Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, hanoi—vietnam contribution of polycyclic aromatic hydrocarbons (PAHs) and human risk assessment

STOTEN-15803; No of Pages Science of the Total Environment xxx (2014) xxx–xxx Contents lists available at ScienceDirect Science of the Total Environment journal homepage: www.elsevier.com/locate/scitotenv Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic hydrocarbons (PAHs) and human risk assessment Le Huu Tuyen a,b, Nguyen Minh Tue a,b, Go Suzuki c, Kentaro Misaki a, Pham Hung Viet b, Shin Takahashi a,d,⁎, Shinsuke Tanabe a a Center for Marine Environmental Studies (CMES), Ehime University, 2-5 Bunkyo-cho, Matsuyama, Japan Research Centre for Environmental Technology and Sustainable Development (CETASD), Hanoi University of Science, 334 Nguyen Trai Street, Hanoi, Viet Nam c Center for Material Cycles and Waste Management Research, National Institute for Environmental Studies (NIES), 16-2 Onogawa, Tsukuba, Japan d Center of Advanced Technology for the Environment, Agricultural Faculty, Ehime University, 3-5-7 Tarumi, Matsuyama, Japan b H I G H L I G H T S • • • • First assessment of total AhR-mediated toxic activities in road dust using DR-CALUX PAHs known as carcinogens were found at high levels in Hanoi PAHs contributed only 0.8–76% to AhR agonist activities in road dust Exposure to PAHs in road dust may pose high cancer risk for Hanoi residents a r t i c l e i n f o Article history: Received 15 November 2013 Received in revised form 23 January 2014 Accepted 23 January 2014 Available online xxxx Keywords: Ah-receptor Dioxins PAHs CALUX Road Dust Vietnam a b s t r a c t Dioxin-Responsive Chemical-Activated LUciferase gene eXpression assay (DR-CALUX) was applied to assess the total toxic activity of the mixture of PAHs and related compounds as well as dioxin-related compounds in road dust from urban areas of Hanoi, Vietnam Road dust from Hanoi contained significantly higher DR-CALUX activities (3 to 39, mean 20 ng CALUX-TEQ/g dw) than those from a rural site (2 to 13, mean ng CALUX-TEQ/g dw) The total concentrations of 24 major PAHs (Σ24PAHs) in urban road dust (0.1 to 5.5, mean 2.5 μg/g dw) were also times higher than those in rural road dust (0.08 to 1.5, mean 0.4 μg/g dw) Diagnostic ratios of PAHs indicated vehicular engine combustion as the major PAH emission source in both sites PAHs accounted for 0.8 to 60% (mean 10%) and to 76% (mean 20%) of the measured CALUX-TEQs in road dust for Hanoi the rural site, respectively Benzo[b]-/benzo[k]fluoranthenes were the major TEQ contributors among PAHs, whereas DRCs contributed b0.1% to CALUX-TEQs for both rural and urban sites These results suggest TEQ contribution of other aryl hydrocarbon receptor agonists in road dust Significant PAH concentrations in urban dust indicated high mutagenic and carcinogenic potencies Estimated results of incremental life time cancer risk (ILCR) indicated that Vietnamese populations, especially those in urban areas such as Hanoi, are potentially exposed to high cancer risk via both dust ingestion and dermal contact This is the first study on the exposure risk of AhR agonists, including PAHs and DRCs, in urban road dust from a developing country using a combined bio-chemical analytical approach © 2014 Elsevier B.V All rights reserved Introduction Industrialization, urbanization and high economic growth in recent years are accompanied by degradation of environmental quality in developing countries, where the pollution of the ambient air in large cities ⁎ Corresponding author at: Center of Advanced Technology for the Environment, Agricultural Faculty, Ehime University, 3-5-7 Tarumi, Matsuyama, Japan Tel./fax: +81 89 927 8171 E-mail address: shint@agr.ehime-u.ac.jp (S Takahashi) has become a major concern Located in the Red River delta, Hanoi, Vietnam's capital is one of the biggest cities in Asia with over 3.4 million inhabitants in the city proper and its urban districts Hanoi urban area consists of urban districts that cover 918.5 km2 in a total of 3344 km2 of the Hanoi metropolitan area Among the six thickly populated Asian cities including Vietnam (Hanoi), Bandung (Indonesia), Bangkok (Thailand), Beijing (China), Chennai (India), and Manila (Philippines), Hanoi has been reported as the most polluted city by dust with the mean of atmospheric particulate matter in the range of 18–168 (PM2.5) and 33–262 μg m−3 (PM10 − 2.5) and frequently exceeded the 0048-9697/$ – see front matter © 2014 Elsevier B.V All rights reserved http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx corresponding 24-h U.S EPA standards (65 and 150 μg m− 3) (Oanh et al., 2006) The hourly average of suspended particulate matter in Hanoi has been reported by yet another study as 0.4–1.5 mg/m3 exceeding the Vietnamese standard of 0.2 mg/m3 (Saksena et al., 2006) According to the Ministry of Natural Resources and Environment, the dust density on Hanoi streets and roads