Novel halogenated flame retardants in Canadian human milk from the MIREC study (2008-2011).

Novel halogenated flame retardants (NHFRs) have been developed to replace those brominated flame retardants that have been restricted due to their persistence, bioaccumulation potential and toxicity, therefore, it is important to determine whether these replacement products are present at detectable concentrations in Canadians. NHFRs were measured in human milk samples (n = 541) collected from across Canada between 2008 and 2011, which is the first pan-Canadian dataset for these chemicals in human milk. Among the 15 measured NHFRs and eight methoxy-polybrominated diphenyl ethers (MeO-PBDEs), nine NHFRs and two MeO-PBDEs (6-MeO-PBDE 47 and 2-MeO-PBDE 68) were detected at a frequency of more than 9%. Despite benzene, 1,1'-(1,2-ethanediyl)bis [2,3,4,5,6-pentabromo-]/decabromodiphenylethane [DBDPE] being detected less frequently than the other observed NHFRs, its relative contribution to the sum of nine NHFRs was important when it was present. The maximum ΣNHFR concentration in Canadian human milk was 6930 pg g-1 lipid while the maximum ΣMeO-PBDEs was 1600 pg g-1 lipid. While most NHFR concentrations were significantly correlated with each other, no relationships between maternal age, parity or pre-pregnancy BMI were identified with ΣNHFR concentrations in the milk. In contrast, maternal age was significantly correlated with ΣMeO-PBDE concentrations (r = 0.237, p < 0.001). ΣNHFR concentrations were similarly not related to maternal education, although ΣMeO-PBDE concentrations were found to be higher in milk from women who had graduated from trade schools relative to the other education levels considered. NHFR detection frequency and concentrations observed in the Canadian human milk seem to align well with Europe.

Differences in mirex [dechlorane] and dechlorane plus [syn- and anti-] concentrations observed in Canadian human milk.

As part of the pan-Canadian Maternal-Infant Research on Environmental Chemicals (MIREC) study, human milk samples were collected between 2008 and 2011, and analyzed for mirex, an organochlorine insecticide and flame retardant, in addition to dechlorane plus (syn- and anti-DDC-CO), the flame retardant replacement for mirex. Mirex was analyzed separately, using a method for the analysis of existing organochlorine insecticides, while the presence of DDC-CO isomers was determined using a method developed for the detection of emerging flame retardants. Mirex was detected in all samples analyzed (n = 298), while syn- and anti-DDC-CO were present in 61.0% and 79.5% of the samples, respectively (n = 541). Mirex concentrations have declined in human milk since the 1990s. Since this is the first pan-Canadian dataset reporting DDC-CO concentrations in human milk, no temporal comparisons can be made. Maternal age was correlated with concentrations of both compounds although parity did not impact concentrations of either analyte. Given the presence of this relatively recently identified flame retardant (DDC-CO) in human milk from women across Canada, studies to identify dominant sources of this compound are critical. Despite low concentrations of environmental chemicals in human milk from Canadian women, Health Canada supports breastfeeding of infants because of the important health benefits to both the mothers and their infants.

The influence of precursors and treatment process on the formation of Iodo-THMs in Canadian drinking water.

