Ecotoxicology and Environmental Safety

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1 Ecotoxicology and Environmental Safety 74 (2011) Contents lists available at ScienceDirect Ecotoxicology and Environmental Safety journal homepage: Perfluorinated chemicals in blood of residents in Wenzhou, China Wei Zhang a, Zhenkun Lin a, Mingyue Hu a, Xuedong Wang a, Qingquan Lian b, Kuangfei Lin c, Qiaoxiang Dong a,n, Changjiang Huang a,n a Institute of Watershed Science and Environmental Ecology, Wenzhou Medical College, Wenzhou , P.R. China b The Second Affiliated Hospital of Wenzhou Medical college, Wenzhou , P.R. China c School of Resources and Environmental Engineering, East China University of Science and Technology/State Environmental Protection Key Laboratory of Environmental Risk Assessment and Control on Chemical Process, Shanghai , P.R. China article info Article history: Received 29 August 2010 Received in revised form 25 April 2011 Accepted 26 April 2011 Available online 12 May 2011 Keywords: Perfluorinated compounds HPLC/MS Exposure Serum Infertile Wenzhou abstract Perfluorinated compounds (PFCs) are persistent organic pollutants ubiquitously distributed in the environment and human populations. Here we report PFC concentrations in the residents of Wenzhou City, which is characterized as the Footwear Capital of China. Specifically, fifty serum samples collected from workers in a leather factory, fifty-five umbilical cord serum samples and fifteen serum samples from infertile men were analyzed. PFOS was one of the most frequently detected PFCs and showed the highest level. The mean serum levels of PFOS and PFOA of workers and infertile males were higher than the cord serum. PFOS concentration in cord serum increased with increase in age of the mother. Gender differences were significant both in worker serum samples and umbilical cord samples with higher levels found in males/male fetuses. Our findings suggested that PFOS, PFOA and PFHxS were widely distributed in Wenzhou residents, but occupational exposure was not the main source for workers. & 2011 Elsevier Inc. All rights reserved. 1. Introduction Perfluorinated compounds (PFCs) form a diverse group of chemicals with surface-active properties, which have been manufactured for more than 50 years. Fluorine has the strongest inductive electron-withdrawing power, forming a strong carbon fluorine (C F) covalent bond, which makes most PFCs stable in the environment and resistant to hydrolysis, photolysis and biodegradation (Beach et al., 2006). Due to their surface active characteristics, PFCs have been employed as surfactants and surface protectors in paper, food packages, leather, carpets, upholstery and fabric, and many other applications (EPA, 2000), which resulted in the global occurrence of these substances in air, water, sediment and sludge, as well as various wildlife species inhabiting not only locations close to pollution sources but also remote areas (Bossi et al., 2005; Senthilkumar et al., 2007; Dreyer and Ebinghaus, 2009). PFCs have also been found in food, drinking water and indoor dust (Skutlarek et al., 2006; Tittlemier et al., 2007; Björklund et al., 2009). Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are the two main PFC products. They are also the terminal degradation end-products of other PFCs, and have frequently been detected with high concentrations in environmental samples. The n Corresponding authors. Fax: þ addresses: [email protected] (Q. Dong), [email protected] (C. Huang). major manufacturer, 3M, in USA announced that they would phase out PFOS-based production in 2002 due to concerns about their environmental persistence and their potential biological effects; there were ubiquitous low-level distribution in humans including children (EPA, 2000). As a consequence, PFOS showed a 60% decline while PFOA decreased by 25% in plasma samples of American Red Cross blood donors collected in 2006 compared to serum samples collected in (Olsen et al., 2008). Despite this, recent report also indicated that serum PFOA did not change in low-exposed areas in East Asia even after the phase-out by 3M (Harada et al., 2010). This difference may be due to the large variations in the PFC exposure sources among different regions. Thus, PFCs are still widespread in the human body and the environment. More recently, significant adverse effects of PFCs on human health have been revealed. Children who were born to mothers with high PFOS levels were found to have delayed development in the age at which they start sitting without support (Fei et al., 2008). PFOS and PFOA concentrations in cord serums were found to be negatively correlated with infant birth weight and size (Apelberg et al., 2007b). Same negative correlation between PFOS in female infants and birth weight was found in another study in Japan (Washino et al., 2009). High PFCs levels were found to be negatively associated with the total sperm counts among men in Denmark (Joensen et al., 2009). Wenzhou is located in the southeast of Zhejiang province. The city is known as the Footwear Capital of China as it has /$ - see front matter & 2011 Elsevier Inc. All rights reserved. doi: /j.ecoenv