is in the range of 20–40 g/m2/ year, especially with higher values on the ring roads with figures up to 100–400 g/m2/year whereas 10 g/m2/year is the normal average in developed countries The traffic in Hanoi has been reported as the most important contributor of particulate matter emission (Oanh et al., 2006) A previous study on the traffic volume has been performed using video camera recording in the year 2004 and the results showed that there were around 10,000 motorbikes passed a selected point per hour This traffic volume has reached over 18,000 motorcycles in 2005 Additionally, the number of cars and light trucks has also increased (Yen et al., 2007) In highway, the motorcycles' density is 9100 vehicles per hour Meanwhile the corresponding values are 13,600 and 3500 vehicles per hour for the arterial and residential areas, respectively (Oanh et al., 2012) Moreover, diesel vehicles were reported as the highest contributors of particulate matter in Hanoi (Oanh, 2009) Air particle pollution by vehicles has been associated with polycyclic aromatic hydrocarbons (PAHs), a major source of which is vehicle exhaust, and high levels of PAHs in road dust from Asian developing countries were reported (Wang et al., 2009; Boonyatumanond et al., 2007; Lee et al., 2001) Road dust is also a sink for complex mixtures of traffic-related pollutants (Keller and Lamprecht, 1995) and may contain aliphatic hydrocarbons, PAHs and their derivatives, as well as other organic substances (Bodzek et al., 1993; Yassaa et al., 2001; Lee et al., 2001) However, contaminations by traffic-related pollutants including PAHs in Hanoi road dust and related human health risk have not been investigated comprehensively PAHs are well-known for their carcinogenic, mutagenic and teratogenic characteristics and they can cause various toxic impacts to human and wildlife (Behnisch et al., 2001; Sjiigren et al., 1996; Poland et al., 1982; Senft et al., 2002; Nebert et al., 2004) Their toxicities usually involve a common mechanism such as binding to the aryl hydrocarbon receptor (AhR), induction of AhR-related genes and subsequent transformation to toxic metabolites (Behnisch et al., 2001) AhR has been associated with tumor-promotion and enhanced oxidative stress (Sjiigren et al., 1996; Poland et al., 1982; Senft et al., 2002; Nebert et al., 2004) Not only PAHs but also many of their derivatives occurring in ambient environment, such as methylated and oxygenated compounds, have been reported to transactivate AhR (Trilecová et al., 2011; Sonneveld et al., 2007; Sovadinová et al., 2006) Moreover, some other chemicals often occur in the environment at low concentrations compared with PAHs, but with a high AhR-mediated activity (e.g dioxinrelated compounds) It is important to know not only concentration levels of organic compounds in the environment but also their toxicities to evaluate the integrated risk for human health effects and environmental risk assessment Regarding this matter, reporter gene assays such as DRCALUX have been known as a useful method to evaluate the AhRmediated toxicities of contaminants in the environment (Behnisch et al., 2003; Machala et al., 2001) Therefore, an approach that aims to establish a causal link between chemical substances and biological effects in environmental samples is necessary to assess the total toxic activity of the mixture of PAHs and related compounds released from traffic-related processes, in addition to the conventional chemical analysis of PAHs This study analyzed road dust collected from Hanoi, a city with the highest levels of traffic-related air pollution in East Asia (Oanh et al., 2006) and Duong Quang as a rural reference site, to investigate the contamination status by AhR agonists including not only the well-known PAHs and dioxin-related compounds (DRCs) but also other potential compounds The AhR-mediated toxic activities were evaluated using Chemical-Activated LUciferase gene eXpression (CALUX) assays PAHs and DRCs were also analyzed to determine their contribution to the overall toxic activities Finally, US-EPA incremental lifetime cancer risk model were applied to identify the human health risk for PAH exposure to road dust via ingestion and dermal contact Material and methods 2.1 Sampling site description In recent years, as a result of its economic growth, infrastructure in Hanoi, capital of Vietnam has seen considerable change Though urban and suburban road extension has been marginal, traffic is generally increasing, leading to higher traffic density and congestion According to the Transport Police Department of Hanoi, registered motorcycles in Hanoi are increasing at ~13.5% per year and car ownership at ~10% a year Duong Quang, a mostly agricultural commune located at approximately 50 km to the east of Hanoi in My Hao district, Hung Yen province, was chosen as the reference site The population of this commune was approximately 6500, and motorbikes were