The National Survey of Disinfection By-Products and Selected Emerging Contaminants investigated the formation of various disinfection by-products and contaminants in 65 water treatment systems (WTSs) across Canada. Results for six iodo-trihalomethanes (iodo-THMs) are reported in this paper. The participating water treatment systems included large, medium and small systems using water sources and treatment processes which were representative of Canadian drinking water. Five water samples (source water, treated water and three water samples along the distribution system) were collected from each treatment system, both under winter and summer conditions. Samples were stabilized, shipped cold and analysed for six iodo-THMs (dichloroiodomethane-DCIM; dibromoiodomethane-DBIM; bromochloroiodomethane-BCIM; chlorodiiodomethane-CDIM; bromodiiodomethane-BDIM and triiodomethane or iodoform-TIM), using a SPME-GC-ECD method developed in our laboratory (MDLs from 0.02 µg/L for iodoform to 0.06 µg/L for bromodiiodomethane). Concentrations of relevant precursors like dissolved organic carbon (DOC), bromide, iodide and total iodine, as well as other water quality parameters, were also determined. Detailed information about the treatment process used at each location was recorded using a questionnaire. The survey showed that one or more iodo-THMs were detected at 31 out of 64 water treatment systems (WTSs) under winter conditions and in 46 out of 64 WTSs under summer conditions (analytical results from one site were excluded due to sampling challenges). Total iodo-THM concentrations measured during this survey ranged from 0.02 µg/L to 21.66 µg/L. The highest total iodo-THM concentration was measured in WTS 63 where all six iodo-THMs were detected and iodoform was present in the highest concentration. The highest iodo-THM formation was found to occur in treatment systems where water sources had naturally occurring ammonium as well as high bromide, high iodide and/or total iodine concentrations. In two such water systems the total concentration of iodo-THMs exceeded the concentration of regulated THMs.

Dioxins/furans and PCBs in Canadian human milk: 2008-2011.

Human milk was collected between 2008 and 2011 as part of the Maternal - Infant Research on Environmental Chemicals (MIREC) study that was initiated to establish Canadian national estimates of maternal and infant exposure to a broad suite of environmental contaminants (e.g., persistent organic pollutants [POPs], trace elements, phthalates, etc.). Among the 1017 human milk samples collected, 298 were analysed for polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs) and polychlorinated biphenyls (PCBs). World Health Organization (WHO) toxic equivalency concentrations (WHO TEQ2005) for PCDD/F+dioxin-like (DL) PCB ranged from 2.2pg TEQ2005 g-1 lipid to 27pg TEQ2005 g-1 lipid. The relative contribution of PCDDs to the overall WHO TEQ2005 (PCDD/F+DL PCB) has decreased from earlier investigations into POP levels in Canadian human milk. Significantly higher PCB concentrations were observed in milk from women born in Europe relative to those born in Canada (p<0.001), in contrast to results for the PCDD/Fs (p=0.496). Age was found to significantly impact milk ∑PCB concentrations (p=0.018), with elevated concentrations observed in milk from women >30years relative to those <30years of age. While this trend was also observed for the PCDD/Fs, this relationship was impacted by parity. WHO TEQ2005 concentrations were significantly higher in milk from primiparous women (p=0.019) and those >30years relative to those <30years of age (p<0.001). No significant differences were associated with education level or pre-pregnancy body mass index. PCB and PCDD/F concentrations have continued to decline in Canadian human milk since the last sampling of human milk was performed.

2- and 3-Monochloropropanediols in paper products and their transfer to foods.

The occurrence of 2- and 3-monochloropropanediol (MCPDs) in selected paper products sold on the Canadian market and the transfer of 3-MCPD from those products to beverages was probed. Products included coffee filters, tea bags, disposable paper hot beverage cups and milk packaged in paperboard containers. The occurrence MCPDs in coffee and tea filters on the German market was investigated as well. Furthermore, the presence of MCPDs in paper towels on the German market was also investigated. Analytes were determined by stable isotope dilution analysis GC-MS in SIM mode. 3-MCPD was detected in most paper products with levels ranging from a few nanograms in tea bags to a few micrograms in white coffee filters. Milk containers' paperboard contained 3-MCPD at about 500-1500 ng g(-1); however, 3-MCPD was detected in milk only in smaller containers, 237-500 ml (likely due to a lower volume/surface ratio) at levels of about 1 ng g(-1). Out of three disposable hot beverage paper cups tested (of 237-473 ml capacity), paperboard of two contained 3-MCPD at 632-792 ng g(-1), and 3-MCPD was detected in leachate from those paper cups at levels of about 16 ng per cup. 3-MCPD was detected in all paper towels at levels of 42-2466 ng g(-1). 2-MCPD was detected in many paper products (from Canadian and German sources) and in all the towels tested at levels varying from about 0.5-10% of that of 3-MCPD.

