (146 days); third exposure period: December 2016-March 2017 (92 days). We found amounts of 4-ADNT in mussel tissue ranging from 2.40 ±â€¯2.13 to 7.76 ±â€¯1.97 ng/(g mussel wet weight). Neither TNT nor 2-ADNT could be detected. Considering seasonal differences, orientation and distances of the moorings to the mine mound no correlation between levels in mussel tissue was evident.


Assuntos
Mytilus edulis/metabolismo , Trinitrotolueno/farmacocinética , Poluentes Químicos da Água/farmacocinética , Compostos de Anilina/farmacocinética , Animais , Ecotoxicologia/métodos , Substâncias Explosivas/metabolismo , Substâncias Explosivas/farmacocinética , Alemanha , Mytilus edulis/efeitos dos fármacos , Estações do Ano , Trinitrotolueno/análise , Trinitrotolueno/metabolismo , Poluentes Químicos da Água/metabolismo
7.
Mar Pollut Bull ; 135: 397-410, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30301051

RESUMO

Large amounts of ammunition containing 2,4,6-trinitrotoluene (TNT) and other substances were dumped in the Baltic Sea after WWII. Considering progressive corrosion processes, studying the transformation of TNT occurring in the environment constitutes an important aspect of a possible associated risk. This study focused on the transformations of TNT in simulated conditions of the Baltic Sea bottom sediment. Methods of analysis of TNT and selected products of its transformations were developed for that purpose. The developed methods allowed for the determination of selected compounds below 1 ng/g. Systematic monitoring of TNT transformations in the environment of the bottom sediment was performed. This allowed for the determination of the kinetics of TNT degradation and identification of degradation reaction products. Based on the obtained results, the TNT decay half-time in conditions present in the Baltic Sea was estimated to be 16.7 years for the abiotic environment and 5.6 for the biotic environment.


Assuntos
Cromatografia Gasosa-Espectrometria de Massas/métodos , Sedimentos Geológicos/análise , Trinitrotolueno/análise , Poluentes Químicos da Água/análise , Oceano Atlântico , Biodegradação Ambiental , Substâncias Explosivas/análise , Substâncias Explosivas/metabolismo , Cromatografia Gasosa-Espectrometria de Massas/instrumentação , Cinética , Espectrometria de Massas em Tandem/métodos , Trinitrotolueno/metabolismo , Poluentes Químicos da Água/metabolismo
8.
Phys Chem Chem Phys ; 20(17): 12157-12165, 2018 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-29683158

RESUMO

The explosive 2,4,6-trinitrotoluene (TNT) is a highly toxic pollutant. Biodegradation is inevitably one of the most cost-effective and enviromentally friendly means of removing TNT pollution. However, the aromatic derivatives from the reduction of nitro groups by several classic enzymes are still toxic. Besides the reduction of nitro groups, pentaerythritol tetranitrate reductase (PETNR) offers a potential route to ring fission and complete degradation of TNT through the pathway of the Meisenheimer complex. This work is devoted to deeply understand the essence of the Meisenheimer pathway and mainly focus on the crucial hydrogen-transfer reaction by means of molecular dynamics (MD) simulations. We obtain three valuable findings. Firstly, the parallel π-π stacking between TNT and the flavin mononucleotide (FMN) cofactor is a precondition. The key residue controlling this conformation is His181. Although His184 does not interact with TNT, the mutation from His184 to Asn184 would abolish the π-π structure. Secondly, the data of the empirical valence bond (EVB) show that the Meisenheimer pathway is predominant because its activation barrier is 6.7 kcal mol-1 far less than that of nitro reduction (26.6 kcal mol-1). Finally, based on the results of thermodynamic integration (TI), the type of transferred hydrogen is also ensured, that is, the H anion (H-) for the Meisenheimer complex and the H radical (H˙) for nitro reduction. Our findings provide an exhaustive understanding for the first hydrogen transfer reaction that has a decisive effect on two competing pathways, and help in searching for and designing new enzymes that can effectively degrade TNT.