2 1788 W. Zhang et al. / Ecotoxicology and Environmental Safety 74 (2011) approximately 10,000 footwear enterprises and about 500,000 employees work in the shoe and leather-related industries (Wang, 2006). Fluorinated repellent has been used in these industries; however, its application may have declined since 2008 due to the restrictions posed on PFOS by the European Union (Cheng et al., 2009). The present study was intended to estimate the PFCs background exposure of workers in the leather factories as this industry sector represents a significant portion of the general population in Wenzhou. We also examined the PFC levels in blood samples from umbilical cords, given the possible adverse effects of PFCs posted on infants as described above. Additionally, male infertility has become a significant social problem in this region lately with an average rate ranging from 8% to 12% based on local hospital estimates (Hongshan Ge, per. commun.). Environmental pollution with various persistent organic pollutants is suspected to be one of the main causative factors. Thus, we also initiated a preliminary examination on PFC concentrations in blood samples of infertile men from local hospital. 2. Materials and methods sample loading. The cartridge was washed with 1 ml 40% methanol/water solution, and then dried completely using a vacuum pump. Target analytes were eluted with 6 ml methanol, and evaporated to dryness under a stream of high purity nitrogen gas. Samples were reconstituted in 1 ml of methanol and filtered through 0.2 mm nylon filter (to remove suspended materials and insoluble particles) into a plastic-lined glass vial prior to analysis Instrumental analysis Concentrations of perfluorooctane sulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonic acid (PFHxS) and nonafluorobutanesulfonic acid (PFBS) in human serum samples were analyzed by Agilent 1100 HPLC MS (Bruker Ion Trap HCT). A 10 ml aliquot of extract was injected onto a ZORBAX SB-C18 column (2.1 mm 150 mm, 5 mm, endcapped) with 10 mm ammonium acetate and methanol as the mobile phases starting at 5% methanol with a flow rate of 400 ml/min. The gradient was increased to 60% methanol at 3 min, and 90% at 10 min before reverting to 5% methanol at 13 min; each procedure lasts for 18 min. Column temperature was maintained at 35 1C. The detection system was an Ion Trap-MS operated in the negative ionization MS mode. The following selected ions were monitored: PFOS, m/z 499; PFOA, m/z 413; PFHxS, m/z 399 and PFBS, m/z 299. Mass spectrometer operated in smart mode with parameters as follows: capillary voltage was 3500 V; nebulizer pressure was 25 psi, drying gas (N 2 ) flow rate was 7.0 L/min and temperature was 350 1C Standards and reagents Heptadecaflurooctanesulfonic acid potassium salt (PFOSKZ98% chemical purity), perfluorooctanoic acid (PFOA, 96%), nonafluoro-1-butane-sulfonic acid potassium salt (PFBS, 98%), Tridecafluorohexane-1-sulfonic acid potassium salt (PFHxSZ98%) and ammonium acetate (499%, HPLC grade) were purchased from Sigma-Aldrich Chemical (St. Louis, MO, USA). HPLC grade methanol and acetonitrile (filtered by 0.2 um filter) were purchased from Merck KgaA (Darmstadt, Germany). Deionized (DI) water was obtained from UPWS-1-20T and contained no detectable PFCs. We used newborn bovine serum from Invitrogen (Carlsbad, CA, USA) as the surrogate matrix Samples Blood samples were obtained from fifty manufacture workers, fifteen infertile men and fifty-five umbilical cords between December 2008 and August 2009 in Wenzhou, East China. The average age of workers was 36 ranging from 19 to 47, of mothers (umbilical cord blood samples) was 28 ranging from 20 to 37 and of infertile men was 29 ranging from 24 to 42. All participants in this study reside within the Wenzhou metropolis area. Umbilical cord blood samples were randomly collected from mothers encompassing a wide range of occupations such as farmers, teachers, doctors, business women and workers. Men were diagnosed as infertile mainly based on sperm density (less than 20 million per ml), motility (less than 50% motile sperm) and morphology (less than 14% sperm with normal forms of head, midpiece or tail). The causative factors of all these infertile men are unknown. Blood samples of 10 ml were collected from each subject by venipuncture and kept in the cleaned polypropylene (PP) tubes that were pre-rinsed with methanol. Age, gender, worker s task