Ecotoxicity characterization of dinitrotoluenes and some of their reduced metabolites

In the present study, the toxic effects of 2,4-dinitrotoluene (2,4-DNT), 2,6-dinitrotoluene (2,6-DNT) and a selection of their respective metabolites were examined and compared to 2,4,6-trinitrotoluene (TNT) using the 15-min Microtox (Vibrio fischen) and 96-h freshwater green alga (Selenastrum capricomutum) growth inhibition tests. All of the compounds tested were less toxic than TNT. Using the Microtox assay, 2,6-DNT was more toxic than 2,4-DNT and the order of toxicity for 2,6-DNT and its metabolites was: 2,6-DNT > or = 2A-6NT > 2,6-DAT; whereas that for 2,4-DNT was: 4A-2NT > 2A-4NT > 2,4-DNT > 2,4-DAT. For the algal test, 2,4-DNT was more toxic than 2,6-DNT and the order of toxicity for 2,4-DNT and its metabolites was: 2,4-DNT > 2,4-DAT approximately equal to 4A-2NT = 2A-4NT. The order of toxicity for 2,6-DNT and its reduced metabolites using the algal test was very similar to the Microtox bioassay. These results demonstrate that the reduced metabolites of 2,6-DNT tested in this study were less toxic than that of the parent compound, but certain partially reduced metabolites of 2,4-DNT can be more toxic than the parent molecule. These data put into question the general hypothesis that reductive metabolism of nitro-aromatics is associated with a sequential detoxification process.     

Phytotoxicity and phytoremediation of 2,6-dinitrotoluene using a model plant, Arabidopsis thaliana

Biochemical and genetic studies of xenobiotic metabolism in the model plant Arabidopsis have significant potential in providing information for phytoremediation. This paper presents the toxicity of 2,6-dinitrotoluene (2,6-DNT) to Arabidopsis under axenic conditions, the fate and transformation of 2,6-DNT after uptake by the plant, and the effect of a putative glutathione S-transferase (GST), which is highly induced by 2,4,6-trinitrotoluene (TNT) in the previous study, on the detoxification of 2,6-DNT. 2,6-DNT had toxic effects on the growth of Arabidopsis based on whole seedling as well as root growth assays. Using [U- 14C]2,6-DNT, the recovery was over 87% and less than 2% accounted for the mineralization of 2,6-DNT in axenic liquid cultures during the 14d of exposure. About half (48.3%) of the intracellular radioactivity was located in the root tissues in non-sterile hydroponic cultures. 2-Amino-6-nitrotoluene (2A6NT) and two unknown metabolites were produced as transformation products of 2,6-DNT in the liquid media. The metabolites were further characterized by proton NMR spectra and the UV-chromatograms when the plant was fed with either 2,6-DNT or 2A6NT. In addition, polar unknown metabolites were detected at short retention times from radiochromatograms of plant tissue extracts. The GST gene of the wild-type of Arabidopsis in response to 2,6-DNT was induced by 4.7-fold. However, the uptake rates and the tolerance at different concentrations of 2,6-DNT and TNT were not significantly different between the wild-type and the gst mutant indicating that induction of the GST gene is not related to the detoxification of 2,6-DNT.     

Toxicity and bioaccumulation of reduced TNT metabolites in the earthworm Eisenia andrei exposed to amended forest soil

