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Residue pro files of organohalogen compounds in human serum frome-waste recycling sites in North Vietnam: Association with thyroid hormone levels a Center for Marine Environmental Studies

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Residue pro files of organohalogen compounds in human serum from

e-waste recycling sites in North Vietnam: Association with thyroid

hormone levels

a

Center for Marine Environmental Studies (CMES), Ehime University, Bunkyo-cho 2-5, Matsuyama 790-8577, Japan

b

Centre for Environmental Technology and Sustainable Development, Hanoi University of Science, 334 Nguyen Trai, Hanoi, Vietnam

c

Center of Advanced Technology for the Environment, Faculty of Agriculture, Ehime University, Tarumi 3-5-7, Matsuyama 790-8566, Japan

a r t i c l e i n f o

Article history:

Received 11 August 2014

Received in revised form

7 January 2015

Accepted 8 January 2015

Available online 6 February 2015

Keywords:

e-waste

PCBs

PBDEs

Hydroxylated metabolites

Thyroid hormones

a b s t r a c t This study demonstrated the contamination levels of polychlorinated biphenyls (PCBs), hydroxylated PCBs (OH-PCBs), polybrominated diphenyl ethers (PBDEs), methoxylated PBDEs (MeO-PBDEs), hydro-xylated PBDEs (OH-PBDEs), and bromophenols (BPhs), and their relationships with thyroid hormones (THs), in the serum of human donors from an e-waste recycling site and a rural site in Hung Yen province, Vietnam Occupationally related exposure was indicated by significantly higher residue levels of PCBs, OH-PCBs, PBDEs, and BPhs in the serum of donors from the e-waste recycling site (median: 420, 160, 290, and 300 pg g1wet wt, respectively) than those in the serum of donors from the rural site (median: 290,

82, 230, and 200 pg g1wet wt, respectively) On the other hand, levels of OH-/MeO-PBDEs were sig-nificantly higher in serum of donors from the reference site (median: 160 and 20 pg g1wet wt, re-spectively) than in those from the e-waste recycling site (median: 43 and 0.52 pg g1wet wt, respec-tively) In addition, we implemented stepwise generalized linear models to assess the association be-tween the levels of TH and PCBs, PBDEs, and their related compounds In females, we found positive associations of PCBs and OH-PCB concentrations with total thyroxine, free thyroxine, total triiodothyr-onine, and free triiodothyrtriiodothyr-onine, and a negative association with thyroid-stimulating hormone con-centrations

& 2015 Elsevier Inc All rights reserved

1 Introduction

Electrical waste and electronic equipment, also known as

e-waste, refers to end-of-life products encompassing information

communication devices, consumer electronics, and household

appliances (UNEP, 2005) These e-wastes contain many hazardous

substances, such as heavy metals and persistent organohalogen

compounds, which can be released into the environment (SVCT,

2002) In addition, recent recycling activities of e-waste in

Viet-nam are considered a problem Our research group has recently

suggested that Vietnamese workers at an e-waste site are

occu-pationally exposed to polychlorinated biphenyls (PCBs),

poly-brominated diphenyl ethers (PBDEs), dioxin-like compounds, and

trace elements during recycling activities (Noguchi et al., 2014;

Tue et al., 2010a,2010b,2013) Inappropriate methods of e-waste

recycling or dismantling are related to increased environmental emissions of toxic chemicals that may affect human health The thyroid hormone (TH) system is an important endocrine target for many organohalogen contaminants It is well-known that TH plays a critical role in regulating biological processes, such

as growth, metabolism, and neurodevelopment (Gelfand et al.,

1987;Zoeller, 2003) The metabolism of PCBs and PBDEs result in the formation of a relatively large number of hydroxylated PCBs (OH-PCBs), hydroxylated PBDE congeners (OH-PBDE), and bro-mophenols (BPhs), which are associated with perturbed thyroid homeostasis and neurodevelopmental deficits (Cheek et al., 1999; Meerts et al., 2002)

Some isomers of OH-PBDEs, BPhs and methoxylated PBDEs (MeO-PBDEs) have been found in humans and various animals (Eguchi et al., 2012;Malmberg et al., 2005;Malmvarn et al., 2008; Marsh et al., 2004;Mizukawa et al., 2013;Nomiyama et al., 2011a; Wan et al., 2009) Among MeO-PBDEs, two abundant isomers (6MeO-BDE47 and 2′MeO-BDE68) have been found to be natural

Contents lists available atScienceDirect

journal homepage:www.elsevier.com/locate/envres Environmental Research

http://dx.doi.org/10.1016/j.envres.2015.01.007

0013-9351/& 2015 Elsevier Inc All rights reserved.

n Corresponding author Fax: þ81 89 927 8196.

