Journal of J Occup Health 2004; 46: 49Ð59 Occupational Health

Perfluorooctanoate and Perfluorooctane Sulfonate Concentrations in Surface Water in

Norimitsu SAITO1, Kouji HARADA2, Kayoko INOUE2, Kazuaki SASAKI1, Takeo YOSHINAGA2 and Akio KOIZUMI2

1Research Institute for Environmental Sciences and Public Health of and 2Department of Health and Environmental Sciences, University Graduate School of Medicine, Japan

Abstract: Perfluorooctanoate and Perfluorooctane exposed to PFOA through drinking water ingestion. Sulfonate Concentrations in Surface Water in (J Occup Health 2004; 46: 49–59) Japan: Norimitsu SAITO, et al. Institute for Environmental Sciences and Public Health of Iwate Key words: PFOA, PFOS, Surface water, area, Prefecture—Perfluorooctanoate (PFOA) and Drinking water contamination perfluorooctane sulfonate (PFOS) are synthetic surfactants used in Japan. An epidemiological study Perfluorooctanoate (PFOA) is a synthetic surfactant of workers exposed to PFOA revealed a significant used in a variety of industrial applications1). It is also increase in prostate cancer mortality. A cross-sectional formed through the degradation or metabolism of certain study of PFOA-exposed workers showed that PFOA other manmade fluorochemical products1). perturbs sex hormone homeostasis. We analyzed their PFOA has been reported to cause diverse toxic effects concentrations in surface water samples collected from 2, 3) all over Japan by LC/MS with a solid phase extraction in laboratory animals including primates . An method. The lowest limits of detection (LOD) (ng/L) epidemiological study of workers exposed to PFOA were 0.06 for PFOA and 0.04 for PFOS. The lowest revealed a significant increase in prostate cancer mortality4). limits of quantification (LOQ) (ng/L) were 0.1 for both A cross-sectional study of PFOA-exposed workers showed analytes. The levels [geometric mean (GM); geometric that PFOA perturbs sex hormone homeostasis5), but recent standard deviation (GS)] (ng/L) of PFOA and PFOS in long-term follow-up studies on the workers could not the surface waters were GM (GS): 0.97 (3.06) and 1.19 confirm the earlier adverse effects6Ð8). (2.44) for -Tohoku (n=16); 2.84(3.56) and 3.69 There is evidence that perfluorooctane sulfonate (PFOS), (3.93) for Kanto (n=14); 2.50 (2.23) and 1.07 (2.36) for which is an analogue chemical of PFOA, is globally Chubu (n=17); 21.5 (2.28) and 5.73 (3.61) for Kinki distributed in humans9, 10) as well as wild life11, 12). In (n=8); 1.51 (2.28) and 1.00 (3.42) for Chugoku (n=9); contrast, contaminations with PFOA have been observed 1.93 (2.40) and 0.89 (3.09) for Kyushu- (n=15). 12, 13) The GM of PFOA in Kinki was significantly higher than in wild animals specific to some geographic areas . in other areas (ANOVA p<0.01). Systematic searches Similarly, geographical differences in serum PFOA levels of Yodo and Kanzaki Rivers revealed two highly have been reported in non-occupationally exposed contaminated sites, a public-water-disposal site for persons5, 10). PFOA and an airport for PFOS. The former was Recent ecological investigations have shown that one estimated to release 18 kg of PFOA/d. PFOA in of the ecological sources of PFOA and PFOS is discharge drinking water in Osaka city [40 (1.07) ng/L] was waters from manufacturers14). Moody et al.15) recently significantly higher than in other areas. The present reported that perfluorinated surfactants, including PFOA study confirms that recognizable amounts of PFOA are and PFOS, which were present in some formulations of released in the Osaka area and that people are fire-fighting foams were detected in ground water at U.S. Air Force Bases after cessation of their use. We have also shown that there is a large variation in PFOS contamination Received Sep 2, 2003; Accepted Oct 20, 2003 16) Correspondence to: A.Koizumi, Department of Health and levels in Japan . Geological differences may be