Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 7-Day Trial for You or Your Team.

Learn More →

What’s in the Pool? A Comprehensive Identification of Disinfection By-products and Assessment of Mutagenicity of Chlorinated and Brominated Swimming Pool Water

What’s in the Pool? A Comprehensive Identification of Disinfection By-products and Assessment of... Research What’s in the Pool? A Comprehensive Identification of Disinfection By-products and Assessment of Mutagenicity of Chlorinated and Brominated Swimming Pool Water 1 2 3,4,5,6 7 7 Susan D. Richardson, David M. DeMarini, Manolis Kogevinas, Pilar Fernandez, Esther Marco, 7 7 8 8 9 10 Carolina Lourencetti, Clara Ballesté, Dick Heederik, Kees Meliefste, A. Bruce McKague, Ricard Marcos, 3,4 7 3,4,5 Laia Font-Ribera, Joan O. Grimalt, and Cristina M. Villanueva 1 2 National Exposure Research Laboratory, U.S. Environmental Protection Agency, Athens, Georgia, USA; National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, USA; 3 4 Centre for Research in Environmental Epidemiology, Barcelona, Spain; Municipal Institute of Medical Research, Hospital del 5 6 Mar, Barcelona, Spain; CIBER Epidemiología y Salud Pública, Barcelona, Spain; Medical School, University of Athens, Greece; 7 8 Department of Environmental Chemistry, Institute of Environmental Assessment and Water Research, Barcelona, Spain; Institute for Risk Assessment Sciences, Division for Environmental Epidemiology, Utrecht University, Utrecht, the Netherlands; CanSyn Chem. Corp., Toronto, Ontario, Canada; Grup de Mutagènesi, Departament de Genètica i de Microbiologia, Edifici Cn, Universitat Autònoma de Barcelona, Bellaterra, Cerdanyola del Vallès, Spain with asthma and other respiratory effects in Background : Swimming pool disinfectants and disinfection by-products (DBPs) have been linked Olympic swimmers and pool workers, and less to human health effects, including asthma and bladder cancer, but no studies have provided a com - clearly with recreational adult swimmers and prehensive identification of DBPs in the water and related that to mutagenicity. children (Goodman and Hays 2008; Jacobs o Bjectives : We performed a comprehensive identification of DBPs and disinfectant species in waters et al. 2007; Stav and Stav 2005; Weisel et al. from public swimming pools in Barcelona, Catalonia, Spain, that disinfect with either chlorine or bro- 2009). However, the mechanisms are poorly mine and we determined the mutagenicity of the waters to compare with the analytical results. understood, and it is not known with certainty Methods : We used gas chromatography/mass spectrometry (GC/MS) to measure trihalomethanes whether trichloramine or other volatile pool in water, GC with electron capture detection for air, low- and high-resolution GC/MS to compre- DBPs are responsible. hensively identify DBPs, photometry to measure disinfectant species (free chlorine, monochloro- Despite the public health relevance, only amine, dichloramine, and trichloramine) in the waters, and an ion chromatography method to a few studies, most rather recent, have inves- measure trichloramine in air. We assessed mutagenicity with the Salmonella mutagenicity assay. tigated the chemistry and potential health results : We identified > 100 DBPs, including many nitrogen-containing DBPs that were likely effects of swimming pool water (Weisel et al. formed from nitrogen-containing precursors from human inputs, such as urine, sweat, and skin 2009; Zwiener et al. 2007). A complete cells. Many DBPs were new and have not been reported previously in either swimming pool or chemical characterization of DBPs in indoor drinking waters. Bromoform levels were greater in brominated than in chlorinated pool waters, swimming pools has not been reported. The but we also identified many brominated DBPs in the chlorinated waters. The pool waters were only mutagenicity study of swimming pool mutagenic at levels similar to that of drinking water (~ 1,200 revertants/L-equivalents in strain water reported that organic extracts from TA100–S9 mix). three public indoor pools in Victoria, British c onclusions : i Th s study identie fi d many new DBPs not identie fi d previously in swimming pool or drinking water and found that swimming pool waters are as mutagenic as typical drinking waters. Address correspondence to S.D. Richardson, National k e y w o r d s : bromination, bromine, chlorination, chlorine, DBPs, disinfection by-products, Exposure Research Laboratory, U.S. Environmental mutagenicity, swimming pools, Salmonella, water. Environ Health Perspect 118:1523–1530 Protection Agency, 960 College Station Rd., Athens, GA 30605 USA. Telephone: (706) 355-8304. Fax: (2010). doi:10.1289/ehp.1001965 [Online 12 September 2010] (706) 355-8302. E-mail: richardson. [email protected] Supplemental Material is available online (doi:10. 1289/ehp.1001965 via http://dx.doi.org/). Disinfection by-products (DBPs) represent air due to continuous disinfection and con- We thank T. Lewis, G. Crumley, C. Lindell, a ubiquitous exposure in developed coun- stant organic load from bathers (e.g., urine, J. Hartmann, and H. Fink for their assistance with tries. DBPs are formed by the reaction of dis- sweat, cosmetics, skin cells, and hair) (Kim extractions, derivatizations, and analyses. We thank infectants (e.g., chlorine, chloramines, ozone, et al. 2002; LaKind et al. 2010). One of the A. Kligerman for his help with statistical analyses, and or chlorine dioxide) with natural organic most prevalent DBPs in chlorinated swim- A. Kligerman and L. Claxton for helpful comments on the manuscript. matter and/or bromide/iodide, and they are ming pools is THMs (Aggazzotti and Predieri This research was supported by the U.S. Environ - an unintended consequence of trying to kill 1986; Beech et al. 1980; Judd and Jeffrey mental Protection Agency (EPA) intra mural research pathogens in drinking water and swimming 1995), with average concentrations ranging program and Spanish grants SAF2005-07643- pools. More than 600 DBPs have been iden- from 16 µg/L (Santa Marina et al. 2009) to C03-01 (Plan Nacional) and CP06/00341 (Fondo tified in drinking water, and many of them 132 µg/L (Chu and Nieuwenhuijsen 2002). de Investigación Sanitaria). C.M.V. and L.F.-R. have, are mutagenic or carcinogenic (Richardson Given the high nitrogen content of organic respectively, a contract and a predoctoral fellowship by the Instituto de Salud Carlos III (CP06/00341, 1998; Richardson et al. 2007). This complex matter from bathers, nitrogenated species FI06/00651). C.L. acknowledges a grant from the mixture of DBPs includes volatile and skin- such as haloacetonitriles, nitrosamines, and Agreement between Santander-Central Hispano and permeable DBPs, such as trihalomethanes chloramines are found in swimming pool Consejo Superior de Investigaciones Científicas. (THMs) and haloketones (Erdinger et al. water (Héry et al. 1995; Kim et al. 2002; The manuscript has been reviewed in accordance 2004: Xu and Weisel 2004, 2005). Inhalation Walse and Mitch 2008; Zwiener et al. 2007). with the U.S. EPA’s peer and administrative review and dermal absorption, which are the primary Chronic exposure to DBPs through differ - policies and approved for publication. Mention of trade names or commercial products does not consti- routes of exposure to DBPs during swim- ent routes has been associated with an increased tute endorsement or recommendation for use by the ming, leads to higher blood levels of THMs risk for bladder cancer (International Agency U.S. EPA. than do oral exposures (Ashley et al. 2005; for Research on Cancer 2004; Villanueva et al. A.M. is employed by CanSyn Chem. Corp., Toronto, Haddad et al. 2006; Leavens et al. 2007). 2004, 2007). Trichloramine and other volatile Ontario, Canada. The authors declare they have no Swimming pools constitute environ- chemicals in swimming pools are respiratory actual or potential competing financial interests. ments with high levels of DBPs in water and irritants; pool attendance has been associated Received 18 January 2010; accepted 8 June 2010. | | Environmental Health Perspectives • volume 118 number 11 November 2010 1523 Richardson et al. Columbia (Canada), were mutagenic in for THM measurements were quenched with pool (16 July and 15 October 2007). Samples Salmonella TA100 (Honer et al. 1980). The 5 mg sodium thiosulfate and stored at 4°C (28 L each) were collected using 2-L Teflon authors found that acidified extracts eluted until analysis on the same day. Chloroform, bottles (headspace-free) and were shipped with ether were more mutagenic in the pres- bromodichloro methane, dibromochloro - overnight in coolers with icepacks to the U.S. ence of metabolic activation (rat liver S9) than methane, and bromoform were measured Environmental Protection Agency laboratory without S9; however, nonacidified extracts using purge-and-trap gas chromatography/ in Athens, Georgia (USA). Water samples eluted with acetone were mutagenic only in mass spectrometry (GC/MS) (Tekmar 3100, were extracted immediately upon arrival using the absence of S9. One genotoxicity study of Voyager MS; ThermoFisher, Waltham, MA, the XAD resin process of Richardson et al. swimming pool water reported that the water USA) following the method described by (2008) [for further details, see Supplemental and its fractions induced DNA damage in Lourencetti et al. (2010). Sixty-eight samples Material (doi:10.1289/ehp.1001965)]. The Hep-G2 cells (comet assay) and that most of were collected from the chlorinated pool final extract was divided for comprehensive the genotoxicity was in the lower-molecular- and 12 from the brominated pool for these GC/MS analysis (1.0 mL, equivalent to 20 L weight DBP fraction (Glauner et al. 2005). quantita tive analyses. water) and mutagenicity analysis (0.4 mL, Another study using the comet assay showed Indoor air samples to measure THMs equivalent to 8 L water, or 20,000×). that pool water was more genotoxic than the were collected with a pump located 60 cm Comprehensive GC/MS analyses. Half source tap water and that the type of disinfec- above the floor and 1.5 m from the pool bor- of the 1.0-mL extract was derivatized with tant and illumination conditions altered the der. Air was pumped (7 mL/min) for 20 min diazomethane [see Supplemental Material genotoxicity (Liviac et al. 2010). through a Tenax TA cartridge (1.8 g; Supelco, (doi:10.1289/ehp.1001965)] to enable the The present study involves an investiga- Sigma-Aldrich, St. Louis, MO, USA). Quality identification of haloacids (through their cor- tion in Barcelona, Spain, where we examined control was assured by daily calibration of the responding methyl esters); the other half was 49 healthy nonsmoking volunteers before and pump. Chloroform, bromodichloromethane, analyzed directly for other DBPs. after swimming in public swimming pools dibromochloromethane, and bromoform were Comprehensive GC/MS analyses were per- treated with either chlorine or bromine to determined through an automatic thermal formed on a high-resolution magnetic sector evaluate personal exposure and a range of bio- desorption unit (ATD 400; Perkin-Elmer, mass spectrometer (Autospec; Waters, Inc., markers of genotoxicity and respiratory dam- Madrid, Spain) coupled to a GC-electron cap- Milford, MA, USA) equipped with an Agilent age (Font-Ribera et al. 2010; Kogevinas et al. ture detector (Perkin-Elmer). Sixty-eight air model 6890 gas chromatograph (Agilent, Santa 2010). To complement the exposure assess- samples were collected from the chlorinated Clara, CA, USA) and operated at an accelerat- ment, we evaluated the mutagenicity of the pool, and 12 from the brominated pool. ing voltage of 8 kV and source temperature pool waters in the Salmonella mutagenicity Trichloramine was measured in pool of 200°C, in both low-resolution (1,000) and assay and screened for DBPs, comprehensively air samples by pumping air (1.2 L/min) for high-resolution (10,000) modes. Injections of identifying most DBPs detected and quan- 115 min, within 1 m from the water and at 1 µL extract were introduced via a split/splitless tifying a few targeted DBPs and disinfectant a height of 60 cm from the o fl or level, using injector (in splitless mode) onto a GC column species (THMs, chlorine, monochloramine, a method described originally by Héry et al. (DB-5, 30-m × 0.25-mm i.d. 0.25-µm film dichloramine, and trichloramine) in the pool (1995). Trichloramine was captured on two thickness; J&W Scientific/Agilent, Santa Clara, waters and in the air phase above the water 37-mm quartz-fiber filters, one of which was CA, USA). The GC temperature program con - (THMs and trichloramine). In this article placed as a backup filter, both impregnated with sisted of an initial temperature of 35°C (4 min) we present a comprehensive identification 500 mL of a solution of diarsenic tri oxide (4 g/L and an increase at 9°C/min to 285°C (held for of DBPs and disinfectant species in the pool As O ), sodium carbonate (40 g/L Na CO ), 30 min). Transfer lines were held at 280°C, 2 3 2 3 waters and compare the species formed in and glycerol (40 mL/L C H O ). These fil- and the injection port at 250°C. 3 8 3 chlorinated versus brominated pool water with ters were placed in a sampling