Dioxin-like activity of brominated dioxins as individual compounds or mixtures in in vitro reporter gene assays with rat and mouse hepatoma cell lines.

In vitro reporter gene assays detecting dioxin-like compounds have been developed and validated since the middle 1990's, and applied to the determination of dioxin-like activities in various samples for their risk management. Data on characterizing the potency of individual brominated dioxins and their activity in mixture with chlorinated dioxins are still limited on the cell-based assay. This study characterized the dioxin-like activities of the 32 brominated dioxins, such as polybrominated dibenzo-p-dioxins, polybrominated dibenzofurans (PBDFs), coplanar polybrominated biphenyls, mixed halogenated dibenzo-p-dioxins and dibenzofurans (PXDFs), as a sole component or in a mixture by DR-CALUX (dioxin-responsive chemically activated luciferase expression) using the rat hepatoma H4IIE cell line and XDS-CALUX (xenobiotic detection systems-chemically activated luciferase expression) assays using the mouse hepatoma H1L6.1 cell line. The 2,3,7,8-TCDD-relative potencies (REPs) of most of the brominated dioxins were within a factor of 10 of the WHO toxicity equivalency factor (WHO-TEF) for the chlorinated analogues. The REPs of a few PXDFs were an order of magnitude higher than the corresponding WHO-TEFs, indicating their toxicological importance. Results with reconstituted mixtures suggest that the activity of brominated and chlorinated dioxins in both CALUX assays was dose-additive. Thus, obtained results indicated the applicability of the CALUX assays as screening tools of brominated dioxins together with their chlorinated analogues.

Test chamber and forensic microscopy investigation of the transfer of brominated flame retardants into indoor dust via abrasion of source materials.

Brominated flame retardants (BFRs) have been detected in indoor dust in many studies, at concentrations spanning several orders of magnitude. Limited information is available on the pathways via which BFRs migrate from treated products into dust, yet the different mechanisms hypothesized to date may provide an explanation for the range of reported concentrations. In particular, transfer of BFRs to dust via abrasion of particles or fibers from treated products may explain elevated concentrations (up to 210 mg g(-1)) of low volatility BFRs like decabromodiphenyl ether (BDE-209). In this study, an indoor dust sample containing a low concentration of hexabromocyclododecane, or HBCD, (110 ng g(-1) ΣHBCDs) was placed on the floor of an in-house test chamber. A fabric curtain treated with HBCDs was placed on a mesh shelf 3 cm above the chamber floor and abrasion induced using a stirrer bar. This induced abrasion generated fibers of the curtain, which contaminated the dust, and ΣHBCD concentrations in the dust increased to between 4020 and 52 500 ng g(-1) for four different abrasion experiment times. The highly contaminated dust (ΣHBCD at 52 500 ng g(-1)) together with three archived dust samples from various UK microenvironments, were investigated with forensic microscopy techniques. These techniques included Micro X-ray fluorescent spectroscopy, scanning emission microscopy coupled with an energy dispersive X-ray spectrometer, Fourier transform infrared spectroscopy with further BFR analysis on LC-MS/MS. Using these techniques, fibers or particles abraded from a product treated with BFRs were identified in all dust samples, thereby accounting for the elevated concentrations detected in the original dust (3500 to 88 800 ng g(-1) ΣHBCD and 24 000 to 1,438 000 ng g(-1) for BDE-209). This study shows how test chamber experiments alongside forensic microscopy techniques, can provide valuable insights into the pathways via which BFRs contaminate indoor dust.

Sorption and desorption of 17ß-estradiol to natural sediment.

The sorption and desorption of 17ß-estradiol (E2) to various natural sediment were investigated. First, the quantitative solvent-water partition indices were measured. Significant differences were found between the n-octanol-water partition coefficient (K(OW)) and the n-hexane-water partition coefficient (K(HW)) of E2. The value of K(HW) (Log K(HW) = 0.07) is lower than those of two to four ring polyaromatic hydrocarbons (PAHs), while the value of K(OW) (Log K(OW) = 3.99) and that of organic matter-water partition coefficient (K(OC)) onto humic acid (Log K(OC) = 4.30) were similar to those of the PAHs. Five natural sediments of various characteristics and origins were selected for sorption and desorption experiments. Linear isotherms were obtained for sorption and desorption. The equilibrium partitioning coefficients of E2 were well-correlated with their values of weight fraction of organic carbon in sediments (f(OC)). Results suggest that E2 is sorbed mainly onto the organic portion of sediments and that its sorption coefficient can be estimated from K(OW) and f(OC), as in the case of non-polar PAHs. However, because of its polarity, the sorption mechanism of E2 onto sediments cannot be explained solely by the hydrophobic interaction.