attributable 17, 18) Environmental Sciences, Kyoto University Graduate School of to area-specific sources and traffic density . Medicine, 606-8501 Kyoto, Japan The major aims of the present study were twofold. (e-mail: [email protected]) First, the PFOA concentrations were determined in 50 J Occup Health, Vol. 46, 2004 surface water in Japan. This would provide a nationwide Hanshin area (Amagasaki and Kobe cities)], and in profile of the PFOA concentration levels. Since surface Tohoku district (Morioka, Sendai and Yokote cities). To water is the major source of drinking water in Japan, minimize the possibility of sample contamination, information on the concentration levels of surface water containers were thoroughly rinsed with methanol and is important for characterizing the exposure of the deionized water prior to use. Samples were stored at population. The second aim was to find sources of PFOA, room temperature (22°C) prior to analysis. which are associated with geographic differences in PFOA concentration levels. Identified sources would suggest a Field quality samples large industrial production of PFOA or other related A field matrix spike (FMS) and a field matrix blank materials as well as environmental leakages in specific (FMB) were prepared when we collected water samples areas. In the present study, PFOS levels were also in Iwate and Osaka areas. Sixteen samples of two-L determined to delineate differences in the contamination bottles were collected at each sampling site. Eight of the profiles between PFOS and PFOA. sixteen bottles were spiked with analytes for FMS and the other eight bottles were analyzed without spiking for Methods and Materials FMB. FMSs collected in Iwate were spiked with PFOA Standards and PFOS at 1 ng/L in the field. FMSs collected in the Perfluorooctanois acid (98% purity) was obtained from Osaka area were spiked with PFOA at 20 or 200 ng/L Aldrich (Milwaukee, WI) and potassium salt of PFOS and PFOS at 20 ng/L in the field. (98% purity) was obtained from Fluka (Milwaukee, WI). Field spike control samples (FSC), deionized water HPLC-grade methanol was purchased from Wako Pure spiked with amounts of PFOA and PFOS corresponding Chemicals (Osaka, Japan). Presep-C Agri cartridges to the counterpart FMS, were also prepared. We also (Solid phase: Styrene divinylbenzene polymethacrylate prepared deionized water samples without spikes as field on a polyethylene housing) (Presep-C, 220 mg cartridge) blanks. were also purchased from Wako Pure Chemicals (Osaka, Japan). Solid phase extraction Waters including deionized water, were contaminated We split a two-L sample into two one-L preparations. with PFOS16), and deionized water was only used after Individual one-L preparations were processed separately. passage through a Presep-C Agri cartridge to remove the First, they were filtered through glass fiber filters (1.0 residual PFOA and PFOS in this study. We confirmed µm¿) (ADVANTEC GA 100, 55 mm ¿, ADVANTEC, that the concentration of PFOA or PFOS in the methanol , Japan) to remove sediments and biota as used was less than the detection limit. previously reported16). Subsequently, they were passed through a membrane filter (Millipore JAWPO4700, 47 Water sampling mm ¿, pore size 1.0 µm)(Millipore, Tokyo, Japan). Water samples were collected from rivers, coastal sea Samples were passed through a Presep-C Agri column at water and tap water. For all sampling, a two-L sample a flow rate of 10 mL/min using a Waters Concentrator was collected in polyethylene terephthalate disposable System (Concentrator Plus, Waters, Tokyo, Japan). containers with narrow-mouth bottle tops and screw caps Presep-C cartridges were then eluted with 1.5 mL of as previously reported (Saito et al.16) We collected tap methanol and concentrated at room temperature under waters in the Osaka area [Kyoto city, Osaka city and nitrogen gas flow to 1 mL for the LC/MS analyses16).