cassette with For qualitative identifications, mass spectra the corresponding mutagenicity of the waters. a 37-mm cellulose support filter and a 37-mm of unknown compounds in the finished and Teflon filter to prevent chloride from airborne raw water concentrates were subjected initially Materials and Methods water droplets from being captured in the to library database searching (using the 2005 Sampling. Water samples were collected from sampler. Impregnated filters were de sorbed in NIST Mass Spectral Library database; National two large public swimming pools in Barcelona, 10-mL ultra-high quality, ultrapure water (spe- Institute of Standards and Technology, Spain. One pool (33 × 25 × 2 m in size) used cific conductivity, 17.8 MΩ/cm at 25°C), soni- Gaithersburg, MD). However, many DBPs chlorine (sodium hypochlorite) for disinfec - cated for 30 min, and centrifuged for 15 min were not present in the library database; in tion, after sand filtration; the other pool at 3,000 × g after sampling. Trichloramine those cases, and also where a library match was (20.9 × 13.2 × 1.3 m in size) used bromine was reduced to chloride and subsequently ana- insufficient to offer a tentative identification, ( 1 - b r o m o - 3 - c hl o r o - 5 , 5 - d i m e t h y l - 2 , 4 - lyzed by ion chromatography (Dionex DX100; high-resolution MS was used to provide empir- imidazolidinedione) for disinfection, after sand Dionex BV, Bavel, the Netherlands; AS14A ical formulas for molecular ions and fragments. and granulated carbon filtration. Floor-to- guard and AS14 highly selective anion column Mass spectra were also interpreted extensively ceiling height was 10 m and 5 m in the chlori- with self-regenerating suppressor; conductivity to provide tentative structural identifications. nated and brominated pools, respectively. detector; flow rate , 1.0 mL/min). Six samples When possible, pure standards were obtained Quantitative analyses. Free chlorine, were collected from the chlorinated pool, and to confirm identifications through a match of monochloramine, dichloramine, trichloramine, three from the brominated pool. GC retention times and mass spectra. and THMs were measured in composite pool Preparation of water extracts and con- Chemical standards. Chemical DBP stan- water samples (1 L) collected from four differ - centrates. For comprehensive GC/MS analy- dards were either synthesized (CanSyn Chem. ent locations. Free chlorine, monochloramine, ses and mutagenicity testing, pool water Corp., Toronto, ON, Canada) or purchased dichloramine, and trichloramine were meas- samples were collected at approximately at the highest level of purity (Sigma-Aldrich, ured immediately using the N,N-diethyl-p- noon on five different sampling events for Milwaukee, WI, USA). The synthesis of phenylenediamine (DPD) method with a the chlorinated pool (7 and 24 May, 11 (E)- and (Z)-bromochlorobutenedioic acid portable photometer (DINKO Instruments, June, and 17 September 2007) and two dif- is presented in Supplemental Material Barcelona, Spain). Water samples (40 mL) ferent sampling events for the brominated (doi:10.1289/ehp.1001965). | | 1524 volume 118 number 11 November 2010 • Environmental Health Perspectives DBPs in and mutagenicity of swimming pool water Mutagenicity assays. The 20,000× in the DMSO control. The positive control OCH ] suggested the presence of a carboxy- XAD/ethyl acetate extracts described above for all strains was sodium azide at 3 µg/plate. lic acid methyl ester in the structures, with a were solvent-exchanged into dimethyl sulfox- We calculated linear regressions, slope molecular ion of m/z 256/258/260. Further, ide (DMSO; Burdick and Jackson, Muskegon, values, the standard error of the slopes, and the m/z 256/258/260 isotopic pattern was MI, USA) and diluted to 10,000× and r values of the dose–response curves and then indicative of one bromine and one chlorine 1,000×. We performed the standard plate- compared the regression lines between strains atom, matching the calculated theoretical pat- incorporation Salmonella (Ames) mutagen- RSJ100 (GST+) and TPT100 (GST–) to tern [see Supplemental Material, Figure S2 icity assay (Maron and Ames 1983) in the obtain p-values using Statgraphics Centurion (doi:10.1289/ehp.1001965)]. This pattern base-substitution strain TA100 (hisG46 rfa XVI (Statpoint Technologies, Inc., Warrenton, results from the overlap of the two natural 79 81 ΔuvrB, pKM101), obtained from B.N. Ames, VA, USA). The model tests the null hypothesis isotopes of bromine ( Br and Br) with the Children’s Hospital Oakland Research that the slopes are equal; we set α = 0.05 for two natural isotopes of chlorine ( Cl and Institute (Oakland, CA, USA). We also tested the F-test with 2 degrees of freedom. Cl). This information suggested a tenta- the extracts in Salmonella strain RSJ100, which tive structural identification of bromochlo- Results expresses the rat GSTT1 gene, and its control robutenedioic acid dimethyl ester, with a strain TPT100. These strains are homologous DBPs. Table 1 lists levels of free chlorine, monoisotopic molecular mass of 256 Da. to TA100 except that they do not contain chloramines, and THM species in the pool Exact mass data provided by high-resolu- the pKM101 plasmid and either do or do water and air. Although we did not detect tion MS supported this empirical formula not express GSTT1 (Thier et al. 1993). These trichloramine in the pool waters, we did n fi d (C H O ClBr). The observed exact mass of 6 6 4 strains were obtained from F.P. Guengerich mean levels of 0.29 and 0.08 mg/m in the the stronger molecular ion isotopic peak (m/z (Vanderbilt University, Nashville, TN, USA). chlorinated and brominated pool air, respec- 257.9116) was within 0.0002 Da of the theo- We did not use S9 mix because we assumed tively, indicating that most of it volatilized retical mass (m/z 257.9118). This supported that pool water was similar to drinking water, from the water into the air (Font-Ribera et al. the general structure of bromochlorobutene- and drinking water extracts are most muta- 2010). We identified > 100 DBPs compre- dioic acid dimethyl ester; however, the exact genic in the absence of S9 mix (Takanashi hensively in the pool waters [Table 2; see also isomer assignments could not be made by MS et al. 2009). Supplemental Material, Figure S1 (doi:10.1289/ data alone because the spectra were too simi- Extracts were tested up to 100 µL/plate ehp.1001965)], including a large number of lar, which is often the case for isomers. Two over a dose range of 0.01–0.3 L-equivalents haloacids, halomethanes, haloacetonitriles, structural isomers are possible for this empiri- (L-eq)/plate based on doses used for drink- haloaldehydes, haloketones, halonitromethanes, cal formula, (Z) and (E), representing cis and ing water (DeMarini et al. 1995) and a dose- haloamides, haloalcohols, and halophenols. All trans isomers, respectively (Figure 1A). range–finding study. Because of limited of these contained either bromine or chlorine; Fortunately, we observed both compounds amounts of samples available for testing in all we detected no iodinated DBPs. Most DBPs in most of the pool water concentrates, so it just three bacterial strains, only two samples from have not been reported previously for swim- remained to be determined which specific iso - the chlorinated pool (C4 and C5) and two ming pool waters, and many were not present mer represented each GC/MS chromatographic from the brominated pool (B1 and B2) were in the mass spectral library database. peak. To make this determination, we synthe- evaluated for mutagenicity, each at one plate The identification of ( E)- and (Z)-bromo- sized the two possible isomers [see Supplemental per dose in single experiments. We incubated chloro butenedioic acid (in their corresponding Material (doi:10.1289/ehp.1001965)] and con- the plates for 3 days at 37°C, counted the methyl ester forms) illustrates how we identi- firmed by a match of the GC retention time colonies [revertants (rev)] on an automatic fied unknown DBPs. They eluted at different and mass spectra that the (Z) isomer is the first colony counter, and calculated linear regres- retention times (Figure 1A) but exhibited sim- peak at 16.8 min and the (E) isomer is the sec- sions over the linear portion of the dose–re- ilar mass spectra (indicative of isomeric struc- ond peak at 16.9 min in the pool water extracts sponse curves to determine the mutagenic tures), each containing m/z 256/258/260, (Figure 1A). potencies (rev/L-eq). We defined a positive 225/227/229, and 59 (Figure 1B). The Mutagenicity. Table 3 and Figure 2 result as a dose-related response with two or loss of 31 Da (typically OCH ) at m/z 225 show the mutagenicity data for two sam- more times the number of revertants observed and the presence of m/z 59 [typically C(O) ples from the chlorinated pool and two Table 1. Free chlorine, chloramine, and THM levels in the swimming pools. Chlorinated pool Brominated pool Chemical and concentration Mean ± SD Minimum Maximum n Mean ± SD Minimum Maximum n Water Free chlorine (mg/L) 1.28 ± 0.43 0.52 2.35 68 0.50 ± 0.16 0.32 0.7 4 Monochloramine (NH Cl) (mg/L) 0.29 ± 0.11 0.10 0.64 68 0.27 ± 0.03 0.24 0.3 4 Dichloramine (NHCl ) (mg/L) 0.38 ± 0.14 < 0.01 0.65 68 < 0.01 < 0.01 < 0.01 4 Trichloramine (NCl ) (mg/L) < 0.10 < 0.10 < 0.10 68 < 0.10 < 0.10 < 0.10 4 Chloroform (CHCl ) (µg/L) 15.4 ± 3.5 8.4 20.8 68 0.2 ± 0.1 0.1 0.3 12 Bromodichloromethane (CHCl Br) (µg/L) 14.2 ± 4.2 9.3 26.8 68 0.4 ± 0.2 0.2 0.7 12 Dibromochloromethane (CHClBr ) (µg/L) 12.8 ± 4.4 6.5 22.6 68 2.4 ± 0.2 2.1 2.7 12 Bromoform (CHBr ) (µg/L) 7.2 ± 3.2 3.0 16.5 68 57.2 ± 4.4 52.0 64.3 12 Total THMs (µg/L) 49.6 ± 10.6 35.2 75.2 68 60.2 ± 4.7 54.4 67.2 12 Air Trichloramine (NCl ) (mg/m ) 0.29 ± 0.10 0.17 0.43 6 0.08 ± 0.01 0.07 0.10 3 Chloroform (CHCl ) (µg/m ) 32.1 ± 11.9 11.9 61.6 68 4.4 ± 2.3 1.7 9.4 12 Bromodichloromethane (CHCl Br) (µg/m ) 14.9 ± 4.5 7.5 23.4 68 2.9 ± 1.0 1.7 4.8 12 Dibromochloromethane (CHClBr ) (µg/m ) 14.0 ± 4.2 6.1 26.2 68 7.3 ± 1.3 6.1 9.7 12 Bromoform (CHBr ) (µg/m ) 11.0 ± 4.6 4.4 22.6 68 74.9 ± 17.6 53.3 101.4 12 Total THMs (µg/m ) 72.1 ± 20.7 44.0 124.9 68 89.5 ± 21.9 63.1 124.7 12 | | Environmental Health Perspectives • volume 118 number 11 November 2010 1525 Richardson et al. from the brominated pool in strain TA100. highest doses (0.04 and 0.05 L-eq/plate). sample B1 was significantly more mutagenic All of the samples were mutagenic in strain Table 4 shows the slopes, r values, and in the GSTT1-expressing strain relative to the TA100 except for sample C5, which was the standard errors of the slopes for these data; nonexpressing strain (Tables 3 and 4). This only sample that showed toxicity—based the average mutagenic potency of the three indicates that some portion of the mutagenic on a reduction of rev/plate in TA100 at the mutagenic samples was 1,190 rev/L-eq. Only activity of sample B1 in strain RSJ100 was due Table 2. DBPs identified in pool waters. Sample Sample DBP C1 C2 C3 C4 C5 B1 B2 DBP C1 C2 C3 C4 C5 B1 B2 Haloalkanes Halonitriles Chloroform x x x x x x x Bromoacetonitrile x x Bromodichloromethane x x x x x x x Dichloroacetonitrile x x x x x Dibromochloromethane x x x x x x x Bromochloroacetonitrile x x x x x x Bromoform x x x x x x x Dibromoacetonitrile x x x x x Dibromomethane x x x x x x x Trichloroacetonitrile x Bromotrichloromethane Haloketones Dibromodichloromethane x Bromopropanone x x 1,1,2-Trichloroethane x x 1,1-Dichloropropanone x x Haloacetic acids 1-Bromo-1-chloropropanone x x Chloroacetic acid x x x x 1,1-Dibromopropanone x x Bromoacetic acid x x x x x 1,3-Dibromopropanone x x Dichloroacetic acid x x x x x 1,1,1-Trichloropropanone x x x x x Bromochloroacetic acid x x x x x x x 1,1,3-Trichloropropanone x x x x Dibromoacetic acid x x x x x x 1-Bromo-1,1-dichloropropanone x x x Trichloroacetic acid x x x x x x x 1,1,1-Tribromopropanone x x x Bromodichloroacetic acid x x x x x x x 1,1,3,3-Tetrachloropropanone x x x x x Dibromochloroacetic acid x x x x x x x 1,1-Dibromo-3,3-dichloropropanone Tribromoacetic acid x x x x x x x Pentachloropropanone x x Other haloacids Dichlorofurandione x x 3-Bromopropenoic acid x 1-Chloro-2-butanone x x 2,2-Dichloropropanoic acid x x x x x 1-Bromo-2-butanone x x 3,3-Dichloropropenoic acid x x x x x Tetrachlorohydroquinone x x x cis-2,3-Bromochloropropenoic acid x x x x x x Halonitromethanes trans-2,3-Bromochloropropenoic acid x x x x x x Dibromonitromethane x x x x x 2,3-Dibromopropanoic acid x x x x x Haloamides cis-2,3-Dibromopropenoic acid x x x x Dichloroacetamide x x x trans-2,3-Dibromopropenoic acid x x x Bromochloroacetamide x x 3,3-Dibromopropenoic acid x x x x Dibromoacetamide x x x x x x x Trichloropropenoic acid x x x x x x x Bromodichloroacetamide x 2-Bromo-3,3-dichloropropenoic acid x x x x x x x Dibromochloroacetamide x x x (E)-3-Bromo-2,3-dichloropropenoic acid x x x x x x x Tribromoacetamide x ( Z)-3-Bromo-2,3-dichloropropenoic acid x x x x x x x Haloalcohols 2,2-Dichlorobutanoic acid x x x x 2,2,2-Trichloroethanol x cis-Bromobutenoic acid x x x x x 1,1,1-Trichloropropanol x x x trans-Bromobutenoic acid x x x Other halogenated DBPs 2,2-Dichlorobutenoic acid x 3-Chlorobenzeneacetonitrile x 2,3-Dibromobutenoic acid x x 