the main transport vehicles 2.2 Sample collection We collected 24 road dust samples from Hanoi urban area and samples from Duong Quang in January 2011 (Fig S1) Each dust sample of approximately 300 g was collected from an area of 20–50 m in length and 0.5 m in width at the side of the road using straw broom After collection, the samples were preserved at −25 °C until analysis 2.3 Sample pre-treatment and extraction Road dust samples were air-dried, and then sieved through a 500 μm stainless sieve to remove coarse particles Two grams of the sample was extracted with an acetone/hexane mixture and then toluene using a rapid solvent extractor (SE100, Mitsubishi Chemical Analytech, Japan) according to a previously reported method (Tue et al., 2010) A 0.2 g-equivalent portion of the crude extract was then concentrated, solvent-exchanged into 0.1 ml biochemical-grade dimethyl sulfoxide (DMSO) and stored at °C for in vitro determination of AhR-mediated activities using DR-CALUX assay The remaining extract was used for chemical analysis of PAHs Every set of seven samples was accompanied with a procedural blank 2.4 DR-CALUX The AhR-inducing potencies of the crude extracts were determined by the DR-CALUX assay This method assays utilize the rat hematoma cell line H4IIE (BioDetection Systems, The Netherlands) stably transfected with the firefly luciferase gene containing a multimerized dioxin response element in front of a minimal promoter All assays were performed following the BioDetection Systems' protocol described elsewhere (Suzuki et al., 2004, 2006) Briefly, 80,000 cells/well were seeded on 96well plates After 24 h of incubation at 37 °C and 5% CO2, the cells were treated with exposure medium contained DMSO solutions (0.8% DMSO in final wells) of either the reference standard 2,3,7,8-tetrachlorobenzop-dioxin (TCDD, to 37.5 nM) or the samples (diluted by a factor of to 1000) After another 24 h of incubation, the cells were subjected to luminescence measurement The AhR agonist activities were derived from the diluted samples with similar response to 1–3 pM TCDD (usually 300 to 1000 time dilution), and expressed in amounts of TCDD equivalent (CALUX-TEQ) per gram dry weight (dw) Each experiment was done in triplicate No significant effect on cell viability was observed in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (Suzuki et al., 2013) for these diluted samples 2.5 Chemical analysis The extract for PAH analysis was spiked with deuterated PAH surrogate standards and cleaned-up using 1.2% deactivated alumina Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx chromatography (4 g, eluted with 80 ml of dichloromethane/hexane 1:1 v/v), activated silica gel chromatography (4 g, eluted with 80 ml of dichloromethane/hexane 5:95 v/v) and gel permeation chromatography (described in Tue et al., 2010), and then finally spiked with chrysen-d12 as internal standard Naphthalene (Nap), acenaphthylene (Acy), acenaphthene (Ace), fluorene (Flu), phenanthrene (Phe), anthracene (Ant), fluoranthene (Fluh), pyrene (Pyr), benzo[c]phenanthrene (B[c] Ph), cyclopenta[c,d]pyrene (CPP), benz[a]anthracene (B[a]A), chrysene (Chy), benzo[b]-, benzo[k]-, and benzo[j]fluoranthene (B[b]F, B[k]F and B[j]F), 7,12-dimethylbenz[a]anthracene (DMBA), benzo[e]- and benzo [a]pyrene (B[e]P and B[a]P), 3-methylcholanthrene (MCA), indeno [1,2,3-c,d]pyrene (I(IDP + B[g]P), dibenz[a,h]anthracene (DBA), benzo [g,h,i]perylene (B[g]P), dibenzo[a,h]-, dibenzo[a,i]-, dibenzo[a,l]pyrene (DB[ah]P, DB[ai]P and DB[al]P were determined using a gas chromatograph (Agilent 5975C Series) connected to a mass spectrometer with electron-impact ionization DRCs such as polychlorinated and polybrominated dibenzo-p-dioxins/dibenzofurans (PCDDs/Fs and PBDDs/Fs) and dioxin-like PCBs (DL-PCBs) were analyzed using a separate extract according to a method described elsewhere (Tue et al., 2010) 2.6 Cancer risk assessment The incremental lifetime cancer risk (ILCR), adopted from U.S EPA (1991) and modified in a previous study (Chen and Liao, 2006; Wang et al., 2011; Peng et al., 2011), was used to quantitatively estimate the risk of exposure to PAHs in road dusts Eqs (1) to (2) was applied to evaluate cancer risk of adults and children exposed to PAHs via ingestion and dermal contact, respectively ILCRsIngestion ¼ ILCRsDermal ¼  pffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi CS  CSFIngestion  BW=70  IRIngestion  EF  ED BW  AT  106 Table AhR-mediated, mutagenic and carcinogenic potencies Congeners Nap Acy Ace Flu Phe Ant Fluh Pyr B[c]Ph CPP B[a]A Chy B[b]F + B[k]F B[j]F DMBA B[a]P B[e]P MCA IDP DBA B[g]P DB[ah]P DB[ai]P DB[al]P AhR-mediated potency Mutagenic potency Carcinogenic potency Relative to TCDD Relative to B[a]P Relative to B[a]P – – – – 1.3E−06a – 2.3E−08b 0.00033c 4.5E−07b 6.53E−07b 0.000143a 0.00033c 0.013c 0.00037b 5.4E−06d 0.00053b 5.2E−07c – 0.002487a 0.0043c – 0.000071b 0.00017b 4.9E−06b – 0.00056 – – – – – – 0.05 