Dioxin related compounds in breast milk of women from vietnamese e waste recycling sites levels, toxic equivalents and relevance of non dietary exposure

Ecotoxicology and Environmental Safety 106 (2014) 220–225 Contents lists available at ScienceDirect Ecotoxicology and Environmental Safety journal homepage: www.elsevier.com/locate/ecoenv Dioxin-related compounds in breast milk of women from Vietnamese e-waste recycling sites: Levels, toxic equivalents and relevance of non-dietary exposure Nguyen Minh Tue a,b, Kana Katsura a, Go Suzuki c, Le Huu Tuyen a,b, Takumi Takasuga d, Shin Takahashi a,e,n, Pham Hung Viet b, Shinsuke Tanabe a a Center for Marine Environmental Studies (CMES), Ehime University, 2-5 Bunkyo-cho, Matsuyama 790-8577, Japan Centre for Environmental Technology and Sustainable Development (CETASD), Hanoi University of Science, 334 Nguyen Trai, Hanoi, Vietnam Center for Material Cycles and Waste Management Research, National Institute for Environmental Studies (NIES), 16-2 Onogawa, Tsukuba 305-8506, Japan d Shimadzu Techno-Research, Inc., Nishinokyo-Shimoaicho, Nakagyo-ku, Kyoto, Japan e Center of Advanced Technology for the Environment, Faculty of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama 790-8566, Japan b c art ic l e i nf o a b s t r a c t Article history: Received 17 January 2014 Received in revised form 28 April 2014 Accepted 29 April 2014 Although informal e-waste recycling sites (EWRSs) are hotspots of both polychlorinated and polybrominated dibenzo-p-dioxins/dibenzofurans (PCDD/Fs and PBDD/Fs), human exposure to the latter has not been studied in details This study investigated the accumulation levels and profiles of dioxin-related compounds (DRCs) in breast milk samples from women living in two Vietnamese EWRSs and estimated the intake contribution from e-waste-related exposure Screening results using Dioxin-Responsive Chemically Activated LUciferase gene eXpression assay (DR-CALUX) showed higher dioxin-like (DL) activities in samples from the EWRS Bui Dau than in those from the EWRS Trang Minh and a reference site (2.3–10 vs 1.7–4.8 and 0.60–5.7 pg CALUX-TEQ/g lipid, n ¼ 10, and 9, respectively) Chemical analysis results of selected samples show that the WHO-TEQ levels of PCDD/Fs, DL-PCBs and PBDD/Fs in EWRS samples were not significantly higher than in those from the reference site (0.22–7.4 vs 1.1–3.0 pg/g lipid) and within the Vietnamese background range, but women involved in recycling accumulated higher concentrations of PCDFs (13–15 vs 2.3–8.8 pg/g lipid) and PBDFs (1.1–1.5 vs o1.1 pg/g lipid) By comparing the DRC profile in milk of these women with the reported profile in house dust from the same site, dust ingestion was estimated to contribute most of the intake for tetraBDF, 37 per cent to 55 per cent for penta– octaCDFs, but less than twenty per cent for PCDDs and DL-PCBs, and 26 per cent for total WHO-TEQs The DL activities in some EWRS milk samples were not fully explained by chemical data, suggesting contribution from unidentified compounds The estimated WHO-TEQ intake doses for breastfed infants (1.3–33 pg/kg/d) mostly exceeded the tolerable value, especially for those living in the EWRSs; and unidentified DRCs might increase further the dioxin-related health risk & 2014 Elsevier Inc All rights reserved Keywords: CALUX Breast milk Brominated dioxin E-waste Vietnam Introduction Rapid growth in the amount of waste electrical and electronic equipment (WEEE), also known as e-waste, and continuing informal recycling activities of these materials pose serious risks of environmental pollution and human health impacts (Zhang et al., 2012) Primitive e-waste processing methods such as acidstripping and open