2,6-Dichloro-4-methylphenol x x x 2-Chloro-3-methylbutanoic acid x x x x x 2-Bromo-4-chlorophenol x Chlorophenylacetic acid x x Trichlorophenol x x x x x 3,5-Dibromobenzoic acid x Bromodichlorophenol x x x x Tribromopropenoic acid x Tribromophenol x x x Halodiacids 2-Bromo-4-chloro-6-methylphenol x x x x cis-Bromobutenedioic acid x x x x x x x Dibromomethylphenol x x trans-Bromobutenedioic acid x x x x x 2,4-Dibromo-1-methoxybenzene x x cis-Dichlorobutenedioic acid x x x 2,3,4-Trichlorobenzeneamine x x x trans-Dichlorobutenedioic acid x x Dibromochloroaniline x cis-Bromochlorobutenedioic acid x x x x x x 2-Bromo-4-chloroanisole x x x trans-Bromochlorobutenedioic acid x x x x x x x 3,4,5-Tribromo-1H-pyrazole x cis-Dibromobutenedioic acid x x x x x x x 2,6-Dibromo-4-nitrophenol x (E)-2-Chloro-3-methylbutenedioic acid x x 2,6-Dibromo-4-nitrobenzeneamine x x (E)-2-Bromo-3-methylbutenedioic acid x Nonhalogenated DBPs/contaminants Haloaldehydes Propionamide x Dichloroacetaldehyde x x Benzaldehyde x x x x x x x Bromochloroacetaldehyde x x x Benzoic acid methyl ester x Dibromoacetaldehyde x x x x x Benzeneacetonitrile x x Trichloroacetaldehyde (chloral hydrate) x x x x x Phthalic acid x x x Bromodichloroacetaldehyde x x x x Diethylphthalate x Dibromochloroacetaldehyde x x x x x Benzophenone x Tribromoacetaldehyde x x x x x 3-Bromo-4-methoxybenzaldehyde x x x x x x Samples C1–C5 represent five samples from the chlorinated pool; B1 and B2 represent two samples from the brominated pool. “X” indicates that a particular DBP was identified in that sample. DBPs shown in italics were confirmed through the analysis of authentic standards; all others should be considered tentative identifications. | | 1526 volume 118 number 11 November 2010 • Environmental Health Perspectives DBPs in and mutagenicity of swimming pool water to the presence of DBPs that were activated those for drinking water, which are much concentrations of mono- and dichloramine by GSTT1, such as the brominated THMs more extensive (Richardson et al. 2007). In reported in Table 1 are consistent with chlo- (DeMarini et al. 1997; Pegram et al. 1997). addition, the pool water composition and rine oxidation of continuous supplies of small mutagenicity reported here can be used to amounts of nitrogen compounds coming from Discussion better understand the reported health effects urine, sweat, skin, and other human residues. Most analytical studies of pool water have of swimming pool water, such as asthma, irri- These levels are also similar to those reported measured only a few targeted DBPs, primarily tation of eyes/throat/skin, and bladder cancer by other researchers who used membrane- chloroform and other THMs. Consequently, (Weisel et al. 2009; Zwiener et al. 2007). introduction MS, which does not have issues this study expands considerably our knowl- Nitrogen-containing DBPs. In general, with interferences from organic chloram- edge of the chemical composition and muta- we observed more nitrogen-containing DBPs ines (Shang and Blatchley 1999; Weaver genicity of swimming pool water beyond the (N-DBPs) in these pool water samples than are et al. 2009). chemical analysis of two outdoor pools by found typically in chlorinated drinking water. The N-DBPs, including chloramines, were Zwiener et al. (2007) and the studies on pool For example, we found a greater number of not surprising to find because pool waters have water mutagenicity (Honer et al. 1980) and haloamides, halonitriles, haloanilines, haloani- a greater contribution of nitrogen-containing genotoxicity (Glauner et al. 2005; Liviac et al. soles, and halonitro-compounds than typically precursors due to human inputs. Because 2010). We found a greater number of DBPs found in drinking water, and several chemicals chloramines are known to cause eye irritation in the chlorinated and brominated indoor within these families have not been reported and other problems, pool operators generally pools studied here than have been found in previously in drinking water. In addition, we try to add enough chlorine to get beyond the chlorinated outdoor pools (Zwiener et al. detected mono- and dichloramine in the pool “break point,” such that these chloramines 2007), which was not surprising, consider- waters (means of 0.29 and 0.38 mg/L, respec- are destroyed, leaving residual chlorine (Ford ing that DBPs can be volatilized or photo- tively, for mono- and dichloramine in the 2007; World Health Organization 2000). lyzed (Lekkas and Nikolaou 2004) in outdoor chlorinated pool, and a mean of 0.27 mg/L However, the amount of chlorine needed to settings. In addition, although most people for monochloramine in the brominated pool). reach “break point” is also dependent on other assume that chlorine levels in swimming pools Because DPD analysis of chloramines can- amines in the water. As we observed in this are much higher than in chlorinated drinking not differentiate organic from inorganic forms study, this goal is not always achieved because water, the mean level of free chlorine (1.28 of these compounds, it is possible that these of continuous human inputs and rapid reac- and 0.50 mg/L in the chlorinated and bromi- levels are overestimated by the occurrence of tions forming chloramines. A few other nated pools, respectively) was similar to that organic chloramines in the swimming pool N-DBPs also have been reported in swimming found typically in drinking water. waters. Model studies with batch experiments pool waters, including organic chloramines Because little is known regarding the show that survival of chloramines depends (Li and Blatchley 2007), and nitrosamines mutagenicity and DBP composition of swim- on the chlorine:nitrogen ratio (Jafvert and (Walse and Mitch 2008), several of which are ming pool water, we compared our data with Valentine 1992). Considering this, the low carcinogenic. C OCH Br 3 Cl Br C C (E ) CH O C (Z ) 3 Cl CH O C C OCH O O 16.55 16.60 16.65 16.70 16.75 16.80 16.85 16.90 16.95 17.00 17.05 17.10 17.15 Time (min) (Z )-Bromochlorobutenedioic acid dimethyl ester m/z 225 m/z 197 Molecular mass = 256 Br Cl OO CH O CC CC OCH 3 3 m/z 177 +• m/z 59 256 258 77 177 59 221 43 182 64 89 93 133 179 197 57 69 73 99 131 230 68 78 127 199 201 220 252 260 268 83 140141 149 48 52 120 160 165 173 211 254 261 35 45 114 233 242 272 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 m/z Figure 1. (A) GC/MS chromatogram showing (Z)- and (E)-2-bromo-3-chlorobutenedioic acid dimethyl ester isomers. (B) Electron ionization mass spectrum for (Z)-2-bromo-3-chlorobutenedioic acid dimethyl ester. | | Environmental Health Perspectives • volume 118 number 11 November 2010 1527 Relative abundance Percent Richardson et al. Comparison of brominated versus chlo- Salmonella TA100 (without S9 mix), com- Our finding that some of the mutagenic rinated pool waters. Bromoform levels were pared with our finding of an average of 1,190 activity of one sample (B1) from the bromi- much higher in the pools treated with bromine rev/L-eq in the two indoor pools in Barcelona. nated pool water was due to activation by versus chlorine, but interestingly, other DBPs However, their solvent-extraction method GSTT1 suggests the presence of compounds and their levels were similar in brominated was considerably different from ours, involv - that are activated to mutagens by this enzyme, versus chlorinated pools, likely owing to the ing ether and acetone, whereas we used ethyl such as brominated THMs (DeMarini et al. high levels of bromide present already in acetate. Although a direct comparison of the 1997; Pegram et al. 1997), some methylene Barcelona source waters (Ventura and Rivera data is not possible, our study confirms their dihalides and bifunctional butanes (Thier et al. 1985) that feed into drinking water treat- pioneering work from three decades ago show- 1995), and/or 1,1-dichloropropene (Granville ment and further swimming pool treatment ing that swimming pool water is mutagenic. et al. 2005). Our chemical analysis (Table 1) (Judd and Jeffrey 1995). In addition, when In general, extracts of drinking water showed that sample B1 had high concen- THMs are compared on a molar basis, the induce an average of 1,100 rev/L-eq in trations of brominated THMs, especially chlorinated pool actually contained somewhat Salmonella strain TA100 (without S9 mix) bromoform. The high cytotoxicity and lack higher levels of total THMs (mean, 306 nM) (Takanashi et al. 2009); however, values as of mutagenicity of sample C5 may reflect the than the brominated pool (mean, 242 nM); high as approximately 15,000 rev/L-eq fact that the concentration of chloroform was this was possibly due to the carbon filtration have been reported (Egorov et al. 2003). 30% higher in this sample than in sample C4. used at the brominated pool that was not used Concentration methods such as reverse osmo- Perhaps the higher concentration of chloro- at the chlorinated pool. sis recover levels of mutagenic activity lower form, which is cytotoxic but not mutagenic, Mutagenicity. In the only other muta- than those recovered by XAD resin (Claxton produced the observed cytotoxicity, preventing genicity study of swimming pool water, Honer et al. 2008), which is why we used XAD to detection of mutagenic activity of the other et al. (1980) found that three public indoor prepare extracts of pool water. Our finding DBPs present in sample C5. As reviewed by pools in Victoria, British Columbia (Canada), that the pool water mutagenicity was similar Richardson et al. (2007), many other DBPs produced approximately 20,000 rev/L-eq in to that of drinking water may reflect the fact in drinking water that we have now identified that the levels of mutagenic DBPs in the pool in pool waters are known to be mutagenic Table 3. Mutagenicity of pool waters in Salmonella. waters were similar to those in drinking water, and/or carcinogenic, including the haloacetic despite the differences in the levels of specific acids, halonitromethanes, haloamides, halo- Strain (rev/plate) classes of DBPs described above in pool versus acetonitriles, and unregulated haloacids (Plewa Sample TPT100 RSJ100 drinking water. et al. 2008a, 2008b; Richardson et al. 2007). L-eq/plate (GST–) (GST+) TA100 C4 0 16, 19 10, 14 127, 115, 111 750 200 C4 C5 0.05 27 28 158 0.075 39 10 200 0.1 52 60 238 0.15 44 57 264 1,800 ± 636 rev/L-eq 0.2 69 81 343 0.3 87 70 495 r = 0.80 1,247 ± 70 rev/L-eq p = 0.105 C5 2 50 r = 0.98 0 9, 19, 12 6, 6, 5 75, 83, 94 p < 0.001 0.01 24 8 128 0 0 0.02 23 18 134 0.0 0.1 0.2 0.3 0.4 0.00 0.01 0.02 0.03 0.04 0.03 21 27 142 a a a L-eq/plate L-eq/plate 0.04 21 23 115 a a a 0.05 12 25 109 B1 B2 B1 0 27, 18, 20 9, 8, 5 130, 128 0.01 14 14 132 0.025 16 7 137 0.05 16 19 164 1,257 ± 149 rev/L-eq 1,067 ± 41 rev/L-eq 0.075 6 15 130 2 2 0.1 19 26 294 r = 0.91 r = 0.99 p < 0.001 p < 0.001 0.15 19 28 274 0.2 26 42 407 0 0 0.3 33 54 471 0.0 0.1 0.2 0.3 0.4 0.00 0.05 0.10 0.15 0.20 0.25 B2 0 16, 19 10, 14 127, 115, 111 L-eq/plate L-eq/plate 0.05 19 24 182 Figure 2. Mutagenicity in Salmonella TA100–S9 of two samples each from the chlorinated (C4, C5) and 0.075 29 28 199 (B1, B2) brominated pools. Data in each curve are from Table 3 and represent a single experiment performed 0.1 30 26 225 with one plate per dose. Slope (mutagenic potency) is rev/L-eq ± SE of the slope. 0.15 37 34 290 a a 0.2 32 34 330 a a a 0.3 26 30 373 Table 4. Mutagenic potencies of pool water samples in GST– and GST+ strains of Salmonella. The average rev/plate for the positive control, sodium Rev/L-eq ± SE (r ) azide (3 µg/plate), was 910 for TPT100, 519 for RSJ100, Sample TPT100 (GST–) RSJ100 (GST+) p-Value and 645 for TA100. The average rev/plate for the solvent blank (2 L-eq/plate) was 10 for RSJ100 and 128 for TA100; C4 228.2 ± 27 (0.93) 357.9 ± 95 (0.78) 0.131 it was not tested in TPT100. C5 500.0 ± 346 (0.68) 730.0 ± 128 (0.94) 0.508 Numbers were outside of the linear range of the dose B1 54.8 ± 20 (0.51) 159.1 ± 22 (0.90) 0.000 response and were not used to calculate the linear B2 136.0 ± 26 (0.90) 136.0 ± 28 (0.89) 0.194 regressions for potency values (Figure 2, Table 4). | | 1528 volume 118 number 11 November 2010 • Environmental Health Perspectives Rev/plate Rev/plate Rev/plate Rev/plate DBPs in and mutagenicity of swimming pool water In addition to the mix of mutagenic DBPs et al. 2010). These findings are especially rele- International Agency for Research on Cancer, Working Group on the Evaluation of Carcinogenic Risks to Humans. 