6.9 0.08 0.01 0.36 0.26 – – – 0.31 0.29 0.19 1.4 3.6 24 – – – – 0.0005 0.0005 0.05 0.001 0.023 0.02 0.005 0.03 0.15 0.05 – 0.002 – 0.1 1.1 0.02 0.1 Mutagenic potencies cited from Durant et al (1996); Carcinogenic Potencies cited from Larsen and Larsen (1998) a Cited from Behnisch et al (2003) b Cited from Machala et al (2001) c Cited from Suzuki (2005, unpublished) d Cited from Marvanová et al (2008) ð1Þ  pffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi CS  CSFDermal  BW=70  SA  AF  ABS  EF  ED BW  AT  106 ð2Þ where CS is the B[a]P-equivalent concentration of PAHs in dust (μg kg− 1, calculated using equivalency factors in Table 1), CSFIngestion and CSF Dermal , are carcinogenic slope factors of B[a]P (7.3, and 25 mg kg− d− , respectively (Knafla et al., 2006; US EPA, 1994, 2011)), BW is the body weight (average 57.7 and 16.8 kg for adults and children, respectively (Walpole et al., 2012; Dang et al., 2010)), AT is the average life span (70 years), EF is the exposure frequency (180 days year− (Ferreira-Baptista and De Miguel, 2005)), ED is the exposure duration (30 and years, respectively (US EPA, 2004)), IRIngestion is the dust intake rate (50 and 100 mg day− for adults and children, respectively (US EPA, 2011)), AF is the dermal adherence factor (0.07 and 0.2 mg cm− h− for adults and children, respectively (US EPA, 2004)), SA is the dermal surface exposure (5700 and 2800 cm2 day− for adults and children, respectively (US EPA, 2004)), and ABS is the dermal adsorption fraction (0.13 (US EPA, 2004)) The total risks were the sum of risks associated with all exposure routes Results and discussion 3.1 CALUX-TEQs AhR-mediated activities were detected in all the road dust samples analyzed with DR-CALUX (see examples of dose–response curves in Supplementary Fig S2) Significant higher CALUX-TEQs in road dust were found for Hanoi than for the rural site Duong Quang (3 to 39, mean 20 ng/g dw compared with to 13, mean ng/g dw, p b 0.005) (Fig 1) When compared with the results of in vitro AhR-mediated activities reported in a number of studies on environmental samples, the CALUX-TEQs in Hanoi road dust were higher than those of Vietnamese settled house dust (median: 12 ng CALUX-TEQ/g dw) (Tue et al., 2012) and of flood-resuspended dust in Germany (7 ng TEQ/g dw, Ethoxyresorufin-O-deethylase assay) (Wölz et al., 2010) The CALUXTEQs in Duong Quang and Hanoi road dust were 14 and 56 times higher than that of a household sewage sludge compost crude extract from Japan (0.36 ng CALUX-TEQ/g dw) (Suzuki et al., 2004), suggesting that road dust in Vietnam, especially from the urban site, contained large amounts of AhR agonists (Fig 1) 3.2 PAH concentrations 2.7 Statistical analysis The R software packages version 2.15.2 were used to perform statistical analyses The Wilcoxon rank sum test was used to examine the significance of the difference in levels between Hanoi and Duong Quang Pearson's correlation analysis was applied to check the relationships between PAH concentrations and toxicity levels (log-transformed values) A p value of b 0.05 is considered as indicating statistical significance Significantly higher concentrations of Σ24PAHs were found in Hanoi road dust (0.1 to 5.5, mean 1.5 μg/g dw) than in rural road dust (0.08 to 1.4, mean 0.4 μg/g dw) B[b]F, B[k]F, I(IDP + B[g]P, Chy, B[a]A, and B[j]F, most potent AhR agonists among PAHs in in vitro assay, were found at three fold higher concentrations in the urban site than in the rural site whereas DBA, also known as high AhR-mediated activity response in in vitro assay, was found in urban road dust at slightly higher level than in rural road dust Concentrations of PAHs recommended for carcinogenic Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 CALUX-TEQ (ng/g) L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx 40 35 30 25 20 15 a b 10 c d Vietnam (Urban RD) Vietnam (Rural RD) Vietnam (E-waste HD) Vietnam (Rural HD) Germany (Flood Dust) Japan (HSS Compost) Fig Concentration of CALUX-TEQ in different samples from various regions (gray bars: this study, white bars: cited from (a,bTue et al., 2012, cWölz et al., 2010 and dSuzuki et al., 2004), RD: road dust, HD: house dust, HSS: household sewage sludge screening by the European Union and the US Environmental Protection Agency (EU- and EPA-PAHs) in Hanoi road dust were among the highest reported for surface dust from various regions (Table 2) The B[a]P concentrations in Hanoi road dust were 2, and 24 times higher than those in dust from Duong Quang, Tehran—Iran and Cairo—Egypt, respectively, and comparable to those in dust from Beijing—China (Wang et al., 2010; Hassanien and Abdel-Latif, 2008; Saeedi et al., 2012) 3.3 Profiles and potential sources of PAHs The proportions of PAHs in Hanoi road dust were in the order of Fluh N Pyr N Phe N Chy N B[b]F + B[k]F N B[g]P N Ant N B[e]P N Nap N I(IDP + B[g]P N B[a]P N B[a]A N B[j]F N Flu whilst the in rural road dust was Phe N Fluh