Table 1. An optimized analytical condition for PFOA and PFOS by LC/MS

HPLC MS

Instrument Agilent 1100 Instrument Agilent 1100MSD SL Column Zorbax XDB C-18 (2.1 × 150 mm) Ionization ESI

Mobile phase A: CH3CN Nebulizer N2 (50 psig)

B: 10 mM CH3COONH4/H2O Drying gas N2 (10.0 L/min, 350°C) Linear gradient; 35%A to 45%A by 2%A/min Polarity Negative Fragmentor 100 V for PFOA and 200 V for PFOS Flow rate 0.2 mL/min. Vcap 4000 V - - Oven temp 40°C SIM (ion) C8F17SO3 : 499, FSO3 : 99 for PFOS - - Injection volume 10.0 µLC7F15CO2 : 413, C7F15 : 369 for PFOA Norimitsu SAITO, et al.: Surface Water Contamination with PFOA and PFOS in Japan 51

Fig. 1. Two typical LC/MS chromatograms of PFOA and PFOS in Iwate Sea water (1 ng/L)(A) and deionized water (0.1 ng/L)(B).

- LC/MS and quantification PFOS ion 499 (C8F 17SO3 ) were monitored for The methanol extracts (10 µL injection volume of the quantification. The monitoring revealed a major peak 1 mL extract) were chromatographed by HPLC at a flow with a retention time of 7.4 min for PFOA and 13.3 min rate of 0.2 mL/min as shown in Table 1. The total runtime for PFOS. To avoid interference and ensure complete - was 20 min, without any equilibration time between selectivity, the fragment of PFOA, ion 369 (C7F15 ) and - samples. We employed gradient conditions in the mobile the fragment of PFOS, ion 99 (FSO3 ) were also phase: the concentration of CH3CN (A) in 10 mM monitored.

CH3COONH4 buffer (B) started at 35%, then increased Throughout this study, the mean of two one-L to 45% at 2%/min for 5 min, and was then maintained at preparations from a two-L sample was calculated for 45% until 20 min (Table 1). The column temperature quantification of each sampling. was maintained at 40°C. The PFOA and PFOS standards were mixtures of linear Validation study and branched isomers (approximately 80% linear). Precision and accuracy were evaluated by determining Because isomerically separated standards for PFOA and PFOA and PFOS in quality control samples. In order to PFOS are not available, it was assumed that the response assay the quality control samples, three calibration curves factors for branched and linear isomers are equivalent spanning concentrations from 0 ng/L to 150 ng/L were and that the standard mixture is representative of that prepared along with the samples. The calibration curves identified in the samples. used for quantification, consisting of six points covering Mass spectra were taken on an LC/MS system equipped 0 ng/L to 2.0 ng/L and 5 ng/L to 150 ng/L in deionized with an orthogonal spray interface, employing water, were plotted using a linear fit. The curves were electrospray ionization in the negative mode. The not forced through zero. fragmentor voltages were 100 V for PFOA and 200 V for The precision of the method at each concentration was PFOS and V cap voltages were 4000 V for both analytes expressed as a coefficient of variation (CV), by (Table 1). The nebulizer pressure was 50 psig and the calculating the standard deviation (SD) as a percentage drying N2 gas flow rate was 10.0 L/min. The selected of the mean calculated concentration, while the accuracy ion monitoring (SIM) mode was employed for of the assay was determined by expressing the mean quantification of analytes (Table 1). calculated concentration as a percentage of the added In the selected negative-ion mode monitoring of ions, concentration. - the fragment ions for PFOA ion 413 (C7F15CO2 ) and for The percentage extraction recovery of PFOA and PFOS 52 J Occup Health, Vol. 46, 2004 at four different concentrations (0.5,5,50 and 150 ng/L) n=3) were also analyzed by LC/MS/MS (Applied was determined by samples dissolved in one-L deionized Biosystems API3000TM, Tokyo, Japan) for confirmation water. These samples were prepared and eluted as other of PFOA and PFOS under optimal analytical conditions9). - surface water samples, resulting in ×1,000 concentration. The 7.4-min peak had four daughter ions [ions 369 (C7F15 - - - They were calibrated against the samples dissolved in ), 219 (C4F9 ), 169 (C3F7 ) and 119 (C2F5 )], confirming methanol at (0.5,5,50 and 150 µg/L). The limit of that it corresponded to PFOA (data not shown)9). The 13.3- detection (LOD), defined as the lowest concentration that min peak had the parent ion, m/z 499, and three daughter - - - the analytical process can reliably differentiate from ions [ions 130 (CF2SO3 ), 99 (FSO3 ) and 80 (SO3 )], background levels, was considered to be three-fold larger confirming that it corresponded to PFOS (data not shown)9). than the signal-to-noise (S/N) ratio. The lowest limit of Based on confirmation by LC/MS/MS, we concluded quantification (LOQ) was defined as the lowest that LC/MS has sensitivity and specificity of PFOS and concentration with a CV<10%. PFOA to a degree similar to LC/MS/MS. Although multiple fragment ions were monitored, quantifications Statistics were based on two ions, ion 413 for PFOA and ion 499 Means and standard deviations were calculated. When for PFOS by LC/MS. indicated, the geometric means (GM) and geometric standard deviations (GSD) were calculated. The data Calibration curve, accuracy and precision for LC/MS were analyzed with SAS software (Version 8.2; SAS The extracted standard curves obtained in the range Institute Inc., Cary, NC). A p<0.05 was considered 0Ð150 ng/L showed excellent linearity with a coefficient significant. of correlation that was invariably greater than 0.999. Figure 2 shows the calibration curves relating the Results integrated peak area to the added concentrations of PFOA Chromatographic identification of PFOA and PFOS and PFOS on three different days. Figure 1 shows typical chromatographic patterns of The accuracy and precision were determined by PFOA and PFOS in a sea water sample spiked at 1.0 ng/ evaluating an extracted curve of ten samples of deionized L and a deionized water sample spiked at 0.1 ng/L. water spiked at four different levels for a total of 40 Portions of the surface water samples (river samples, samples per analyte. The results for all analytes at each