2004. identified in the pool water, many other DBPs vant with regard to a case–control study by Some drinking-water disinfectants and contaminants, have not yet been studied for health effects, Cantor et al. (2010) in this issue that identifies including arsenic. IARC Monogr Eval Carcinog Risks Hum and no doubt, many other DBPs remain to an enhanced risk for bladder cancer associated 84:1–477. Jacobs JH, Spaan S, van Rooy GB, Meliefste C, Zaat VA, be identified that also may contribute to the with DBP exposure among people with geno- Rooyackers JM, et al. 2007. Exposure to trichloramine and observed mutagenicity of swimming pool types that metabolize various DBPs. Further respiratory symptoms in indoor swimming pool workers. water. In this regard, the study by Glauner research on a wide array of swimming pools Eur Respir J 29:690–698. Jafvert CT, Valentine RL. 1992. Reaction scheme for the chlorina- et al. (2005), which found that the low- under various conditions of maintenance and tion of ammoniacal water. Environ Sci Technol 26:577–586. molecular-weight fraction of extracts from use are warranted based on the limited but Judd S, Jeffrey JA. 1995. Trihalomethane formation during indoor and outdoor pools in Germany was developing data now available on the chemi- swimming pool water disinfection using hypobromous and hypochlorous acids. Water Res 29:1203–1206. the most potent of all fractions for inducing cal composition and health risks of swimming Kim H, Shim J, Lee S. 2002. Formation of disinfection by- DNA damage in mammalian cells (using the pool water. products in chlorinated swimming pool water. Chemosphere comet assay), suggests that the low- molecular- 46:123–130. weight DBPs may be most responsible for Refe Rences K o g e v i n a s M , V i l l a n u e v a C M , F o n t - R i b e r a L , L i v i a c D , Bustamante M, Espinoza F, et al. 2010. Genotoxic effects the genotoxic effect of swimming pool water. in swimmers exposed to disinfection by-products in indoor Aggazzotti G, Predieri G. 1986. Survey of volatile halogenated Also using the comet assay, Liviac et al. (2010) swimming pools. Environ Health Perspect 118:1531–1537. organics (VHO) in Italy: levels of VHO in drinking waters, found that pool water was more genotoxic LaKind JS, Richardson SD, Blount BC. 2010. The good, the surface water and swimming pools. Water Res 20:959–963. bad, and the volatile: can we have both healthy pools and than the source tap water; a similar analysis Ashley DL, Blount BC, Singer PC, Depaz E, Wilkes C, Gordon S, healthy people? Environ Sci Technol 44:3205–3210. et al. 2005. Changes in blood trihalomethane concentra- using the Ames assay would help in charac- Leavens TL, Blount BC, DeMarini DM, Madden MC, Valentine JL, tions resulting from differences in water quality and water terizing the relative mutagenicity of drinking Case MW, et al. 2007. Disposition of bromodichloro methane use activities. Arch Environ Occup Health 60:7–15. in humans following oral and dermal exposure. Toxicol Sci versus pool water. Sources of mutagens unique Beech JA, Diaz R, Ordaz C, Palomeque B. 1980. Nitrates, chlo- 99:432–445. rates, and trihalomethanes in swimming pool water. Am J to pool water could include active agents Lekkas TD, Nikolaou AD. 2004. Degradation of disinfection Public Health 70:79–82. in sunscreens, which can be transformed to byproducts in drinking water. Environ Engin Sci 21:493–506. Cantor KP, Villanueva CM, Silverman DT, Figueroa JD, Real FX, Li J, Blatchley ER III. 2007. Volatile disinfection byproduct mutagens by exposure to free chlorine under Garcia-Closas M, et al. 2010. Polymorphisms in GSTT1, formation resulting from chlorination of organic-nitrogen GSTZ1, and CYP2E1, disinfection by-products, and risk conditions similar to swimming pool water precursors in swimming pools. Environ Sci Technol of bladder cancer in Spain. Environ Health Perspect (Nakajima et al. 2009). 41:6732–6739. 118:1545–1550. Liviac D, Wagner ED, Mitch WA, Altonji MJ, Plewa MJ. 2010. Our limited data indicate that the muta- Chu H, Nieuwenhuijsen MJ. 2002. Distribution and determinants Genotoxicity of water concentrates from recreational of trihalomethane concentrations in indoor swimming genic potencies of chlorinated versus bromi- pools after various disinfection methods. Environ Sci pools. Occup Environ Med 59:243–247. nated pool waters were similar, as were the Technol 44:3527–3532. Claxton LD, Pegram R, Schenck, KM, Simmons JE, Warren SH. dose ranges over which the pool waters were Lourencetti C, Ballester C, Fernandez P, Marco E, Prado C, 2008. Integrated disinfection by-products research: Periago JF, et al. 2010. New method for determination of tri- Salmonella mutagenicity of water concentrates disinfected mutagenic, approximately 0.1–0.3 L-eq/plate halo methanes in exhaled breath: applications to swimming by chlorination and ozonation/postchlorination. J Toxicol (Figure 2). In contrast, the typical dose range pool and bath environments. Anal Chim Acta 662:23–30. Environ Health A 71:1187–1194. for drinking water mutagenicity is 0.3–1.5 M a r o n D M , A m e s B N . 1 9 8 3 . R e v i s e d m e t h o d s f o r t h e D e M a r i n i D M , A b u - S h a k r a A , F e l t o n C F , P a t t e r s o n K S , Salmonella mutagenicity test. Mutat Res 113:173–215. L-eq/plate (DeMarini et al. 1995). This differ - Shelton ML. 1995. Mutation spectra in Salmonella of chlori- Nakajima M, Kawakami T, Niino T, Takahashi Y, Onodera S. nated, chloraminated, or ozonated drinking water extracts: ence reflects the considerably higher toxicity 2009. Aquatic fate of sunscreen agents octyl-4-methoxy- comparison to MX. Environ Mol Mutagen 26:270–285. of swimming pool water relative to drinking cinnamate and octyl-4-dimethylaminobenzoate in model DeMarini DM, Shelton ML, Warren SH, Ross TM, Shim SY, swimming pools and the mutagenic assays of their chlori- water, with the highest testable mutagenic Richard AM, et al. 1997. Glutathione S-transferase- nation byproducts. J Health Sci 55:363–372. mediated induction of GC→AT transitions by halomethanes dose of pool water being the lowest mutagenic Pegram RA, Andersen ME, Warren SH, Ross TM, Claxton LD. in Salmonella. Environ Mol Mutagen 30:440–447. dose of drinking water. 1997. Glutathione S-transferase-mediated mutagenicity of E g o r o v A I , T e r e s c h e n k o A A , A l t s h u l L M , V a r t i a i n e n T , trihalomethanes in Salmonella typhimurium: contrasting Samsonov D, LaBrecque B, et al. 2003. Exposures to drink- results with bromodichloromethane and chloroform. Toxicol Conclusions ing water chlorination by-products in a Russian city. Int J Appl Pharmacol 144:183–188. Hyg Environ Health 206:539–551. We identified > 100 DBPs in two indoor Plewa MJ, Muellner MG, Richardson SD, Fasano F, Buettner KM, Erdinger L, Kuhn KP, Kirsch F, Feldhues R, Frobel T, Nohynek B, Woo Y-T, et al. 2008a. Occurrence, synthesis, and mamma- pools, including a prevalence of N-DBPs, et al. 2004. Pathways of trihalomethane uptake in swim- lian cell cytotoxicity and genotoxicity of haloacetamides: an ming pools. Int J Hyg Environ Health 207:571–575. likely formed from nitrogen-containing pre- emerging class of nitrogenous drinking water disinfection Font-Ribera L, Kogevinas M, Zock J-P, Gómez FP, Barreiro E, cursors from human inputs. This study pro - byproducts. Environ Sci Technol 42:955–961. Nieuwenhuijsen MJ, et al. 2010. Short-term changes in vides the most comprehensive analysis to date Plewa MJ, Wagner ED, Muellner MG, Hsu KM, Richardson SD. respiratory biomarkers after swimming in a chlorinated 2008b. Comparative mammalian cell toxicity of N-DBPs and pool. Environ Health Perspect 118:1538–1544. of DBPs in swimming pool waters, as well as a C-DBPs. In: By-products in Drinking Water: Occurrence, Ford RL, ed. 2007. Certified Pool-Spa Operator Handbook. clear demonstration of their mutagenicity. In Formation, Health Effects and Control (Karanfil T, Krasner Colorado Springs, CO:National Swimming Pool Foundation. addition, many DBPs we identified are new SW, Westerhoff P, Xie Y, eds). ACS Symposium Series 995. G l a u n e r T , Z w i e n e r C , W a l d m a n n P , F r i m m e l F H . 2 0 0 5 . Washington, DC:American Chemical Society, 36–50. and have not been reported previously in pool Swimming pool water—fractionation and genotoxicologi- Richardson SD. 1998. Drinking Water Disinfection By-products. cal characteri zation of organic constituents. Water Res waters. Bromoform levels were much greater In: The Encyclopedia of Environmental Analysis and 39:4494–4502. in the brominated versus chlorinated pools. Remediation, Vol 3 (Meyers RA, ed). New York:Wiley, Goodman M, Hays S. 2008. Asthma and swimming: a meta- 1398–1421. Compared with previous research on outdoor analysis. J Asthma 45:639–649. Richardson SD, Plewa MJ, Wagner ED, Schoeny R, DeMarini DM. Granville CA, Ross MK, Tornero-Velez R, Hanley NM, Grindstaff pools, we found a much greater number of 2007. Occurrence, genotoxicity, and carcino genicity of RD, Gold A, et al. 2005. Genotoxicity and metabolism of the DBPs in these indoor pools. regulated and emerging disinfection by-products in drink- source-water contaminant 1,1,dichloropropene: activation ing water: a review and roadmap for research. Mutat Res The mutagenicity of these pool waters was by GSTT1-1 and structure-activity considerations. Mutat 636:178–242. Res 572:98–112. similar to that of drinking water, indicating Richardson SD, Thruston AD Jr, Krasner SW, Weinberg HS, Haddad S, Tardif GC, Tardif R. 2006. Development of physiologi- that the levels of mutagenic DBPs are simi- Miltner RJ, Schenck KM, et al. 2008. Integrated dis infection cally based toxicokinetic models for improving the human by-products mixtures research: comprehensive characteri- lar in both waters. Subjects who swam in the indoor exposure assessment to water contaminants: zation of water concentrates prepared from chlorinated trichloroethylene and trihalomethanes. J Toxicol Environ mutagenic, chlorinated pool water evaluated and ozonated/postchlorinated drinking water. J Toxicol Health A 69:2095–2136. in this study had increases in geno toxicity bio- Environ Health A 71:1165–1186. Héry M, Hecht G, Gendre JC, Hubert G, Rebuffaud I. 1995. markers that were associated with the con- S a n t a M a r i n a L , I b a r l u z e a J , B a s t e r r e c h e a M , G o ñ i F , Exposure to chloramines in the atmosphere of indoor Ulibarrena E, Artieda J, et al. 2009. Indoor air and bathing swimming pools. Ann Occup Hyg 39:427–439. centrations of brominated THMs, but not water pollution in indoor swimming pools in Guipuzcoa Honer WG, Ashwood-Smith MJ, Warby C. 1980. Mutagenic chloroform, in their exhaled breath (Kogevinas (Spain). Gac Sanit 23:115–120. activity of swimming-pool water. Mutat Res 78:137–144. | | Environmental Health Perspectives • volume 118 number 11 November 2010 1529 Richardson et al. Shang C, Blatchley ER III. 1999. Differentiation and quantifica - Ventura F, Rivera J. 1985. Factors influencing the high content environmental exposures at swimming pools: state of the tion of free chlorine and inorganic chloramines in aqueous of brominated trihalomethanes in Barcelona’s water supply science and research recommendations. Environ Health solution by MIMS. Environ Sci Technol 33:2218–2223. (Spain). Bull Environ Contam Toxicol 35:73–81. Perspect 117:500–507. Stav D, Stav M. 2005. Asthma and whirlpool baths. N Engl J Villanueva CM, Cantor KP, Cordier S, Jaakkola JJK, King WD, W o r l d H e a l t h O r g a n i z a t i o n . 2 0 0 0 . G u i d e l i n e s f o r S a f e Med 353:1635–1636. Lynch CF, et al. 2004. Disinfection byproducts and bladder Recreational-Water Environments, Vol. 2, Swimming Takanashi H, Kishida M, Nakajima T, Ohki A, Akiba M, Aizawa T. cancer. A pooled analysis. Epidemiology 15:357–367. Pools, Spas and Similar Recreational-Water Environments. 2009. Surveying the mutagenicity of tap water to elicit the Villanueva CM, Cantor KP, Grimalt JO, Malats N, Silverman D, Geneva:World Health Organization. effects of purification processes on Japanese tap water. Tardon A, et al. 2007. Bladder cancer and exposure to Xu X, Weisel CP. 2005. Dermal uptake of chloroform and halo ketones Chemosphere 77:434–439. water disinfection by-products through ingestion, bathing, during bathing. J Exp Anal Environ Epidemiol 15:289–296. Thier R, Müller M, Taylor JB, Pemble SE, Ketterer B, Guengerich showering and swimming pool attendance. Am J Epidemiol Xu W, Weisel CP. 2004. Human respiratory uptake of chloro form FP. 1995. Enhancement of bacterial mutagenicity of bifunc- 165:148–156. and haloketones during showering. J Exp Anal Environ tional alkylating agents by expression of mammalian gluta- Walse SS, Mitch WA. 2008. Nitrosamine carcinogens also swim Epidemiol 15:6–16. thione S-transferase. Chem Res Toxicol 8:465–472. in chlorinated pools. Environ Sci Technol 42:1032–1037. Zwiener C, Richardson SD, DeMarini DM, Grummt T, Glauner T, Thier R, Taylor JB, Pemble SE, Humphreys WG, Persmark M, Weaver WA, Li J, Wen Y, Johnston J, Blatchley MR, Blatchley ER Frimmel FH. 2007. Drowning in disinfection byproducts? Ketterer B, et al. 1993. Expression of mammalian glutathi- III. 2009. Volatile disinfection by-product analysis from chlo- Assessing swimming pool water. Environ Sci Technol one S-transferase 5-5 in Salmonella typhimurium TA1535 rinated indoor swimming pools. Water Res 43:3308–3318. 41:363–372. leads to base-pair mutations upon exposure to dihalo- Weisel CP, Richardson SD, Nemery B, Aggazzotti G, Baraldi E, methanes. Proc Natl Acad Sci USA 90:8576–8580. Blatchley ER III, et al. 2009. Childhood asthma and | | 1530 volume 118 number 11 November 2010 • Environmental Health Perspectives http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Environmental Health Perspectives Pubmed Central

What’s in the Pool? A Comprehensive Identification of Disinfection By-products and Assessment of Mutagenicity of Chlorinated and Brominated Swimming Pool Water

Download PDF
8 pages

Loading next page...