Water-quality monitoring technique for dioxins during dredging using on-site solid phase extraction with graphitic carbon and analysis with DR-CALUX.

In Japan, water-quality monitoring with the environmental dioxin standard of 1 pg-TEQ/L at basic monitoring points set in a border between operation-area water and normal water is necessary for removal operations for bottom sediments containing dioxins. This paper presents a technique for combining on-site graphitic carbon (GrC) solid-phase extraction and bioassay method for the simplification and facilitation of water-quality monitoring. Using this method, we conducted experiments at an actual sediment removal site. Comparative analyses using high-resolution GC/MS (HRGC-HRMS) between the dioxin concentrations detected using a GrC solid-phase extraction kit and the official analytical method yielded similar concentrations and distributions of isomers for detected PCDD/Fs and DL-PCBs. The GrC solid-phase extraction method is inferred to have capacity equal to official analytical method extraction. Toxicity equivalent concentrations estimated using DR-CALUX correlate with equivalent concentrations with HRGC/HRMS. No false positive or false negative result was obtained for the standard value 1 pg-TEQ/L.

Bioassay monitoring of waste PCB samples during chemical destruction treatments.

Capacitor oil samples (PCBs > 90%wt) were treated in a bench scale experiment to investigate the destruction of PCBs during chemical destruction processes (a catalytic hydrodechlorination treatment with palladium carbon and additional treatment with potassium tert-butyloxide). Using those results, this study confirmed the decrease of PCBs and other undesirable dioxin-like compounds such as PCDD/Fs in treated samples during the treatment. Dioxin-responsive chemical-activated luciferase expression (DR CALUX) AhR reporter gene bioassay was used to evaluate dioxin-like activity in the samples. During the treatment, the efficiency for PCB capacitor oil was around 99.99% or more in WHO-TEQ and CALUX-TEQ, whereas the sum of PCBs was reduced at a resulting efficiency of >99.9999%. In this study, a new cleanup procedure for separating PCBs from the mineral oil matrix was also developed for DR CALUX. The procedure consists of dimethylsulphoxide partitioning followed by silica gel-44% sulphuric acid reflux treatment and activated carbon chromatography. With the cleanup, CALUX-TEQ values were in good agreement with WHO-TEQ values and were as much as 3.3 times higher than WHO-TEQs for untreated/treated PCB-containing insulating oil samples. The DR CALUX results of mineral oil samples containing various PCB concentrations of 0.5-50 mg/kg (corresponding WHO-TEQs: 0.012-1.2 microg-TEQ/g) also correlated well with WHO-TEQs (CALUX-TEQ/WHO-TEQ ratio = 1.0-3.0), which was consistent with the theoretical quantification limit of the CALUX. These results supported the validity of the proposed clean-up method.

The use of enzyme-linked immunosorbent assays (ELISA) for the determination of pollutants in environmental and industrial wastes.

Twelve enzyme-linked immunosorbent assays (ELISA), for the determination of surfactants [linear alkylbenzene sulfonates (LAS), alkyl ethoxylates (AE), and alkylphenol ethoxylates (APE)], endocrine disruptors [alkylphenol (AP), AP + APE, and bisphenol A (BPA)], estrogens [17beta-estradiol (E2), estrone (El), estrogen (ES: El + E2 + estriol (E3)), 1 7alfa-ethynylestradiol (EE2)], dioxins and polychlorinated biphenyls (PCBs), were validated on environmental water and industrial wastes. The lowest quantification limits of these ELISAs were 0.05 microg/L (BPA, E2, El, ES and EE2), 2 microg/L (AE), 3 microg/L (dioxins and PCBs), 5 microg/L (AP, AP + APE) and 20 microg/L (LAS and APE). To apply these ELISAs to environmental or industrial waste samples, simple and appropriate pre-treatment methods were also developed for each ELISA. With optimized pre-treatments, the values of ELISAs were well co-related, in all cases, to those of instrumental analytical methods such as liquid chromatography (HPLC), liquid chromatography-tandem mass spectrometry (LC-MS/MS), and high-resolution gas chromatography mass spectrometry (HR-GC-MS), etc.