Fig. 2. Regression curves for deionized water on three different days relating the peak response to the added concentrations of PFOA and PFOS. For all curves, correlation coefficients were greater than 0.999. Norimitsu SAITO, et al.: Surface Water Contamination with PFOA and PFOS in Japan 53

Table 2. Accuracy of the present analytical method determined by evaluating extracted samples of deionized water spiked at four different concentrations

PFOA% PFOS% ng/L No Concentration ± SD CV Concentration ± SD CV

0.5 10 98.9 ± 4.9 5.0 101.1 ± 1.6 1.6 510 98.3 ± 1.5 1.5 103.1 ± 4.9 4.8 50 10 102.2 ± 3.5 3.4 104.2 ± 1.4 1.3 100 10 99.3 ± 4.4 4.4 101.4 ± 3.8 3.7

Table 3. Determination of extracted efficiency determined by evaluating extracted samples of deionized water

PFOA % extraction PFOS % extraction ng/L No efficiency ± SD CV efficiency ± SD CV

0.5 10 98.9 ± 4.8 4.85 105.5 ± 3.4 3.22 51098.1 ± 5.7 5.81 92.0 ± 5.4 5.87 50 10 92.6 ± 4.4 4.75 100.5 ± 6.5 6.47 100 10 91.5 ± 3.7 4.04 99.8 ± 4.8 4.81

Table 4. Results of analyses of quality control samples

FMS FMB PFOA PFOS PFOA PFOS Sampling Site Spiked Spiked No (ng/L) Recovery % (ng/L) Recovery % No (ng/L) (ng/L)

Iwate River (Site 4 in Fig. 3) 8 1 98.9 ± 4.8 1 105.5 ± 3.4 8 0.54 ± 0.06 0.46 ± 0.01 Iwate Sea (Iwate Coastal Sea water) 8 1 98.1 ± 5.7 1 92.0 ± 5.4 8 0.16 ± 0.02

Iwate River (Site 4 in Fig. 3) 3 1 98.9 ± 30 1 101.4 ± 2.0 3

FMB and field blanks: Neither PFOA or PFOS was added to the original solution.

of the four concentration levels, along with the standard Although we did not determine the lowest level to meet deviation for analysis and CV, are summarized in Table the criteria as CV<10%, the lowest point of the calibration 2. For all analytes, the precision as indicated by CV was curve, 0.1 ng/L, was assumed to be LOQ, of which the within 10%. CV was less than 7% for both analytes. Differences in When field samples were concentrated by a factor of concentrations of PFOA and PFOS in two one-L 1000 in a preconcentration step, the LOD was considered preparations for each sample were all within 10%. to be 0.06 ng/L for PFOA and 0.03 ng/L for PFOS. The extraction efficiency was determined by evaluating 54 J Occup Health, Vol. 46, 2004 extracts of deionized water prepared at four levels (0.5, Ina River flows into Kanzaki River at K9. Ina River 5, 50, 100 ng/L, n=10 for each concentration) (Table 3). collects water discharge from the Osaka International PFOA recoveries ranged from 92 to 99%, and PFOS Airport. The width of Ina River is 100 m at K10. There recoveries ranged from 92 to 106%. is a canal between Kanzaki River and (K15). The upstream region of Kanzaki River is named the Ai Determination of PFOA and PFOS in surface waters in River. The width of is 50 m at the Aigawa Japan Ryuiki water disposal site (A5). Disposed water is Eight FMS and FMB samples and three FSC samples discharged at A5 at a flow rate of 270,000 m3/d. Samples were collected per site in Osaka and Iwate fields. The were collected from various sites as shown in Fig. 4. results are shown in Table 4. All FMSs were within 10% The PFOA concentration was high along the foreshores of the expected values. The recoveries of FSCs were of (K1, K3, K5, K6 and Y1) (Table 6). The also within 10% of the expected values. We analyzed 12 concentrations increased along Kanzaki River and Ai field blanks, and no analytes were detected above the River, reaching a maximum at the mouth of the Aigawa LOD (Table 4). Ryuiki water disposal site (A5), where 67,000 ng/L of The concentrations of PFOA and PFOS in surface water PFOA was recorded. If we assume that 270,000 m3/d is samples from rivers and seas collected from all over Japan discharged from A5, the total PFOA discharged from this (Fig. 3) were determined and the results are summarized site was estimated to be 18 kg/d (67,000 ng/L × 270,000 in Table 5. m3/d × 10Ð12 × 103). A log-linear relationship was found between PFOA and PFOS concentrations in Yodo River were high at water PFOS concentrations in Japan. The PFOA concentrations discharge sites (Y9, Y15, Y16). The concentrations in in surface water (Y) were significantly correlated with Kanzaki River were increased from K6 to K12 (Table 6). PFOS concentrations (X) as expressed by Y=1.8 × 100.717X The Osaka International Airport is upstream of K12. The (r=0.60, p<0.01, N=85). systematic search revealed a highly contaminated site of Both PFOA and PFOS concentrations were greater in PFOS (O4 ) (Table 6). Kinki district than in other districts (ANOVA, p<0.01). The coastal sea water samples also showed a similar trend: Tap water contamination levels in the Osaka area the PFOA and PFOS concentrations were highest in the Several cities collect water from Yodo River (K15) Koshien Coast (Hyogo). (Hanshin area, and Osaka city). The water supply for Hanshin area is a mixture from various sources of water Sources of PFOA and PFOS contamination in the Osaka including Yodo River. In contrast, Osaka city derives its area water supply mostly from Yodo River at sites downstream To search for sources of PFOA, we systematically of Y8. collected surface water samples from Yodo River and The concentrations of PFOA and PFOS are summarized Kanzaki River. Yodo River runs from to Osaka Bay, and the width is 300 m downstream of Y10 (Fig. 4) with a flow rate of 600,000 m3/h. Kanzaki River (100,000 m3/h at K9) runs in parallel with the Yodo River.