N Pyr N B[b]F + B[k]F N Nap N Chy N B[g]P N B[a]P N Flu N Ant N B[a]A N B[e]P N I(IDP + B[g]P N B[j]F (Fig 2) Such difference in the proportions of PAHs between the urban Hanoi and rural road dusts may be due to their emission sources The compound ratios that have the same molar mass are assumed to have similar physicochemical properties Therefore, ratios in between specific PAHs could be used to identify and characterize the emission sources The ratios of Phe/(Phe + Ant) (mean 0.87 for both sites), B[a]A/(B[a]A + Chy) (mean 0.27 and 0.3 for urban and rural sites, respectively) and B[a] P/B[g]P (0.53 and 0.86 for urban and rural sites, respectively) indicate that a major source of PAHs in Vietnamese road dust was fuel combustion in vehicle engines (Phe/(Phe + Ant) N 0.7, Guo, 2003), B[a]A/(B[a]A + Chy) = 0.22–0.55, Martuzevicius et al., 2011), B[a]P/B[g]P = 0.42–1.36 (Kume et al., 2007; Katsoyiannis et al., 2011) rather than petrogenic source (B[a]A/(B[a]A + Chy) b 0.2, Martuzevicius et al., 2011) The ratios of Flu/(Flu + Pyr) (mean 0.12 and 0.28 for the urban and rural sites, respectively) suggest that these compounds were from combustion of gasoline (b 0.5, Ravindra et al., 2008) rather than diesel (N0.5, Ravindra et al., 2008), whereas the ratios of Pyr/B[a]P (5.2 and 2.6, respectively) and I(IDP + B[g]P/B[g]P (0.53 and 0.62 for urban and rural sites, respectively) suggest high contribution of diesel combustion (Pyr/B[a]P ≈ 10, IND/B[g]P ≈ 1, Ravindra et al., 2008) in addition to gasoline combustion (Pyr/B[a]P ≈ 1, Ravindra et al., 2008, IND/B[g]P = 0.27–0.4, Ravindra et al., 2008) for the presence of these higher-ring PAHs in road dust In addition, the ratios of Fluh/(Fluh + Pyr) ranged from 0.44 to 0.76 (mean 0.52, urban) and 0.47 to 0.6 (mean 0.55, rural), B[a]A/(B[a]A + Chy) ranged from 0.17 to 0.58 (mean 0.3, Table Concentrations (ranges and arithmetic means) of EPA-/EU-PAHs, their theoretical-TEQs (Theo-TEQs), mutagenic equivalents (Theo-MEQs), carcinogenic equivalents (Theo-CEQs) and AhR-mediated activity (CALUX-TEQs) in road dust from various regions (ng/g dw) PAHs Priority list IARC classification Hanoi Beijinga Cairob Duong Quang Tehranc Nap Acy Ace Flu Phe Ant Fluh Pyr B[c]Ph CPP B[a]A Chy B[b]F + B[k]F B[j]F DMBA B[a]P B[e]P MCA IDP DBA B[g]P DB[ah]P DB[ai]P DB[al]P Total PAHs CALUX-TEQs Theo-TEQs Theo-MEQs Theo-CEQs EPA EPA EPA EPA EPA EPA EPA EPA 2B – 3 3 3 2B 2A 2B 2B 2B 2B – – 2B 2A 2B 2B 2A 43 (0.30–188) 11 (0.60–58) 3.6 (0.15–25) 29 (3.4–123) 190 (20–669) 68 (1.6–297) 276 (15–1153) 230 (12–646) 4.4 (ND–29) 3.2 (ND–22) 55 (2.3–308) 120 (8.4–535) 140 (7.1–1080) 33 (1.5–171) 0.80 (ND–10) 57 (3.8–271) 65 (1.9–365) 0.80 (ND–8.5) 54 (3.7–224) 2.7 (ND–26) 97 (6.1–285) 9.2 (ND–50) 1.6 (ND–38) 0.5 (ND–8.9) 1500 (127–5542) 20 (2.6–39) 2.1 (0.10–15) 208 (14–856) 117 (6.5–546) 94 39 109 17 155 98 NA NA 42 105 115 NA NA 94 NA NA 2.0 5.0 31 NA NA NA 922 NA 1.6 148 129 21.5 2.7 NA 13.6 403.7 6.9 68.2 165.6 NA NA 2.8 9.1 3.5 NA NA 2.4 NA NA NA NA NA NA NA NA 700 NA 0.10 4.0 7.0 23 (1.2–51) 3.6 (0.80–8.4) 0.6 (ND–1.1) 12 (3.3–20) 60 (16–133) 15 (ND–70) 54 (10–176) 44 (7.4–147) 4.1 (ND–20) 1.1 (ND–5.1) 19 (2.0–97) 40 (5.2–142) 49 (7.0–208) 12 (ND–59) 1.2 (ND–2.4) 23 (3.1–99) 23 (2.3–110) 0.050 (ND–0.30) 18 (2.4–86) 2.1 (ND–7.1) 23 (3.3–79) 0.80 (ND–2.6) ND ND 430 (76–1400) (1.9–13) 0.70 (0.10–3.1) 65 (9.2–279) 41 (6.1–166) 48 41 70 13 20 NA NA 10 20 NA NA NA NA 27 15 30 NA NA NA 325 NA 0.30 29 29 EU EPA, EU EPA, EU EPA, EU EU EPA, EU EPA, EU EPA, EU EPA, EU EU EU EU ND: not detected, NA: not analyzed a Cited from Wang et al (2010) b Cited from Hassanien and Abdel-Latif (2008) c Cited from Saeedi et al (2012) Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx Hanoi Duong Quang 30 25 20 15 ce Fl u Ph e A nt Fl uh Py r B[ c] Ph C PP B[ a] A C B[ hy b] F+ B[ k] B[ F j]F D M BA B[ a] P B[ e] P M C A ID P D BA B[ g D ]P B[ ah D ]P B[ D ]P B[ al ]P cy A N ap 10 A Proposition (%) 40 35 Fig Profiles of PAHs in road dust urban) and 0.18 to 0.57 (mean 0.3, rural) confirming traffic-related origin of PAHs, predominantly from gasoline emissions (Fluh/(Fluh + Pyr) = 0.4–0.6, Tsapakis and Stephanou, 2003; Ravindra et al., 2008, B [a]A/(B[a]A + Chy) = 0.4–0.6, Ravindra et al., 2008) Interestingly, ratios of B[a]P/(B[a]P + B[e]P) were 0.46 to 0.6 in average, indicating fresh vehicle emissions (approximately 0.5, Oliveira et al., 2011) (Table S1) The results of calculation of the I(IDP + B[g]P/(I(IDP + B [g]P + B[g]P) and B[a]P/B[g]P, plotted in Fig 3, showed that vehicular traffic was the major contributor of PAHs in Hanoi whereas mixed sources were the contributors in Duong Quang 3.4 Contribution of PAHs to the AhR-mediated toxicity Toxic contributions of PAHs in the samples were evaluated by comparison of CALUX-TEQs measured with DR-CALUX and total theoretical TEQs (Theo-TEQs) Theo-TEQ of a compound was calculated as the product of its concentration and its TCDD-relative potency (REP) in CALUX assays reported in previous studies (Behnisch et al., 2003; Machala et al., 2001; Marvanová et al., 2008) Significantly higher Theo-TEQs were observed in urban sites which are almost three times higher than those in rural site (Table 3) To compare Theo-TEQs between the present study and other reports, the PAH concentrations from previous studies were also converted to Theo-TEQs by multiplying them with respective REPs The highest levels of Theo-TEQs were found in Hanoi followed by Beijing, and the Theo-TEQs in Duong Quang (rural site) were higher than those in Tehran and Cairo (urban sites) Abundant contributor for Theo-TEQs was B[b] + B[k]F for all regions (Table 3) As shown in Hanoi Fig 4A, Theo-TEQs in road dust accounted for 0.8 to 60% of the CALUXTEQs (mean 9.8%) for Hanoi and 2.0 to 76% (mean 19.9%) for Duong Quang The principal contributors as potent AhR agonists were B[b]F + B[k]F (8% and 17% for urban and rural sites, respectively), I(IDP + B[g]P (0.7% and 1.2%), Pyr (0.4% for both sites), Chy (0.2 and 0.4%), and B[a]P (0.1 and 0.3%) (Table S2) Among the PAHs analyzed in this study, B[b]F + B[k]F were also major contributors of TheoTEQs (Fig 4B & Table S3) B[b]F + B[k]F were also major contributors of Theo-TEQs in other regions (44 to 91 % total Theo-TEQs) except in Cairo (Egypt) where Pyr contributed equally to B[b]F + B[k]F These results indicate the necessity to use effect-based bioassays in addition to chemical analysis to avoid missing non-target toxic contributors in risk assessment of human exposure to toxic chemicals in environment As shown in Fig 5, significant correlations were observed between total PAHs and Theo-TEQs (Pearson's ρ = 0.96, p b 0.005 for both sites) Total PAHs and CALUX-TEQs also correlated for the urban site (ρ = 0.63, p b 0.001), suggesting that major contributors of toxic activities may share the same sources with PAHs In contrast, no such correlation was found in the rural site (p = − 0.6), which can be explained by variable AhR-mediated potencies of different PAH congeners present in samples (Koppen et al., 2001) 3.5 Concentration of dioxin-related compounds and their contributions to the AhR-mediated toxicity Concentrations of total PCDD/Fs found in the urban site ranged from 40 to 170, mean 90 pg/g dw (0.60 pg WHO-TEQ/g dw, based on toxic Duong Quang 1.0 Coal Combustion Mixed Source IDP/(IDP + B[g]P) 0.5 Traffic Emission Mixed Source 0.0 0.0 0.9 1.5 B[a]P/B[g]P Fig Plot of IDP/(IDP + B[g]P) and B[a]P/B[g]P in road dust Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx Table Theo-TEQs calculated from PAHs in road dust of various regions (ng/g) PAHs Hanoi Beijinga Cairob Duong Quang Tehranc Phe Fluh Pyr B[c]Ph CPP B[a]A Chy B[b]F + B[k]F B[j]F DMBA B[a]P B[e]P IDP DBA DB[ah]P DB[ai]P DB[al]P Total 2.5 × 10−4 6.3 × 10−6 7.7 × 10−2 2.0 × 10−6 2.1 × 10−6 8.0 × 10−3 3.9 × 10−2 1.8 1.2 × 10−2 4.5 × 10−6 3.0 × 10−2 3.4 × 10−5 1.3 × 10−1 1.2 × 10−2 6.6 × 10−4 2.7 × 10−4 2.2 × 10−6 2.1 1.4 × 10−4 3.6 × 10−6 3.2 × 10−2 NA NA 6.0 × 10−3 3.5 × 10−2 1.5 NA NA 5.0 × 10−2 NA 5.0 × 10−3 2.2 × 10−2 NA NA NA 1.6 5.3 × 10−4 1.6 × 10−6 5.5 × 10−2 NA NA 4.0 × 10−4 3.0 × 10−3 4.6 × 10−2 NA NA 1.3 × 10−3 NA NA NA NA NA NA 0.10 7.8 × 10−5 1.2 × 10−6 1.4 × 10−2 1.9 × 10−6 7.0 × 10−7 3.1 × 10−3 1.3 × 10−2 6.3 × 10−1 4.2 × 10−3 6.7 × 10−6 1.2 × 10−2 1.2 × 10−5 4.6 × 10−2 9.2 × 10−3 5.9 × 10−5 ND ND 0.70 9.1 × 10−5 3.0 × 10−7 6.6 × 10−3 NA NA 1.4 × 10−3 6.6 × 10−3 1.2 × 10−1 NA NA 3.2 × 10−3 NA 6.7 × 10−2 6.5 × 10−2 NA NA NA 0.30 ND: not detected, NA: not analyzed a Cited from Wang et al (2010) b Cited from Hassanien and Abdel-Latif (2008) c Cited from Saeedi et al (2012) equivalency factors from Van den Berg et al., 2006, see Table S4) was slightly higher than those in the rural site which ranged from 50 to 70, mean 50 pg/g dw (0.30 pg WHO-TEQ/g dw), whereas concentrations of DL-PCBs were 28 times higher (970 to 110,000, mean 17,000 pg/g dw or 3.1 pg WHO-TEQ/g dw vs 230 to 600, mean 400 pg/g dw or 0.20 pg WHO-TEQ/g dw) (Table S5) Concentrations of brominated dioxins (PBDD/Fs) in urban dust ranged from 50 to 1100, mean 400 pg/g dw (2.1 pg WHO-TEQ/g dw) were also significantly higher than those in rural dust which ranged from nd to 260, mean 80 ng/g dw (0.50 pg WHO-TEQ/g dw) (Table S6) No significant difference was observed between the proportions of DRCs in the urban and rural sites (Fig S3) Among PCDD/Fs, PCDDs were major compounds, particularly OCDD and HpCDDs, whereas HpBDFs and OBDF were the predominant among PBDD/Fs (Fig S4) Among DL-PCBs, CB118 had the highest concentrations followed by CB105 In order to evaluate the contributions of individual and total DRCs we also used REPs reported in previous studies (detail in Theo-TEQs Rural site The theoretical mutagenic and carcinogenic equivalents of a PAH were calculated by multiplying the concentrations in road dust by appropriate mutagenic and carcinogenic potencies relative to B[a]P (Durant