and sperm counts (in infertile men) were obtained simultaneously. Serum was separated from the red blood cells and other cellular components by centrifugation at 2500 rpm for 15 min after coagulation. Supernatant was transferred to another pre-cleaned polypropylene tube, and stored at 20 1C in a freezer until analysis. All the containers and wares were prerinsed with distilled water followed by methanol; teflon and glass containers were avoided. The ethics committee of Wenzhou Medical College approved the design of this study, and informed consent was obtained from all participants Fluorochemical analysis (sample extraction) The serum samples were extracted using solid-phase extraction (SPE) and analyzed by a liquid chromatography (LC) system coupled to a single mass spectrometer (MS). The method was adopted from Yeung et al. (2009) with minor modifications. In detail, refrigerated samples were allowed to warm to room temperature, vortex mixed for 15 s and then 0.5 ml serum was transferred to a 15 ml polypropylene centrifuge tube. Samples were added with 1.5 ml acetonitrile, sonicated for 15 min and centrifuged at 4500 rpm for 15 min. The supernatant was transferred to a new tube; 8 ml distilled water was added for diluting the acetonitrile concentration, and then was transferred to Waters (Milford, MA, USA) Oasis hydrophilic lipophilic balance (HLB 200 mg, 6 cc) cartridge for solid phase extraction. HLB cartridge was firstly pre-conditioned by passing through the cartridge in the sequence of 6 ml of methanol and 6 ml distilled water with the rate of 2 drops/s. The cartridge was prevented from drying at all times during 2.5. Quality assurance and control Methanol, water and the nylon mesh filter were evaluated as sources of target analytes background levels before sample analysis. Double blanks (containing only mobile phase) were injected prior to running each type of sample and once every ten samples to monitor for potential PFC contamination from the instrument. Serum blank and operational blank of 0.5 ml distilled water were included within the same run to check possible method contamination. Levels of the target compounds in the blank samples were less than LOQ. Quality control (QC) and continuing calibration verification were prepared with a base of newborn bovine serum pool, which was considered to be an acceptable surrogate matrix for human serum (Haug et al., 2009). Lower (10 mg/l), moderate (50 mg/l) and high QC (200 mg/l) materials were prepared in duplicate and analyzed along with the serum samples using the same method to assure the accuracy and reliability of the data. Results within 725% of the theoretical value were considered acceptable to proceed with sample analysis. Matrix spiked with 50 ng/ml target compounds was analyzed to evaluate the matrix effect on the recovery efficiency, as well as measure the precision of each analyte. The background concentrations of the analytes in the sample matrix were determined and the measured values in the matrix spike duplicates were corrected for background concentrations. Eight random selected serum samples were reanalyzed at the end of the study to verify the stability of the method Quantitative analysis Determination of PFC concentration was performed using external calibration curves. Five-point calibration curves were plotted from the limit of quantification to 1000 ng/ml. Calibration curves demonstrated good linearity with a coefficient of correlation greater than 0.99 for each analyte. The lowest concentration at which the analytical process can be reliably differentiated from the background level was expressed either as the limit of detection (LOD) or the limit of quantification (LOQ). LOD was defined as three-fold larger than the signal-tonoise ratio, and LOQ was defined as ten-fold larger than the signal-to-noise ratio. Analytes were considered to be positively identified if retention times were within 2.5% of the standards, and the peak area was ten times greater than the adjacent baseline peak-to-peak noise. Quantization of samples was performed based on the relative response of each analyte to the extracted calibration curves described above. Concentrations were not adjusted for the purity of standards or recovery as calibration standards followed the same extraction procedure as samples Method performance Six randomly selected human serum samples spiked with target analytes were used to assess the recoveries. Internal contents were subtracted from the results. Blanks were free of detectable concentrations of all PFCs in this study. The accuracy and precision were showed in Table 1. Calibration curves showed good linearity (R in each target). Duplicate measurements of eight serum samples showed good repeatability for PFOS (R 2 ¼0.90; Po0.01) and PFOA (R 2 ¼0.81; Po0.01).