Fig. 3. A map of Japan and the sampling sites. Norimitsu SAITO, et al.: Surface Water Contamination with PFOA and PFOS in Japan 55

Table 5. PFOA and PFOS concentrations in surface water in Japan

Site1) 2) Name Prefecture City/Town District PFOA (ng/L) PFOS (ng/L) Sampling date

1 Kusiro River Hokkaido Kushiro Hokkaido-Tohoku 0.40 1.90 2003.3.10 2 Aomori Rokkasho Hokkaido-Tohoku 0.66 2.63 2003.3.11 3 Aomori Hokkaido-Tohoku 0.59 0.60 2003.3.11 4 Takamatsu Pond Iwate Morioka Hokkaido-Tohoku 3.77 1.15 2003.3.13 5 Akita Honjo Hokkaido-Tohoku 0.55 1.40 2003.3.14 6 Yamagata Murayama Hokkaido-Tohoku 1.04 3.24 2003.3.15 7 Mogami River Yamagata Murayama Hokkaido-Tohoku 4.22 1.77 2003.3.15 8 Aka River Yamagata Hokkaido-Tohoku 0.47 2.82 2003.3.15 9 Kitagami River Miyagi Kahoku Hokkaido-Tohoku 3.58 3.27 2003.3.16 10 Naruse River Miyagi Onoda Hokkaido-Tohoku 0.10 0.90 2003.3.16 11 Shiroishi River Miyagi Shiroishi Hokkaido-Tohoku 0.83 0.58 2003.3.16 12 Fukushima Kouriyama Hokkaido-Tohoku 1.40 4.62 2003.3.17 13 Natsui River Fukushima Iwaki Hokkaido-Tohoku 1.10 0.45 2003.3.17 14 Abukuma River Fukushima Fukushima Hokkaido-Tohoku 0.89 0.39 2003.3.17 15 Suriage River Fukushima Fukushima Hokkaido-Tohoku 2.55 0.25 2003.3.17 16 Matsu River Fukushima Fukushima Hokkaido-Tohoku 2.95 0.58 2003.3.17

GM (GSD) 0.97 (3.06) 1.19 (2.44) 17 Naka River Tochigi Batou Kanto 1.67 0.33 2003.3.18 18 Kido River Tochigi Ujiie Kanto 3.87 4.98 2003.3.18 19 Gunma Maebashi Kanto 3.24 3.53 2003.3.19 20 Gunma Higashimurayama Kanto 3.03 3.31 2003.3.19 21 Tone River Gunma Ohizumi Kanto 6.40 1.78 2003.3.19 22 Ara River Saitama Kanto 7.56 19.88 2003.3.20 23 Ara River Saitama Kawaguchi Kanto 14.46 19.61 2003.3.20 24 Saitama Saitama Kanto 5.65 18.44 2003.3.20 25 Tokyo Okutama Kanto 0.40 1.74 2003.3.17 26 Tama River Tokyo Oume Kanto 0.33 1.00 2003.3.17 27 Tama River Tokyo Hamura Kanto 0.36 2.42 2003.3.17 28 Tama River Tokyo Akishima Kanto 2.55 0.72 2003.3.17 29 Tama River Kanagawa Kawasaki Kanto 15.08 31.42 2003.3.19 30 Kanagawa Kawasaki Kanto 4.89 3.66 2003.3.19

