Anaerobic degradation of benzene, toluene, ethylbenzene, and xylene compounds by Dechloromonas strain RCB

Dechloromonas strain RCB has been shown to be capable of anaerobic degradation of benzene coupled to nitrate reduction. As a continuation of these studies, the metabolic versatility and hydrocarbon biodegradative capability of this organism were investigated. The results of these revealed that in addition to nitrate, strain RCB could alternatively degrade benzene both aerobically and anaerobically with perchlorate or chlorate [(per)chlorate] as a suitable electron acceptor. Furthermore, with nitrate as the electron acceptor, strain RCB could also utilize toluene, ethylbenzene, and all three isomers of xylene (ortho-, meta-, and para-) as electron donors. While toluene and ethylbenzene were completely mineralized to CO2, strain RCB did not completely mineralize para-xylene but rather transformed it to some as-yet-unidentified metabolite. Interestingly, with nitrate as the electron acceptor, strain RCB degraded benzene and toluene concurrently when the hydrocarbons were added as a mixture and almost 92 microM total hydrocarbons were oxidized within 15 days. The results of these studies emphasize the unique metabolic versatility of this organism, highlighting its potential applicability to bioremediative technologies.     

Anaerobic degradation of benzene, toluene, ethylbenzene, and o-xylene in sediment-free iron-reducing enrichment cultures

Monoaromatic hydrocarbons such as benzene, toluene, ethylbenzene, and xylene (BTEX) are widespread contaminants in groundwater. We examined the anaerobic degradation of BTEX compounds with amorphous ferric oxide as electron acceptor. Successful enrichment cultures were obtained for all BTEX substrates both in the presence and absence of AQDS (9,10-anthraquinone-2,6-disulfonic acid). The electron balances showed a complete anaerobic oxidation of the aromatic compounds to CO2. This is the first report on the anaerobic degradation of o-xylene and ethylbenzene in sediment-free iron-reducing enrichment cultures.     

Substrate interactions during the biodegradation of benzene,toluene, ethylbenzene,and xylene (BTEX) hydrocarbons by the fungus Cladophialophora sp. strain T1

The soil fungus Cladophialophora sp. strain T1 (= ATCC MYA-2335) was capable of growth on a model water-soluble fraction of gasoline that contained all six BTEX components (benzene, toluene, ethylbenzene, and the xylene isomers). Benzene was not metabolized, but the alkylated benzenes (toluene, ethylbenzene, and xylenes) were degraded by a combination of assimilation and cometabolism. Toluene and ethylbenzene were used as sources of carbon and energy, whereas the xylenes were cometabolized to different extents. o-Xylene and m-xylene were converted to phthalates as end metabolites; p-xylene was not degraded in complex BTEX mixtures but, in combination with toluene, appeared to be mineralized. The metabolic profiles and the inhibitory nature of the substrate interactions indicated that toluene, ethylbenzene, and xylene were degraded at the side chain by the same monooxygenase enzyme. Our findings suggest that soil fungi could contribute significantly to bioremediation of BTEX pollution.     

Bioremediation of benzene,toluene, ethylbenzene,xylenes-contaminated soil: a biopile pilot experiment

Aims: In this study, we evaluated the removal efficiency of fuel hydrocarbons from a jet fuel contaminated area using bioaugmentation treatment in biopile. Methods and Results: The hydrocarbon analysis of the sample revealed total hydrocarbons mainly constituted by benzene, toluene, ethylbenzene, xylenes (BTEX) and heavy aliphatic hydrocarbons. Enrichments of soil sample were performed with BTEX, pristane and fuel JP-5, respectively, selected hydrocarbon-degrading strains, namely Acinetobacter sp., Pseudomonas sp. and Rhodococcus sp. Three hundred litres of culture containing 10⁸ cell ml⁻¹ of each strain and nutrients sprayed on the biopile allowed a removal of 90% of total hydrocarbons in 15 days. Bioremediation process was monitored by observation of the respiration rate and the bacterial abundance and GC-MS analysis. Conclusions: The efficiency of the treatment in the biopile was considerable. The assessment of microbial activity during the experiment is necessary for interventions targeted to improve environmental parameters such as humidity, temperature, pH and nutrients for optimization of the bioremediation process. Significance and Impact of the Study: A better knowledge of microbial successions at oil-polluted sites is essential for environmental bioremediation. Data obtained in biopile study improve our understanding of processes occurring during oil pollution.     