Soils contaminated with 2,4,6-trinitrotoluene (TNT) and TNT primary reduction products have been found to be toxic to certain soil invertebrates, such as earthworms. The mechanism of toxicity of TNT and of its by-products is still not known. To ascertain if one of the TNT reduction products underlies TNT toxicity, we tested the toxicity and bioaccumulation of TNT reduction products. 2-Amino-4,6-dinitrotoluene (2-ADNT), 4-amino-2,6-dinitrotoluene (4-ADNT), 2,4-diamino-6-nitrotoluene (2,4-DANT) and 2,6-diamino-4-nitrotoluene (2,6-DANT) were tested separately in adult earthworms (Eisenia andrei) following a 14-d exposure to amended sandy loam forest soil. TNT, 4-ADNT, and 2-ADNT were lethal to earthworms (14-d LC(50) were: 580, 531 and 1088 micromol kg(-1), or 132, 105 and 215 mgkg(-1) dry soil, respectively) and gave the following order of toxicity: 4-ADNT>TNT>2-ADNT. Exposure to 2,4-DANT and to 2,6-DANT caused no mortality at 600 micromol kg(-1) or 100 mgkg(-1) dry soil. We found that all four TNT reduction products accumulated in earthworm tissues and 2-ADNT reached the highest levels at 3.0+/-0.3 micromol g(-1) tissue. The 14-d bioaccumulation factors were 5.1, 6.4, 5.1 and 3.2 for 2-ADNT, 4-ADNT, 2,4-DANT and 2,6-DANT, respectively. Results also suggest that some TNT metabolites are at least as toxic as TNT and should be considered when evaluating the overall toxicity of TNT-contaminated soil to earthworms.     

Experimental evidence for in situ natural attenuation of 2,4- and 2,6-dinitrotoluene in marine sediment

Dinitrotoluenes (DNTs) are widely used in the manufacturing of explosives and propellants hence causing contamination of several terrestrial and aquatic environments. The present study describes biotransformation of 2,4-DNT and 2,6-DNT in marine sediment sampled from a shipwreck site near Halifax Harbour. Incubation of either 2,4-DNT or 2,6-DNT in anaerobic sediment slurries (10% w/v) at 10 degrees C led to the reduction of both DNTs to their corresponding diaminotoluene (2,4-DAT and 2,6-DAT) via the intermediary formation of their monoamine derivatives (ANTs). The production of diaminotoluene was enhanced in the presence of lactate for both DNT isomers. Using [(14)C]-2,4-DNT less than 1% mineralization was observed as determined by liberated (14)CO(2). Sorption of DNTs, ANTs, and DATs was thus investigated to learn of their fate in marine sediments. Under anaerobic conditions, sorption followed the order: DNTs (K(d)=8.3-11.7lkg(-1))>ANTs (K(d)=4.5-7.0lkg(-1))>DATs (K(d)=3.8-4.5lkg(-1)). Incubation of 2,4-DAT in aerobic sediment led to rapid disappearance from the aqueous phase. LC/MS analysis of the aqueous phase and the acetone sediment extract showed the formation of azo- and hydrazo-dimers and trimers, as well as unidentified polymers. Experiments with radiolabelled 2,4-DAT showed a mass balance distributed as follows: 22% in the aqueous phase, 24% in acetone extracts, and 50% irreversibly bound to sediment. We concluded that DNT in anoxic marine sediment can undergo in situ natural attenuation by reduction to DAT followed by oxidative coupling to hydrazo-oligomers or irreversible binding to sediment.     

Identification of oxidized TNT metabolites in soil samples of a former ammunition plant

Water extracts of soil samples of the former ammunition plant “Tanne” near Clausthal-Zellerfeld, Lower Saxony, Germany, were investigated for highly polar oxidized 2,4,6-trinitrotoluene (TNT) metabolites. 0.4 to 9.0 mg/kg dry soil 2,4,6-trinitrobenzoic acid (TNBA) and 5.8 to 544 mg/kg dry soil 2-amino-4,6-dinitrobenzoic acid (2-ADNBA) were found. In addition to the oxidized metabolites, TNT, 4- and 2-aminodinitrotoluene (4- and 2-ADNT), and 2,4-dinitrotoluene (2,4-DNT) were extractable with water. Most interestingly, in one sample, 2-ADNBA represented the main contaminant. The origin of the oxidized nitroaromatics is unknown at this time. They might be generated chemically or photochemically. Furthermore, a biological synthesis seems possible.     