GM (GSD) 2.84 (3.56) 3.69 (3.93) 31 Niigata Ojiya Chubu 1.96 6.04 2003.3.15 32 Shinano River Niigata Niigata Chubu 3.74 0.91 2003.3.15 33 Niigata Mikawa Chubu 1.53 0.87 2003.3.15 34 Agano River Niigata Niigata Chubu 2.86 0.62 2003.3.15 35 Niigata Jyouetsu Chubu 3.50 2.00 2003.3.15 36 Seki River Niigata Itakura Chubu 3.63 1.96 2003.3.15 37 Tenryu River Shizuoka Hamakita Chubu 2.04 0.42 2003.3.16 38 Kiku River Shizuoka Kikukawa Chubu 2.36 0.41 2003.3.16 39 Ohi River Shizuoka Ooigawa Chubu 2.28 0.73 2003.3.16 40 Shizuoka Shizuoka Chubu 0.28 0.86 2003.3.16 41 Shizuoka Shibakawa Chubu 5.08 1.98 2003.3.16 42 Aichi Toyohashi Chubu 1.64 2.32 2003.3.13 43 Yasaku River Aichi Okazaki Chubu 2.79 0.96 2003.3.13 44 Shonai River Aichi Nagoya Chubu 1.50 0.24 2003.3.13 45 Gifu Kaizu Chubu 16.28 3.88 2003.3.13 46 Gifu Kasamatsu Chubu 2.17 0.46 2003.3.13 47 Gifu Kaizu Chubu 3.88 1.33 2003.3.13

GM (GSD) 2.50 (2.23) 1.07 (2.36) 56 J Occup Health, Vol. 46, 2004

(continued)

Site1) 2) Name Prefecture City/Town District PFOA (ng/L) PFOS (ng/L) Sampling date

48 Kino River Wakayama Iwade Kinki 2.14 1.45 2003.3.17 49 Shiga Yasu Kinki 10.34 4.12 2003.3.19 50 Yodo River Osaka Higashiyodogawa Kinki 140.56 9.29 2003.3.12 51 Yodo River Osaka Miyakojima Kinki 24.42 10.43 2003.3.12 52 Osaka Suminoe Kinki 41.60 18.01 2003.3.12 53 Ina River Hyogo Kawanishi Kinki 4.88 3.81 2003.3.13 54 Ina River Hyogo Kawanishi Kinki 5.70 0.78 2003.3.13 55 Ina River Hyogo Amagasaki Kinki 456.41 37.32 2003.3.13

GM (GSD) 21.15 (6.16)** 5.73 (3.61) 56 Takanashi River Okayama Soujya Chugoku 1.41 0.87 2003.3.11 57 Okayama Okayama Chugoku 8.11 0.80 2003.3.11 58 Okayama Kumayama Chugoku 1.00 0.67 2003.3.11 59 Oota River Hiroshima Hiroshima Chugoku 1.22 0.65 2003.3.13 60 Hiroshima Fukuyama Chugoku 0.68 0.82 2003.3.13 61 Gouno River Hiroshima Miyoshi Chugoku 1.56 0.42 2003.3.13 62 Tottori Kurayoshi Chugoku 1.66 0.55 2003.3.10 63 Kii River Tottori Izumo Chugoku 3.18 25.10 2003.3.10 64 Sanami River Yamaguchi Houfu Chugoku 0.51 0.69 2003.3.12

GM (GSD) 1.51 (2.28) 1.00 (3.42) 65 Onnga River Fukuoka Kaho Kyushu-Shikoku 0.20 1.51 2003.3.21 66 Onnga River Fukuoka Nakama Kyushu-Shikoku 1.71 0.96 2003.3.21 67 Fukuoka Kurume Kyushu-Shikoku 2.64 1.72 2003.3.21 68 Kase River Saga Mikitsuki Kyushu-Shikoku 1.29 0.29 2003.3.19 69 Matsuura River Saga Ouchi Kyushu-Shikoku 1.61 0.94 2003.3.19 70 Yamakuni River Ohita Yamakuni Kyushu-Shikoku 1.38 0.35 2003.3.11 71 Yamakuni River Ohita Nakatsu Kyushu-Shikoku 3.28 0.55 2003.3.11 72 Motoake River Nagasaki Isahaya Kyushu-Shikoku 0.95 0.57 2003.3.13 73 Tokushima Tokushima Kyushu-Shikoku 2.78 0.59 2003.3.14 74 Doki River Kagawa Marukame Kyushu-Shikoku 13.82 14.86 2003.3.15 75 Hiji River Ehime Oosu Kyushu-Shikoku 2.78 3.53 2003.3.16 76 Shigenobu River Ehime Shigenobu Kyushu-Shikoku 2.50 0.30 2003.3.16 77 Niyodo River Kouchi Tosa Kyushu-Shikoku 2.60 2.40 2003.3.18 78 Kouchi Kubokawa Kyushu-Shikoku 2.46 0.40 2003.3.18 79 Mononobe River Kouchi Noichi Kyushu-Shikoku 1.35 0.24 2003.3.18