3 W. Zhang et al. / Ecotoxicology and Environmental Safety 74 (2011) Statistical analysis A Mann-Whitney U-test was used to assess the effects of gender and age on PFCs concentrations in the blood samples. The Kruskal Wallis test was used to assess the effect of work type. A Spearman rank correlation analysis was used to examine possible correlations among the contents of studied PFCs, and between PFCs and sperm parameters in the samples. Reanalyzed samples were treated with fit linear analysis. The conventional 5% cut off was used to report results as statistically significant. Because of their low frequency of detection, PFBS was not included in the statistical analysis. Samples with concentrations lower than LOQ were computed as 1/2 of LOQ. All the statistical analyses were performed by the software of SPSS Results Samples were scanned for four PFCs; the mean PFOS concentration was found to be the highest, followed by PFOA and PFHxS (Table 2). PFOS and PFOA were detected in most samples (498% for PFOS and 494% for PFOA) while PFHxS was found in less than 50% of the samples. Only two samples had PFBS concentrations above LOQ. Mean PFOS and PFOA levels in serum samples from workers were higher than those found in umbilical cord blood samples or infertile men, while PFHxS was the highest in cord serum. The PFOA and PFHxS had the lowest concentrations in serum from infertile men. There was higher frequency of high levels of PFOS than PFOA in serum samples from workers, but the same trend was not observed for cord serums (Fig. 1). Frequency of high level PFOS (e.g., 415 mg/l) in workers was higher than that in cord serum. Significant correlations between PFOS and PFOA were only found in cord serum samples (P¼0.02). There were no correlations between PFC concentration and sperm density ( /ml with a range from 0 to /ml) of samples from the infertile men (P40.05). PFOS and PFOA in serum samples from workers and umbilical cords were also analyzed after grouping the data based on age (o or Z30 yr) and gender (Table 3). There were no significant differences in PFOS or PFOA concentrations between the two age groups (P40.05). However, weak positive correlation was found between cord serum PFOS levels and mother s age (Rho¼0.09, Po0.05). Gender difference was significant for serum PFOS in workers and umbilical cords as male samples were significantly higher than female samples (Fig. 2). There was no gender effect for PFOA (Table 3). To determine whether PFCs concentrations in serum samples were related to a particular job assignment in the leather factory, data from the 50 workers were further classified into three groups: workers specialized on making chemical solutions, coating the leather and gluing multiple layers of leather together. However, no significant difference was found in any of the PFC concentrations among these three groups (P40.05; Table 4). 4. Discussion 4.1. PFCs in adult serum Our findings revealed that PFCs in serum samples from workers in the leather factory are the modest, and are not related to any particular type of job assignments, suggesting no occupational Table 1 Method performance characteristics of standard-spiked matrix. Analyte Range (mg/l) Accuracy (%) Intra-assay Inter-assay (SE) (SE) 50 mg/l 10 mg/l 50 mg/l 200 mg/l PFOS PFOA PFHxS PFBS Correlation coefficient (40.99). Fig. 1. Frequency distribution of PFOS and PFOA in serum samples from workers (gray bars) and umbilical cords (dark bars). Table 2 PFC concentrations (ng/ml) in serum samples. Sample type Statistics PFOS PFOA PFHxS PFBS Workers (n¼50) N a Range Mean Median Umbilical cord (n¼55) N a Range Mean Median Infertile man (n¼15) N a Range Mean Median a Number of samples with values4lowest OQ.