E-mail address: keinomi@agr.ehime-u.ac.jp (K Nomiyama).

Environmental Research 137 (2015) 440–449

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products (Teuten et al., 2005) Demethylation of MeO-PBDEs by

cytochrome P450 (CYP) can result in the formation of more

OH-PBDEs than through the metabolism of parent OH-PBDEs (Wan et al.,

2009) On the other hand, an in vitro study has shown that

ex-posure of human liver microsomes to BDE 99 led to the generation

of OH-PBDEs and BPhs (Stapleton et al., 2009), and an in vivo study

has shown that exposure of rats and mice to PBDEs led to the

generation of OH-PBDEs (Malmberg et al., 2005; Morck et al.,

2003;Qiu et al., 2007)

In recent years, scientists have paid considerable attention to

potential risk to environmental and human health of both natural

and anthropogenic halogenated phenolic compounds; however,

few studies have reported exposure risk of PCBs, PBDEs and their

hydroxylated/methoxylated derivatives in Asian developing

countries, especially for donors of e-waste recycling sites In the

present study, we aimed to determine serum concentrations of

PCBs, OH-PCBs, PBDEs, MeO-PBDEs, OH-PBDEs, and BPhs in

hu-man donors at an e-waste recycling site and a rural site in

northern Vietnam

On the other hand, it was reported that iodide uptake is

es-sential for the production of THs (Bianco et al., 2002), while

per-chlorate and thiocyanate can competitively inhibit iodide uptake

by the thyroid gland via the sodium/iodide symporter, reducing

the synthesis of TT3and TT4(Dohan et al., 2007;Tonacchera et al.,

2004) However, a few studies focused on the associations among

organohalogen contaminants, anions and THs (Alvarez-Pedrerol

et al., 2009;Hisada et al., 2013;Lopez-Espinosa et al., 2009)

Thus, we also examined the relationships between serum

concentrations of THs and these organohalogen compounds and

their metabolites to assess the effects on TH homeostasis in

Viet-namese populations, considering various potential co-factors, such

as donor characteristics and levels of perchlorate, thiocyanate and

iodide analyzed in our previous research (Eguchi et al., 2014)

2 Materials and methods

2.1 Collection of serum samples

Human serum samples were collected from donors who were

workers at an e-waste recycling site of Bui Dau (n¼77) and donors

who were residents from the rural area of Duong Quang (n¼34) in

Vietnam, during 2010 and 2011 (Fig S1) These 111 donors were

informed beforehand about the purpose of the study at local

government health stations where volunteers registered their

consent to participate, and they consented to participation in our

study All the participants were randomly selected without

arbi-trary criteria Informed consent was obtained from all donors, and

this study was approved by the Ethical Committee of Ehime

Uni-versity, Japan Demographic, health, and diet information were

collected through personal interviews (Table 1) All samples were

shipped to Ehime University, Japan, on gel ice and stored at

25 °C in the environmental specimen bank for global monitoring

(es-BANK:http://esbank-ehime.com) (Tanabe, 2006) until

chemi-cal analysis

2.2 Chemicals

The certified standards used for the identification and

quanti-fication of PCBs, OH-PCBs, PBDEs, OH-PBDEs, MeO-PBDEs, and

BPhs are described in theSupplementary information

2.3 Thyroid hormone analysis

THs in serum were analyzed by following the

electro-chemiluminescence immunoassay (ECLIA) procedures reported in

a previous study (Kunisue et al., 2011) Thyroid-stimulating hor-mone (TSH), total triiodothyronine (TT3), total thyroxine (TT4), free

T3 (FT3), and free T4 (FT4) were measured using Elecsyss kits (Roche Diagnostics, Mannheim, NY, USA) and Modular Analytics E170 systems (Hitachi Ltd., Tokyo, Japan) Analysis of THs in serum samples was commissioned to SRL Inc (Tokyo, Japan)

The expected reference values in euthyroid humans are within 0.270–4.2μU mL1 for TSH, 0.80–2.0 ng mL1 for TT3, 45–

117 ng mL1for TT4, 2.0–4.4 pg mL1for FT3, and 9.7–17 pg mL1

for FT4(Roche Diagnostics GmbH, 2008)