GM (GSD) 1.93 (2.40) 0.89 (3.09) AThe Sea of Kushiro Hokkaido Kushiro Hokkaido-Tohoku 1.90 2.12 2003.4.10 BMutsu Bay Aomori Noheji Hokkaido-Tohoku 2.09 0.61 2003.4.12 C Honjo Marina Akita Honjyo Hokkaido-Tohoku 2.13 0.87 2003.4.13 D Funabashi Coast Chiba Chiba Kanto 32.21 2.58 2003.3.17 E Pacific Ocean Aichi Toyohashi Chubu 11.52 0.65 2003.3.13 F Koshien Coast Hyogo Nishinomiya Kinki 447.74 27.69 2003.3.13

**: Significantly higher than other areas by ANOVA (p<0.01), 1) For sites, refer to Fig. 3., 2) The mean of the duplicate one-L preparations for each sampling site Norimitsu SAITO, et al.: Surface Water Contamination with PFOA and PFOS in Japan 57

Fig. 4. A map of Osaka area and the sampling sites.

Table 6. Systematic search for the source of contaminations in Osaka area

Site1) 2) PFOA PFOS Site1) 2) PFOA PFOS (ng/L) (ng/L) Sampling date (ng/L) (ng/L) Sampling date

Y1 562.8 6.5 2003.4.11 K1 215 4.1 2003.4.15 Y2 463.1 5.8 2003.4.11 K2 41 11.0 2003.4.15 Y3 313.3 5.8 2003.4.11 K3 153 3.3 2003.4.15 Y4 271.1 6.3 2003.4.11 K4 45 1.4 2003.4.15 Y5 46.8 13.3 2003.4.11 K5 122 2.5 2003.4.15 Y6 104.0 4.9 2003.4.11 K6 1,040 10.9 2003.4.15 Y7 31.1 11.0 2003.4.11 K7 1,270 18.2 2003.4.15 Y8 31.8 10.1 2003.4.11 K8 1,690 15.7 2003.4.15 Y9 146.8 26.9 2003.4.11 K9 3,750 23.3 2003.4.15 Y10 33.7 7.3 2003.4.11 K10 430 28.8 2003.4.15 Y11 11.3 3.4 2003.4.11 K11 81 9.4 2003.4.15 Y12 18.5 2.6 2003.4.11 K12 48 86.2 2003.4.15 Y13 80.9 27.7 2003.4.11 K13 4,220 14.1 2003.4.15 Y14 30.8 7.5 2003.4.11 K14 7,990 13.9 2003.4.15 Y15 64.9 24.8 2003.4.11 K15 101 7.0 2003.4.15 Y16 39.7 24.1 2003.4.11 K16 531 6.2 2003.4.15 Y17 6.6 1.7 2003.4.11 A1 19,400 11.7 2003.4.15 Y18 6.8 2.6 2003.4.11 A2 24,080 9.1 2003.4.15 Y19 4.5 2.5 2003.4.11 A3 39,500 8.3 2003.5.9 Y20 9.3 3.6 2003.4.11 A4 42,950 6.1 2003.5.9 Y21 7.5 1.5 2003.4.11 A5 67,000 13.0 2003.5.9 O1 39.2 96.0 2003.3.10 A6 124 1.9 2003.5.9 O2 40.8 9.6 2003.3.10 A7 76 1.8 2003.5.9 O3 36.4 57.2 2003.3.10 A8 3,750 20.2 2003.5.9 O4 56.6 526.0 2003.3.10 O5 64.4 73.2 2003.3.10 O6 55.0 10.2 2003.3.10 O7 41.2 30.4 2003.3.10

1) For sites, refer to Fig. 4, 2) The mean of the duplicate one-L preparations for each sampling site 58 J Occup Health, Vol. 46, 2004