4 1790 W. Zhang et al. / Ecotoxicology and Environmental Safety 74 (2011) Table 3 The median serum concentrations (ng/ml) of PFOS and PFOA in workers and umbilical cord grouped by sex and age. Group Analyte Gender Age (yr) Male Female P-value o P-value Worker PFOS (42) 6.42 (8) (15) (35) PFOA 3.49 (42) 3.51 (8) (15) 3.50 (35) Umbilical cord a PFOS 5.16 (33) 4.11 (22) (38) 4.42 (17) PFOA 5.56 (33) 5.22 (22) (38) 5.63 (17) a Fetal sex and maternal age were used; values within the parenthesis indicate sample size. PFOS (µg/l) p<0.05 Worker Serum sample p<0.05 Umbilical cord Fig. 2. The PFOS concentrations (mean7se) in serum samples collected from general workers and umbilical cord grouped by sex (male dark bars; female gray bars). Table 4 PFCs concentrations (ng/ml) in serum samples of workers in the leather factory with different job assignments. Job type Statistics PFOS PFOA PFHxS PFBS Solution preparation (n¼17) N a Range Mean Median Coating (n¼27) N a Range Mean Median Gluing (n¼6) N a Range Mean Median a Number of samples with values4lowest OQ. exposure of PFCs in this sampling population. One possible reason could be due to the PFOS restriction posted by the European Union, which was to be effective in June 2008 and limit PFOS use as a substance in preparation to be less than 0.005% by mass, in semifinished product or articles to be less than 0.1% by mass, and for textiles and coated materials to be less than 1 mg/m 2 (EU Directive 2006/122/EC). As EU constitutes one of the major exporting markets for the shoe and leather industries in this region, manufactories may have decreased the use of PFOS to meet this compliance. In fact, the local government of Wenzhou solicited a research initiative on PFOS substitutes in Other factors contributing to the modest levels of PFCs in blood samples from workers in the leather factory may include better personal protection through the use of gloves and masks, and more rigorous restrictions in production area where food consumption is not allowed. Our study also confirmed that PFCs were ubiquitous in adult human blood; PFOS and PFOA were the two mostly detected targets. Mean PFCs level in the present study was in the range of concentrations reported in other cities of China (Fig. 3) or other nations such as southern Bavaria (Fromme et al., 2007). All of the PFCs levels in the present study were lower than residents of the American continent where median concentration of PFOS was 31.1 mg/l, of PFOA was 11.6 mg/l and of PFHxS was 2.0 mg/l (Calafat et al., 2006). In particular, PFOS and PFOA from occupational exposure in USA (3M) were about 100 times higher than the current study (Olsen et al., 2003). However, PFOS concentrations in the present study were higher than those found in India, Sri Lanka and Japan, and PFOA concentrations were higher than those found in Vietnam and Korea (Kannan et al., 2004; Guruge et al., 2005; Harada et al., 2010). When compared with other cities in China, PFCs in Wenzhou were comparable to that of Beijing, higher than PFCs in Nanjing and lower than that of Shengyang (Yeung et al., 2008; Liu et al., 2009). The discrepancies among different cities may mainly be attributed to different industry sectors and their specific exposure sources and distribution routes. For example, the predominant heavy industries in Shenyang may explain the high PFC concentrations in its residents, while the relative high PFCs detected in Wenzhou could be due to the industry sectors such as shoes, textiles, costumes and small electric appliances. Distinct regional differences in PFCs concentration and distributions has also been reported in other parts of the world, for example, PFHxS was found to be the highest in human blood in southeast Queensland of Australia (Toms et al., 2009) PFCs in cord serum Cord serum was found to be contaminated with PFOS, PFOA and PFHxS in the present study. In particular, PFHxS was higher than those from workers and infertile men. The PFOS concentration in our study was similar to those found in Canada ( ng/ml) (Monroy et al., 2008), but higher than studies reported in Japan ( ng/ml) (Inoue et al., 2004) and USA (4.9 ng/ml) (Apelberg et al., 2007a). PFHxS level was relatively low compared to that reported in Canada ( ng/ml) (Monroy et al., 2008). The PFOA concentration in our study was higher than those reported in Shengyang, China ( ng/ ml) (Jin et al., 2004); USA (1.6 ng/ml) (Apelberg et al., 2007a); and Germany (3.4 ng/ml) (Midasch et al., 2007).This latter study