Assuntos
Hidrogênio/química , Simulação de Dinâmica Molecular , Oxirredutases/metabolismo , Trinitrotolueno/metabolismo , Biodegradação Ambiental
9.
J Hazard Mater ; 349: 79-90, 2018 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-29414755

RESUMO

Citrobacter sp. has been shown to degrade 2,4,6-trinitrotoluene (TNT). However, the mechanism of its TNT biodegradation is poorly understood. An integrated proteome and transcriptome analysis was performed for investigating the differential genes and differential proteins in bacterial growth at the onset of experiments and after 12 h treatment with TNT. With the RNA sequencing, we found a total of 3792 transcripts and 569 differentially expressed genes (≥2 fold, P < 0.05) by. Genes for amino acid transport, cellular metabolism and stress-shock proteins were up-regulated, while carbohydrate transport and metabolism were down-regulated. A total of 42 protein spots (≥1.5 fold, P < 0.05) showed differential expression on two-dimensional gel electrophoresis and these proteins were identified by mass spectrometry. The most prominent proteins up-regulated were involved in energy production and conversion, amino acid transport and metabolism, posttranslational modification, protein turnover and chaperones. Proteins involved in carbohydrate transport and metabolism were down-regulated. Most notably, we observed that nemA encoding N-ethylmaleimide reductase was the most up-regulated gene involved in TNT degradation, and further proved that it can transform TNT to 4-amino-2,6-dinitrotoluene (4-ADNT) and 2-amino-4,6-dinitrotoluene (2-ADNT). This study highlights the molecular mechanisms of Citrobacter sp. for TNT removal.


Assuntos
Citrobacter/metabolismo , Poluentes do Solo/metabolismo , Trinitrotolueno/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biodegradação Ambiental , Citrobacter/genética , Oxirredutases/genética , Oxirredutases/metabolismo , Proteoma , Transcriptoma
10.
Aquat Toxicol ; 194: 176-184, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29197231

RESUMO

Organ specific uptake and depuration, and biological effects in Atlantic salmon (Salmo salar) exposed to 2, 4, 6-trinitrotoluene (TNT) were studied. Two experiments were conducted, the first using radiolabeled TNT (14C-TNT, 0.16mg/L) to study uptake (48h) and depuration (48h), while the second experiment focused on physiological effects in fish exposed to increasing concentrations of unlabeled TNT (1µg-1mg/L) for 48h. The uptake of 14C-TNT in the gills and most of the organs increased rapidly during the first 6h of exposure (12h in the brain) followed by a rapid decrease even though the fish were still exposed to TNT in the water. The radioactivity in the gall bladder reached a maximum after 55h, 7h after the transfer to the clean water. A high concentration of 14C-TNT in the gall bladder indicates that TNT is excreted through the gall bladder. Mortality (2 out of 14) was observed at a concentration of 1mg/L, and the surviving fish had hemorrhages in the dorsal muscle tissue near the spine. Analysis of the physiological parameters in blood from the high exposure group revealed severe effects, with an increase in the levels of glucose, urea and HCO3, and a decrease in hematocrit and the levels of Cl and hemoglobin. No effects on blood physiology were observed in fish exposed to the lower concentrations of TNT (1-100µg/L). TNT and the metabolites 2-amino-4,6-dinitrotoluene (2-ADNT) and 4-amino-2,6-dinitrotoluene (4-ADNT) were found in the muscle tissue, whereas only 2-ADNT and 4-ADNT were found in the bile. The rapid excretion and estimated bioconcentration factors (range of 2-18 after 48h in gills, blood, liver, kidney, muscle and brain) indicated a low potential for bioaccumulation of TNT.