2.4 Measurements of organic contaminants in serum Serum samples (1–2 g) were denatured with 1 mL of 6 M hy-drochloric acid (HCl), and then spiked with the following surrogate internal standards: 13C12-labeled OH-PCBs (4′OH-CB29, 4′OH-CB61, 4′OH-CB107, 4′OH-CB120, 4′OH-CB159, 4′OH-CB172, and 4OH-CB187),13C12-labeled PCBs (CB28, 52, 95, 101, 105, 118, 138,

153, 156, 167, 170, 178, 180, 189, 194, 202, 206, 208, and 209),13C12 -labeled PBDEs (BDE28, 47, 99, 153, 183, 197, 207, and 209),13C12 -labeled OH-PBDEs (6OH-BDE47, 6′OH-BDE99, and 6′OH-BDE100), and 13C6-labeled BPhs (2,4,6-BPh; 2,3,4,6-BPh; and penta-BPh) The samples were then extracted according to QuEChERS method (Plossl et al., 2006), which entailed the following steps Each sample was poured into a 50 mL fluoroethylenepropylene cen-trifugation tube; added to this were aliquots of 2-propanol (2 mL), methyl tert-butyl ether (MTBE)/hexane (10 mL, 5:5 v/v, Wako, Osaka), and a mixture of MgSO4, NaCl, sodium citrate, and sodium hydrogen citrate sesquihydrate (4, 1, 1, 0.5 g, respectively, Agilent Tokyo), and the tube was shaken vigorously for 1 min by hand After centrifuging the tube at 3000 rpm for 5 min, the supernatant was transferred to a mini-centrifuge tube (15 mL) containing a mixture of C18 and MgSO4(150 mg each, Agilent, Tokyo), mixed for 1 min, and centrifuged at 3000 rpm for 5 min

After transferring the extract to a glass flask, 1 M potassium hydroxide (KOH) in 50% ethanol/water was added and the mixture was shaken for 10 min This organic–alkaline partition process was repeated twice, and the alkaline phases were combined The or-ganic phase was concentrated and passed through an activated silica gel column (Wakogels DX, Wako, Osaka), where PCBs, PBDEs, and MeO-PBDEs were eluted with 80 mL of 5% di-chloromethane (DCM)/hexane (DCM, Wako, Osaka) and were

Table 1 Characteristics of donors from e-waste recycling site and reference site in Vietnam.

e-waste recycling site (Bui Dau)

Reference site (Doung Quang)

Consumption of fish o1 times/

week

1–3 times/

week

Consumption of meat and egg

1–3 times/

week

4–6 times/

week

a Arithmetic mean7Standard deviation.

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concentrated for analysis by high-resolution gas chromatography

(GC; Agilent 6890N) mass spectrometry (MS; JEOL JMS-800D)

(HRGC/HRMS) The alkaline phase was acidified (pH 2) with

sul-furic acid, then OH-PCBs, OH-PBDEs, and BPhs were extracted

twice with 50% MTBE/hexane (60 mL, 5:5, v/v) The organic phases

were passed through deactivated silica gel and then eluted with

50% DCM/hexane OH-PCBs, OH-PBDEs, and BPhs were methylated

by using trimethylsilyldiazomethane (Tokyo Chemical Industry,

Tokyo) Finally, the derivatized solution was passed through an

activated silica gel packed column (WakogelsS1, Wako, Osaka),

eluted with 140 mL of 10% DCM/hexane, and then concentrated

2.5 Quality assurance and quality control

PCBs, OH-PCBs, PBDEs, MeO-PBDEs, OH-PBDEs, and BPhs were

quantified using an isotope dilution method for the corresponding

13C-labeled internal standards The recoveries for these internal

standards in this analytical procedure were within 61–121%, 58–

120%, 43–108%, 79–112%, and 67–116% for OH-PCBs, OH-PBDEs,

BPhs, PCBs, and PBDEs, respectively The signal to noise ratios (S/N)

were higher than 10, and the deviation of ion intensity ratios was

within 15% of those of the standard compounds Procedural blanks

were analyzed simultaneously with every batch of four samples to

check for interferences or contamination from solvent and

glass-ware For each analysis, the mean procedural blank value was used

for subtraction After the blank subtraction, the limit of quanti

fi-cation (LOQ) was defined as the amount of target compound that

resulted in an S/N of 10:1 Only 2,4,6-BPh and BDE-209 were

de-tected in the blank samples, with blank values of the average

concentration values measured in the serum samples being

around 10% and 1%, respectively For quality assurance and control,

our laboratory participated in an inter-calibration exercise

orga-nized by the National Institute of Standards and Technology using

the Standard Reference Material 1958 (NIST SRM 1958; PCBs,

PBDEs, dioxin/furan, pesticides, and PFCs in human serum)

