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Fast remediation of coal-tar-related compounds in biofilm bioreactors.

The biological degradation of complex mixtures of recalcitrant substances is still a major challenge in environmental biotechnology and the remediation of coal-tar constitutes one such problem area. Biofilm bioreactors offer many advantages and may be successfully used for this purpose. Two stirred-tank reactors and one packed-bed reactor were tested in a continuous mode. Continuous cultivation allows microbial selection to take place whilst adhesive growth provides a high degradation capacity and process stability. The reactors were inoculated with mixed microbial populations to favour complete metabolism and to prevent metabolite accumulation and substrate inhibition effects. Phenol, o-cresol, quinoline, dibenzofuran, acenaphthene and phenanthrene were used as model contaminants and constituted the sole energy and carbon sources. The hydraulic retention time (HRT) was initially set to 2.5 days for a period of several months to allow the establishment of a stable biofilm and was then gradually decreased. All the compounds were found to be degraded by more than 90% at HRT of 3 h or more. Neither substrate inhibition nor metabolite accumulation effects were observed. The stirred-tank configuration was found to be the most efficient for use with high loads. No improvement in the degradation capacity could be achieved by increasing the biofilm surface in these reactors, illustrating that the limiting factor may be the mass transfer limitations rather than the availability of the biofilm surface. Finally, anaerobic treatment was successfully achieved, confirming the potential for remediation of contaminated sites under anaerobic conditions, providing that alternative electron acceptors are present.

Acenaphthenes↗

The effects of chronic tobacco smoke exposure from high-tar cigarettes on the phagocytic and killing capacity of polymorphonuclear cells of mice.

Mice were exposed to high-tar (HT) (16 mg tar/cigarette) filtered cigarettes for up to 32 weeks. Intraperitoneal caseinate induction of polymorphonuclear (PMN) cells in both tobacco smoke (TS)-exposed and age-matched control mice provided the source of cells which enabled an assessment to be made in vitro of both their phagocytic and degradative properties. Radioactive Staphylococcus aureus were opsonized with either immune serum or heat-inactivated (complement deficient) immune serum for the tests. Two strains of mice were used in all trials. The PMN cells from BALB/c strain mice were less affected by TS exposure than those of C57 Black mice in both phagocytic and degradative capacities when the S. aureus were opsonized with immune serum. However, heat inactivation of complement within the serum reduced the differences between TS-exposed animals and age-matched controls in both strains of mice. The treatment effects were seen at all times tested from 3 days to 32 weeks of TS exposure. The results suggest that fc-receptor site activity of PMN cells is not significantly affected by TS exposure but that complement interactions within the phagocytic process are significantly suppressed. Whether this is due directly to complement receptors or metabolic associations of phagocytosis of the complement/opsonin/antigen complex is unclear.

Animals↗

Pyridine and other coal tar constituents as inhibitors of potato polyphenol oxidase: a non-animal model for neurochemical studies.

Potato polyphenol oxidase activity was strongly and noncompetitively inhibited by the "Perov mixture" of coal tar components and by pyridine alone, while phenol competitively inhibited the enzyme. These two inhibitors are structural components of the parkinsonogenic neurotoxin N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). By extension, dopamine and neuromelanin synthesis in the brain may be influenced by the inhibitory effects of such compounds upon the copper-dependent steps of tyrosine metabolism. The non-animal model used in this study may represent an alternative to the use of animal tissues in neurodegenerative disease research.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗

Cytogenetic activities of tobacco particulate matter (TPM) derived from a low to middle tar British cigarette.

Tobacco particulate matter (TPM) derived from an experimental low to middle tar cigarette was tested for its cytogenetic activity upon a low passage number Chinese hamster pulmonary cell line. Examination of the mitotic profiles (after one cell cycle) revealed no interference by the agent with mitotic spindle formation and/or function. However, complete chromosomes (or parts of them) were seen to dislocate from the mitotic spindles. Such an event was probably the result of the chromosome aberrations, substantial numbers of which were observed in second division cells, or through a process of centromere inactivation. In second division cells there was a reduction in the number of diploid cells accompanied by an increase in both hypodiploidy and polyploidy and there was also a non-dose-related increase in endoreduplication. The results demonstrate that TPM was capable of inducing both structural and numerical chromosome aberrations in cultured mammalian cells.

Animals↗

Mutagenicity of coal tar paints used in drinking water distribution systems.