Analysis of benzene,toluene, ethylbenzene and m-xylene in biological samples from the general population

A method for the determination of benzene, toluene, ethylbenzene and xylene in blood and urine of people not occupationally exposed to solvents is described. The headspace technique combined with gas chromatography with a mass spectrometer detector is used. The sensitivity of recent mass spectrometers is good enough to furnish reliable results also in biological samples collected from the general population. No treatment for concentrating solvents present in the blood or urine is necessary. The main features of the method are easy preparation of biological samples, small volumes (7 ml), good repeatability and linearity in the range of interest. The limits of detection in blood were 16, 43, 22 and 52 ng/l for benzene, toluene, ethylbenzene and m-xylene respectively. Slightly greater sensitivity was found for urine samples. The results obtained in biological samples from 25 woodworkers not occupationally exposed to BTEX (15 non-smokers and 10 smokers) are comparable to those obtained by other investigators.     

Construction of a bacterial consortium for the biofiltration of benzene,toluene and xylene emissions

On equal parts of benzene, toluene and p-xylene (BTX), a stable bacterial consortium was enriched for removal of BTX vapours from air. As demonstrated by gas chromatographic monitoring, this consortium removed all three BTX components but was able to grow only on benzene and/or toluene. A Pseudomonas putida strain, PPO1, isolated from this consortium behaved in an identical manner. When immobilized on a porous peat/perlite column, both the consortium and the PPO1 isolated removed all three BTX components from metered air streams. However, due to the accumulation of products from the incompletely metabolized p-xylene, the removal rates were unsatisfactory and declined further with time. P. putida ATCC 33015 bearing the TOL plasmid was capable of growing on toluene, on para- and on meta- xylene isomers, but not on benzene. When the PPO1 and ATCC 33015 strains were immobilized, in equal parts, on peat/perlite columns a much improved and sustainable removal of all three BTX components was observed at the rate of 40–50 g/h. m3 filter bed. Due to the dominance of the ring-hydroxylating pathways over the TOL pathway, the classical enrichment approach did not result in a consortium capable of the sustained removal of all BTX components. However, a rationally formulated consortium consisting of members with complementary metabolic abilities was capable of this task and should be of use both in industrial emission control and in soil venting operations.     

The mutagenic effect of benzene, toluene and xylene studied by the SCE technique.

In experiments in vitro, neither benzene, toluene nor xylene changed the number of sister-chromatid exchanges (SCEs) or the number of chromosomal aberrations in human lymphocytes. Toluene and xylene caused a significant cell growth inhibition which was not observed with benzene in the same concentrations.     

Simultaneous high-performance liquid chromatographic determination of urinary metabolites of benzene, nitrobenzene, toluene, xylene and styrene

A high-performance liquid chromatographic method is described for the simultaneous determination of six urinary metabolites of several aromatic chemicals: phenol (from benzene), hippuric acid (from toluene), 3-methylhippuric acid (from xylene), mandelic and phenylglyoxylic acid (from styrene) and 4-nitrophenol (from nitrobenzene). Reversed-phase liquid chromatography was performed in an isocratic mode at 1 ml/min on a 5-μm C₁₈ column using two mobile phases: (A) acetonitrile—1% phosphoric acid (10:90); (B) acetonitrile—1% phosphoric acid (30:70). Phase A separates the six metabolites well, but phase B allows to a more rapid and reproducible simultaneous determination of phenolic compounds than phase A. For these compounds a prior enzymic hydrolysis step using Helix pomatia juice is performed to hydrolyse their sulphate and glucuronate conjugates. The reproducibility and the specificity are both excellent. Furthermore, the method is rapid, economical and easily automated. The proposed method appears very suitable for the routine monitoring of workers exposed to these chemicals on the basis of the biological threshold limit values.     

Simultaneous determination of benzene, toluene, ethylbenzene, and xylenes in urine by thermal desorption–gas chromatography

The determination of metabolites of benzene, toluene, ethylbenzene, and xylenes in urine has been used to assess human exposure to these compounds. The analyses of urine samples for these metabolites are tedious and time consuming. The determination of unmetabolized individual compounds in urine has been studied previously with some success. A simultaneous determination of several unmetabolized VOC compounds in urine by thermal desorption–gas chromatography was conducted to assess the exposure of smokers and nonsmokers to these compounds. The method of thermal desorption–GC was sensitive enough to detect a significant difference in exposure levels due to the contribution of light smoking in the environmentally-exposed group.     