et al., 1996; Larsen and Larsen, 1998), and the total values were expressed as Theo-MEQs and Theo-CEQs, respectively The Theo-MEQs were 10 to 278 ng/g dw (mean 65 ng/g dw) in the rural site and 14 to 858 ng/g dw (mean 208 ng/g dw) in urban site (Table S7) Contributions by individual PAHs to mutagenic potencies was in the order of B[a]P (27%) N B[b]F + B [k]F (24%) N CPP (10%) N I(IDP + B[g]P (9%) N B[g]P (7%) N B[j]F (5%) N others (6%) for urban site and B[a]P (35%) N B[b]F + B[k] F (27%) N CPP (11%) N B[g]P (9%) N I(IDP + B[g]P (8%) N B[j]F (4%) N others (17%) for rural site (Fig 4C) The Theo-CEQs in rural site ranged from to 166 ng/g dw with a mean value of 40 ng/g dw whereas this value ranged from to 546 ng/g dw with a mean of 117 ng/g dw for urban site (Table S8) Contributions by individual PAHs to carcinogenic potencies are in order of B[a]P (48%) N B[b]F + B[k]F (17%) N Fluh (12%) N DBA (6%) N I(IDP + B[g]P (5%) N Chy (3%) N others (5%) for urban site and B[a]P (57%) N B[b]F + B[k]F (18%) N Fluh (7%) N I(IDP + B[g]P (5%) N Chy (3%) N DBA (3%) N others (12%) for rural site (Fig 4D) Significant mutagenic and carcinogenic potencies found in this study should be considered in view of potential health risk and B[b]F+B[k]F Rural site 80.1 9.8 0% 3.6 Mutagenic and carcinogenic equivalents of PAHs and their contribution Unexplained Activity 19.9 Urban site Table S4, Olsman et al., 2007; Behnisch et al., 2003) to calculate the theoretical TEQ of DRCs Contributions of DRCs to CALUX-TEQs were in the range between 0.004 and 0.08% (mean 0.03%) for rural site and between 0.006 and 0.25% (mean 0.06%) for urban site Contributions of brominated dioxins (PBDD/Fs) to CALUX-TEQs were very small (b0.005%) compared to those of PCDD/Fs and DL-PCBs in both rural and urban sites The major contributor of DRCs in rural site was 1,2,3,4,6,7,8-HpCDD (15 to 61%, mean 39%) and CB126 (9 to 23%, mean 15%), and in urban site they were CB126 (20 to 61%, mean 33%), 1,2,3,4,6,7,8-HpCDD (2 to 28%, mean 15%) and 2,3,4,7,8-PeCDF (ND to 31%, mean 12%) Considering these results on theoretical TEQs of DRCs and major PAHs, occurrence of unidentified potential AhR agonists in the dust samples can be suggested Further studies are necessary to determine other related compounds (e.g., oxygenated and methylated PAHs) which have been reported to transactivate AhR signaling pathway (Trilecová et al., 2011) 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% Pyr Rural site 35 Urban site 27 0% B[b]F+B[k]F CPP B[g]P 27 IDP B[j]F 10 0% Others Others 5.6 2.8 7.6 4.8 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% B[a]P Rural site 17 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% C B[j]F B 11 24 B[a]P 82.0 A B[a]P Chy 84.4 Urban site 90.2 IDP B[b]F+B[k]F Fluh DBA 57 Urban site 18 48 0% IDP 18 Chy Others 12 12 5 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% D Fig Contribution of Theo-TEQs to CALUX-TEQs (A), individual PAHs to Theo-TEQs (B), individual PAHs to mutagenic equivalents (Theo-MEQs) (C), and individual PAHs to carcinogenic equivalents (Theo-CEQs) (D) Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx Theo-TEQs (ng/g) 1.0 0.5 A 0.2 Theo-TEQs (ng/g) 10.0 2.0 5.0 2.0 1.0 0.5 B 0.2 0.1 0.1 100 200 500 1000 200 1000 2000 5000 Total PAHs (ng/g) 12 10 C CALUX-TEQs (ng/g) CALUX-TEQs (ng/g) Total PAHs (ng/g) 500 20 10 D 100 200 500 1000 200 Total PAHs (ng/g) 500 1000 2000 5000 Total PAHs (ng/g) Fig Relationships between total PAHs and Theo-TEQs (A: rural site, Pearson's ρ = 0.96, p b 0.005; B: urban site, Pearson's ρ = 0.96, p b 0.001) and relationship between total PAHs and CALUX-TEQs (C: rural site, Pearson's ρ = −0.3, p N 0.1; D: urban site, Pearson's ρ = 0.63, p b 0.001) cancer risk will be evaluated in health risk assessment section DB[al]P and B[b]F + B[k]F were the major mutagenic contributors in Vietnamese road dust, similar to the results calculated for Beijing and Greater-Cairo, whereas I(IDP + B[g]P and B[a]P were the major mutagenic PAHs in Tehran The major carcinogenic PAHs found in Vietnamese road dust were B[a]P and B[b]F + B[k]F, whereas in Cairo road dust they were B[a]P and Fluh, and in Tehran were B[a]P and B[g] P Strong positive correlations were observed between pairs of TheoTEQs/Theo-MEQs and Theo-TEQs/Theo-CEQs (correlation coefficients ranged from 0.95 to 0.98, p b 0.001, Fig S5) which can be explained by the fact that PAHs with higher AhR-mediated activities also tend to have higher mutagenic and carcinogenic potencies In summary, the levels of AhR-mediated toxicity and carcinogenicity calculated for PAHs in Hanoi road dust were comparable to those of a heavily air particlepolluted region (Beijing) and significantly higher than those of Duong Quang, Cairo and Tehran (Table 2) In contrast, mutagenic equivalent levels in Hanoi were significantly higher than those in Beijing, Duong Quang, Tehran and Cairo (Table 2) health risk (Liao and Chiang, 2006) The acceptable