Phytotoxicity of nitroaromatic energetic compounds freshly amended or weathered and aged in sandy loam soil

The toxicities of 2,4,6-trinitrotoluene (TNT), 1,3,5-trinitrobenzene (TNB), 2,4-dinitrotoluene (2,4-DNT), and 2,6-dinitrotoluene (2,6-DNT) to terrestrial plants alfalfa (Medicago sativa L.), Japanese millet (Echinochloa crusgalli L.), and perennial ryegrass (Lolium perenne L.) were determined in Sassafras sandy loam soil using seedling emergence, fresh shoot, and dry mass measurement endpoints. A 13-week weathering and aging of energetic materials in soils, which included wetting and drying cycles, and exposure to sunlight of individual soil treatments, was incorporated into the study design to better reflect the soil exposure conditions in the field than toxicity determinations in freshly amended soils. Definitive toxicity tests showed that dinitrotoluenes were more phytotoxic for all plant species in freshly amended treatments based on EC20 values for dry shoot ranging from 3 to 24mgkg(-1) compared with values for TNB or TNT ranging from 43 to 62mgkg(-1). Weathering and aging of energetic materials (EMs) in soil significantly decreased the toxicity of TNT, TNB or 2,6-DNT to Japanese millet or ryegrass based on seedling emergence, but significantly increased the toxicity of all four EMs to all three plant species based on shoot growth. Exposure of the three plant species to relatively low concentrations of the four compounds initially stimulated plant growth before the onset of inhibition at greater concentrations (hormesis).     

Toxicological responses of red-backed salamanders (Plethodon cinereus) to subchronic soil exposures of 2,4-dinitrotoluene

Dinitrotoluenes are used as propellants and in explosives by the military and as such have been found at relatively high concentrations in the soil. To determine whether concentrations of 2,4-dinitrotoluene (2,4-DNT) in soil are toxic to amphibians, 100 red-backed salamanders (Plethodon cinereus) were exposed to either 1500, 800, 200, 75 or 0mg 2,4-DNT/kg soil for 28 days and evaluated for indicators of toxicity. Concentrations of 2,4-DNT were less than targets and varied with time. Most salamanders exposed to concentrations exceeding 1050 mg/kg died or were moribund within the first week. Salamanders exposed to soil concentrations exceeding 345 mg/kg lost >6% of their body mass though no mortality occurred. Overt effects included a reduction in feed consumption and an increase in bucco-pharyngeal oscillations in salamanders. These results suggest that only high soil concentrations of 2,4-DNT have the potential to cause overtly toxic effects in terrestrial salamanders.     

Biotransformation of 2,4,6-trinitrotoluene (TNT) by enchytraeids (Enchytraeus albidus) in vivo and in vitro

2,4,6-Trinitrotoluene (TNT) is toxic to soil invertebrates, but little is known about its toxicokinetic behavior in soil. Tissue residue analysis was used to evaluate whether the presence of TNT and its reduced metabolites in soil invertebrates was due to uptake of these compounds from the soil into the organism, or due to microbial transformation of TNT associated with the organism followed by uptake. Adult white potworms (Enchytraeus albidus) were exposed to non-lethal concentrations of TNT in amended artificial soil for 21 d, or to TNT in solution for 20 h. Soil exposure studies confirmed earlier reports that TNT was transformed in enchytraeids in vivo to 2- and 4-aminodinitrotoluenes. However, enchytraeid exposure to TNT in solution led to the additional presence of 2,4-diaminonitrotoluene as well as 2- and 4- hydroxyamino-dinitrotoluenes and azoxy-compounds, suggesting that TNT can be metabolized in vivo in the absence of soil. Incubation of unexposed enchytraeid homogenates with TNT led to a protein-dependent appearance of these metabolites in vitro after > or =16 h incubation. Cellular fractionation studies indicated that most of this activity resided in the 8000 x g pellet, and was completely inhibited by broad-spectrum antibiotics. These studies demonstrate that enchytraeids can transform TNT in vivo and in vitro, at least in part, by bacteria associated with the host organism.     