Cytoenzymatic studies on neutrophils in workers having contact with organic solvents containing benzene, toluene and xylene

In 106 workers (47 women and 59 men) being in professional contact with organic solvents containing benzene and its homologues during 1 to 122 months the cytochemical examination of peripheral blood neutrophils has been performed. The patterns of neutrophil functional activation have been noted expressed in increased activities of acid phosphatase and beta-glucuronidase, increased NBT reduction and diminished glycogen reserves. Those changes were accompanied by diminished peroxidase and alkaline phosphatase activities. The stimulated NBT reduction, elevated in majority of workers, exhibited negative correlation with the exposure time what indicates the practical value of that test monitoring the biological effects of professional contact with the solvents.     

Kinetics of aerobic biodegradation of benzene and toluene in sandy aquifer material

Monod's equation adequately described aerobic biodegradation rates of benzene and toluene by the microbial population of a sandy aquifer when these compounds were initially present at concentrations lower than 100 mg/l each. Concentrations higher than 100 mg/l were inhibitory, and no benzene or toluene degradation was observed when these compounds were initially present at 250 mg/l each. The Monod coefficients were calculated as k = 8.3 g-benzene/g-cells/day and Ks = 12.2 mg/l for benzene, and k = 9.9 g-toluene/g-cells/day and Ks = 17.4 mg/l for toluene. Specific first-order coefficients would be 0.68 l/mg.day for benzene and 0.57 l.mg.day for toluene.     

Anaerobic degradation of toluene and xylene by aquifer microorganisms under sulfate-reducing conditions.

Toluene and the three isomers of xylene were completely mineralized to CO2 and biomass by aquifer-derived microorganisms under strictly anaerobic conditions. The source of the inoculum was gasoline-contaminated sediment from Seal Beach, Calif. Evidence confirming that sulfate was the terminal electron acceptor is presented. Benzene and ethylbenzene were not degraded under the experimental conditions used. Successive transfers of the mixed cultures that were enriched from aquifer sediments retained the ability to degrade toluene and xylenes. Greater than 90% of 14C-labeled toluene or 14C-labeled o-xylene was mineralized to 14CO2. The doubling time for the culture grown on toluene or m-xylene was about 20 days, and the cell yield was about 0.1 to 0.14 g of cells (dry weight) per g of substrate. The accumulation of sulfide in the cultures as a result of sulfate reduction appeared to inhibit degradation of aromatic hydrocarbons.     

Anaerobic degradation of toluene and xylene by aquifer microorganisms under sulfate-reducing conditions.

Toluene and the three isomers of xylene were completely mineralized to CO2 and biomass by aquifer-derived microorganisms under strictly anaerobic conditions. The source of the inoculum was gasoline-contaminated sediment from Seal Beach, Calif. Evidence confirming that sulfate was the terminal electron acceptor is presented. Benzene and ethylbenzene were not degraded under the experimental conditions used. Successive transfers of the mixed cultures that were enriched from aquifer sediments retained the ability to degrade toluene and xylenes. Greater than 90% of 14C-labeled toluene or 14C-labeled o-xylene was mineralized to 14CO2. The doubling time for the culture grown on toluene or m-xylene was about 20 days, and the cell yield was about 0.1 to 0.14 g of cells (dry weight) per g of substrate. The accumulation of sulfide in the cultures as a result of sulfate reduction appeared to inhibit degradation of aromatic hydrocarbons.     

Degradation of benzene, toluene, ethylbenzene, and xylenes (BTEX) by the lignin-degrading basidiomycete Phanerochaete chrysosporium.

Degradation of the BTEX (benzene, toluene, ethylbenzene, and o-, m-, and p-xylenes) group of organopollutants by the white-rot fungus Phanerochaete chrysosporium was studied. Our results show that the organism efficiently degrades all the BTEX components when these compounds are added either individually or as a composite mixture. Degradation was favored under nonligninolytic culture conditions in malt extract medium, in which extracellular lignin peroxidases (LIPs) and manganese-dependent peroxidases (MNPs) are not produced. The noninvolvement of LIPs and MNPs in BTEX degradation was also evident from in vitro studies using concentrated extracellular fluid containing LIPs and MNPs and from a comparison of the extents of BTEX degradation by the wild type and the per mutant, which lacks LIPs and MNPs. A substantially greater extent of degradation of all the BTEX compounds was observed in static than in shaken liquid cultures. Furthermore, the level of degradation was relatively higher at 25 than at 37 degrees C, but pH variations between 4.5 and 7.0 had little effect on the extent of degradation. Studies with uniformly ring-labeled [14C]benzene and [14C]toluene showed substantial mineralization of these compounds to 14CO2.     