level is equal or lower than 10− (Chiang et al., 2009) Our estimated results suggest that children and adults in both study sites in Vietnam are exposed to high potential carcinogenic risk via both dust ingestion and dermal contact pathways Cancer risk levels via ingestion in urban sites (1.3 × 10−4 and 1.5 × 10−4 for children and adults, respectively) were somewhat higher than those in rural sites (4.7 × 10−5 and 5.1 × 10−5 for children and adults, respectively) Similarly, cancer risk levels via dermal contact in urban sites (3.3 × 10−4 and 5.2 × 10−4 for children and adults, respectively) were also somewhat higher than those in rural sites (1.2 × 10−4 and 1.8 × 10−4) (Table S9, Fig 6) The result of cancer risk assessment obtained in this study also raises the concern over the potential effect of ambient air contaminated by PAHs on the occurrence of common diseases related to urban air pollution in Hanoi urban areas, where higher rates of respiratory and skin disease cases were observed compared with sub-urban areas (Hung, 2010), and such effect needs immediate attention Conclusions 3.7 Health risk assessment To assess human health risk by exposure to PAHs, incremental life time cancer risk (ILCR) model was used as a tool to assess human cancer risk The ILCR model was developed to quantitatively estimate the exposure risk for road dust PAHs based on U.S EPA standard model This model could be applied to evaluate cancer risk for human who are exposed to urban dust via two pathways — ingestion and dermal contact with dust particles (Wang et al., 2011; Peng et al., 2011) Carcinogenic potencies relative to B[a]P (Larsen and Larsen, 1998), carcinogenic slope factor (CSF), and probabilistic risk assessment framework were applied to estimate cancer risk incurred from exposure to PAHs via these two pathways Probabilistic risk assessment for personal exposure to carcinogenic PAHs showed that an ILCR between 10−6 and 10−4 indicates potential risk, whereas ILCR greater than 10−4 suggests high potential To our knowledge, this is the first report on the determination and toxicant contributors' assessment of PAHs and DRCs in road dust from Hanoi, a sub-tropical Asian metropolitan area, by DR-CALUX assay and confirmation by chemical analysis Profiles of PAHs determined by chemical analysis indicated that fossil fuel combustion is the major PAH source in not only urban dust but also rural dust Particularly, it could have been released from the incomplete diesel and gasoline combustion in vehicles An examination of toxic contribution in this study indicated that PAHs are major contributor of overall AhR-mediated activities in road dust with smaller contribution of DRCs The differences between CALUX-TEQ and Theo-TEQs in road dust samples for both sites indicated occurrence of unknown potential AhR agonists in the road dust It could be PAH derivatives such as oxygenated PAHs, nitrogenated PAH, or methylated PAHs Approaches in this study are very useful for determination of overall toxicity of road dust as well as evaluation of specific chemical components in Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total Environ (2014), http://dx.doi.org/10.1016/j.scitotenv.2014.01.086 L.H Tuyen et al / Science of the Total Environment xxx (2014) xxx–xxx 1.0E-02 Ingestion Dermal Cancer Risk Levels Children Adults 1.0E-03 1.0E-04 1.0E-05 1.0E-06 Rural site Urban site Rural site Urban site Fig Estimated values of incremental life time cancer risk environmental samples whereas further studies are required to determine unidentified potential agonists in the contaminated road dust The results from this study also suggests that further studies on the status of contaminant and toxic effect assessment of PAHs and PAH derivatives in ambient air (particulate and gas phases) in Hanoi should be carried out Acknowledgments This study was partly supported by Grants-in-Aid for Scientific Research (A: 25257403) from Japan Society for the Promotion of Science (JSPS) and the Environment Research and Technology Development Fund (K123001 and 3K133010) from the Ministry of the Environment, Japan The award of a JSPS postdoctoral fellowship to N.M.T (P 13072) is also acknowledged Appendix A Supplementary data Supplementary data to this article can be found online at http://dx doi.org/10.1016/j.scitotenv.2014.01.086 References Behnisch PA, Hosoe K, Sakai S Bioanalytical screening methods for dioxins and dioxin-like compounds a review of 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Contribution of polycyclic aromatic. .. the determination and toxicant contributors' assessment of PAHs and DRCs in road dust from Hanoi, a sub-tropical Asian metropolitan area, by DR-CALUX assay and confirmation by chemical analysis Profiles... Please cite this article as: Tuyen LH, et al, Aryl hydrocarbon receptor mediated activities in road dust from a metropolitan area, Hanoi—Vietnam: Contribution of polycyclic aromatic , Sci Total