The aim of this study was to evaluate the mutagenicity of coal tar paints used for coating drinking water tanks and pipes, as a preliminary screening for potential genotoxic hazards associated with leaching of mutagens into drinking water during water storage and distribution. To this end, the Salmonella/microsome assay was performed on different fractions of two paints. The fractions were obtained using different fractionation procedures (a sequential solvent extraction and an acid-base fractionation) for removing the presence of inhibitory components. Both fractionation procedures confirmed an extraordinarily high mutagenicity in both paints, with metabolic activation, much higher than the mutagenicity of the unfractionated paints. The acid-base fractionation was more time-consuming but gave higher mutagenicity recoveries and provided information as to the general nature of the genotoxic constituents, which were concentrated in the neutral fractions. On the other hand, the sequential solvent extraction by sonication was a shorter and simpler method and permitted to reveal the presence of direct-acting mutagens. It is concluded that the application of the Salmonella/microsome assay coupled with both fractionation methods may give complementary and confirmatory data on the genotoxic properties of these coal based paints, as a screening of the potential mutagenic/carcinogenic hazards derived from these materials used in drinking water distribution systems.

Coal Tar↗

Genetic monitoring of aluminum workers exposed to coal tar pitch volatiles.

A group of 50 workers exposed to coal tar pitch volatiles (CTPV) in an aluminum reduction plant and a group of 50 non-exposed workers were selected to evaluate the genotoxic effects of CTPV exposure. A battery of tests was performed on 3 different body fluids; urine, blood and semen. Urine samples were evaluated for mutagenic constituents using the Ames/Salmonella assay. Cultured lymphocytes from blood samples were used to perform cytogenetic analysis. Semen samples were used to measure sperm count, percent abnormal sperm morphology and frequency of sperm carrying double fluorescent bodies (2-F). 14 of 28 (50%) exposed workers and 7 of 36 (19.4%) non-exposed workers had mutagenic urine. This difference was significant (p less than 0.01). Among the non-smokers a significantly higher percentage of workers who were exposed had positive urine (36%) compared to the non-exposed workers (5%) (p less than 0.05). Among the exposed group, more mechanics had mutagenic urine than did other types of workers. Overall chromosome aberration rates were similar in both exposed and non-exposed workers. Among exposed workers a significant inverse correlation (p less than 0.05) between age and chromatid aberration rate was observed. Results of semen analysis failed to detect differences between exposed and non-exposed workers. Results of these tests lend support to a battery approach to genetic monitoring and suggest a link between exposure to CTPV and genotoxic effects. Detection of exposure to mutagens at an early time offers an opportunity for disease prevention by the reduction of exposure.

Adult↗

Effect of ascorbic acid prophylaxis on the frequency of chromosome aberrations in the peripheral lymphocytes of coal-tar workers.

The possible anti-mutagenic potential of prophylactically administered ascorbic acid (AA) preparations was studied in a group of 35 coal-tar workers occupationally exposed to PAH and benzene. The effectiveness of AA prophylaxis was assessed by differences in the frequency of chromosome aberrations in peripheral blood lymphocytes of the exposed workers examined before and after a 3-month treatment with AA at the daily doses of 1.0 g for 5 days a week. In the exposed group the cytogenetic analysis of peripheral blood lymphocytes revealed a significant drop in the frequency of aberrant cells (AB.C.) from the initial 5.07% AB.C. (0.0657 B/C, breaks per cell) to 1.77% AB.C. (0.0197 B/C). In a group of matching controls the frequency of AB.C. was 1.50% (0.0170 B/C) and 1.45% (0.0180 B/C), respectively. The study showed that the risk of genetic injury assessed by the frequency of chromosome aberrations in peripheral blood lymphocytes was substantially reduced after AA prophylaxis.

Adult↗

Prostaglandin precursors in plasma phospholipids of patients with psoriasis: effects of treatment with coal tar.

Plasma phospholipids of patients with psoriasis have significantly reduced levels of dihomogammalinolenic acid (20:3n-6), arachidonic acid (20:4n-6) and adrenic acid (22:4n-6), the precursors of the 1, 2 and homo-2 series of prostaglandins (PGs). Concentrations of the 3 series PG precursor, eicosapentaenoic acid (20:5n-3) were normal. Hospital treatment with a coal tar regime produced a rise in 20:3n-6 to levels which were significantly above normal.