burning of wires for metal retrieval, heating circuit boards for dismantling, and chipping and melting of plastics not only facilitate the release of hazardous chemicals contained in n Corresponding author at: Center of Advanced Technology for the Environment, Faculty of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama 790-8566, Japan Fax: ỵ81 89 946 9980 E-mail address: shint@agr.ehime-u.ac.jp (S Takahashi) http://dx.doi.org/10.1016/j.ecoenv.2014.04.046 0147-6513/& 2014 Elsevier Inc All rights reserved e-waste (e.g toxic metals, brominated flame retardants (BFRs), polychlorinated biphenyls (PCBs), etc.), but also generate dioxinrelated compounds (DRCs)—including polychlorinated dibenzo-pdioxins/dibenzofurans (PCDD/Fs), their brominated (PBDD/Fs) and mixed brominated/chlorinated homologues (PXDD/Fs)—as secondary pollutants (Tue et al., 2013b) The occurrence of PBDD/Fs in EWRSs has been associated with high content of BFRs, especially polybrominated diphenyl ethers (PBDEs), in e-waste plastics (Ma et al., 2009; Tue et al., 2010) Technical PBDE formulations have been found to contain PBDFs as impurities (Hanari et al., 2006) PBDD/Fs can be generated from these precursors not only during waste incineration (Duan et al., 2011; Weber and Kuch, 2003), as is the case with the more well known PCDD/Fs, but also through degradation at o300 1C (Weber and Kuch, 2003) or under natural light (Kajiwara et al., 2008) N.M Tue et al / Ecotoxicology and Environmental Safety 106 (2014) 220–225 PBDD/Fs have been found in soil, dust and air of Asian e-waste recycling sites (EWRSs) at concentrations greater than those of PCDD/Fs (Tue et al., 2013b) Considering that PBDD/Fs can produce dioxin-like (DL) toxic effects by activating the aryl hydrocarbon receptor (AhR) signalling pathway with similar relative effect potencies (REPs) to those of PCDD/Fs, and have recently been recommended for inclusion in the World Health Organisation Toxicity Equivalency Factor (WHO-TEF) concept (van den Berg et al., 2013), it is important to include PBDD/Fs in the assessment of human exposure and potential risks related to DRCs released from informal e-waste recycling Environmental contamination with PCDD/Fs in EWRSs resulted in high accumulation in human via soil/dust ingestion, inhalation and consumption of contaminated local food, as reported in various studies in China (reviewed by Chan and Wong, 2013) For PBDD/Fs, although high exposure levels from non-dietary sources have been estimated (Tue et al., 2013b), information on the levels of these DRCs in EWRS residents and their diet is notably lacking Such information will be a useful addition to the scarce data on PBDD/Fs in human, currently available only for small numbers of adipose tissue and breast milk samples (in the range of a few picogram per gram lipid; Choi et al., 2003; Ericson Jogsten et al., 2010; Croes et al., 2013) The present study investigated breast milk samples as bioindicators for human exposure to DRCs in two Vietnamese EWRSs, where our previous survey found relatively high PBDF concentrations in house dust (median 23– 49 ng/g; Tue et al., 2010) We aimed to elucidate the accumulation levels and profiles of DRCs including PBDD/Fs as well as the relevance of non-dietary exposure for human intake, to evaluate the contribution of various DRCs to the total DL toxicity levels in breast milk and to assess the risk for breastfeeding infants Using a combined in vitro bioassay/instrumental analysis approach, we used the Dioxin-Responsive Chemically Activated LUciferase gene eXpression (DR-CALUX) assay as a tool for screening and measuring the total DL toxicity of all potential DRCs present in the breast milk samples 221 2.3 