Bio/chemical analysis of dioxin-like compounds in sediment samples from Osaka Bay, Japan.

The combinatorial bio/chemical investigation of sediments (six surface samples and one core sample) from Osaka Bay, Japan was conducted to clarify the horizontal and vertical distribution profiles of persistent organic pollutants in the sediments. Concentrations of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), polychlorinated biphenyls (PCBs), polybrominated dibenzo-p-dioxins and dibenzofurans (PBDD/Fs), and polybrominated diphenylethers (PBDEs) were determined by chemical analysis and compared to bioassay results using H4IIE-luc/Dioxin Responsive-Chemical Activated LUciferase eXpression (DR-CALUX). For surface sediments, World Health Organization-toxicity equivalent (WHO-TEQ) values ranged from 1.8 to 92 pg g(-1) dry weight and the bioassay-TEQ (CALUX-TEQ) values (3.7-140 pg g(-1) dry weight) yielded significant correlation with them (r2 = 0.96). On the other hand, correlation between both TEQs (for WHO-TEQ, 5.5-47 and for CALUX-TEQ, 27-76 pg g(-1) dry weight) for core samples was not so good (r2 = 0.46). Comparing the vertical profiles of CALUX-TEQ and WHO-TEQ, they were different in that WHO-TEQ reached the maximum in the 1957 core section, while CALUX-TEQ reached in the 1984 core section. CALUX-TEQ values were 1-5-fold more than WHO-TEQ values in all the surface and core samples. CALUX-TEQ values were calculated for PBDE and PBDD/F concentrations, employing their CALUX toxicity equivalent factors (CALUX-TEFs). The estimated CALUX-TEQ values obtained for the brominated compounds could explain for 11% on average (range 4.7-31%) of the experimentally obtained CALUX-TEQ values in the investigated surface sediments.

The Bacillus subtilis rec-assay: a powerful tool for the detection of genotoxic substances in the water environment. Prospect for assessing potential impact of pollutants from stabilized wastes.

The Bacillus subtilis rec-assay has been specially developed to detect genotoxicity in environmental water samples. The rationale of the B. subtilis rec-assay is based on the relative difference of survival of a DNA repair-recombination proficient strain and its deficient strain, which is interpreted as genotoxicity. This assay method can be very powerful in that it has higher sensitivity for the detection of mutagens in highly polluted waters than other bacterial mutation assays. Hydrophobic fractions from various environmental waters were fractionated by using XAD-2 resins and assayed, targeting the detection of organic genotoxic substances. Genotoxic response was detected in most of them, which revealed that many unknown micropollutants with genotoxicity occur in public water bodies. Positive response was also detected from a treated municipal solid waste (MSW) landfill leachate. Genotoxicity remaining in the treated effluent suggests that genotoxic micropollutants may pass through conventional water treatment processes such as activated sludge treatment process. Without proper control of waste quality and landfill facilities, waste landfill could be a heavy pollution source.

Indirubin and indigo are potent aryl hydrocarbon receptor ligands present in human urine.

Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates genes involved in xenobiotic metabolism, cellular proliferation, and differentiation. Numerous xenobiotic and biological compounds are known to interact with AhR, but it remains an orphan receptor, because its physiological ligand is unknown. We identified AhR ligands in human urine using a yeast AhR signaling assay and then characterized their properties. Two ligands, indirubin and indigo, were both present at average concentrations of approximately 0.2 nm in the urine of normal donors. Indirubin was also detected in fetal bovine serum and contributed half of the total AhR ligand activity. The activities of indirubin and indigo were comparable with or more potent than that of the archetypal ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin, in yeast AhR activation assays. We suggest that the endogenous levels and potencies of indirubin and indigo are such that they activate AhR-mediated signaling mechanisms in vivo.