Assuntos
Compostos de Anilina/análise , Bile/química , Salmo salar/metabolismo , Trinitrotolueno/metabolismo , Poluentes Químicos da Água/metabolismo , Animais , Biomarcadores Ambientais , Feminino , Masculino
11.
Biosens Bioelectron ; 102: 150-156, 2018 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-29128717

RESUMO

The olfactory system is a natural biosensor since its peripheral olfactory sensory neurons (OSNs) respond to the external stimuli and transmit the signals to the olfactory bulb (OB) where they are integrated and processed. The axonal connections from the OSNs expressing about 1000 different types of odorant receptors are precisely organized and sorted out onto 1800 glomeruli in the OB, from which the olfactory information is delivered to and perceived by the central nervous system. This process is carried out with particularly high sensitivity, specificity and rapidity, which can be used for explosive detection. Biomimetic olfactory biosensors use various biological components from the olfactory system as sensing elements, possessing great commercial prospects. In this study, we utilized the genetically labeled murine M72 olfactory sensory neurons with the green fluorescent protein (GFP) as sensing components and obtained long-term in vivo electrophysiological recordings from the M72 OSNs by implanting the microelectrode arrays (MEAs) into the behaving mouse's OB. The electrophysiological responses showed high reliability, reproducibility and specificity for odor detection, and particularly, the high sensitivity for the detection of odorants that contain benzene rings. Furthermore, our results indicated that it can detect trinitrotoluene (TNT) in liquid at a concentration as low as 10-5M and can distinguish TNT from other chemicals with a similar structure. Thus our study demonstrated that the in vivo biomimetic olfactory system could provide novel approaches to enhancing the specificity and increasing working lifespan of olfactory biosensors capable of detecting explosives.


Assuntos
Técnicas Biossensoriais/instrumentação , Odorantes/análise , Neurônios Receptores Olfatórios/metabolismo , Receptores Odorantes/metabolismo , Animais , Técnicas Biossensoriais/métodos , Eletrodos Implantados , Desenho de Equipamento , Substâncias Explosivas/análise , Substâncias Explosivas/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Masculino , Camundongos Transgênicos , Microeletrodos , Modelos Moleculares , Bulbo Olfatório/citologia , Bulbo Olfatório/fisiologia , Receptores Odorantes/genética , Trinitrotolueno/análise , Trinitrotolueno/metabolismo
12.
FEMS Microbiol Lett ; 365(1)2018 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-29228170

RESUMO

Trinitrotoluene (TNT) is an explosive chemical generally used for military, civil and industrial purposes. Therefore, TNT residue can be found in soil and ground water as an environmental pollutant. The environmental control of TNT pollution has become a critical issue because of its potential toxicity and carcinogenicity. The aim of this study is to evaluate the cyto-toxicological effects of TNT after bioremediation. Citrobacter sp. YC4 is able to utilize TNT as a sole nitrogen source. Citrobacter sp. YC4 cells grown in medium with TNT as the sole nitrogen source (TNT-N) were able to rapidly degrade TNT, in contrast to cells grown in Luria Bertani medium as determined by resting cell suspension. The concentration of TNT decreased from 100 to 0 ppm within 10 h in the solution containing TNT mixed with TNT-N-grown YC4. The cytotoxicity of TNT and its degradation products generated by TNT-N-grown YC4 were assessed by WST-1-based cell cytotoxicity assays. Our results showed that the cytotoxic potential of solutions containing TNT decreased almost to the level of the control after a 1-h incubation with TNT-N-grown YC4 cells. The rapid conversion of TNT into possibly less toxic products by Citrobacter sp. YC4 proposes a bioremediation prospection.