 
/lp/pubmed-central/what-s-in-the-pool-a-comprehensive-identification-of-disinfection-by-0aiMBSu92B

References

References for this paper are not available at this time. We will be adding them shortly, thank you for your patience.

Publisher
Pubmed Central
ISSN
0091-6765
eISSN
1552-9924
DOI
10.1289/ehp.1001965
Publisher site
See Article on Publisher Site

Abstract

Research What’s in the Pool? A Comprehensive Identification of Disinfection By-products and Assessment of Mutagenicity of Chlorinated and Brominated Swimming Pool Water 1 2 3,4,5,6 7 7 Susan D. Richardson, David M. DeMarini, Manolis Kogevinas, Pilar Fernandez, Esther Marco, 7 7 8 8 9 10 Carolina Lourencetti, Clara Ballesté, Dick Heederik, Kees Meliefste, A. Bruce McKague, Ricard Marcos, 3,4 7 3,4,5 Laia Font-Ribera, Joan O. Grimalt, and Cristina M. Villanueva 1 2 National Exposure Research Laboratory, U.S. Environmental Protection Agency, Athens, Georgia, USA; National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, USA; 3 4 Centre for Research in Environmental Epidemiology, Barcelona, Spain; Municipal Institute of Medical Research, Hospital del 5 6 Mar, Barcelona, Spain; CIBER Epidemiología y Salud Pública, Barcelona, Spain; Medical School, University of Athens, Greece; 7 8 Department of Environmental Chemistry, Institute of Environmental Assessment and Water Research, Barcelona, Spain; Institute for Risk Assessment Sciences, Division for Environmental Epidemiology, Utrecht University, Utrecht, the Netherlands; CanSyn Chem. Corp., Toronto, Ontario, Canada; Grup de Mutagènesi, Departament de Genètica i de Microbiologia, Edifici Cn, Universitat Autònoma de Barcelona, Bellaterra, Cerdanyola del Vallès, Spain with asthma and other respiratory effects in Background : Swimming pool disinfectants and disinfection by-products (DBPs) have been linked Olympic swimmers and pool workers, and less to human health effects, including asthma and bladder cancer, but no studies have provided a com - clearly with recreational adult swimmers and prehensive identification of DBPs in the water and related that to mutagenicity. children (Goodman and Hays 2008; Jacobs o Bjectives : We performed a comprehensive identification of DBPs and disinfectant species in waters et al. 2007; Stav and Stav 2005; Weisel et al. from public swimming pools in Barcelona, Catalonia, Spain, that disinfect with either chlorine or bro- 2009). However, the mechanisms are poorly mine and we determined the mutagenicity of the waters to compare with the analytical results. understood, and it is not known with certainty Methods : We used gas chromatography/mass spectrometry (GC/MS) to measure trihalomethanes whether trichloramine or other volatile pool in water, GC with electron capture detection for air, low- and high-resolution GC/MS to compre- DBPs are responsible. hensively identify DBPs, photometry to measure disinfectant species (free chlorine, monochloro- Despite the public health relevance, only amine, dichloramine, and trichloramine) in the waters, and an ion chromatography method to a few studies, most rather recent, have inves- measure trichloramine in air. We assessed mutagenicity with the Salmonella mutagenicity assay. tigated the chemistry and potential health results : We identified > 100 DBPs, including many nitrogen-containing DBPs that were likely effects of swimming pool water (Weisel et al. formed from nitrogen-containing precursors from human inputs, such as urine, sweat, and skin 2009; Zwiener et al. 2007). A complete cells. Many DBPs were new and have not been reported previously in either swimming pool or chemical characterization of DBPs in indoor drinking waters. Bromoform levels were greater in brominated than in chlorinated pool waters, swimming pools has not been reported. The but we also identified many brominated DBPs in the chlorinated waters. The pool waters were only mutagenicity study of swimming pool mutagenic at levels similar to that of drinking water (~ 1,200 revertants/L-equivalents in strain water reported that organic extracts from TA100–S9 mix). three public indoor pools in Victoria, British c onclusions : i Th s study identie fi d many new DBPs not identie fi d previously in swimming pool or drinking water and found that swimming pool waters are as mutagenic as typical drinking waters. Address correspondence to S.D. Richardson, National k e y w o r d s : bromination, bromine, chlorination, chlorine, DBPs, disinfection by-products, Exposure Research Laboratory, U.S. Environmental mutagenicity, swimming pools, Salmonella, water. Environ Health Perspect 118:1523–1530 Protection Agency, 960 College Station Rd., Athens, GA 30605 USA. Telephone: (706) 355-8304. Fax: (2010). doi:10.1289/ehp.1001965 [Online 12 September 2010] (706) 355-8302. E-mail: richardson. [email protected] Supplemental Material is available online (doi:10. 1289/ehp.1001965 via http://dx.doi.org/). Disinfection by-products (DBPs) represent air due to continuous disinfection and con- We thank T. Lewis, G. Crumley, C. Lindell, a ubiquitous exposure in developed coun- stant organic load from bathers (e.g., urine, J. Hartmann, and H. Fink for their assistance with tries. DBPs are formed by the reaction of dis- sweat, cosmetics, skin cells, and hair) (Kim extractions, derivatizations, and analyses. We thank infectants (e.g., chlorine, chloramines, ozone, et al. 2002; LaKind et al. 2010). One of the A. Kligerman for his help with statistical analyses, and or chlorine dioxide) with natural organic most prevalent DBPs in chlorinated swim- A. Kligerman and L. Claxton for helpful comments on the manuscript. matter and/or bromide/iodide, and they are ming pools is THMs (Aggazzotti and Predieri This research was supported by the U.S. Environ - an unintended consequence of trying to kill 1986; Beech et al. 1980; Judd and Jeffrey mental Protection Agency (EPA) intra mural research pathogens in drinking water and swimming 1995), with average concentrations ranging program and Spanish grants SAF2005-07643- pools. More than 600 DBPs have been iden- from 16 µg/L (Santa Marina et al. 2009) to C03-01 (Plan Nacional) and CP06/00341 (Fondo tified in drinking water, and many of them 132 µg/L (Chu and Nieuwenhuijsen 2002). de Investigación Sanitaria). C.M.V. and L.F.-R. have, are mutagenic or carcinogenic (Richardson Given the high nitrogen content of organic respectively, a contract and a predoctoral fellowship by the Instituto de Salud Carlos III (CP06/00341, 1998; Richardson et al. 2007). This complex matter from bathers, nitrogenated species FI06/00651). C.L. acknowledges a grant from the mixture of DBPs includes volatile and skin- such as haloacetonitriles, nitrosamines, and Agreement between Santander-Central Hispano and permeable DBPs, such as trihalomethanes chloramines are found in swimming pool Consejo Superior de Investigaciones Científicas. (THMs) and haloketones (Erdinger et al. water (Héry et al. 1995; Kim et al. 2002; The manuscript has been reviewed in accordance 2004: Xu and Weisel 2004, 2005). Inhalation Walse and Mitch 2008; Zwiener et al. 2007). with the U.S. EPA’s peer and administrative review and dermal absorption, which are the primary Chronic exposure to DBPs through differ - policies and approved for publication. Mention of trade names or commercial products does not consti- routes of exposure to DBPs during swim- ent routes has been associated with an increased tute endorsement or recommendation for use by the ming, leads to higher blood levels of THMs risk for bladder cancer (International Agency U.S. EPA. than do oral exposures (Ashley et al. 2005; for Research on Cancer 2004; Villanueva et al. A.M. is employed by CanSyn Chem. Corp., Toronto, Haddad et al. 2006; Leavens et al. 2007). 2004, 2007). Trichloramine and other volatile Ontario, Canada. The authors declare they have no Swimming pools constitute environ- chemicals in swimming pools are respiratory actual or potential competing financial interests. ments with high levels of DBPs in water and irritants; pool attendance has been associated Received 18 January 2010; accepted 8 June 2010. | | Environmental Health Perspectives • volume 118 number 11 November 2010 1523 Richardson et al. Columbia (Canada), were mutagenic in for THM measurements were quenched with pool (16 July and 15 October 2007). Samples Salmonella TA100 (Honer et al. 1980). The 5 mg sodium thiosulfate and stored at 4°C (28 L each) were collected using 2-L Teflon authors found that acidified extracts eluted until analysis on the same day. Chloroform, bottles (headspace-free) and were shipped with ether were more mutagenic in the pres- bromodichloro methane, dibromochloro - overnight in coolers with icepacks to the U.S. ence of metabolic activation (rat liver S9) than methane, and bromoform were measured Environmental Protection Agency laboratory without S9; however, nonacidified extracts using purge-and-trap gas chromatography/ in Athens, Georgia (USA). Water samples eluted with acetone were mutagenic only in mass spectrometry (GC/MS) (Tekmar 3100, were extracted immediately upon arrival using the absence of S9. One genotoxicity study of Voyager MS; ThermoFisher, Waltham, MA, the XAD resin process of Richardson et al. swimming pool water reported that the water USA) following the method described by (2008) [for further details, see Supplemental and its fractions induced DNA damage in Lourencetti et al. (2010). Sixty-eight samples Material (doi:10.1289/ehp.1001965)]. The Hep-G2 cells (comet assay) and that most of were collected from the chlorinated pool final extract was divided for comprehensive the genotoxicity was in the lower-molecular- and 12 from the brominated pool for these GC/MS analysis (1.0 mL, equivalent to 20 L weight DBP fraction (Glauner et al. 2005). quantita tive analyses. water) and mutagenicity analysis (0.4 mL, Another study using the comet assay showed Indoor air samples to measure THMs equivalent to 8 L water, or 20,000×). that pool water was more genotoxic than the were collected with a pump located 60 cm Comprehensive GC/MS analyses. Half source tap water and that the type of disinfec- above the floor and 1.5 m from the pool bor- of the 1.0-mL extract was derivatized with tant and illumination conditions altered the der. Air was pumped (7 mL/min) for 20 min diazomethane [see Supplemental Material genotoxicity (Liviac et al. 2010). through a Tenax TA cartridge (1.8 g; Supelco, (doi:10.1289/ehp.1001965)] to enable the The present study involves an investiga- Sigma-Aldrich, St. Louis, MO, USA). Quality identification of haloacids (through their cor- tion in Barcelona, Spain, where we examined control was assured by daily calibration of the responding methyl esters); the other half was 49 healthy nonsmoking volunteers before and pump. Chloroform, bromodichloromethane, analyzed directly for other DBPs. after swimming in public swimming pools dibromochloromethane, and bromoform were Comprehensive GC/MS analyses were per- treated with either chlorine or bromine to determined through an automatic thermal formed on a high-resolution magnetic sector evaluate personal exposure and a range of bio- desorption unit (ATD 400; Perkin-Elmer, mass spectrometer (Autospec; Waters, Inc., markers of genotoxicity and respiratory dam- Madrid, Spain) coupled to a GC-electron cap- Milford, MA, USA) equipped with an Agilent age (Font-Ribera et al. 2010; Kogevinas et al. ture detector (Perkin-Elmer). Sixty-eight air model 6890 gas chromatograph (Agilent, Santa 2010). To complement the exposure assess- samples were collected from the chlorinated Clara, CA, USA) and operated at an accelerat- ment, we evaluated the mutagenicity of the pool, and 12 from the brominated pool. ing voltage of 8 kV and source temperature pool waters in the Salmonella mutagenicity