8,11,14-Eicosatrienoic Acid↗

In vivo induction of aryl hydrocarbon hydroxylase in human scalp hair follicles by topical application of a commercial coal tar preparation.

Five low-dose applications of a commercial coal tar preparation on a small scalp skin region resulted in an induction of aryl hydrocarbon hydroxylase (AHH) activity in freshly isolated human hair follicles. Large but reproducible interindividual differences in AHH-inducibility could be detected. The method offers the opportunity to measure AHH-inducibility, which has been correlated to the risk of developing chemical-induced cancer, in vivo in normal epithelium, a cell-type highly relevant for chemical carcinogenesis. Smoking habits did not have any effect on AHH-activity in freshly isolated hair follicles. Therefore the method potentially permits the identification of persons with high and low genetically determined AHH-inducibility.

Administration, Topical↗

Skin tumor initiating activity of therapeutic crude coal tar as compared to other polycyclic aromatic hydrocarbons in SENCAR mice.

Crude coal tar (CCT), a complex mixture rich in polycyclic aromatic hydrocarbons (PAHs) including carcinogens such as benzo[a]pyrene (BP), is widely used therapeutically in dermatological practice, particularly in combination with ultraviolet radiation in the treatment of chronic dermatoses such as psoriasis. In this study we analyzed the tumor initiating activity of therapeutic CCT preparation (USP) in a two-stage model system (initiation and promotion) in SENCAR mice. The tumorigenicity of CCT was compared with other conventionally studied carcinogenic PAHs; 7,12-dimethylbenz[a]anthracene (DMBA), 3-methylcholanthrene (MCA), benzo[a]pyrene 7,8-diol (BP-7,8-diol) and benzo[a]pyrene (BP). A single topical application of an initiating dose of CCT (20 microliter), DMBA (39 nmol), MCA (746 nmol), BP-7,8-diol (352 nmol) or BP (396 nmol) was followed by twice weekly application of 12-O-tetradecanoylphorbol-13-acetate (TPA) (3.24 nmol). The first tumor appearance in the CCT treated group was at 6 weeks on test as compared with DMBA and MCA (3 weeks) and BP-7,8-diol and BP (4-5 weeks). In a total of 20 animals after 8 weeks on test the DMBA, MCA, BP-7,8-diol, BP and CCT groups of mice showed 457, 176, 106, 76 and 46 tumors, respectively. After 11 weeks 100% of the mice in each experimental group had developed tumors and the number of tumors/mouse was 24.3, 15.0 9.8, 6.6 and 3.3 in the DMBA, MCA, BP-7,8-diol, BP and CCT groups, respectively. These studies provide first evidence that a single topical application of a therapeutic preparation of CCT possesses skin tumor initiating activity.

Administration, Topical↗

Benzo(a)pyrene diol epoxide-I-DNA adduct formation in the epidermis and lung of SENCAR mice following topical application of crude coal tar.

The levels of benzo[a]pyrene diol epoxide-I-deoxyguanosine (BPDE-I-dG) adduct formation in epidermis and lung of SENCAR mice following the topical application of benzo[a]pyrene (BP) alone, crude coal tar (CCT) alone, and the two combined were determined in an enzyme linked immunosorbent (ELISA) assay using monoclonal antibodies. Topical application of two doses of BP (20 micrograms) at 72-h intervals, with sacrifice 24 h later resulted in the formation of 197 fmol and 205 fmol BPDE-I-dG adducts per mg DNA in epidermis and lung, respectively. Topical application of 0.5 ml CCT alone resulted in the formation of 278 fmol and 410 fmol BPDE-I-dG adducts per mg DNA in epidermis and lung, respectively. Simultaneous topical application of 20 micrograms BP and CCT (0.1-0.5 ml) resulted in substantially lower BPDE-I-dG adducts in the epidermis as well as in the lung. Our results suggest that CCT may contain inhibitors of carcinogen-DNA adduct formation and that topical application of CCT produces greater effects on DNA-adduct formation in lung than in epidermis. Thus the cancer-causing potency of the polycyclic aromatic hydrocarbons (PAHs) in CCT may be reduced by other anticarcinogenic constituents present in CCT and systemic absorption of carcinogenic PAHs in CCT applied to skin might have tumorigenic effects in other tissues.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

Detection of free radicals in aqueous extracts of cigarette tar by electron spin resonance.