2.6 Statistical analysis

The statistical analyses were performed with R program, Ver

3.0.1 (The R Foundation for Statistical Computing) and EZR

(Sai-tama Medical Center, Jichi Medical University) (Kanda, 2013),

which is a graphical user interface for R (version 2.13.0) The

Wilcoxon rank-sum test was used to determine the difference in

concentrations of organic compounds in samples collected from

the reference site and from the e-waste recycling site

The distributions of TH, PCBs, PBDEs, and their related

com-pounds were assessed for normal distribution (Shapiro–Wilk test)

FT4and TT4were normally distributed in Bui Dau, Duong Quang,

and mixed locations (Bui Dau and Duong Quang) However, TSH,

FT3, TT3, and all PCB, OH-PCB, PBDE, OH-PBDE, MeO-PBDE, and

BPh congeners were not normally or log-normally distributed in

all locations

We implemented stepwise generalized linear models (GLMs) to

assess the association between levels of TH and those of PCBs,

PBDEs, and their related compounds This model was also used to

examine the influence of other characteristics: perchlorate, iodide,

thiocyanate (Eguchi et al., 2014), cholesterol, triglyceride,γ-GTP,

BMI, age, gender (male¼0, female¼1), living site (reference¼0,

e-waste¼1), meat and egg consumption (1–3 times/week¼0, 4–6

times/week¼1), and fish consumption (o1 times/week¼0, 1–3

times/week¼1) We also determined stepwise GLMs to assess the

associations between PCBs, PBDEs, and their related compounds

and characteristics The results obtained with po0.05 were

con-sidered statistically significant The data for TT4 and FT4 were

normally distributed in serum; hence, these data were analyzed by

GLMs with a Gaussian distribution of the response variable and

identity-link function The concentrations of TSH, TT3, and FT3

were not normally distributed; hence, these data were analyzed by GLMs with a gamma distribution of the response variable and log link function

For any model, the parameters that optimize the approximation

of the likelihood can be found numerically The optimized like-lihoods from different models can then be compared through Akaike’s information criterion (AIC) to maximize the model’s likelihood (Akaike, 1998) Selected models and their parameters are shown inTables S5–S16

3 Results and discussion 3.1 Residue profiles of PCBs and OH-PCBs in human serum Concentrations of PCB and OH-PCB congeners in serum of do-nors from e-waste recycling and reference sites are shown in Table 2 Total PCB levels were significantly higher (po0.05) in donors from the e-waste recycling site (420 pg g1) than from the reference site (290 pg g1) (Table 2) Concentrations of PCBs and OH-PCBs in the present study were lower than those reported in Japan (PCBs: 2900 pg g1, OH-PCBs: 630 pg g1;Nomiyama et al.,

2010), Romania (PCBs: 3100 pg g1, OH-PCBs: 175 pg g1; Dirtu

et al 2010), Belgium (PCBs: 3380 pg g1, OH-PCBs: 310 pg g1; Dirtu et al 2010), and Sweden (PCBs: 3560 pg g1, OH-PCBs

2670 pg g1;Sjodin et al., 2000) but higher than those reported in India (PCBs: 110 pg g1, OH-PCBs: 54 pg g1;Eguchi et al., 2012) Concentrations of total OH-PCBs were significantly higher (po0.05) in donors from the e-waste recycling site (160 vs

88 pg g1; Table 2) At both sites, congener profiles for OH-PCBs were dominated by 4′OH-CB107/4′OH-CB108 followed by 4OH-CB146, 4OH-CB101/120, and 4OH-CB187 (46.5%, 9.5%, 10.5% and 9.0% for Bui Dau and 29.5%, 13.6%, 11.3% and 14.8% for Duong Quang, respectively)

PCB congener profiles were different in each studied popula-tion Dominant PCB congeners in each population were CB153 (17.7% in Bui Dau; 29.3% in Duong Quang) and CB138 (29.3% in Bui Dau; 18.3% in Duong Quang) Levels of less-chlorinated PCB con-geners, including CB28, CB74, CB105, and CB118, were higher at the e-waste recycling site than the reference site, consistent with our previous results on breast milk collected from Bui Dau (Tue

et al., 2010a) Specific exposure to some of these more volatile congeners could be explained by release from old capacitors in e-waste (Tue et al., 2013) reported high levels of CB74 in serum (Freels et al., 2007), and relatively high levels of CB74 and lower levels of CB138 and CB180 were found in serum collected from capacitor manufacturing workers (Wolff et al., 1992)