Trichloramine was measured in pool of 200°C, in both low-resolution (1,000) and assay and screened for DBPs, comprehensively air samples by pumping air (1.2 L/min) for high-resolution (10,000) modes. Injections of identifying most DBPs detected and quan- 115 min, within 1 m from the water and at 1 µL extract were introduced via a split/splitless tifying a few targeted DBPs and disinfectant a height of 60 cm from the o fl or level, using injector (in splitless mode) onto a GC column species (THMs, chlorine, monochloramine, a method described originally by Héry et al. (DB-5, 30-m × 0.25-mm i.d. 0.25-µm film dichloramine, and trichloramine) in the pool (1995). Trichloramine was captured on two thickness; J&W Scientific/Agilent, Santa Clara, waters and in the air phase above the water 37-mm quartz-fiber filters, one of which was CA, USA). The GC temperature program con - (THMs and trichloramine). In this article placed as a backup filter, both impregnated with sisted of an initial temperature of 35°C (4 min) we present a comprehensive identification 500 mL of a solution of diarsenic tri oxide (4 g/L and an increase at 9°C/min to 285°C (held for of DBPs and disinfectant species in the pool As O ), sodium carbonate (40 g/L Na CO ), 30 min). Transfer lines were held at 280°C, 2 3 2 3 waters and compare the species formed in and glycerol (40 mL/L C H O ). These fil- and the injection port at 250°C. 3 8 3 chlorinated versus brominated pool water with ters were placed in a sampling cassette with For qualitative identifications, mass spectra the corresponding mutagenicity of the waters. a 37-mm cellulose support filter and a 37-mm of unknown compounds in the finished and Teflon filter to prevent chloride from airborne raw water concentrates were subjected initially Materials and Methods water droplets from being captured in the to library database searching (using the 2005 Sampling. Water samples were collected from sampler. Impregnated filters were de sorbed in NIST Mass Spectral Library database; National two large public swimming pools in Barcelona, 10-mL ultra-high quality, ultrapure water (spe- Institute of Standards and Technology, Spain. One pool (33 × 25 × 2 m in size) used cific conductivity, 17.8 MΩ/cm at 25°C), soni- Gaithersburg, MD). However, many DBPs chlorine (sodium hypochlorite) for disinfec - cated for 30 min, and centrifuged for 15 min were not present in the library database; in tion, after sand filtration; the other pool at 3,000 × g after sampling. Trichloramine those cases, and also where a library match was (20.9 × 13.2 × 1.3 m in size) used bromine was reduced to chloride and subsequently ana- insufficient to offer a tentative identification, ( 1 - b r o m o - 3 - c hl o r o - 5 , 5 - d i m e t h y l - 2 , 4 - lyzed by ion chromatography (Dionex DX100; high-resolution MS was used to provide empir- imidazolidinedione) for disinfection, after sand Dionex BV, Bavel, the Netherlands; AS14A ical formulas for molecular ions and fragments. and granulated carbon filtration. Floor-to- guard and AS14 highly selective anion column Mass spectra were also interpreted extensively ceiling height was 10 m and 5 m in the chlori- with self-regenerating suppressor; conductivity to provide tentative structural identifications. nated and brominated pools, respectively. detector; flow rate , 1.0 mL/min). Six samples When possible, pure standards were obtained Quantitative analyses. Free chlorine, were collected from the chlorinated pool, and to confirm identifications through a match of monochloramine, dichloramine, trichloramine, three from the brominated pool. GC retention times and mass spectra. and THMs were measured in composite pool Preparation of water extracts and con- Chemical standards. Chemical DBP stan- water samples (1 L) collected from four differ - centrates. For comprehensive GC/MS analy- dards were either synthesized (CanSyn Chem. ent locations. Free chlorine, monochloramine, ses and mutagenicity testing, pool water Corp., Toronto, ON, Canada) or purchased dichloramine, and trichloramine were meas- samples were collected at approximately at the highest level of purity (Sigma-Aldrich, ured immediately using the N,N-diethyl-p- noon on five different sampling events for Milwaukee, WI, USA). The synthesis of phenylenediamine (DPD) method with a the chlorinated pool (7 and 24 May, 11 (E)- and (Z)-bromochlorobutenedioic acid portable photometer (DINKO Instruments, June, and 17 September 2007) and two dif- is presented in Supplemental Material Barcelona, Spain). Water samples (40 mL) ferent sampling events for the brominated (doi:10.1289/ehp.1001965). | | 1524 volume 118 number 11 November 2010 • Environmental Health Perspectives DBPs in and mutagenicity of swimming pool water Mutagenicity assays. The 20,000× in the DMSO control. The positive control OCH ] suggested the presence of a carboxy- XAD/ethyl acetate extracts described above for all strains was sodium azide at 3 µg/plate. lic acid methyl ester in the structures, with a were solvent-exchanged into dimethyl sulfox- We calculated linear regressions, slope molecular ion of m/z 256/258/260. Further, ide (DMSO; Burdick and Jackson, Muskegon, values, the standard error of the slopes, and the m/z 256/258/260 isotopic pattern was MI, USA) and diluted to 10,000× and r values of the dose–response curves and then indicative of one bromine and one chlorine 1,000×. We performed the standard plate- compared the regression lines between strains atom, matching the calculated theoretical pat- incorporation Salmonella (Ames) mutagen- RSJ100 (GST+) and TPT100 (GST–) to tern [see Supplemental Material, Figure S2 icity assay (Maron and Ames 1983) in the obtain p-values using Statgraphics Centurion (doi:10.1289/ehp.1001965)]. This pattern base-substitution strain TA100 (hisG46 rfa XVI (Statpoint Technologies, Inc., Warrenton, results from the overlap of the two natural 79 81 ΔuvrB, pKM101), obtained from B.N. Ames, VA, USA). The model tests the null hypothesis isotopes of bromine ( Br and Br) with the Children’s Hospital Oakland Research that the slopes are equal; we set α = 0.05 for two natural isotopes of chlorine ( Cl and Institute (Oakland, CA, USA). We also tested the F-test with 2 degrees of freedom. Cl). This information suggested a tenta- the extracts in Salmonella strain RSJ100, which tive structural identification of bromochlo- Results expresses the rat GSTT1 gene, and its control robutenedioic acid dimethyl ester, with a strain TPT100. These strains are homologous DBPs. Table 1 lists levels of free chlorine, monoisotopic molecular mass of 256 Da. to TA100 except that they do not contain chloramines, and THM species in the pool Exact mass data provided by high-resolu- the pKM101 plasmid and either do or do water and air. Although we did not detect tion MS supported this empirical formula not express GSTT1 (Thier et al. 1993). These trichloramine in the pool waters, we did n fi d (C H O ClBr). The observed exact mass of 6 6 4 strains were obtained from F.P. Guengerich mean levels of 0.29 and 0.08 mg/m in the the stronger molecular ion isotopic peak (m/z (Vanderbilt University, Nashville, TN, USA). chlorinated and brominated pool air, respec- 257.9116) was within 0.0002 Da of the theo- We did not use S9 mix because we assumed tively, indicating that most of it volatilized retical mass (m/z 257.9118). This supported that pool water was similar to drinking water, from the water into the air (Font-Ribera et al. the general structure of bromochlorobutene- and drinking water extracts are most muta- 2010). We identified > 100 DBPs compre- dioic acid dimethyl ester; however, the exact genic in the absence of S9 mix (Takanashi hensively in the pool waters [Table 2; see also isomer assignments could not be made by MS et al. 2009). Supplemental Material, Figure S1 (doi:10.1289/ data alone because the spectra were too simi- Extracts were tested up to 100 µL/plate ehp.1001965)], including a large number of lar, which is often the case for isomers. Two over a dose range of 0.01–0.3 L-equivalents haloacids, halomethanes, haloacetonitriles, structural isomers are possible for this empiri- (L-eq)/plate based on doses used for drink- haloaldehydes, haloketones, halonitromethanes, cal formula, (Z) and (E), representing cis and ing water (DeMarini et al. 1995) and a dose- haloamides, haloalcohols, and halophenols. All trans isomers, respectively (Figure 1A). range–finding study. Because of limited of these contained either bromine or chlorine; Fortunately, we observed both compounds amounts of samples available for testing in all we detected no iodinated DBPs. Most DBPs in most of the pool water concentrates, so it just three bacterial strains, only two samples from have not been reported previously for swim- remained to be determined which specific iso - the chlorinated pool (C4 and C5) and two ming pool waters, and many were not present mer represented each GC/MS chromatographic from the brominated pool (B1 and B2) were in the mass spectral library database. peak. To make this determination, we synthe- evaluated for mutagenicity, each at one plate The identification of ( E)- and (Z)-bromo- sized the two possible isomers [see Supplemental per dose in single experiments. We incubated chloro butenedioic acid (in their corresponding Material (doi:10.1289/ehp.1001965)] and con- the plates for 3 days at 37°C, counted the methyl ester forms) illustrates how we identi- firmed by a match of the GC retention time colonies [revertants (rev)] on an automatic fied unknown DBPs. They eluted at different and mass spectra that the (Z) isomer is the first colony counter, and calculated linear regres- retention times (Figure 1A) but exhibited sim- peak at 16.8 min and the (E) isomer is the sec- sions over the linear portion of the dose–re- ilar mass spectra (indicative of isomeric struc- ond peak at 16.9 min in the pool water extracts sponse curves to determine the mutagenic tures), each containing m/z 256/258/260, (Figure 1A). potencies (rev/L-eq). We defined a positive 225/227/229, and 59 (Figure 1B). The Mutagenicity. Table 3 and Figure 2 result as a dose-related response with two or loss of 31 Da (typically OCH ) at m/z 225 show the mutagenicity data for two sam- more times the number of revertants observed and the presence of m/z 59 [typically C(O) ples from the chlorinated pool and two Table 1. Free chlorine, chloramine, and THM levels in the swimming pools. Chlorinated pool Brominated pool Chemical and concentration Mean ± SD Minimum Maximum n Mean ± SD Minimum Maximum n Water Free chlorine (mg/L) 1.28 ± 0.43 0.52 2.35 68 0.50 ± 0.16 0.32 0.7 4 Monochloramine (NH Cl) (mg/L) 0.29 ± 0.11 0.10 0.64 68 0.27 ± 0.03 0.24 0.3 4 Dichloramine (NHCl ) (mg/L) 0.38 ± 0.14 < 0.01 0.65 68 < 0.01 < 0.01 < 0.01 4 Trichloramine (NCl ) (mg/L) < 0.10 < 0.10 < 0.10 68 < 0.10 < 0.10 < 0.10 4 Chloroform (CHCl ) (µg/L) 15.4 ± 3.5 8.4 20.8 68 0.2 ± 0.1 0.1 0.3 12 Bromodichloromethane (CHCl Br) (µg/L) 14.2 ± 4.2 9.3 26.8 68 0.4 ± 0.2 0.2 0.7 12 Dibromochloromethane (CHClBr ) (µg/L) 12.8 ± 4.4 6.5 22.6 68 2.4 ± 0.2 2.1 2.7 12 Bromoform (CHBr ) (µg/L) 7.2 ± 3.2 3.0 16.5 68 57.2 ± 4.4 52.0 64.3 12 Total THMs (µg/L) 49.6 ± 10.6 35.2 75.2 68 60.2 ± 4.7 54.4 67.2 12 Air Trichloramine (NCl ) (mg/m ) 0.29 ± 0.10 0.17 0.43 6 0.08 ± 0.01 0.07 0.10 3 Chloroform (CHCl ) (µg/m ) 32.1 ± 11.9 11.9 61.6 68 4.4 ± 2.3 1.7 9.4 12 Bromodichloromethane (CHCl Br) (µg/m ) 14.9 ± 4.5 7.5 23.4 68 2.9 ± 1.0 1.7 4.8 12 Dibromochloromethane (CHClBr ) (µg/m ) 14.0 ± 4.2 6.1 26.2 68 7.3 ± 1.3 6.1 9.7 12 Bromoform (CHBr ) (µg/m ) 11.0 ± 4.6 4.4 22.6 68 74.9 ± 17.6 53.3 101.4 12 Total THMs (µg/m ) 72.1 ± 20.7 44.0 124.9 68 89.5 ± 21.9 63.1 124.7 12 | | Environmental Health Perspectives • volume 118 number 11 November 2010 1525 Richardson et