Assuntos
Biotransformação , Citrobacter/metabolismo , Poluentes do Solo/metabolismo , Trinitrotolueno/metabolismo , Células 3T3 , Animais , Biodegradação Ambiental , Células Hep G2 , Humanos , Camundongos , Nitrogênio/metabolismo , Microbiologia do Solo , Testes de Toxicidade
13.
PLoS One ; 12(12): e0189177, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29281650

RESUMO

Explosives are a common soil contaminant at a range of sites, including explosives manufacturing plants and areas associated with landmine detonations. As many explosives are toxic and may cause adverse environmental effects, a large body of research has targeted the remediation of explosives residues in soil. Studies in this area have largely involved spiking 'pristine' soils using explosives solutions. Here we investigate the fate of explosives present in soils following an actual detonation process and compare this to the fate of explosives spiked into 'pristine' undetonated soils. We also assess the effects of the detonations on the physical properties of the soils. Our scanning electron microscopy analyses reveal that detonations result in newly-fractured planes within the soil aggregates, and novel micro Computed Tomography analyses of the soils reveal, for the first time, the effect of the detonations on the internal architecture of the soils. We demonstrate that detonations cause an increase in soil porosity, and this correlates to an increased rate of TNT transformation and loss within the detonated soils, compared to spiked pristine soils. We propose that this increased TNT transformation is due to an increased bioavailability of the TNT within the now more porous post-detonation soils, making the TNT more easily accessible by soil-borne bacteria for potential biodegradation. This new discovery potentially exposes novel remediation methods for explosive contaminated soils where actual detonation of the soil significantly promotes subsequent TNT degradation. This work also suggests previously unexplored ramifications associated with high energy soil disruption.


Assuntos
Substâncias Explosivas , Poluentes do Solo/análise , Trinitrotolueno/química , Bactérias/metabolismo , Disponibilidade Biológica , Microscopia Eletrônica de Varredura , Porosidade , Microbiologia do Solo , Trinitrotolueno/análise , Trinitrotolueno/metabolismo , Microtomografia por Raio-X
14.
J Environ Qual ; 46(5): 968-974, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28991974

RESUMO

High explosives (HEs) deposited on military ranges can leach through the soil and contaminate groundwater. We examined the transport and fate of HEs in laboratory columns containing soils from two hand grenade bays (Bays C and T) and the impact of organic amendments on biodegradation. Soil characteristics were similar; however, Bay C had somewhat higher clay and organic C. Experimental treatments included addition of crude glycerin and lignosulfonate, and parallel control columns. Experimental results showed extensive 2,4,6-trinitrotoluene (TNT) degradation with minimal leaching, consistent with prior batch microcosm results. Amendment addition enhanced TNT degradation in both Bays C and T compared with controls. Although hexahydro-1,3,5-trinitro-1,3,5-triazine (Royal Demolition Explosive, or RDX) did not biodegrade in prior aerobic batch microcosms, 64 to 77% of RDX biodegraded in untreated soil columns with O present in the mobile soil gas. The RDX biodegradation was likely associated with short-term anoxic conditions or anoxic micro-niches. In nearly saturated Bay C columns, RDX removal increased to >92%. Amendment addition to unsaturated Bay T columns increased RDX removal to >86%. In one column, the soil remained anoxic (O < 5% by volume) for about a year after amendment addition, significantly reducing RDX leaching. Nitroso degradation products were produced equivalent to 9 to 39% of the RDX degraded, with most retained in the soil (9-37%) and 0 to 3% in the effluent. These results demonstrate that RDX biodegradation can occur in soils with measurable O, and that amendment addition can reduce RDX leaching by stimulating anaerobic biodegradation.


Assuntos
Biodegradação Ambiental , Substâncias Explosivas/metabolismo , Triazinas/metabolismo , Trinitrotolueno/metabolismo , Substâncias Explosivas/química , Solo , Microbiologia do Solo , Poluentes do Solo , Triazinas/química , Trinitrotolueno/química
15.
Toxicology ; 390: 117-123, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28899748