Among OH-PCBs, 4OH-CB107/4′OH-CB108 was the dominant metabolite at the reference site, followed by CB146, 4OH-CB187, and 4′OH-CB101/120 Several studies have reported that 4OH-CB107 was the dominant congener in human serum (Dirtu

et al., 2010;Eguchi et al., 2012;Fangstrom et al., 2002;Nomiyama

et al., 2010;Sandau et al., 2000;Sjodin et al., 2000); however, in Swedish and Slovakian populations, 4OH-CB187 was the dominant metabolite (Bergman et al., 1994;Park et al., 2009a) This meta-bolite is probably formed via the metabolism of CB183 and/or CB187, which represent 2% and 5% of the Aroclor 1254 mixture An entirely different profile was reported for wives of Swedish fish-ermen, with 4OH-CB146 being the main congener, followed by 4OH-CB107 and 4OH-CB187 (Weiss et al., 2006) Differences in OH-PCB profiles in human serum may reflect regional variations in using technical PCBs (Breivik et al., 2002a,2002b;Ishikawa et al.,

2007)

We also assessed associations between PCB and OH-PCB con-centrations and various characteristics using stepwise GLMs (Table S3)

A Eguchi et al / Environmental Research 137 (2015) 440–449 442

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Concentrations of PCBs, OH-PCBs, PBDE, OH-PBDEs, MeO-PBDEs, and BPhs ands in serum of donors from the reference and e-waste recycling sites in Vietnam (pg g wet wt).

PCBs

Total PCBs b

OH-PCBs

Total OH-PCBs c

OH-PCBs/PCBs ratio

PBDEs

OH-PBDEs

MeO-PBDEs

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The Concentrations of PCBs and OH-PCBs positively correlated with age and negatively correlated withfish consumption (o1 times/week¼0, 1–3 times/week¼1) Fish consumption has been positively correlated with PCB concentration (Sjodin et al., 2000) However, we observed an opposite trend, unclear reasons; how-ever, there may be some bias related to less people eating more fish (herein, DQ¼6 and BD¼4) Other characteristics were not significantly associated with PCB or OH-PCB concentrations When concentration ratios of OH-PCBs to their possible parent PCB congeners were calculated, ratio of 4OH-CB107þ108/ CB105þ118 was markedly higher in donors from the e-waste re-cycling site than in those from the reference site (Table 2), sug-gesting that CYP1A1 upregulation was because of exposure to in-door dust containing “high levels of dioxin-related compounds released from e-waste recycling (Tue et al., 2010b)

3.2 Residue profiles of PBDEs, MeO-PBDEs, OH-PBDEs, and BPhs in human serum

PBDE, MeO-PBDE, OH-PBDE, and BPh concentrations are shown

inTable 2 PBDE and BPh concentrations were significantly higher

in e-waste recycling workers than in residents at the reference site, according to GLM results (reference¼0, e-waste¼1) (Table S4) Other characteristics did not markedly associate with PBDE or BPh concentrations

Significantly higher levels (po0.05) of PBDEs were found in serum samples from the e-waste recycling site (290 pg g1) than from the reference site (230 pg g1) PBDE concentrations at the e-waste recycling (290 pg g1) and reference sites (230 pg g1) were similar to or higher than in Indian e-waste recycling workers (150 pg g1; Eguchi et al., 2012), Swedish rubber workers (400 pg g1;Thuresson et al., 2005), and electronics dismantlers (210 pg g1; Sjodin et al., 1999), indicating that PBDE levels in Vietnamese subjects were comparable with these occupational workers but lower than e-waste recycling workers in China (930–

4800 pg g1;Jin et al., 2009;Zhao et al., 2010) (Fig 1B)

BDE47 was the dominant congener in all PBDE samples, followed by BDE99 and BDE209 (55%, 30%, and, 7.2%, respec-tively) BDE153 concentrations at each location were relatively lower than other reports These profiles may reflect higher con-sumption of deca-BDE technical mixtures in Asia (Watanabe and Sakai, 2003)