DR-CALUX assay To remove non-persistent compounds such as polyaromatic hydrocarbons (PAHs) which are not considered as DRCs, the extract was treated with sulphuric acid-impregnated silica gel column according to a previously reported method (Tue et al., 2010), then concentrated and solvent-exchanged into 50 mL biochemicalgrade dimethyl sulphoxide (DMSO) for bioassay Dioxin-like activity was measured using DR-CALUX assay with a rat hepatoma cell line having an AhR-regulated luciferase gene construct (H4IIE-luc, BioDetection Systems b.v., The Netherlands) The culture conditions, assay procedures and data analysis followed those used in our previous reports (Suzuki et al., 2010; Tue et al., 2010) All measurements satisfied the criteria of the standard operating procedure regarding the calculated EC50 of 2,3,7,8-CDD (TCDD) (11.3 1.1 pM, n¼ 25) and the maximum induction relative to DMSO control (8.67 1.6) Results were expressed in picogram CALUX TCDD-equivalent (CALUX-TEQ) per gram milk lipid 2.4 Chemical analyses of DRCs Fourteen selected milk extracts containing more than 0.03 pg CALUX-TEQ/g wet weight were spiked with 13C12-labelled surrogates (2,3,7,8-substituted tetra– octaCDD/Fs, tetra–hexaBDD/Fs and DL-PCBs) and then subjected to additional clean-up with multilayer silica gel column (silica, ten per cent AgNO3-impregnated silica, silica, 22 per cent H2SO4-impregnated silica, 44 per cent H2SO4-impregnated silica and silica in bottom-up order) and separation over activated carbonimpregnated silica gel column 13C12-labelled 1,2,3,4-TeCDD, 1,2,3,4,6,9-HxCDF, 1,2,3,4,6,8,9-HpCDF were added to the PCDD/F-PBDD/F fraction and 13C12-CB-111 to the DL-PCB fraction as internal standards 2,3,7,8-substituted PCDD/Fs, PBDD/Fs and DL-PCBs were quantified using a gas chromatograph (HP-6890, Agilent) and a high resolution mass spectrometer (AutoSpec Ultima, Waters) PCDD/Fs and DL-PCBs were separated using a BPX-DXN column (60 m  0.25 mm, SGE, Australia) with a temperature programme of 20 1C/min from 150 to 220 1C, 1C/min to 260 1C, 1C/min to 320 1C, hold min, and a RH-12ms column (60 m  0.25 mm, Inventx, USA) with a programme of 20 1C/min from 150 to 210 1C, 1C/min to 280 1C, 10 1C/min to 320 1C, hold 13 PBDD/Fs were separated using a DB-5MS column (15 m  0.25 mm, Agilent) and a programme of 120 1C for one minute, 20 1C/min to 240 1C, 10 1C/min to 300 1C and hold thirteen minutes All analyses were carried out by an accredited laboratory (Shimadzu Techno-Research Inc., Japan) which frequently participates in worldwide intercalibration studies for PCDD/Fs, DL-PCBs (UNEP, 2005) and PBDD/Fs (Takahashi et al., 2006) The average recoveries of the 13C-labelled surrogates were 80–97 per cent, 66–114 per cent and 95–110 per cent for PCDD/Fs, DL-PCBs and PBDD/Fs, respectively Results were expressed in picogram per gram milk lipid WHO-TEQs were calculated for PCDD/Fs and DL-PCBs using the World Health Organisation toxic equivalency factors (TEFs) (van den Berg et al., 2006) and for PBDD/Fs using TEFs of similarly substituted PCDD/Fs (van den Berg et al., 2013) Material and Methods 2.1 Sample collection This sampling was conducted in three locations in northern Vietnam: two e-waste recycling sites Trang Minh (TM, Kien An district, Hai Phong city) and Bui Dau (BD, My Hao district, Hung Yen province), and a reference site Thach Hoa (TH, Thach That district, suburban Hanoi) Information on the populations and recycling activities of the EWRSs has been provided in our previous reports (Tue et al., 2010, 2013a) Breast milk was chosen as the bioindicator because the large collectable volume and relatively high lipid content are suitable for studying the human accumulation of lipophilic