RESUMO

Since World War I considerable amounts of warfare material have been dumped at sea worldwide, but little is known about the fate of the explosive components in the marine environment. Sea dumped munitions are able to contaminate the surroundings because of the release of explosive chemicals due to corrosion and breaching or by detonation after blast-operations. This implies the risk of accumulation of toxic compounds in human and wildlife food chains. With the help of divers, we performed an active biomonitoring study with transplanted blue mussels (M. edulis) in a burdened area (Kolberger Heide, Germany) with explosive compounds near blast craters over an exposure time of 93days. With this biomonitoring system, we could show that blue mussels accumulate 2,4,6-trinitrotoluene (TNT) and its metabolites 2-amino-4,6-dinitrotoluene (2-ADNT) and 4-amino-2,6-dinitrotoluene (4-ADNT) in their tissues. In all mussels deployed at the ground, we found a body burden with 2-ADNT of 103.75±12.77ng/g wet weight and with 4-ADNT of 131.31±9.53ng/g wet weight. TNT itself has been found in six mussels with an average concentration of 31.04±3.26ng/g mussel wet weight. In the mussels positioned at one meter above the ground no TNT nor 2-ADNT could be detected, but 4-ADNT was found in all samples with an average concentration of 8.71±2.88ng/g mussel wet weight. To the best of our knowledge, this is the first study using blue mussels M. edulis as an active biomonitoring system for TNT and its metabolites 2-ADNT and 4-ADNT in a free field experiment in a burdened area. Moreover, with this system, we unequivocally proved that these toxic explosives accumulate in the marine biota resp. in the marine food chain, thereby posing a possible risk to the marine ecosphere and human health.


Assuntos
Monitoramento Ambiental/métodos , Mytilus edulis/metabolismo , Trinitrotolueno/metabolismo , Poluentes Químicos da Água/metabolismo , Poluição da Água , Compostos de Anilina/metabolismo , Animais , Biomarcadores/metabolismo , Biotransformação , Carga Corporal (Radioterapia) , Cadeia Alimentar , Contaminação de Alimentos , Alemanha , Humanos , Reprodutibilidade dos Testes , Medição de Risco
16.
Plant Mol Biol ; 95(1-2): 99-109, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28762129

RESUMO

KEY MESSAGE: Expression of the bacterial nitroreductase gene, nfsI, in tobacco plastids conferred the ability to detoxify TNT. The toxic pollutant 2,4,6-trinitrotoluene (TNT) is recalcitrant to degradation in the environment. Phytoremediation is a potentially low cost remediation technique that could be applied to soil contaminated with TNT; however, progress is hindered by the phytotoxicity of this compound. Previous studies have demonstrated that plants transformed with the bacterial nitroreductase gene, nfsI have increased ability to tolerate and detoxify TNT. It has been proposed that plants engineered to express nfsI could be used to remediate TNT on military ranges, but this could require steps to mitigate transgene flow to wild populations. To address this, we have developed nfsI transplastomic tobacco (Nicotiana tabacum L.) to reduce pollen-borne transgene flow. Here we have shown that when grown on solid or liquid media, the transplastomic tobacco expressing nfsI were significantly more tolerant to TNT, produced increased biomass and removed more TNT from the media than untransformed plants. Additionally, transplastomic plants expressing nfsI regenerated with high efficiency when grown on medium containing TNT, suggesting that nfsI and TNT could together be used to provide a selectable screen for plastid transformation.


Assuntos
Bactérias/enzimologia , Nitrorredutases/metabolismo , Plastídeos/genética , Tabaco/genética , Trinitrotolueno/metabolismo , Biodegradação Ambiental/efeitos dos fármacos , Vetores Genéticos/metabolismo , Plantas Geneticamente Modificadas , Regeneração/efeitos dos fármacos , Tabaco/efeitos dos fármacos , Tabaco/crescimento & desenvolvimento , Transformação Genética , Trinitrotolueno/toxicidade
17.
Molecules ; 22(8)2017 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-28805729