Decomposition of nitrotoluenes from trinitrotoluene manufacturing process by Electro-Fenton oxidation

Oxidative degradation of dinitrotoluene (DNT) isomers and 2,4,6-trinitrotoluene (TNT) in spent acid was conducted by Electro-Fenton's reagents. The electrolytic experiments were carried out to elucidate the influence of various operating parameters on the performance of mineralization of total organic compounds (TOC) in spent acid, including reaction temperature, dosage of oxygen, sulfuric acid concentration and dosage of ferrous ions. It deserves to note that organic compounds could be completely destructed by Electro-Fenton's reagent with in situ electrogenerated hydrogen peroxide obtained from cathodic reduction of oxygen, which was mainly supplied by anodic oxidation of water. Based on the spectra analyzed by gas chromatograph/mass spectrometer, it is proposed that initial denitration of 2,4,6-TNT gives rise to formation of 2,4-DNT and/or 2,6-DNT, which undergo the cleavage of nitro group into o-mononitrotoluene, followed by denitration to toluene and subsequent oxidation of the methyl group. Owing to the removal of both TOC and partial amounts of water simultaneously, the electrolytic method established is potentially applied to regenerate spent acid from toluene nitration processes in practice.     

Degradation of picric acid and 2,6-DNT in marine sediments and waters: the role of microbial activity and ultra-violet exposure

Bio- and photo-transformation of two munitions and explosives of concern, 2,6-dinitrotoluene (2,6-DNT) and 2,4,6-trinitrophenol (picric acid) were assessed in spiked marine sediments and water. A sandy and a fine-grained sediment, with 0.25% and 1.1% total organic carbon, respectively, were used for biotransformation assessments at 10 and 20 degrees C. Sterilized sediments were used as controls for biotic vs. abiotic transformation. Transformation products were analyzed by HPLC, GC/MS and LC/MS. Biotransformation in sediments started soon after the initial contact of the chemicals with the sediments and proceeded for several months, with rates in the following sequence: fine-grain at 20 degrees C > fine-grain at 10 degrees C > sand at 20 degrees C > sand at 10 degrees C. The biotransformation paths seemed to be similar for all conditions. The major biotransformation product of 2,6-DNT was 2-amino-6-nitrotoluene (2-A-6-NT). 2-Nitrotoluene (2-NT) and other minor components, including N,N-dimethyl-3-nitroaniline, benzene nitrile, methylamino-2-nitrosophenol and diaminophenol, were also identified. After more prolonged incubation these chemicals were replaced by high molecular weight polymers. Several breakdown products of picric acid were identified by GC/MS, including 2,4-dinitrophenol, amino dinitrophenols, 3,4-diamino phenol, amino nitrophenol and nitro diaminophenol. Photo-transformation of 2,6-DNT and picric acid in seawater was assessed under simulated solar radiation (SSR). No significant photolysis of picric acid in seawater was observed for up to 47 days, but photo-transformation of 2,6-DNT began soon after the initial exposure to SSR, with 89% being photo-transformed in 24 h and none remaining after 72 h. High molecular weight chemicals were generated, with mass spectra ranging from molecular weight 200-500 compared to 182 for DNT, and the color of the stock solution changed from clear to orange. Complexity of the mass spectra and mass differences among fragments suggest that multiple polymers were produced and were co-eluting during the LC/MS analyses.     

Acute toxicity assessment of explosive-contaminated soil extracting solution by luminescent bacteria assays

Explosive-contaminated soil is harmful to people’s health and the local ecosystem. The acute toxicity of its extracting solution was tested by bacterial luminescence assay using three kinds of luminescent bacteria to characterize the toxicity of the soil. An orthogonal test L ₁₆ (4⁵) was designed to optimize the soil extracting conditions. The optimum extracting conditions were obtained when the ultrasonic extraction time, ultrasonic extraction temperature, and the extraction repeat times were 6 h, 40 °C, and three, respectively. Fourier transform infrared spectroscopy (FTIR) results showed that the main components of the contaminated soil’s extracting solution were 2,4-dinitrotoluene-3-sulfonate (2,4-DNT-3-SO₃ ⁻); 2,4-dinitrotoluene-5-sulfonate (2,4-DNT-5-SO₃ ⁻); and 2,6-dinitrotoluene (2,6-DNT). Compared with Photobacterium phosphoreum and Vibrio fischeri, Vibrio qinghaiensis sp. Nov. is more suitable for assessing the soil extracting solution’s acute toxicity. Soil washing can remove most of the contaminants toxic to luminescent bacterium Vibrio qinghaiensis sp. Nov., suggesting that it may be a potential effective remediation method for explosive-contaminated soil.