Table 7. PFOA and PFOS concentration levels in tap water1)

PFOA (ng/L) PFOS (ng/L) District Sampling date Prefecture Local Area n GM GSD GM GSD

Osaka area 2003.4.12 Hyogo Hanshin Area 5 12.5B 1.6 1.1B 4.3 2003.4.24 Osaka Osaka City 5 40.0A 1.1 12.0A 1.1 2003.4.21 Kyoto Kyoto City 5 5.4C 1.5 4.9A 2.0 Tohoku area 2003.4.15 Iwate Morioka City 5 0.7D 1.5 0.2B 2.0 2003.4.18 Miyagi Sendai City 5 0.13E 1.3

Hanshin Area: Amagasaki City and Kobe City, 1) Superscript letters indicate the results of Scheffe test after ANOVA. The same supercript letter is not significantly different, but the different letters indicate significant differences (p<0.05). Concentrations less than LOD were assumed to be 0.1 ng/L for calculations. in Table 7. PFOA and PFOS concentrations were highest be addressed. Further researches on workers and residents in the tap water in Osaka city. PFOA concentrations, in are warranted for risk assessment. particular, were significantly higher than in other cities. Acknowledgments: This project was funded by Nissei Discussion Zaidan and also supported by a Health and Labor Sciences The quantification of surface water samples collected Research Grants (15210301). We are grateful to Ms Ami from all over Japan showed surface water contamination Ito, Shigi Kaku and Aiko Tamada for their help in with PFOA and PFOS. We found a large geographical sampling water. difference in their levels. References For PFOS, the levels determined in the present study were in accord with those determined in our previous 1) Federal Register: April 16, 68, Number 73. 2003: study16) suggesting that the contamination profile of PFOS 18626Ð18633. is reproducible. 2) LB Biegel, ME Hurt, SR Frame, JC O’Connor and JC Cook: Mechanisms of extrahepatic tumor induction by The geographical systematic search revealed a highly 18) peroxisome proliferators in male CD rats. Toxicol Sci contaminated site in the airport. In our previous study , 60, 44Ð55 (2001) we identified as a source of contamination a water 3) J Butebhoff,G Costa G, C Elcombe, D Farrar, K discharge site which collected a large amount of disposed Hansen, H Iwai, R Jung, G Jr Kennedy, P Leider, G water discharge from the Yokota Air Force Base. These Olsen and P Thomford: Toxicity of ammonium lines of evidence may suggest that fire-fighting foams, perfluorooctanoate in male cynomolgus monkeys after which contain PFOS and had been used but have been oral dosing for 6 months. Toxicol Sci 69, 244Ð257 banned recently, are still contaminating soil and water (2002) with PFOS even long after their use has ceased, as 4) FD Gilliland and JS Mandel: Mortality among reported by Moody15). employees of a perfluorooctanoic acid production plant. For PFOA, the contamination levels were significantly J Occup Med 35, 950Ð954 (1993) 5) F Gilliland: Fluorocarbonds and human health: studies greater in the Osaka area than in other geographic areas. in an occupational cohort. Ph.D. Thesis Minneapolis, The present study was able to find the highest MN: University of Minnesota; 1992 contamination levels at the Aigawa Ryuiki water disposal 6) FD Gilliland and JS Mandel: Serum perfluorooctanoic site. This site discharges an estimated 18 kg of PFOA acid and hepatic enzymes, lipoproteins, and cholesterol: daily. The levels of contaminations were unexpectedly A study of occupationally exposed men Am J Ind Med higher than those in another report14). High levels of 29, 560Ð568 (1996) contamination with PFOA in the Osaka area warrant 7) GW Olsen, FD Gilliland, MM Burlew, JM Burris, JS further study to find the sources of PFOA in this area. A Mandel and JH Mandel: An epidemiological future mass balance study in the Osaka area will give us investigation of reproductive hormones in men with a clue in terms of sources of PFOA. occupational exposure to perfluorooctanoic acid. J The effects of exposure to PFOA at 40 ng/L through Occup Environ Med 40, 614Ð622 (1998) 8) GW Olsen, JM Burris, MM Burlew and JH Mandel: drinking water ingestion remain unknown, although there Epidemiological assessment of worker serum is a report that PFOA may increase the risk of prostate perfluoroocatne sulfonate (PFOS) and 4) cancer . The fact that more than one million people are perfluorooctanoate (PFOA) concentrations and medical estimated to drink water at this contamination level should Norimitsu SAITO, et al.: Surface Water Contamination with PFOA and PFOS in Japan 59

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