Aqueous extracts of cigarette tar (ACT) autooxidize to produce semiquinone, hydroxyl, and superoxide radicals in air-saturated buffered aqueous solutions. The semiquinone species were detected by direct electron spin resonance (ESR) measurements and identified as o- and p-benzosemiquinone radicals by comparison with the ESR signals of catechol and hydroquinone radicals under similar conditions. The rate of formation of these radicals was dependent on pH. Hydroxyl and superoxide radicals were detected as 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) spin adducts by ESR spin trapping. Superoxide dismutase (SOD) (20 units/ml) inhibited the formation of the superoxide spin adduct of DMPO completely. Addition of Fe2+ to this system increased the ESR signal intensity of hydroxyl radical spin adduct of DMPO three to five times. These results indicate that superoxide and hydroxyl radicals are produced during the autooxidation of hydroquinone- and catechol-related species in ACT.

Cyclic N-Oxides↗

Early specific free radical-related cytotoxicity of gas phase cigarette smoke and its paradoxical temporary inhibition by tar: An electron paramagnetic resonance study with the spin trap DEPMPO.

Electron paramagnetic resonance (EPR) spin trapping studies demonstrated aqueous tar particulate matter (TPM) and gas phase cigarette smoke (GPCS) to behave as different sources of free radicals in cigarette smoke (CS) but their cytotoxic implications have been only assessed in CS due to its relevance to the natural smoking process. Using a sensitive spin trapping detection with 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO), this study compared the respective roles of CS- and GPCS-derived free radicals on smoke-induced cytotoxicity and lipid peroxidation of filtered and unfiltered, machine-smoked experimental and reference cigarettes yielding a wide range of TPM yields. In buffer bubbled with CS the DEPMPO/superoxide spin adduct was the major detected nitroxide. Use of appropriate control experiments with nitric oxide radical (NO*) or carbonyl sulfide, and a computer analysis of spin adduct diastereoisomery showed that the hydroxyl radical (HO*) adduct of DEPMPO seen in GPCS-bubbled was rather related to metal-catalyzed nucleophilic synthesis than to direct HO* trapping. Unexpectedly a protective effect of TPM on murine 3T3 fibroblasts was observed in early (<3h) free radical-, GPCS-induced cell death, and carbon filtering decreased free radical formation, toxicity and lipid peroxidation in three cell lines (including human epithelial lung cells) challenged with GPCS. These results highlight an acute, free radical-dependent, harmful mechanism specific to the GPCS phase, possibly involving NO* chemistry, whose physical or chemical control may be of great interest with the aim of reducing the toxicity of smoke.

Animals↗

Investigation of organic matter dynamics during in-vessel composting of an aged coal-tar contaminated soil using fluorescence excitation-emission spectroscopy.

In-vessel composting of an aged coal-tar contaminated soil from a manufactured gas plant site was investigated over 98days using laboratory-scale in-vessel composting reactors. The composting reactors were operated at 18 different operational conditions using a logistic three-factor factorial design with three temperatures (T=38, 55 and 70 degrees C), four soil to green waste ratios (S:GW; 0.6:1, 0.7:1, 0.8:1 and 0.9:1 on a dry weight basis) and three moisture contents (MC; 40%, 60% and 80%). Excitation-emission matrix (EEM) fluorescence spectroscopy was used to investigate organic matter dynamics in the composting mixture. The results of this investigation indicated that formation of humic substances can be monitored by fluorescence excitation-emission matrix, and provided evidence of progressive mineralization or humification of the composting mixture. Peak excitation wavelength shifts and peak fluorescence intensity can both be used as indicators to monitor the humification or maturation of compost. Finally, the fluorescence index can be applied to investigate the origin of humic substances and fulvic acids, and the humification or maturation of compost.

Benzopyrans↗

Monitoring coal-tar pitch composition changes during air-blowing by gas chromatography.

A series of air-blown coal-tar pitches was studied by GC and GC-MS in order to achieve a deeper understanding of the behaviour of the different pitch components during air-blowing. Compounds present in the parent pitch were identified and quantified and then compared to those present in the air-blown pitches. The compounds observed were identical before and after the treatments, but the concentration of each compound changed with the treatment to a different extent depending on its molecular structure and consequently its reactivity to oxygen. The most reactive compounds were those with a mehylene-bridge in a five member ring, followed by those with a methyl group in their structure.