     

Anaerobic biotransformation of dinitrotoluene isomers by Lactococcus lactis subsp. lactis strain 27 isolated from earthworm intestine

Dinitrotoluenes are widely used as solvents and are intermediates in the synthesis of dyes, explosives, and pesticides. Environmental concerns regarding DNTs have increased due to their widespread use and their discharge into the environment. In this study, the anaerobic biodegradation of four dinitrotoluene isomers, 2,3-, 2,4-, 2,6- and 3,4-DNT, was investigated using Lactococcus lactis subsp. lactis strain 27, which was isolated from the intestines of earthworms. Liquid chromatography/mass spectrometry and NMR spectroscopy showed that L. lactis strain 27 non-specifically reduced the nitro groups on the tested dinitrotoluenes to their corresponding aminonitrotoluenes. L. lactis strain 27, however, did not reduce either sequentially or simultaneously two nitro groups of the dinitrotoluenes, resulting in the formation of the corresponding diaminotoluenes. In vitro formation of dinitroazoxytoluenes suggested the presence of oxygen-sensitive hydroxylaminonitrotoluenes. L. lactis strain 27 was capable of reducing 2,4-, 2,6-, 2,3-, and 3,4-dinitrotoluenes up to 173.6, 66.6, 287.1, and 355 microM, respectively in 12 h incubation. A relatively rapid reduction was observed in the case of the 2,3-, and 3,4-dinitrotoluenes, which have vicinal nitro groups on their arene structure. Non-specific anaerobic reduction of dinitrotoluenes by the intestinal bacterium L. lactis strain 27 differentiated the extent of reduction of DNTs according to the substitutional position of the nitro groups and produced in vitro more toxic dinitroazoxytoluenes, suggesting that anaerobic biotransformation of dinitrotoluenes could increase environmental risk.     

Laccase mediated biodegradation of 2,4-dichlorophenol using response surface methodology

The effects of different environmental parameters, i.e., pH, temperature, time and enzyme concentration on the biodegradation of 2,4-dichlorophenol (2,4-DCP) in aqueous phase was evaluated with laccase from Pleurotus sp. using response surface methodology (RSM) in the present investigation. The Box-Behnken design of experiments was used to construct second order response surfaces with the investigated parameters. It was observed that the maximum degradation efficiency of approximately 98% was achieved at pH 6, temperature of 40 degrees C, time 9h and an enzyme concentration of 8IUml(-1). The adequacy of the model was confirmed by the coefficient of multiple regression, R(2) and adjusted R(2) which were adjudged to be 87.9% and 73.6%, respectively indicating a reasonably good model for practical implementation. Despite the fact that many successful attempts have been taken in the past for biodegradation of 2,4-DCP using whole cells, the present study emphasizes the fastest biodegradation of 2,4-DCP, a potent xenobiotic compound.     