al. from the brominated pool in strain TA100. highest doses (0.04 and 0.05 L-eq/plate). sample B1 was significantly more mutagenic All of the samples were mutagenic in strain Table 4 shows the slopes, r values, and in the GSTT1-expressing strain relative to the TA100 except for sample C5, which was the standard errors of the slopes for these data; nonexpressing strain (Tables 3 and 4). This only sample that showed toxicity—based the average mutagenic potency of the three indicates that some portion of the mutagenic on a reduction of rev/plate in TA100 at the mutagenic samples was 1,190 rev/L-eq. Only activity of sample B1 in strain RSJ100 was due Table 2. DBPs identified in pool waters. Sample Sample DBP C1 C2 C3 C4 C5 B1 B2 DBP C1 C2 C3 C4 C5 B1 B2 Haloalkanes Halonitriles Chloroform x x x x x x x Bromoacetonitrile x x Bromodichloromethane x x x x x x x Dichloroacetonitrile x x x x x Dibromochloromethane x x x x x x x Bromochloroacetonitrile x x x x x x Bromoform x x x x x x x Dibromoacetonitrile x x x x x Dibromomethane x x x x x x x Trichloroacetonitrile x Bromotrichloromethane Haloketones Dibromodichloromethane x Bromopropanone x x 1,1,2-Trichloroethane x x 1,1-Dichloropropanone x x Haloacetic acids 1-Bromo-1-chloropropanone x x Chloroacetic acid x x x x 1,1-Dibromopropanone x x Bromoacetic acid x x x x x 1,3-Dibromopropanone x x Dichloroacetic acid x x x x x 1,1,1-Trichloropropanone x x x x x Bromochloroacetic acid x x x x x x x 1,1,3-Trichloropropanone x x x x Dibromoacetic acid x x x x x x 1-Bromo-1,1-dichloropropanone x x x Trichloroacetic acid x x x x x x x 1,1,1-Tribromopropanone x x x Bromodichloroacetic acid x x x x x x x 1,1,3,3-Tetrachloropropanone x x x x x Dibromochloroacetic acid x x x x x x x 1,1-Dibromo-3,3-dichloropropanone Tribromoacetic acid x x x x x x x Pentachloropropanone x x Other haloacids Dichlorofurandione x x 3-Bromopropenoic acid x 1-Chloro-2-butanone x x 2,2-Dichloropropanoic acid x x x x x 1-Bromo-2-butanone x x 3,3-Dichloropropenoic acid x x x x x Tetrachlorohydroquinone x x x cis-2,3-Bromochloropropenoic acid x x x x x x Halonitromethanes trans-2,3-Bromochloropropenoic acid x x x x x x Dibromonitromethane x x x x x 2,3-Dibromopropanoic acid x x x x x Haloamides cis-2,3-Dibromopropenoic acid x x x x Dichloroacetamide x x x trans-2,3-Dibromopropenoic acid x x x Bromochloroacetamide x x 3,3-Dibromopropenoic acid x x x x Dibromoacetamide x x x x x x x Trichloropropenoic acid x x x x x x x Bromodichloroacetamide x 2-Bromo-3,3-dichloropropenoic acid x x x x x x x Dibromochloroacetamide x x x (E)-3-Bromo-2,3-dichloropropenoic acid x x x x x x x Tribromoacetamide x ( Z)-3-Bromo-2,3-dichloropropenoic acid x x x x x x x Haloalcohols 2,2-Dichlorobutanoic acid x x x x 2,2,2-Trichloroethanol x cis-Bromobutenoic acid x x x x x 1,1,1-Trichloropropanol x x x trans-Bromobutenoic acid x x x Other halogenated DBPs 2,2-Dichlorobutenoic acid x 3-Chlorobenzeneacetonitrile x 2,3-Dibromobutenoic acid x x 2,6-Dichloro-4-methylphenol x x x 2-Chloro-3-methylbutanoic acid x x x x x 2-Bromo-4-chlorophenol x Chlorophenylacetic acid x x Trichlorophenol x x x x x 3,5-Dibromobenzoic acid x Bromodichlorophenol x x x x Tribromopropenoic acid x Tribromophenol x x x Halodiacids 2-Bromo-4-chloro-6-methylphenol x x x x cis-Bromobutenedioic acid x x x x x x x Dibromomethylphenol x x trans-Bromobutenedioic acid x x x x x 2,4-Dibromo-1-methoxybenzene x x cis-Dichlorobutenedioic acid x x x 2,3,4-Trichlorobenzeneamine x x x trans-Dichlorobutenedioic acid x x Dibromochloroaniline x cis-Bromochlorobutenedioic acid x x x x x x 2-Bromo-4-chloroanisole x x x trans-Bromochlorobutenedioic acid x x x x x x x 3,4,5-Tribromo-1H-pyrazole x cis-Dibromobutenedioic acid x x x x x x x 2,6-Dibromo-4-nitrophenol x (E)-2-Chloro-3-methylbutenedioic acid x x 2,6-Dibromo-4-nitrobenzeneamine x x (E)-2-Bromo-3-methylbutenedioic acid x Nonhalogenated DBPs/contaminants Haloaldehydes Propionamide x Dichloroacetaldehyde x x Benzaldehyde x x x x x x x Bromochloroacetaldehyde x x x Benzoic acid methyl ester x Dibromoacetaldehyde x x x x x Benzeneacetonitrile x x Trichloroacetaldehyde (chloral hydrate) x x x x x Phthalic acid x x x Bromodichloroacetaldehyde x x x x Diethylphthalate x Dibromochloroacetaldehyde x x x x x Benzophenone x Tribromoacetaldehyde x x x x x 3-Bromo-4-methoxybenzaldehyde x x x x x x Samples C1–C5 represent five samples from the chlorinated pool; B1 and B2 represent two samples from the brominated pool. “X” indicates that a particular DBP was identified in that sample. DBPs shown in italics were confirmed through the analysis of authentic standards; all others should be considered tentative identifications. | | 1526 volume 118 number 11 November 2010 • Environmental Health Perspectives DBPs in and mutagenicity of swimming pool water to the presence of DBPs that were activated those for drinking water, which are much concentrations of mono- and dichloramine by GSTT1, such as the brominated THMs more extensive (Richardson et al. 2007). In reported in Table 1 are consistent with chlo- (DeMarini et al. 1997; Pegram et al. 1997). addition, the pool water composition and rine oxidation of continuous supplies of small mutagenicity reported here can be used to amounts of nitrogen compounds coming from Discussion better understand the reported health effects urine, sweat, skin, and other human residues. Most analytical studies of pool water have of swimming pool water, such as asthma, irri- These levels are also similar to those reported measured only a few targeted DBPs, primarily tation of eyes/throat/skin, and bladder cancer by other researchers who used membrane- chloroform and other THMs. Consequently, (Weisel et al. 2009; Zwiener et al. 2007). introduction MS, which does not have issues this study expands considerably our knowl- Nitrogen-containing DBPs. In general, with interferences from organic chloram- edge of the chemical composition and muta- we observed more nitrogen-containing DBPs ines (Shang and Blatchley 1999; Weaver genicity of swimming pool water beyond the (N-DBPs) in these pool water samples than are et al. 2009). chemical analysis of two outdoor pools by found typically in chlorinated drinking water. The N-DBPs, including chloramines, were Zwiener et al. (2007) and the studies on pool For example, we found a greater number of not surprising to find because pool waters have water mutagenicity (Honer et al. 1980) and haloamides, halonitriles, haloanilines, haloani- a greater contribution of nitrogen-containing genotoxicity (Glauner et al. 2005; Liviac et al. soles, and halonitro-compounds than typically precursors due to human inputs. Because 2010). We found a greater number of DBPs found in drinking water, and several chemicals chloramines are known to cause eye irritation in the chlorinated and brominated indoor within these families have not been reported and other problems, pool operators generally pools studied here than have been found in previously in drinking water. In addition, we try to add enough chlorine to get beyond the chlorinated outdoor pools (Zwiener et al. detected mono- and dichloramine in the pool “break point,” such that these chloramines 2007), which was not surprising, consider- waters (means of 0.29 and 0.38 mg/L, respec- are destroyed, leaving residual chlorine (Ford ing that DBPs can be volatilized or photo- tively, for mono- and dichloramine in the 2007; World Health Organization 2000). lyzed (Lekkas and Nikolaou 2004) in outdoor chlorinated pool, and a mean of 0.27 mg/L However, the amount of chlorine needed to settings. In addition, although most people for monochloramine in the brominated pool). reach “break point” is also dependent on other assume that chlorine levels in swimming pools Because DPD analysis of chloramines can- amines in the water. As we observed in this are much higher than in chlorinated drinking not differentiate organic from inorganic forms study, this goal is not always achieved because water, the mean level of free chlorine (1.28 of these compounds, it is possible that these of continuous human inputs and rapid reac- and 0.50 mg/L in the chlorinated and bromi- levels are overestimated by the occurrence of tions forming chloramines. A few other nated pools, respectively) was similar to that organic chloramines in the swimming pool N-DBPs also have been reported in swimming found typically in drinking water. waters. Model studies with batch experiments pool waters, including organic chloramines Because little is known regarding the show that survival of chloramines depends (Li and Blatchley 2007), and nitrosamines mutagenicity and DBP composition of swim- on the chlorine:nitrogen ratio (Jafvert and (Walse and Mitch 2008), several of which are ming pool water, we compared our data with Valentine 1992). Considering this, the low carcinogenic. C OCH Br 3 Cl Br C C (E ) CH O C (Z ) 3 Cl CH O C C OCH O O 16.55 16.60 16.65 16.70 16.75 16.80 16.85 16.90 16.95 17.00 17.05 17.10 17.15 Time (min) (Z )-Bromochlorobutenedioic acid dimethyl ester m/z 225 m/z 197 Molecular mass = 256 Br Cl OO CH O CC CC OCH 3 3 m/z 177 +• m/z 59 256 258 77 177 59 221 43 182 64 89 93 133 179 197 57 69 73 99 131 230 68 78 127 199 201 220 252 260 268 83 140141 149 48 52 120 160 165 173 211 254 261 35 45 114 233 242 272 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 m/z Figure 1. (A) GC/MS chromatogram showing (Z)- and (E)-2-bromo-3-chlorobutenedioic acid dimethyl ester isomers. (B) Electron ionization mass spectrum for (Z)-2-bromo-3-chlorobutenedioic acid dimethyl ester. | | Environmental Health Perspectives • volume 118 number 11 November 2010 1527 Relative abundance Percent Richardson et al. Comparison of brominated versus chlo- Salmonella TA100 (without S9 mix), com- Our finding that some of the mutagenic rinated pool waters. Bromoform levels were pared with our finding of an average of 1,190 activity of one sample (B1) from the bromi- much higher in the pools treated with bromine rev/L-eq in the two indoor pools in Barcelona. nated pool water was due to activation by versus chlorine, but interestingly, other DBPs However, their solvent-extraction method GSTT1 suggests the presence of compounds and their levels were similar in brominated was considerably different from ours, involv - that are activated to mutagens by this enzyme, versus chlorinated pools, likely owing to the ing ether and acetone, whereas we used ethyl such as brominated THMs (DeMarini et al. high levels of bromide present already in acetate. Although a direct comparison of the 1997; Pegram et al. 1997), some methylene Barcelona source waters (Ventura and Rivera data is not possible, our study confirms their dihalides and bifunctional butanes (Thier et al. 1985) that feed into drinking water treat- pioneering work from three decades ago show- 1995), and/or 1,1-dichloropropene (Granville ment and further swimming pool treatment ing that swimming pool water is mutagenic. et al. 2005). Our chemical analysis (Table 1) (Judd and Jeffrey 1995). In addition, when In general, extracts of drinking water showed that sample B1 had high concen- THMs are compared on a molar basis, the induce an average of 1,100 rev/L-eq in trations of brominated THMs, especially chlorinated pool actually contained somewhat Salmonella strain TA100 (without S9 mix) bromoform. The high cytotoxicity and lack higher levels of total THMs (mean, 306 nM) (Takanashi et al. 2009); however, values as of mutagenicity of sample C5 may reflect the than the brominated pool (mean, 242 nM); high as approximately 15,000 rev/L-eq fact that the concentration of chloroform was this was possibly due to the carbon filtration have been reported (Egorov et al. 2003). 