Air↗

In-vessel composting--bioremediation of aged coal tar soil: effect of temperature and soil/green waste amendment ratio.

The biodegradation of 16 United States Environmental Protection Agency (USEPA)-listed polycyclic aromatic hydrocarbons (PAHs) present in contaminated soil from a manufactured gas plant site was investigated using laboratory-scale in-vessel composting-bioremediation reactors over 8 weeks. The influence of temperature (T, 38, 55, and 70 degrees C) and soil/green waste ratio (S:GW, 0.6:1, 0.7:1, 0.8:1, and 0.9:1) was investigated. A comparative study using a temperature profile during in-vessel composting-bioremediation to meet current regulatory requirements was also investigated. Temperature and amendment ratio were found to be important operating parameters for PAH removal for in-vessel composting-bioremediation of aged coal tar-contaminated soil. After 8 weeks of continuous treatment, the highest removal of 16 USEPA PAHs was observed at T=38 degrees C and S:GW=0.8:1 (75.2%). Lower removal of 16 USEPA PAHs was observed for temperature profile treatment (60.8%). We recommend that when conventional composting processes using temperature profiles to meet regulatory requirements for pathogen control need to be used, these should start with a prolonged mesophilic stage (6 weeks in this investigation) followed by thermophilic, cooling, and maturation stages.

Biodegradation, Environmental↗

The influence of different temperature programmes on the bioremediation of polycyclic aromatic hydrocarbons (PAHs) in a coal-tar contaminated soil by in-vessel composting.

The biodegradation of 16 US. EPA-listed polycyclic aromatic hydrocarbons (sigma PAHs), with accompanying humification and microbial community structure changes during simulated in-vessel composting-bioremediation of an aged coal-tar-contaminated soil amended with green waste were studied over 56 days. The experimental design compared one constant temperature profile (TC=38 degrees C) with three variable temperature profiles (TP1, TP2 and TP3), including treatment at 70 degrees C to comply with regulatory requirements. Greatest sigma PAHs removal (75.4+/-0.1%; k(1)=0.026 day(-1), R(2)=0.98) occurred at TC=38 degrees C compared to all variable temperature profiles TP1 (62.1+/-11.0%; k(1)=0.016 day(-1), R(2)=0.93), TP2 (71.8+/-8.2%; k(1)=0.021 day(-1), R(2)=0.95) and TP3 (45.3+/-9.7%; k(1)=0.010 day(-1), R(2)=0.91). This study proved that using thermophilic temperatures (70 degrees C) towards the end of in-vessel composting processes (TP2) resulted in greater sigma PAHs removal than using other variable temperature profiles (TP1, TP3), as long as the increase was stepwise via an intermediate temperature (55 degrees C). Phospholipid fatty acid (PLFA) signatures indicated that use of thermophilic temperatures towards the end of the in-vessel composting-bioremediation (TP2) resulted in a higher fungal to bacterial PLFA ratio and a lower Gram-positive to Gram-negative (G(+)/G(-)) bacterial ratio. Fluorescence excitation-emission matrix (EEM) showed the presence of peaks typical of humic-like (Ex/Em wavelength pair approximately 340/460 nm) and fulvic-acid-like (Ex/Em wavelength pair approximately 245/460 nm) substances, indicating mineralization and/or maturation of the compost. Varying the temperature during in-vessel composting to comply with regulatory requirements for pathogen control, promoted contaminant biodegradation, microbial activity and compost maturation.

Bacteria↗

Cigarette tar phenols impede T cell cycle progression by inhibiting cyclin-dependent kinases.

Cigarette smoking causes profound suppression of pulmonary T cell responses, which is associated with increased susceptibility to respiratory tract infections and decreased tumor surveillance. We previously demonstrated that the phenolic compounds in cigarette tar inhibit blastogenesis and interfere with human T cell cycle progression. To identify the mechanism by which cell cycle arrest occurs, we examined the effects of these compounds on cyclin-dependent kinases (Cdk) that control the G0/G1 transition. We found that hydroquinone inhibited induction of Cdk4 and Cdk6 kinase activities by >80%, while catechol and phenol were markedly less potent. HQ did not affect mitogenic induction of the Cdk6 protein, but inhibited expression of cyclin D3 by >90% resulting in a dramatic reduction in proper Cdk6/Cyclin D3 complex formation.

Cell Cycle↗