5 W. Zhang et al. / Ecotoxicology and Environmental Safety 74 (2011) Fig. 3. Concentrations of PFOS (white), PFOA (gray) and PFHxS (dark) in blood samples of people from different cities in China. Data for Nanjing, Jintan, Guiyang, Beijing and Shenyang-2 were from Yeung et al. (2008); data for Fuxin, Jinzhou, Shenyang-1, Anshan, Yingkou, Hulidao and Dalian were from Liu et al. (2009). also reported corresponding maternal serum level for PFOA (2.6 ng/ml) and suggested that PFOA may intensely accumulate in fetus from mother due to their lower body weight. We did not analyze the maternal blood in the present study. A more conclusive relationship between PFCs in mothers and fetus could be obtained through analysis of samples from maternal serum, cord blood and newborn baby blood Gender effect on PFC concentrations Earlier studies on occupational exposure with US workers have showed higher PFOS concentration in adult men than women (Olsen et al., 2003). PFOS was also found to be statistically lower in females than males of the 2005 American Red Cross blood donors (Olsen et al., 2007). The same gender difference of PFOS was also found with the general population in Japan (Harada et al., 2004) and Australia (Toms et al., 2009). However, a study in Spain reported no statistically significant difference of PFOS levels between the males and the females in their serum pools (Ericson et al., 2007). Our study also showed higher PFOS (but not PFOA or PFHxS) in male workers than female workers. Although we suspected that differences in job type may confound the gender effect as a particular job could be dominated by males or females, we did not observe any work related PFC elevation in the present study. Because PFCs were detected in maternal serum and milk (Kärrman et al., 2007), menstrual bleeding, pregnancy and lactation in females are the possible routes for PFOS excretion, and were thus suspected to be the main reason for lower PFOS concentration in females. This was supported by recent findings of a Japanese study where mean serum concentrations of PFOA and PFOS in 20 adult males and 8 postmenopausal females were very similar regardless of age, but higher than that of 20 females who still menstruate regularly (Harada et al., 2005). Studies on the effects of pregnancy have shown decreased levels of PFOS and PFOA in maternal blood samples with increasing parity, suggesting fetal uptake during pregnancy and excretion during lactation (Fei et al., 2007). Interestingly, our study also showed higher PFOS in cord serum from male fetuses than female. To our knowledge, this is the first study to show that fetus sex can influence PFOS level in cord blood. It is unknown whether this phenomenon is also true for populations in other cities or nations. We recommend other researchers to examine this phenomenon in their studies. If this is a consistent phenomenon across many studies/populations, it may suggest that fetus sex could have a differential role in accumulation of different chemicals, and this may help explain some sex-related birth defects Age effect on PFC concentrations Studies were controversial in terms of the age effect on PFC concentrations. We did not find any PFC concentration difference when data were divided into two age groups (o or Z30 years). However, there was a tendency of increasing PFOS level in cord serum along with increase in ages of the mother. Earlier studies on an adult German population in southern Bavaria reported higher PFOA and PFOS in older females than young females, but the same phenomenon was not found in males, suggesting menstrual bleeding served as an important route for excretion of PFCs in females (Fromme et al., 2007). Studies on a Chinese population with the mean ages ranging from 23 to 44 in 9 cities, however, did not show any age-related differences in the concentrations of PFOA, PFOS, PFOSA and PFHxS (Yeung et al., 2006). Elucidation of the effects of age on PFC accumulation, metabolism and excretion in humans is complicated as individuals vary in their life styles. Using experimental animals could be a more appropriate approach while PFC dosing and excretion can be precisely controlled or measured PFCs effect on sperm quality Earlier studies indicated that PFCs were associated with increased serum estradiol and reduced testosterone (Cook et al., 1992). Toxicological studies of PFCs in rats showed an increased incidence of testicular tumors (Biegel et al., 2001). More recently, serum PFC concentrations were reported to have a negative correlation with sperm quality in a Danish study (Joensen et al., 2009). As the present study and others have shown that males tend to accumulate higher PFCs than females, we thus suspect a negative effect of PFCs on sperm quality. However, we did not observe any significant correlation between sperm density and PFC concentration. It is possible that PFCs in serum do not necessary reflect their contents in seminal plasma while sperm quality is most likely affected by PFCs locally in seminal plasma.

6 1792 W. Zhang et al. / Ecotoxicology and Environmental Safety 74 (2011) In fact, previous studies have shown lower PFC concentrations in seminal plasma than those in serum (Guruge et al., 2005). The same study also revealed that seminal plasma accumulates measurable concentrations of various PFCs, for example, the overall detection frequency for PFOS, PFHS and PFOA was 90%. We thus suggest future studies to measure PFCs directly in seminal plasma. Also other sperm parameters such as sperm motility, morphology and membrane surface protein properties should be included to evaluate any possible adverse effect from PFC exposure. More rigorous experimental design should also include matched normal controls with a much larger sample size. 5. Conclusion The present study indicated that PFCs were widespread in residents of Wenzhou with concentrations much lower than the occupational exposure levels. Higher levels of PFOS were found in males than females. 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