RESUMO

Microbial remediation of nitroaromatic compounds (NACs) is a promising environmentally friendly and cost-effective approach to the removal of these life-threating agents. Escherichia coli (E. coli) has shown remarkable capability for the biotransformation of 2,4,6-trinitro-toluene (TNT). Efforts to develop E. coli as an efficient TNT degrading biocatalyst will benefit from holistic flux-level description of interactions between multiple TNT transforming pathways operating in the strain. To gain such an insight, we extended the genome-scale constraint-based model of E. coli to account for a curated version of major TNT transformation pathways known or evidently hypothesized to be active in E. coli in present of TNT. Using constraint-based analysis (CBA) methods, we then performed several series of in silico experiments to elucidate the contribution of these pathways individually or in combination to the E. coli TNT transformation capacity. Results of our analyses were validated by replicating several experimentally observed TNT degradation phenotypes in E. coli cultures. We further used the extended model to explore the influence of process parameters, including aeration regime, TNT concentration, cell density, and carbon source on TNT degradation efficiency. We also conducted an in silico metabolic engineering study to design a series of E. coli mutants capable of degrading TNT at higher yield compared with the wild-type strain. Our study, therefore, extends the application of CBA to bioremediation of nitroaromatics and demonstrates the usefulness of this approach to inform bioremediation research.


Assuntos
Engenharia Metabólica/métodos , Trinitrotolueno/química , Trinitrotolueno/metabolismo , Biodegradação Ambiental , Biotransformação , Catálise , Técnicas de Cultura de Células , Escherichia coli , Cinética , Modelos Biológicos , Fenótipo , Biologia de Sistemas
18.
J Appl Microbiol ; 123(2): 401-413, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28561275

RESUMO

AIMS: To isolate and identify TNT-transforming cultures from explosive-contaminated soils with the ability to produce biosurfactants. METHODS AND RESULTS: Bacteria (pure and mixed cultures) were selected based on their ability to transform TNT in minimum media with TNT as the sole nitrogen source and an additional carbon source. TNT-transforming bacteria were identified by 16S rRNA gene sequencing. TNT transformation rates were significantly lower when no additional carbon or nitrogen sources were added. Surfactant production was enabled by the presence of TNT. Fourteen cultures were able to transform the explosive (>50%); of these, five showed a high transformation capacity (>90%), and six produced surfactants. CONCLUSIONS: All explosive-transforming cultures contained Proteobacteria of the genera Achromobacter, Stenotrophomonas, Pseudomonas, Sphingobium, Raoultella, Rhizobium and Methylopila. These cultures transformed TNT when an additional carbon source was added. Remarkably, Achromobacter spanius S17 and Pseudomonas veronii S94 have high TNT transformation rates and are surfactant producers. SIGNIFICANCE AND IMPACT OF THE STUDY: TNT is a highly toxic, mutagenic and carcinogenic nitroaromatic explosive; therefore, bioremediation to eliminate or mitigate its presence in the environment is essential. TNT-transforming cultures that produce surfactants are a promising method for remediation. To the best of our knowledge, this is the first report that links surfactant production and TNT transformation by bacteria.


Assuntos
Bactérias/isolamento & purificação , Bactérias/metabolismo , Poluentes do Solo/metabolismo , Tensoativos/metabolismo , Trinitrotolueno/metabolismo , Bactérias/classificação , Bactérias/genética , Biodegradação Ambiental , Biotransformação , Carbono/metabolismo , Nitrogênio/metabolismo , Microbiologia do Solo
19.
Ecotoxicol Environ Saf ; 138: 39-46, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28006730