A method for the analysis of multiple novel halogenated flame retardants in cow's milk.

A method was developed for the extraction and analysis of cow's milk to measure 21 halogenated flame retardants (FRs), including individual isomers plus eight methoxy-polybrominated diphenyl ethers (MeO-PBDEs). Extraction was performed using homogenisation with acetone:hexane with size exclusion chromatography followed by adsorption chromatography clean-up. Analysis was undertaken using gas chromatography high-resolution mass spectrometry. The method was validated in fortified cow's milk with FRs and 2-methoxy-PBDE 68 at three levels (low [12.5-1250 pg g(-)(1)], mid [37.5-5000 pg g(-)(1)] and high [400-10 000 pg g(-)(1)]). Additional methoxy-PBDEs were tested at two fortification levels. Isotope dilution was used to correct for losses during sample preparation and average recoveries ranged from 58% (allyl 2,4,6-tribromophenyl ether [ATE]) to 121% (γ-tetrabromoethylcyclohexane (γ-TBECH)). Limits of detection ranged from 0.055 pg g(-)(1) (6-methoxy-PBDE 47) to 38.9 pg g(-)(1) (decabromodiphenylethane (DBDPE)). Matrix effects were overcome through the use of surrogate and performance standards. A single FR (1-bromomethyl-2,3,4,5,6-pentabromobenzene [PBBB]) and two methoxy-PBDEs were detected in commercially available cow's milk collected from local supermarkets in Ottawa, ON, Canada. Detection frequency was < 25% for these compounds and, where present, concentrations were low.

Hair as a biomarker of systemic exposure to polybrominated diphenyl ethers.

The efficacy of using hair as a biomarker for exposure to polybrominated diphenyl ether (PBDE) flame retardants was assessed in humans and an animal model. Paired human hair and serum samples were obtained from adult men and women (n = 50). In parallel, hair, serum, liver, and fat were collected from adult male Sprague-Dawley rats exposed to increasing doses of the PBDE mixture found in house dust for 70 days via the diet. All samples were analyzed by GC-MS for eight common PBDEs: BDE-28, -47, -99, -100, -153, -154, -183, and -209. Paired human hair and serum samples had five congeners (BDE-28, -47, -99, -100, and -154) with significant individual correlations (0.345-0.566). In rat samples, BDE-28 and BDE-183 were frequently below the level of detection. Significant correlations were observed for BDE-47, -99, -100, -153, -154, and -209 in rat hair, serum, liver, and fat across doses, with r values ranging from 0.803 to 0.988; weaker correlations were observed between hair and other tissues when data from the lowest dose group or for BDE-209 were analyzed. Thus, human and rat hair PBDE measurements correlate strongly with those in alternative matrices, validating the use of hair as a noninvasive biomarker of long-term PBDE exposure.

A mesofluidic multiplex immunosensor for detection of circulating cytokeratin-positive cells in the blood of breast cancer patients.

We have recently reported the analytical performance of an immunosensor comprising one mm-scale parallel plate laminar flow chamber and applied to capture MCF7 breast cancer cells (Ehrhart et al., Biosens. Bioelectr. 24, 467, 2008). Herein we present a new multiplex immunosensor embodying four parallel plate laminar flow chambers that fit onto a standard, functionalized, microscopy glass slide. The four surfaces are coated with long alkyl chain spacers of 21-aminohenicosyl trichlorosilane (AHTS) and then grafted with a monoclonal anti-human epithelial cell adhesion molecule (EpCAM) antibody specific of target cells to immobilize. We first demonstrate a significantly (P < 0.01) improved capacity of each of the four flow chambers of the multiplex immunosensor to capture MCF7 cells compared to the previous single chamber device. Second, in addition to an increase of cell immobilization, the multiplex device offers a versatile tool easily grafted with various purified antibodies onto the four surfaces. Third, we obtained high cell capture rate and efficiency of various numbers of MCF7 cells spiked in buffer containing an equal number of background leukocytes. And fourth, we demonstrate isolation efficiency of circulating tumor cells (CTCs) from peripheral blood drawn from a small cohort of patients with localized or metastatic breast cancer. This new multiplex immunosensor could be tested for its potential to capture different subpopulations of CTCs.