Significantly higher (po0.05) OH-PBDE and MeO-PBDE con-centrations were detected at the reference (160 and 20 pg g1, respectively) than at e-waste recycling sites (43 and 0.52 pg g1, respectively), with unclear reasons; however, numbers of children and detailed information on food intake may be important OH-PBDE concentrations found in the present study were generally lower than those found in fetal and maternal serum samples from the United States (290 pg g1; Qiu et al., 2009), serum samples from children living near a waste disposal site in Nicaragua (260 pg g1;Athanasiadou et al., 2008), and serum samples from Japanese volunteers (450 pg g16OH-BDE47;Fujii et al., 2014) but higher than those in donors from India (30 pg g1;Eguchi et al.,

2012) Our results differ compared with previous reports, possibly because of differences in exposure levels caused by dietary habits

Of the 28 OH-PBDE (tri- to octa-) isomers monitored in this study, 6OH-BDE47 and 2′OH-BDE68 were dominant at each location These congeners have been predominantly detected in several marine and terrestrial animals (Malmberg et al., 2005; Mizukawa et al., 2013;Nomiyama et al., 2011a;Wan et al., 2009), suggesting that dietary intake was an primary exposure route for these congeners (Eguchi et al., 2012;Valters et al., 2005;Wan

et al., 2010)

Congener profiles for BPhs were dominated by 2,4,6-BPh (99% and 87% for Duong Quang and Bui Dau, respectively) Total BPh

BPhs 2,4,5-BPh

n Signi

a Concentr

c Including

A Eguchi et al / Environmental Research 137 (2015) 440–449 444

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concentrations were significantly (po0.05) higher at the e-waste

recycling (300 pg g1) than in donors from reference sites

(200 pg g1) The predominance of 2,4,6-tri-BPh, even in donors

living at the reference site (Table 2), may reflect potential for

ex-posure from both dietary and non-dietary indoor sources This

congener has been regarded as a natural product in several marine

organisms (Malmberg et al., 2005;Marsh et al., 2004;Nomiyama

et al., 2011a,2011b;Wan et al., 2009) and is also manufactured at

high volumes (9500 t/year in 2001) for use asflame retardants and

wood preservatives/fungicides (Watanabe and Sakai, 2003)

Contrastingly, 2,3,4,6-tetra-BPh, 2,4,5-tri-BPh,

2,3,5,6-tetra-BPh, and penta-BPh were more prominent at the e-waste recycling

site (4.5%, 2.8%, 2.3%, and 2.2%, respectively) (Table 2) Total levels

of non-2,4,6-BPhs in donors from the e-waste recycling site

(36 pg g1) were significantly higher (po0.05) than at the

re-ference site (median: 0.27 pg g1), indicating that accumulation of

these BPhs is related to exposure to e-waste contaminants

2,4,5-tri-BPh has been detected as a metabolite of BDE99 in human

hepatocytes in vitro (Stapleton et al., 2009), possibly resulting from

simple cleavage at the ether bond Further, three types of BPh

metabolites have been identified in mice blood after exposure to

PBDEs (Qiu et al., 2007) In this study, 2,3,4,6-tetra-BPh and BDE209 showed a weak (but significant) positive correlation (po0.05) Increased exposure to PBDEs from e-waste recycling and/or inhalation from house dust containing these contaminants may have contributed to higher levels of these BPh metabolites (Suzuki et al., 2008)

3.3 Association between thyroid hormone concentrations and PCBs, PBDEs, and related compounds

TH concentrations in most samples were within normal limits However, FT3, TT3, and TT4concentrations were significantly lower

in serum samples from the e-waste recycling site (3.3 pg g1, 1.2 ng g1, and 78 ng g1, respectively) than from the reference site (3.5 pg g1, 1.3 ng g1, and 85 ng g1, respectively;Table 3) Serum TT4and TT3concentrations in females were significantly higher (po0.05) than in males (Table 3) Sex differences may be related to hormone balance Soldin et al (2004) reported that estrogen induced serum thyroxine-binding globulin (TBG), the main transport protein for THs in humans, followed by increased serum TT4 concentration, whereas androgen therapy reduced

Fig 1 Comparison of serum concentrations of (A) PCBs and (B) PBDEs in Vietnamese donors analyzed in this study with previous data References: (1) Eguchi et al (2012) ; (2) Zhao et al (2010) ; (3) Nomiyama et al (2010) ; (4) Dirtu et al (2010) ; (5) Sjodin et al (2000) ; (6) Sjodin et al (1999) ; (7) Thuresson et al (2005) ; and (8) Jin et al (2009)

n Note: (1), (2), (3), (4), (5), (6), (7)¼Median value, (8)¼Mean value (SD was not shown in reference) nn PCB and PBDE concentrations were converted to wet basis using serum lipid levels of 7.9 g L1as applied by Longnecker et al (2003)

Table 3

Concentrations of thyroid hormones (pg g 1 wet wt), γ-GTP (IU L 1 ) and lipids (mg dL 1 ) in serum of donors from reference and e-waste sites, Vietnam.