Fungal laccase-catalyzed degradation of hydroxy polychlorinated biphenyls

Hydroxy polychlorinated biphenyls (hydroxy PCBs) are toxic metabolites of PCBs. Their toxicity such as strong endocrine disruption demands effective remediation methods. Laccases from Trametes versicolor and Pleurotus ostreatus were tested to degrade hydroxy PCBs. Optimum pHs for both enzymes were around 4.0. Laccase from T. versicolor degrades hydroxy PCBs more rapidly than that from P. ostreatus. The enzymatic activities remained little changes in up to 10% organic solvents, but decreased rapidly in more than 10% acetone, acetonitrile or DMSO. Degradation rate constants decreased with increase of chlorination and no degradation was observed with tetra-, penta- and hexa-chloro hydroxy PCBs in non-mediated reactions. However, the tetra- to hexa-chloro hydroxy PCBs were degraded by laccase from T. versicolor in the presence of the mediator 2,2,6,6-tetramethylpiperidine-N-oxy radical. The other mediators, 4-ethyl-2-methoxyphenol, 2,2'-azino-bis(3-ethylbenzthiazoline sulfonic acid) diammonium salt and 1-hydroxybenzotriazole and humic acid, also enhanced degradation of all the hydroxy PCBs except 4-hydroxy-2',3,3',4',5,5'-hexachlorobiphenyl. The results showed that 3-hydroxy biphenyl was more resistant to laccase degradation than 2- or 4-hydroxy analogues. Significant linear-correlations (coefficient of determination, r2 = 0.9097 and 0.8186 for laccases from P. ostreatus and T. versicolor, respectively) were found between the ionization potentials and the removal rate constants of hydroxy PCBs.     

Toxicological properties of thio- and alkylphenols causing flavor tainting in fish from the upper Wisconsin River

EC50 Microtox (5 min, 25 degrees C) assay values for 2-isopropylphenol, 3-isopropylphenol, 4-isopropylphenol, 2,4-diisopropylphenol, 2,5-diisopropylphenol 2,6-diisopropylphenol, 3,5-diisopropylphenol, carvacrol, thymol, thiophenol, and thiocresol ranged from 2 x 10(-2) mM for thymol (least toxic) to 2 x 10(-4) mM for 2,4-diisopropylphenol and 4-isopropylphenol (most toxic).     

Anaerobic incorporation of the radiolabeled explosive TNT and metabolites into the organic soil matrix of contaminated soil after different treatment procedures

Four bioreactor designs were performed to evaluate the level of incorporation of 14C-labeled 2,4,6-trinitrotoluene (TNT) and metabolites into the organic soil matrix of different anaerobically treated contaminated soils. The contaminated soils were amended with molasses slivers (80:20% per weight) as auxiliary substrate to enhance microbial activity. After 5 weeks (bioreactors 1 and 2), 8 weeks (bioreactor 3) and 12 weeks (bioreactor 4) of anaerobic incubation, we determined 41%, 58%, 72%, and 54%, respectively, of the initially applied radioactivity immobilized in various soil fractions. After alkaline hydrolyses of the solvent-extracted soils, low quantities of radiolabel were found in the humic and fulvic acid fractions, whereas the bulk of 14C activity was found to be strongly bound to the humin fraction (solid soil residues). The amounts of solvent extractable radioactivity were 53%, 40%, 16%, and 29% for bioreactors 1, 2, 3, and 4, respectively. The level of TNT transformation at the end of the experiments was within 90-94%. Regarding the results presented in this study, we can assume that there is the possibility of high incorporation levels of TNT metabolites into the soil organic matrix mediated by microbial cometabolism under strictly anoxic conditions.     

Laccase activity in soils: considerations for the measurement of enzyme activity

Laccases (benzenediol: oxygen oxidoreductases, EC 1.10.3.2) are copper-containing enzymes that catalyze the oxidative conversion of a variety of chemicals, such as mono-, oligo-, and polyphenols and aromatic amines. Laccases have been proposed to participate in the transformation of organic matter and xenobiotics as well as microbial interactions. Several laccase assays have been proposed and used in soils. Here, we show that the optimal pH conditions for the laccase substrates 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS, pH 3-5), 2,6-dimethoxyphenol (4-5.5), L-3,4-dihydroxyphenylalanine (DOPA; 4-6), guaiacol (3.5-5), 4-methylcatechol (3.5-5), and syringaldazine (5.5-7.0) are similar between purified laccases from Trametes versicolor and Pyricularia sp. and soil extracts; the substrate affinities of purified enzymes (K(M)) and soil extracts were also similar. The laccase assays showed specificity overlap with tyrosinase and ligninolytic peroxidases when hydrogen peroxide is present. The ABTS oxidation assay is able to reliably detect the presence of 13.5 pg mL(-1) or 0.199×10(-12) mol mL(-1) of T. versicolor laccase, which is three times more sensitive than the 2,6-dimethoxyphenol-based assay and more than 40 times more sensitive than any of the other assays. The low molecular mass soil-derived compounds and the isolated fulvic and humic acids influence the laccase assays and should be removed from the soil extracts before measurements of the enzyme activity are performed.     