Silicate-entrapped porous coatings for preparing high-efficiency solid-phase microextraction sorbents.

We present a novel way to prepare SPME fibers using a silicate entrapment of porous particles, followed by derivatization using classical organosilane chemistry. The fibers provide a good platform for on-fiber derivatization of desired extraction phases while providing porosity necessary for high extractions capacities. The porous network was created using potassium silicate and porous silica particles. Fibers derivatized using n-butyl, n-octyl, n-octadecyl and n-triacontyl groups were shown to extract benzodiazepines successfully. The coatings were determined to have an average thickness of ca. 8 microm, as determined by a scanning electron microscope, permitting equilibrium times as fast as 2 min. The fibers also showed very good ruggedness towards a vast range of solvents and prolonged use. It was determined that greater extraction efficiencies could be obtained using triacontyl as an extraction phase. The C18 and C30 fibers were also found to provide good linearity (>0.99) for the model analytes over two orders of magnitude, with limits of detection in the sub ng mL(-1) levels. C30 fibers were used to establish a correlation between structurally diverse beta-blockers and their literature reported Log P values. The C30 fibers provided a good correlation (R(2)=0.9255) between beta-blockers ranging in hydrophobicity from Log P(literature) 0.16-4.15 and their respective experimentally determined Log K(spme) values.

Development of the space-resolved solid-phase microextraction technique and its application to biological matrices.

To facilitate rapid in situ analyte monitoring within heterogeneous samples, a space-resolved solid phase microextraction (SR-SPME) technique was developed that utilized miniaturized segmented fibers. Initially, a multilayered agarose gel was used to determine the effects of diffusion-based mass transfer and fiber dimension on the space-resolving capability of SPME. For diazepam within agarose gel, the SR-SPME limit of detection was 2.5 ng/mL, with a linear dynamic range up to 500 ng/mL. The efficacy of the SR-SPME technique was further evaluated within diverse biological matrices (onion bulb, fish muscle, and adipose tissues) containing stratified pharmaceutical analytes. Empirically, the results agreed well with established techniques such as microdialysis and liquid extraction, but SR-SPME was simpler to implement, displayed higher spatial resolution, and was more cost-effective than traditional approaches. Additionally, the segmented design of the SPME fibers and stepwise desorption protocols offer potential advantages within high throughput applications.

Immunoaffinity in-tube solid phase microextraction coupled with liquid chromatography-mass spectrometry for analysis of fluoxetine in serum samples.

The inherent selectivity of the antibody was combined with in-tube solid-phase microextraction by immobilization of the antibody into the fused silica capillary. A sensitive, selective, and reproducible immunoaffinity in-tube solid-phase microextraction coupled with liquid chromatography-mass spectrometry (in-tube SPME/LC-MS) method was developed, and validated for fluoxetine analysis in human serum. Important factors in the optimization of in-tube SPME variables, as well as the evaluation of the immunoaffinity capillary capacity are discussed. The in-tube SPME/LC-MS method presented a limit of quantitation of 5.00 ng/mL, and precision intra-assays with RSDs lower than 5%. The response of the in-tube SPME/LC-MS method for fluoxetine was linear over a dynamic range from 5.00 to 50.00 ng/mL, with correlation coefficients better than 0.998. Based on analytical validation it was demonstrated that in-tube SPME/LC-MS method offers high sensitivity, selectivity, and enough reproducibility to permit the quantification of fluoxetine in human serum at therapeutic levels. Thus, the proposed SPME/LC method can be useful tool to determine fluoxetine serum concentrations in patients receiving therapeutic dosages.