contaminants such as DRCs from e-waste recycling Milk samples (n ¼6, 9, and 10 for TH, TM and BD, respectively) were collected in September 2008 with informed consent of the donors All donors were healthy and non-smokers (see supplementary Table S1 for further details) Milk was expressed by the donor or with the help of a midwife into a solvent-precleaned glass bottle with Teflon-lined screw caps All samples were kept with gel ice immediately and then sent within eight hours to our laboratory at Hanoi University of Science and frozen at À 20 1C The frozen samples were later air-transported with gel ice to Ehime University and stored at À 25 1C until analysis 2.2 Sample extraction Approximately 50 g of milk sample was freeze-dried and extracted with an acetone/hexane mixture (1:1 v/v) using a rapid solvent extractor (SE100, Mitsubishi Chemical Analytech, Japan) for 30 at 35 1C and a flow rate of 10 ml/min Five per cent of the extract was used for gravimetric determination of the lipid content, and the remaining extract was subjected to gel permeation chromatography for lipid removal Two portions equivalent to 13.5 and 25 g wet weight were used for DR-CALUX and chemical analysis, respectively A procedural blank was analysed with every set of seven samples Results and discussion 3.1 Dioxin-like activities and accumulation levels of DRCs in human milk All sulphuric acid-treated human milk extracts exhibited DL activities as evidenced by the dose-dependent luciferase induction in the DR-CALUX assay (supplementary Fig S1) The CALUX-TEQ levels were 0.60–10 pg/g lipid in the whole sample set (Table 1) The levels in TM and TH were not different but those from BD were significantly higher (Wilcoxon rank sum test p o0.05), showing 3.2 and 1.7-fold difference in terms of medians compared with TH and TM, respectively The minimum CALUX-TEQ levels in the EWRSs were still the same or higher than the median in TH PCDD/Fs and DL-PCBs were detected in all fourteen samples selected for chemical analysis PBDFs were found in only two samples whereas PBDDs were not detected (Table 1) In general, there was no significant difference in DRCs concentrations for the three study locations However, the two samples from BD recyclers had considerably higher concentrations of PCDFs (13–15 pg/g lipid) and DL-PCBs (6600–7600 pg/g lipid) as compared with TH and TM samples (2.3–8.8 and 430–5000 pg/g lipid, respectively) These were also the only samples with detectable levels of PBDFs (1.1–1.5 pg/g lipid) In general, the most abundant among PCDD/Fs were OCDD and 1,2,3,4,6,7,8-HpCDD (42–56 per cent and 10–13 per cent of total concentrations), and among DL-PCBs were 222 N.M Tue et al / Ecotoxicology and Environmental Safety 106 (2014) 220–225 Table Concentrations (pg/g lipid) of PCDD/Fs, PBDFs, DL-PCBs, WHO-TEQs and CALUX-TEQs in human milk samples from two e-waste recycling sites (TM and BD) and a reference site (TH) in Vietnam Compound TH (n¼4) TM (n ¼5) BD (n¼ 5) Min Max Median Min Max Median Min Max Median 2,3,7,8-TeCDD 1,2,3,7,8-PeCDD 1,2,3,4,7,8-HxCDD 1,2,3,6,7,8-HxCDD 1,2,3,7,8,9-HxCDD 1,2,3,4,6,7,8-HpCDD OCDD Total PCDDs 0.12 0.29 o 0.12 0.27 o 0.12 1.5 5.6 8.4 0.25 0.58 0.39 1.1 0.54 3.9 15 22 0.20 0.35 0.20 0.48 0.23 2.3 11 14 0.074 0.17 0.16 0.24 o0.16 0.98 3.8 5.6 0.45 1.0 0.57 1.5 0.42 3.2 17 23 0.18 0.32 0.19 0.47 0.24 1.1 6.2 8.8 o 0.049 0.049 o 0.097 o 0.097 o 0.097 0.18 0.49 0.72 0.64 0.96 0.59 1.6 0.61 9.8 46 56 0.10 0.15 ND 0.23 ND 1.7 7.5 12 2,3,7,8-TeCDF 1,2,3,7,8-PeCDF 2,3,4,7,8-PeCDF 1,2,3,4,7,8-HxCDF 1,2,3,6,7,8-HxCDF 1,2,3,7,8,9-HxCDF 2,3,4,6,7,8-HxCDF 1,2,3,4,6,7,8-HpCDF 1,2,3,4,7,8,9-HpCDF OCDF Total PCDFs 0.14 0.10 0.53 0.27 0.27 o 0.12 0.16 0.46 o 0.12 o 0.20 2.3 0.35 