Chemically catalyzed uptake of 2,4,6-trinitrotoluene by Vetiveria zizanioides

The efficiency of vetiver grass (Vetiveria zizanioides) in removing 2,4,6-trinitrotoluene (TNT) from aqueous media was explored in the presence of a common agrochemical, urea, used as a chaotropic agent. Chaotropic agents disrupt water structure, increasing solubilization of hydrophobic compounds (TNT), thus, enhancing plant TNT uptake. The primary objectives of this study were to: (i) characterize TNT absorption by vetiver in hydroponic media, and (ii) determine the effect of urea on chemically catalyzing TNT uptake by vetiver grass in hydroponic media. Results showed that vetiver exhibited a high TNT uptake capacity (1.026 mgg(-1)), but kinetics were slow. Uptake was considerably enhanced in the presence of urea, which significantly (p<0.001) increased the 2nd-order reaction rate constant over that of the untreated (no urea) control. Three major TNT metabolites were detected in the roots, but not in the shoot, namely 1,3,5-trinitrobenzene, 4-amino 2,6-dinitrotoluene, and 2-amino 4,6-dinitrotoluene, indicating TNT degradation by vetiver grass.     

Evaluation of biodegradation of nonylphenol ethoxylate and lignin by combining toxicity assessment and chemical characterization

The aerobic biodegradation of commercial nonylphenol ethoxylate (NPE) mixture and alkali lignin was studied using the OECD headspace test accompanied by the simultaneous measurement of ecotoxicity directly from the biodegradation liquors and by the follow-up of the chemical composition of the studied chemicals. NPE degradation was dependent on the inoculum source: approximately 40% of NPE was mineralized into CO₂ during the 4-week experiment when inoculum from Helsinki City wastewater treatment plant (WWTP) was used, and only 12% was mineralized when inoculum from Jyväskylä City WWTP was used. Chemical analyses revealed a shift in the ethoxylate chain length from longer to shorter soon after the beginning of the NPE biodegradation tests. At the same time also toxicity (reverse electron transport assay, RET) and estrogenic activity (human estrogen receptor yeast) measured directly from the biodegradation liquors decreased. In case of alkali lignin, approximately 11% was mineralized in the test and chemical analysis showed in maximum a 30% decrease in lignin concentration. Toxicity of lignin biodegradation liquors started to decrease in the beginning of the test, but became more toxic towards the end of the test again. Especially RET assay proved to be sensitive enough for measuring toxicity changes directly from biodegradation liquors, although a concentrating treatment of the liquors is recommended for a more detailed characterization and identification of toxic metabolites.     

Degradation of 2,4,6-trinitrotoluene by selected helophytes

Four emergent plants (helophytes, synonyms emersion macrophytes, marsh plants, etc.) Phragmites australis, Juncus glaucus, Carex gracillis and Typha latifolia were successfully used for degradation of TNT (2,4,6-trinitrotoluene) under in vitro conditions. The plants took up and transformed more than 90% of TNT from the medium within ten days of cultivation. The most efficient species was Ph. australis which took up 98% of TNT within ten days. The first stable degradation products 4-amino-2,6-dinitrotoluene (4-ADNT) and 2-amino-4,6-dinitrotoluene (2-ADNT) were identified and analysed during the cultivation period. [14C] TNT was used for the detection of TNT degradation products and their compartmentalization in plant tissues after two weeks of cultivation. Forty one percent of 14C was detected as insoluble or bound in cell structures: 34% in roots and 8% in the aerial parts. These results open the perspective of using the above-mentioned plants for the remediation of TNT contaminated waters.