RESUMO

In this study, the bacterial strain Citrobacter youngae strain E4 was isolated from 2,4,6-trinitrotoluene (TNT)-contaminated soil and used to assess the capacity of TNT transformation with/without exogenous nutrient amendments. C. youngae E4 poorly degraded TNT without an exogenous amino nitrogen source, whereas the addition of an amino nitrogen source considerably increased the efficacy of TNT transformation in a dose-dependent manner. The enhanced TNT transformation of C. youngae E4 was mediated by increased cell growth and up-regulation of TNT nitroreductases, including NemA, NfsA and NfsB. This result indicates that the increase in TNT transformation by C. youngae E4 via nitrogen nutrient stimulation is a cometabolism process. Consistently, TNT transformation was effectively enhanced when C. youngae E4 was subjected to a TNT-contaminated soil slurry in the presence of an exogenous amino nitrogen amendment. Thus, effective enhancement of TNT transformation via the coordinated inoculation of the nutrient-responsive C. youngae E4 and an exogenous nitrogen amendment might be applicable for the remediation of TNT-contaminated soil. Although the TNT transformation was significantly enhanced by C. youngae E4 in concert with biostimulation, the 96-h LC50 value of the TNT transformation product mixture on the aquatic invertebrate Tigriopus japonicas was higher than the LC50 value of TNT alone. Our results suggest that exogenous nutrient amendment can enhance microbial TNT transformation; however, additional detoxification processes may be needed due to the increased toxicity after reduced TNT transformation.


Assuntos
Biotransformação/efeitos dos fármacos , Citrobacter/efeitos dos fármacos , Fertilizantes , Poluentes do Solo/metabolismo , Trinitrotolueno/metabolismo , Aminoácidos/farmacologia , Biodegradação Ambiental/efeitos dos fármacos , Carbono/farmacologia , Células Cultivadas , Citrobacter/crescimento & desenvolvimento , Citrobacter/metabolismo , Nitrogênio/farmacologia , Nitrorredutases/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
20.
Environ Toxicol Chem ; 36(5): 1170-1180, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-27791286

RESUMO

The lack of knowledge on the fate of explosive compounds 2,4,6-trinitrotoluene (TNT) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), particularly in marine ecosystems, constrains the application of bioremediation techniques in explosive-contaminated coastal sites. The authors present a comparative study on anaerobic biodegradation and mineralization of 15 N-nitro group isotopically labeled TNT and RDX in organic carbon-rich, fine-grained marine sediment with native microbial assemblages. Separate sediment slurry experiments were carried out for TNT and RDX at 23°C for 16 d. Dissolved and sediment-sorbed fractions of parent and transformation products, isotopic compositions of sediment, and mineralization products of the dissolved inorganic N pool (15 NH4+ ,15 NO3- ,15 NO2- , and 15 N2 ) were measured. The rate of TNT removal from the aqueous phase was faster (0.75 h-1 ) than that of RDX (0.37 h-1 ), and 15 N accumulation in sediment was higher in the TNT (13%) than the RDX (2%) microcosms. Mono-amino-dinitrotoluenes were identified as intermediate biodegradation products of TNT. Two percent of the total spiked TNT-N is mineralized to dissolved inorganic N through 2 different pathways: denitration as well as deamination and formation of NH4+ , facilitated by iron and sulfate reducing bacteria in the sediments. The majority of the spiked TNT-N (85%) is in unidentified pools by day 16. Hexahydro-1,3,5-trinitro-1,3,5-triazine (10%) biodegrades to nitroso derivatives, whereas 13% of RDX-N in nitro groups is mineralized to dissolved inorganic N anaerobically by the end of the experiment. The primary identified mineralization end product of RDX (40%) is NH4+ , generated through either deamination or mono-denitration, followed by ring breakdown. A reasonable production of N2 gas (13%) was seen in the RDX system but not in the TNT system. Sixty-eight percent of the total spiked RDX-N is in an unidentified pool by day 16 and may include unquantified mineralization products dissolved in water. Environ Toxicol Chem 2017;36:1170-1180. © 2016 SETAC.


Assuntos
Substâncias Explosivas/metabolismo , Sedimentos Geológicos/química , Triazinas/metabolismo , Trinitrotolueno/metabolismo , Biodegradação Ambiental , Substâncias Explosivas/química , Marcação por Isótopo , Isótopos de Nitrogênio/química , Análise de Componente Principal , Triazinas/química , Trinitrotolueno/química , Água/química
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