0.27 1.6 0.89 0.86 o 0.23 0.58 1.2 0.29 0.88 6.1 0.23 0.16 0.76 0.61 0.52 ND 0.24 0.89 0.11 0.45 4.2 0.15 0.099 0.32 0.20 0.30 o0.15 0.16 0.35 o0.15 o0.27 2.4 0.32 0.35 2.5 1.8 1.8 o 0.16 0.79 1.2 o 0.16 0.49 8.8 0.22 0.16 0.90 0.58 0.50 ND 0.27 0.59 ND ND 2.9 0.098 0.074 0.17 o 0.15 o 0.15 o 0.097 o 0.097 o 0.15 o 0.097 o 0.16 0.34 1.7 2.4 3.6 2.1 2.3 0.39 1.2 16 2.8 39 61 0.15 0.21 0.70 0.46 0.52 0.26 0.80 1.2 0.21 ND 13 2,3,7,8-TeBDF Total PBDFs o 0.60 ND o 1.1 ND ND ND o0.74 ND o 0.80 ND ND ND o 0.49 ND 1.5 1.5 ND ND CB-77 CB-81 CB-126 CB-169 o 2.2 o 1.2 2.8 1.5 2.7 1.6 10 3.9 1.1 0.6 5.2 2.4 o1.1 o1.6 1.2 o1.6 3.7 3.5 15 7.2 1.8 ND 5.3 2.4 o 5.6 o 2.7 0.74 o 1.3 9.5 7.0 21 6.7 ND ND 2.3 1.3 Non-ortho PCBs CB-105 CB-114 CB-118 CB-123 CB-156 CB-157 CB-167 CB-189 4.4 74 28 380 5.6 150 32 38 8.6 18 470 74 1800 27 350 70 110 25 9.3 330 47 1200 15 250 54 76 17 1.2 74 9.4 270 4.0 47 11 16 3.5 29 790 160 3000 47 600 130 200 45 9.7 320 45 1200 19 210 43 80 15 0.74 37 4.4 130 2.0 22 4.7 7.6 1.5 44 1300 270 4300 72 1100 230 250 64 3.6 180 28 570 11 120 26 36 5.9 Mono-ortho PCBs WHO-TEQ (no PBDFs) WHO-TEQ (with PBDFs) CALUX-TEQ CALUX-TEQa 750 1.1 1.1 1.0 0.60 2900 3.3 3.3 4.5 5.7 2000 1.7 1.7 1.7 1.7 430 0.72 0.72 2.4 1.7 5000 4.8 4.8 4.8 4.8 2000 1.8 1.8 3.2 3.2 210 0.22 0.22 3.3 2.3 7600 5.9 6.1 10 10 980 1.4 1.4 7.8 5.3 ND: not detected a whole sample set (n¼6, 9, 10 for TH, TM and BD, respectively) PCB-118, À 105 and À 156 (58–61 per cent, 15–18 per cent, 12–15 per cent) (Supplementary materials Fig S2) Several samples from BD had larger proportions of PCDFs (24–54 per cent of total PCDD/Fs) Higher concentrations of DRCs, especially PCDFs which are often indicative of combustion sources (Minh et al., 2003), in breast milk of some donors living in BD indicate possible exposure through their involvement in informal recycling activities such as e-waste burning and/or melting Accumulation in human tissues reflects environmental contamination by DRCs in BD as evidenced by their high concentrations in indoor dust reported previously (Tue et al., 2010) Despite similar indoor DRC contamination levels reported in TM and BD (Tue et al., 2010), milk samples from TM did not have significantly higher DRC concentrations than those from TH (Table 1), which could be explained by TM donors being primarily non-recyclers (Table S1) and less likely to be exposed to DRCs from recycling Overall, the human milk PCDD/F-TEQ levels in these EWRSs were within the Vietnamese background range (0.8–8.2 pg/g lipid), and the highest level was 2–3 times lower than the geometric means reported for Agent Orange-sprayed areas (7.56–10.89 pg/g lipid; The Tai et al., 2011) The highest PCDDFỵPCB-TEQ level in BD was 68 times lower than in women exposed to DRCs from open waste burning in India (average 39–50 pg/g; Someya et al., 2010) and also lower than in women living at the EWRS Luqiao in China (average 21.0 and 12.6 pg/g lipid for PCDD/F- and PCB-TEQ, respectively; Chan et al., 2007; Zhang et al., 2010) Compared with the CALUX-TEQ results from a limited number of studies investigating in vitro DL activities in human milk (median 15, 15 and 13 pg/g lipid in Hong Kong, China and Belgium, respectively; Hui et al., 2007; Leng et al., 2009; Croes et al., 2013), the only other study to date, the levels in this study were lower, which is consistent with the low concentrations of DRCs in Vietnamese human milk in general (The Tai et al., 2011) This study is the first to report the detection of PBDFs in breast milk of women involved in informal e-waste recycling TeBDF was detected only in milk samples from BD recyclers with elevated concentrations of total PBDEs (11–19 ng/g lipid vs 0.2–2.8 ng/g lipid