Reference area: Duong Quang (n¼34) e-waste recycling area: Bui Dau (n¼77)

Mean SD a Min 1st quartile Median 3rd quartile Max Mean SD a Min 1st quartile Median 3rd quartile Max

a

SD standard deviation.

n Significantly higher than in the other group (po0.05: Wilcoxon rank sum test).

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Table 4

Coefficients of multiple linear regression of thyroid hormone levels with contaminant levels as controlling factors n

All donors

Total OH-PCBs a

Male

Female

Total PCBs c

Total BPhs d

Total concentrations: refer to Table 1 for further details.

n These models report the contaminants–thyroid hormone association after adjusting for perchlorate, iodide, thiocyanate, cholesterol, triglyceride, γ-GTP, BMI, age, gender, living site, consumption of meat and egg and consumption of marine fish Blank cells indicate factors eliminated by stepwise procedures.

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serum TBG and TT4levels Our results revealed that females had a

significant positive correlation between FT4and most PCB and

OH-PCB congeners (r¼0.23–0.56, po0.05), but only CB138 and CB153

(r¼0.21–0.27, po0.05), positively correlated with FT4 in males

(Table 4) TT3, FT3, and TT4had a significant positive association

with hepta-chlorinated PCBs and some OH-PCB congeners in

fe-male serum samples: TT3 with CB187, FT3 with 4OH-CB79 and

4OH-CB172, and TT4 with 4OH-CB101/120 TSH demonstrated

significant negative correlation with CB74, CB118, and the sum of

the non-2,4,6-BPhs in female serum samples One possible

me-chanism involved in TH homeostasis disruption is the competitive

binding of OH-PCBs and BPhs to TH transport protein transthyretin

(TTR) found in blood (Cheek et al., 1999;Londono et al., 2010;Park

et al., 2009a, 2009b) Furthermore, binding affinity of OH-PCBs,

OH-PBDEs, and BPhs with TTR has been comparable to that of THs

(Marchesini et al., 2008;Ucan-Marin et al., 2010) However, serum

TH concentrations did not significantly correlate with PBDEs,

OH-PBDEs, 2,4,6-BPh, or total BPhs

We observed significant positive correlations between PCB and

OH-PCB concentrations and FT4and negative correlations between

lower-chlorinated PCB concentrations and TSH in females;

how-ever, few compounds significantly correlated with THs in males

(Tables 4,S5–S16) Our results also suggest that gender differences

exist regarding the effects of organochlorines on THs Previous

studies also found associations between POPs and THs in women

Chlorinated POPs, including PCBs and levels of T3and T4,

nega-tively correlated in females, particularly pregnant women

(Chevrier et al., 2008;Koopmanesseboom et al., 1994;Takser et al.,

2005)

In vivo studies with rats have reported that TT4and TT3levels

significantly decreased in Aroclor 1254- and 4OH-CB107-exposed

groups, whereas FT4 levels did not significantly change (Meerts

et al., 2002) Many epidemiological studies examining associations

between PCBs and thyroid function have reported no association

with FT4(Hagmar et al., 2001;Hallgren et al., 2001;Meeker et al.,

2007;Sala et al., 2001) However, some studies reported positive

and negative associations with FT4 It has also been reported that

DLCs have altered thyroid hormone homeostasis (Chevrier et al.,

2008;Koopmanesseboom et al., 1994;Takser et al., 2005) e-waste

recycling workers from Bui Dau are occupationally exposed to

dioxin-like compounds and trace elements during recycling

ac-tivities (Noguchi et al., 2014;Tue et al., 2010a,2010b,2013)

Ex-posure to dioxin-like compounds positively associated with FT4

(Calvert et al., 1999), indicating that exposure to dioxin-like

com-pounds was reflected by decreasing FT4levels Workers exposed to

PCBs and chlorinated napthalenes at an electrical capacitor

man-ufacturing facility in Illinois (Persky et al., 2001) and fish

con-sumers with exposure to organochlorines living near the Great

Lakes (Turyk et al., 2006) had decreased TSH levels TH production

is controlled by a complex mechanism of positive and negative

regulation Although TSH stimulates TH secretion, TH suppresses

TSH Feedback between these two hormones allows TH levels to be

maintained within a narrow dose range (Vandenberg et al., 2012)