Isolation and characterization of a thermotolerant bacterium Ralstonia sp. strain PHS1 that degrades benzene, toluene, ethylbenzene, and o-xylene

A thermotolerant bacterium, designated as PHS1, was isolated from a hot spring in Pohang, Korea, on the basis of its ability to grow on benzene, toluene, ethylbenzene, and xylenes (BTEX) as a sole carbon source. Strain PHS1 is a gram-negative, rod-shaped aerobe and grows optimally at 42 degrees C and pH 7.2. According to 16 S rDNA analysis, strain PHS1 showed highest similarity to Ralstonia eutropha (previously named Alcaligenes eutrophus). Unlike its closest known Ralstonia species, however, strain PHS1 was able to utilize toluene, ethylbenzene, o-xylene, and both m- and o-cresol. The degradation of o-xylene by strain PHS1 is particularly important, since o-xylene is a compound of considerable environmental interest, owing to its recalcitrance; and very few microorganisms have been reported to utilize o-xylene as a sole carbon source. It was found that strain PHS1 transformed o-xylene to 2,3-dimethylphenol through direct oxygenation of the aromatic ring. The unique properties of strain PHS1, such as thermotolerance and the ability to degrade o-xylene, may have important implications for the treatment of BTEX-contaminated industrial effluents.     

The influence of creosote compounds on the aerobic degradation of toluene

The inhibiting effect of 14 typical creosote compounds on the aerobic degradation of toluene was studied in batch experiments. Four NSO-compounds (pyrrole, 1-methylpyrrole, thiophene, and benzofuran) strongly inhibited the degradation of toluene. When the NSO-compounds were present together with toluene, little or no degradation of toluene was observed during 16 days of incubation, compared with a total removal of toluene within 4 days when the four compounds were absent. Indole (an N-compound) and three phenolic compounds (phenol, o-cresol, and 2,4-dimethylphenol) also inhibited the degradation of toluene, though the effect was much weaker that of the four NSO-compounds. O-xylene, p-xylene, naphthalene and 1-methylnaphthalene seemed to stimulate the degradation even though the influence was very weak. No effects of benzothiophene (an S-compound) and quinoline (an N-compound) were observed. Benzofuran (an O-compound) was identified as the compound that most inhibited the degradation of toluene. An effect could be detected even at low concentrations (40 g/l).Abbreviations bf benzofuran - bt benzothiophene - dmp 2,4-dimethylphenol - GC gas chromatograph - ind indole - mnap 1-methylnaphthalene - MAH monoaromatic hydrocarbons - mpyr 1-methylpyrrole - nap naphthalene - o-cre o-cresol - o-xyl o-xylene - phe phenol - pyr pyrrole - p-xyl p-xylene - tol toluene - thi thiophene - qui quinoline     

The influence of creosote compounds on the aerobic degradation of toluene

The inhibiting effect of 14 typical creosote compounds on the aerobic degradation of toluene was studied in batch experiments. Four NSO-compounds (pyrrole, 1-methylpyrrole, thiophene, and benzofuran) strongly inhibited the degradation of toluene. When the NSO-compounds were present together with toluene, little or no degradation of toluene was observed during 16 days of incubation, compared with a total removal of toluene within 4 days when the four compounds were absent. Indole (an N-compound) and three phenolic compounds (phenol, o-cresol, and 2,4-dimethylphenol) also inhibited the degradation of toluene, though the effect was much weaker that of the four NSO-compounds. O-xylene, p-xylene, naphthalene and 1-methylnaphthalene seemed to stimulate the degradation even though the influence was very weak. No effects of benzothiophene (an S-compound) and quinoline (an N-compound) were observed. Benzofuran (an O-compound) was identified as the compound that most inhibited the degradation of toluene. An effect could be detected even at low concentrations (40 μg/l).