for other donors, details not shown) and of PCDFs, suggesting that their exposure sources involved e-waste plastics containing PBDEs Higher brominated PBDFs were likely to accumulate to a certain extent, because Pe-, Hx-, Hp- and OBDFs were approximately 2, 10, 90, 50 times more abundant than TeBDF in BD house N.M Tue et al / Ecotoxicology and Environmental Safety 106 (2014) 220–225 dust (Tue et al., 2010) However, only the latter could be detected in this study, possibly due to the low instrumental sensitivity for the heavier congeners (LOD ¼ 3, 5, 20, and 50 pg/g lipid, respectively) and/or their low uptake/accumulation in the human body PeBDFs have been detected in a pooled human milk sample from Belgium at a similar concentration with that of TeBDF (0.6 vs 0.7 pg/g lipid; Croes et al., 2013) Nevertheless, the concentrations of TeBDF in milk samples from BD recyclers were comparable to those of TeCDF and higher than those of TeCDD (Table 1, Supplementary materials Fig S3), indicating exposure of these donors to both PCDFs and PBDFs at similar levels during their involvement with e-waste recycling 3.2 Relevance of different DRCs to total dioxin-like activities The largest WHO-TEQ contribution in all milk samples was from CB-126 (17–34 per cent) Other important contributors were 1,2,3,7,8-PeCDD (11–34 per cent) and 2,3,4,7,8-PeCDF (12–28 per cent) For BD samples, WHO-TEQs of PCDFs were higher than those of PCDDs (29–50 per cent vs 20–35 per cent) When detected, TeBDF contributed about two per cent of total WHO-TEQs, in agreement with the low WHO-TEQ contribution of TeBDF and PeBDFs in human adipose tissues from Sweden and human milk from Belgium (0.2–2 per cent; Ericson Jogsten et al., 2010; Croes et al., 2013) As shown in Table 1, the measured CALUX-TEQs were consistently higher than the respective WHO-TEQs This could be explained in part by the difference between the in vitro REP values of DRCs in DR-CALUX and their WHO-TEFs (Behnisch et al., 2003) The largest change in the total calculated TEQ values of the milk samples by replacing WHO-TEQs with REP-derived theoretical CALUX-TEQs (Theo-TEQs) were for PCDFs (40–110 per cent increase), followed by PCDDs (2–70 per cent increase) and DL-PCBs (4–10 per cent decrease) Detailed Theo-TEQ values and ratios between the total Theo-TEQs and the respective measured CALUX-TEQs are shown in Fig In four samples, the total TheoTEQs exceeded the CALUX-TEQs (ratio¼ 1.5–2.6), possibly because of low recovery of DRCs during clean-up of these samples for Theo−TEQ (pg/g lipid) 10 Total Theo−TEQ/CALUX−TEQ ratio PBDFs PCBs PCDFs PCDDs TH TM BD 2.5 2.0 1.5 1.0 223 bioassay (not adjusted with surrogate standards as was the case with chemical analysis) or because of antagonistic effects from unknown compounds In five other samples, Theo-TEQs and CALUX-TEQs were comparable (ratio¼ 0.7–1.2) For the remaining five, all from EWRSs, Theo-TEQs of known DRCs accounted for only 6–58 per cent of the measured activity Unexplained DL activity has been commonly observed in previous studies with sulphuric acid-treated house dust extracts, where identified PCDD/Fs, DL-PCBs, PBDD/Fs and polychlorinated naphthalenes explained less than 50 per cent of the activity measured with DR-CALUX in most cases (Suzuki et al., 2010; Tue et al., 2010, 2013a) Contaminants suspected as responsible for these unexplained activities include potentially persistent AhR agonists such as non-2,3,7,8-substituted PBDFs (and unidentified PXDFs in the case of EWRSs) (Tue et al., 2010), halogenated polyaromatic hydrocarbons (Horii et al., 2009) as well as unknown compounds not eluting with DRCs during clean-up (Tue et al., 2013a) However, unlike reported in studies