Although sample size was limited in this study, negative feedback

effect may have been detected following low-dose exposure to

PCBs and their metabolites using applicable statistical analyses

Some PCBs, OH-PCBs, and BPhs significantly correlated with

FT3, TT3, TT4, and TSH, respectively, in the present study

Asso-ciations between organohalines and TT4and/or TT3were reported

to be significantly negative in half of the studies cited in a review

(Abdelouahab et al., 2008); however, remaining studies found no

association with these hormones An in vivo study showed that

PCBs reduced serum FT4 and TT4 in a dose-dependent manner

(Martin and Klaassen, 2010) In this study, residual levels of PCBs

and OH-PCBs in serum were one order lower than those of

epi-demiological studies (1700–6700 pg (Hagmar et al., 2001;Hallgren

et al., 2001;Meeker et al., 2007;Sala et al., 2001)), and three to four orders lower than with in vitro TTR inhibition assays (IC50: OH-PCBs 7.3–32.7 nM;Marchesini et al., 2008)

Previous studies have demonstrated that OH-PBDEs compete with THs for binding sites on human plasma proteins Based on in vitro studies using human cells, IC50range for OH-PBDEs was from

22–110 nM for TTR and 100–870 nM for TBG, depending on the congener (Marchesini et al., 2008) OH-PBDE concentrations in serum from Vietnamese populations were two to three orders of magnitude lower than the IC50value for TT4–TTR binding activity There were no marked associations between THs and PBDEs or OH-PBDEs in this study

Several researches have also reported that associations be-tween PBDEs and TT3, TT4, FT3, FT4, and TSH were not clear (Mazdai et al., 2003; Roze et al., 2009) A recent study showed negative associations between PBDE levels and TT3and FT3in cord blood (Lin et al., 2011) and positive associations between PBDEs and TSH levels in pregnant women (Zota et al., 2011) Another study reported positive associations between PBDE and OH-PBDE concentrations and TSH (Zota et al., 2011)

PCBs, PBDEs, and their metabolites may disrupt TH homeostasis

in pregnant women and negatively affect neurological develop-ment of fetuses (Roze et al., 2009;Herbstman et al., 2010) Thus, detailed investigations on their exposure effects for pregnant women and the health effects for neonates are required

In this study, we used a generalized linear model to examine associations between TH and other compounds, including per-chlorate, iodide, and thiocyanate (Eguchi et al., 2014;Tables 4,S5– S16) Consequently, iodide and thiocyanate levels were found to contribute to TH variations (Tables S5–S16) Iodide is essential for

TH production (Bianco et al., 2002) Thiocyanate can competitively inhibit iodide uptake by the thyroid gland via the sodium/iodide symporter, reducing TT3 and TT4 synthesis (Dohan et al., 2007; Tonacchera et al., 2004) It has been suggested that iodide and thiocyanate were covariates in the association between PCBs and OH-PCBs with THs, indicating that analysis of these anions is ne-cessary to elucidate effects of TH disruption

In summary, we determined serum concentrations of organo-halogen compounds and their metabolites and examined asso-ciations with THs in Vietnamese populations PCB, OH-PCB, PBDE, and BPh concentrations in serum samples from the e-waste re-cycling site were significantly higher than in those from the rural site, indicating occupationally related exposure to these com-pounds Concentrations of these contaminants were lower in Vietnam than in the US and European countries; however, PCB and OH-PCB levels correlated with serum TH concentrations in Viet-namese females Therefore, detailed investigations on exposure effects in females are required; particularly those focused on pregnant women and health effects in developing newborns

Acknowledgment This study was supported by Grants-in-Aid for Young Scientists (A) (No 25701014), Grants-in-Aid for Scientific Research (A) (No 25257403) and Grant-in-Aid for Research Activity start-up (No 26881003) from the Japan Society for the Promotion of Science (JSPS), Japan, the Global Center of Excellence (COE) Program of the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan, and the Environment Research and Technology Development Fund (K2311 and 3K133001) from the Japanese Ministry of the Environment We also acknowledge the JSPS Re-search Fellowships for Young Scientists (DC1 and PD) in Japan that were provided to Dr A Eguchi (Nos 22-6331 and 25-6617)

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Appendix A Suplementary information

Supplementary data associated with this article can be found in

the online version athttp://dx.doi.org/10.1016/j.envres.2015.01.007

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