on house dust, unexplained DL activities were found only in about one third of the milk samples in this study, suggesting that some of these unidentified AhR agonists may also accumulate in human tissues, but with lower persistence or uptake efficiency than those of DRCs 3.3 Relevance of non-dietary DRC exposure for occupationally exposed women Although diet is generally considered as the principal human exposure route of DRCs such as PCDD/Fs and DL-PCBs (Liem et al., 2000), PBDFs detected in some donors in BD were likely from nondietary intake Dust ingestion has been estimated to contribute up to 88 per cent of the intake of PBDEs, which are precursors of PBDFs, for e-waste recycling workers in TM and BD (Tue et al., 2013c) Moreover, low PBDF levels in the diet were expected for these EWRSs because the main food supplies were from non-ewaste neighbouring communes and contained low levels of PBDEs (Tue et al., 2013c) The occurrence of PBDFs in food has been investigated by only a few studies, where their abundance relative to PBDEs in fish and shellfish was in the range of three orders of magnitude lower (Fernandes et al., 2008; Skinner, 2011) but could reach the per cent range in seafood from south-eastern China (Miyake et al., 2008) In the absence of any recent information on the dietary exposure of DRCs in Vietnam, the difference in the profiles of individual DRCs in human milk and those previously reported in house dust (Tue et al., 2010) can provide useful insight regarding exposure routes Assuming that TeBDF in breast milk of BD donors occupationally involved in e-waste recycling was solely from dust ingestion (100 per cent), the contribution of this exposure route to their intake of a DRC can be estimated from the ratio between its TeBDF-relative abundance in dust and in milk, its relative persistence and uptake efficiency (see Supplementary materials) As shown in Fig 2, dust ingestion was estimated as an important contributor to the PCDF intake of these donors (37–55 per cent for PeCDFs, HxCDFs and OCDF) For PCDDs and DL-PCBs which are uncharacteristic of informal e-waste recycling, dust ingestion contributed only twenty per cent or less Overall, dust ingestion was estimated to contribute 26 per cent of the total WHO-TEQ intake for occupationally exposed women 0.5 3.4 DRC exposure of infants through breastfeeding 0.0 Fig REP20-based theoretical CALUX-TEQs (Theo-TEQs) of PCDD/Fs, DL-PCBs and PBDFs in human milk from a suburban (TH) and two e-waste recycling sites (TM and BD) (top) and comparison with the measured CALUX-TEQs (bottom); two rightmost bars representing data for BD e-waste recyclers Daily intake doses (DIs) of DRCs for infants from breast milk were estimated (Fig 3) assuming an average daily milk consumption of 700 g/d and an average infant body weight of kg (van Oostdam et al., 1999) Although the levels of DRCs in breast milk of 224 N.M Tue et al / Ecotoxicology and Environmental Safety 106 (2014) 220–225 exposure doses for infants living in TM and BD (median DI 9.5 and 7.4 pg WHO-TEQ/kg/d, respectively) would increase by 1.3 and 4.1 folds with the inclusion of the DL activities from unknown compounds (Fig 3), estimated for some milk samples by multiplying the total WHO-TEQ of known DRCs with the unexplained/ explained CALUX-TEQ ratio (Section 3.2) However, it is still necessary to identify these unknown AhR agonists and to determine whether they can cause significant in vivo effects Abundance (median vs TeBDF) Intake contribution from dust (%) 5668 14 100 House dust Human milk 12 10 4 Conclusions 80 60 40 20 WHOTEQ TeBDF DLPCBs OCDF HpCDFs HxCDFs PeCDFs TeCDF OCDD HpCDD HxCDDs PeCDD TeCDD Fig Abundance of DRCs relative to 2,3,7,8-TeBDF (median ratios) in human milk of e-waste-exposed women (this study) and in house dust (Tue et al., 2010) from BD (top) and estimated contribution of dust ingestion to the total intake of DRCs (bottom) 40 WHO−TEQ With unknown DRCs Daily dose (pg TEQ/kg/d) 30 The results of the present study confirm that residents of Vietnamese EWRSs were exposed to DRCs released from informal e-waste recycling People involved in recycling activities in Bui Dau village were likely to be more exposed to PCDFs and PBDFs, and ingestion of contaminated dust was an important exposure route of these compounds Among PBDFs, only TeBDF was detected in human milk, but at similar or higher concentrations compared with those of the chlorinated homologue TeCDD/Fs The results of DR-CALUX analysis indicate that some milk samples collected from the EWRSs may have contained unidentified compounds with dioxin-like activities The total WHO-TEQ levels found in human milk of the present study remained within the Vietnamese background range, indicating relative low exposure levels at the current scale of e-waste recycling Nevertheless, exposure to DRCs from e-waste via dust was found as significant contributor of WHO-TEQ in breast milk of recyclers and the estimated WHO-TEQ intake doses for breastfed infants in the EWRSs were up to an order of magnitude higher than the WHO tolerable dose It should be noted that data from the small sample size of the present study may not reflect wholly the situation of human exposure to DRCs in the EWRSs The EWRS donors were also limited to lactating mothers who were likely to participate less actively in recycling activities than other recyclers and were possibly less exposed Proper management is necessary to avoid escalating environmental and human contamination with the growing e-waste business at these EWRSs Acknowledgments 20 This study was supported in part by Grants-in-Aid for Scientific Research (A: 25257403), Grants-in-Aid for Young Scientists (A: 23681011) from Japan Society for the Promotion of Science (JSPS), the Environment Research and Technology Development Fund (K123001 and 3K133010) from the Japanese Ministry of the Environment The award of a JSPS postdoctoral fellowship to N.M.T (P 13072) is also acknowledged 10 Appendix A Supplementary materials TH TM BD Fig Estimated daily intake doses of PCDD/Fs, DL-PCBs, PBDFs (in WHO-TEQ) and unknown DRCs (based on the percentage of unexplained DL activity in Fig 2) for breastfeeding infants in a suburban (TH) and two e-waste recycling sites (TM and BD); whiskers representing extreme values within 1.5-time the interquartile ranges from the lower and upper quartiles this study were not higher than the Vietnamese background, the range of estimated DIs in terms of WHO-TEQ was still higher than the range of tolerable daily dose recommended by the WHO (1–4 pg/kg/d; van Leeuwen et al., 2000), suggesting that the exposure levels from breastfeeding were high enough for potential DRC-related adverse health effect Involvement 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Environ Sci Technol 46, 10861–10867  ... Contamination of indoor dust and air by polychlorinated biphenyls and brominated flame retardants and relevance of non-dietary exposure in Vietnamese informal e-waste recycling sites Environ Int... 2010 Evaluation of dioxin-like activities in settled house dust from Vietnamese e-waste recycling sites: relevance of polychlorinated/brominated dibenzo-pdioxin/furans and dioxin-like PCBs Environ... from informal e-waste recycling People involved in recycling activities in Bui Dau village were likely to be more exposed to PCDFs and PBDFs, and ingestion of contaminated dust was an important exposure