30% higher in this sample than in sample C4. used at the brominated pool that was not used Concentration methods such as reverse osmo- Perhaps the higher concentration of chloro- at the chlorinated pool. sis recover levels of mutagenic activity lower form, which is cytotoxic but not mutagenic, Mutagenicity. In the only other muta- than those recovered by XAD resin (Claxton produced the observed cytotoxicity, preventing genicity study of swimming pool water, Honer et al. 2008), which is why we used XAD to detection of mutagenic activity of the other et al. (1980) found that three public indoor prepare extracts of pool water. Our finding DBPs present in sample C5. As reviewed by pools in Victoria, British Columbia (Canada), that the pool water mutagenicity was similar Richardson et al. (2007), many other DBPs produced approximately 20,000 rev/L-eq in to that of drinking water may reflect the fact in drinking water that we have now identified that the levels of mutagenic DBPs in the pool in pool waters are known to be mutagenic Table 3. Mutagenicity of pool waters in Salmonella. waters were similar to those in drinking water, and/or carcinogenic, including the haloacetic despite the differences in the levels of specific acids, halonitromethanes, haloamides, halo- Strain (rev/plate) classes of DBPs described above in pool versus acetonitriles, and unregulated haloacids (Plewa Sample TPT100 RSJ100 drinking water. et al. 2008a, 2008b; Richardson et al. 2007). L-eq/plate (GST–) (GST+) TA100 C4 0 16, 19 10, 14 127, 115, 111 750 200 C4 C5 0.05 27 28 158 0.075 39 10 200 0.1 52 60 238 0.15 44 57 264 1,800 ± 636 rev/L-eq 0.2 69 81 343 0.3 87 70 495 r = 0.80 1,247 ± 70 rev/L-eq p = 0.105 C5 2 50 r = 0.98 0 9, 19, 12 6, 6, 5 75, 83, 94 p < 0.001 0.01 24 8 128 0 0 0.02 23 18 134 0.0 0.1 0.2 0.3 0.4 0.00 0.01 0.02 0.03 0.04 0.03 21 27 142 a a a L-eq/plate L-eq/plate 0.04 21 23 115 a a a 0.05 12 25 109 B1 B2 B1 0 27, 18, 20 9, 8, 5 130, 128 0.01 14 14 132 0.025 16 7 137 0.05 16 19 164 1,257 ± 149 rev/L-eq 1,067 ± 41 rev/L-eq 0.075 6 15 130 2 2 0.1 19 26 294 r = 0.91 r = 0.99 p < 0.001 p < 0.001 0.15 19 28 274 0.2 26 42 407 0 0 0.3 33 54 471 0.0 0.1 0.2 0.3 0.4 0.00 0.05 0.10 0.15 0.20 0.25 B2 0 16, 19 10, 14 127, 115, 111 L-eq/plate L-eq/plate 0.05 19 24 182 Figure 2. Mutagenicity in Salmonella TA100–S9 of two samples each from the chlorinated (C4, C5) and 0.075 29 28 199 (B1, B2) brominated pools. Data in each curve are from Table 3 and represent a single experiment performed 0.1 30 26 225 with one plate per dose. Slope (mutagenic potency) is rev/L-eq ± SE of the slope. 0.15 37 34 290 a a 0.2 32 34 330 a a a 0.3 26 30 373 Table 4. Mutagenic potencies of pool water samples in GST– and GST+ strains of Salmonella. The average rev/plate for the positive control, sodium Rev/L-eq ± SE (r ) azide (3 µg/plate), was 910 for TPT100, 519 for RSJ100, Sample TPT100 (GST–) RSJ100 (GST+) p-Value and 645 for TA100. The average rev/plate for the solvent blank (2 L-eq/plate) was 10 for RSJ100 and 128 for TA100; C4 228.2 ± 27 (0.93) 357.9 ± 95 (0.78) 0.131 it was not tested in TPT100. C5 500.0 ± 346 (0.68) 730.0 ± 128 (0.94) 0.508 Numbers were outside of the linear range of the dose B1 54.8 ± 20 (0.51) 159.1 ± 22 (0.90) 0.000 response and were not used to calculate the linear B2 136.0 ± 26 (0.90) 136.0 ± 28 (0.89) 0.194 regressions for potency values (Figure 2, Table 4). | | 1528 volume 118 number 11 November 2010 • Environmental Health Perspectives Rev/plate Rev/plate Rev/plate Rev/plate DBPs in and mutagenicity of swimming pool water In addition to the mix of mutagenic DBPs et al. 2010). These findings are especially rele- International Agency for Research on Cancer, Working Group on the Evaluation of Carcinogenic Risks to Humans. 2004. identified in the pool water, many other DBPs vant with regard to a case–control study by Some drinking-water disinfectants and contaminants, have not yet been studied for health effects, Cantor et al. (2010) in this issue that identifies including arsenic. IARC Monogr Eval Carcinog Risks Hum and no doubt, many other DBPs remain to an enhanced risk for bladder cancer associated 84:1–477. Jacobs JH, Spaan S, van Rooy GB, Meliefste C, Zaat VA, be identified that also may contribute to the with DBP exposure among people with geno- Rooyackers JM, et al. 2007. Exposure to trichloramine and observed mutagenicity of swimming pool types that metabolize various DBPs. Further respiratory symptoms in indoor swimming pool workers. water. In this regard, the study by Glauner research on a wide array of swimming pools Eur Respir J 29:690–698. Jafvert CT, Valentine RL. 1992. Reaction scheme for the chlorina- et al. (2005), which found that the low- under various conditions of maintenance and tion of ammoniacal water. Environ Sci Technol 26:577–586. molecular-weight fraction of extracts from use are warranted based on the limited but Judd S, Jeffrey JA. 1995. Trihalomethane formation during indoor and outdoor pools in Germany was developing data now available on the chemi- swimming pool water disinfection using hypobromous and hypochlorous acids. Water Res 29:1203–1206. the most potent of all fractions for inducing cal composition and health risks of swimming Kim H, Shim J, Lee S. 2002. Formation of disinfection by- DNA damage in mammalian cells (using the pool water. products in chlorinated swimming pool water. Chemosphere comet assay), suggests that the low- molecular- 46:123–130. weight DBPs may be most responsible for Refe Rences K o g e v i n a s M , V i l l a n u e v a C M , F o n t - R i b e r a L , L i v i a c D , Bustamante M, Espinoza F, et al. 2010. Genotoxic effects the genotoxic effect of swimming pool water. in swimmers exposed to disinfection by-products in indoor Aggazzotti G, Predieri G. 1986. Survey of volatile halogenated Also using the comet assay, Liviac et al. (2010) swimming pools. Environ Health Perspect 118:1531–1537. organics (VHO) in Italy: levels of VHO in drinking waters, found that pool water was more genotoxic LaKind JS, Richardson SD, Blount BC. 2010. The good, the surface water and swimming pools. Water Res 20:959–963. bad, and the volatile: can we have both healthy pools and than the source tap water; a similar analysis Ashley DL, Blount BC, Singer PC, Depaz E, Wilkes C, Gordon S, healthy people? Environ Sci Technol 44:3205–3210. et al. 2005. Changes in blood trihalomethane concentra- using the Ames assay would help in charac- Leavens TL, Blount BC, DeMarini DM, Madden MC, Valentine JL, tions resulting from differences in water quality and water terizing the relative mutagenicity of drinking Case MW, et al. 2007. Disposition of bromodichloro methane use activities. Arch Environ Occup Health 60:7–15. in humans following oral and dermal exposure. Toxicol Sci versus pool water. Sources of mutagens unique Beech JA, Diaz R, Ordaz C, Palomeque B. 1980. Nitrates, chlo- 99:432–445. rates, and trihalomethanes in swimming pool water. Am J to pool water could include active agents Lekkas TD, Nikolaou AD. 2004. Degradation of disinfection Public Health 70:79–82. in sunscreens, which can be transformed to byproducts in drinking water. Environ Engin Sci 21:493–506. Cantor KP, Villanueva CM, Silverman DT, Figueroa JD, Real FX, Li J, Blatchley ER III. 2007. Volatile disinfection byproduct mutagens by exposure to free chlorine under Garcia-Closas M, et al. 2010. Polymorphisms in GSTT1, formation resulting from chlorination of organic-nitrogen GSTZ1, and CYP2E1, disinfection by-products, and risk conditions similar to swimming pool water precursors in swimming pools. Environ Sci Technol of bladder cancer in Spain. Environ Health Perspect (Nakajima et al. 2009). 41:6732–6739. 118:1545–1550. Liviac D, Wagner ED, Mitch WA, Altonji MJ, Plewa MJ. 2010. Our limited data indicate that the muta- Chu H, Nieuwenhuijsen MJ. 2002. Distribution and determinants Genotoxicity of water concentrates from recreational of trihalomethane concentrations in indoor swimming genic potencies of chlorinated versus bromi- pools after various disinfection methods. Environ Sci pools. Occup Environ Med 59:243–247. nated pool waters were similar, as were the Technol 44:3527–3532. Claxton LD, Pegram R, Schenck, KM, Simmons JE, Warren SH. dose ranges over which the pool waters were Lourencetti C, Ballester C, Fernandez P, Marco E, Prado C, 2008. Integrated disinfection by-products research: Periago JF, et al. 2010. New method for determination of tri- Salmonella mutagenicity of water concentrates disinfected mutagenic, approximately 0.1–0.3 L-eq/plate halo methanes in exhaled breath: applications to swimming by chlorination and ozonation/postchlorination. J Toxicol (Figure 2). In contrast, the typical dose range pool and bath environments. Anal Chim Acta 662:23–30. Environ Health A 71:1187–1194. for drinking water mutagenicity is 0.3–1.5 M a r o n D M , A m e s B N . 1 9 8 3 . R e v i s e d m e t h o d s f o r t h e D e M a r i n i D M , A b u - S h a k r a A , F e l t o n C F , P a t t e r s o n K S , Salmonella mutagenicity test. Mutat Res 113:173–215. L-eq/plate (DeMarini et al. 1995). This differ - Shelton ML. 1995. Mutation spectra in Salmonella of chlori- Nakajima M, Kawakami T, Niino T, Takahashi Y, Onodera S. nated, chloraminated, or ozonated drinking water extracts: ence reflects the considerably higher toxicity 2009. Aquatic fate of sunscreen agents octyl-4-methoxy- comparison to MX. Environ Mol Mutagen 26:270–285. of swimming pool water relative to drinking cinnamate and octyl-4-dimethylaminobenzoate in model DeMarini DM, Shelton ML, Warren SH, Ross TM, Shim SY, swimming pools and the mutagenic assays of their chlori- water, with the highest testable mutagenic Richard AM, et al. 1997. Glutathione S-transferase- nation byproducts. J Health Sci 55:363–372. mediated induction of GC→AT transitions by halomethanes dose of pool water being the lowest mutagenic Pegram RA, Andersen ME, Warren SH, Ross TM, Claxton LD. in Salmonella. Environ Mol Mutagen 30:440–447. dose of drinking water. 1997. Glutathione S-transferase-mediated mutagenicity of E g o r o v A I , T e r e s c h e n k o A A , A l t s h u l L M , V a r t i a i n e n T , trihalomethanes in Salmonella typhimurium: contrasting Samsonov D, LaBrecque B, et al. 2003. Exposures to drink- results with bromodichloromethane and chloroform. Toxicol Conclusions ing water chlorination by-products in a Russian city. Int J Appl Pharmacol 144:183–188. Hyg Environ Health 206:539–551. We identified > 100 DBPs in two indoor Plewa MJ, Muellner MG, Richardson SD, Fasano F, Buettner KM, Erdinger L, Kuhn KP, Kirsch F, Feldhues R, Frobel T, Nohynek B, Woo Y-T, et al. 2008a. Occurrence, synthesis, and mamma- pools, including a prevalence of N-DBPs, et al. 2004. Pathways of trihalomethane uptake in swim- lian cell cytotoxicity and genotoxicity of haloacetamides: an ming pools. Int J Hyg Environ Health 207:571–575. likely formed from nitrogen-containing pre- emerging class of nitrogenous drinking water disinfection Font-Ribera L, Kogevinas M, Zock J-P, Gómez FP, Barreiro E, cursors from human inputs. This study pro - byproducts. Environ Sci Technol 42:955–961. Nieuwenhuijsen MJ, et al. 2010. Short-term changes in vides the most comprehensive analysis to date Plewa MJ, Wagner ED, Muellner MG, Hsu KM, Richardson SD. respiratory biomarkers after swimming in a chlorinated 2008b. Comparative mammalian cell toxicity of N-DBPs and pool. Environ Health Perspect 118:1538–1544. of DBPs in swimming pool waters, as well as a C-DBPs. In: By-products in Drinking Water: Occurrence, Ford RL, ed. 2007. Certified Pool-Spa Operator Handbook. clear demonstration of their mutagenicity. In Formation, Health Effects and Control (Karanfil T, Krasner Colorado Springs, CO:National Swimming Pool Foundation. addition, many DBPs we identified are new SW, Westerhoff P, Xie Y, eds). ACS Symposium Series 995. G l a u n e r T , Z w i e n e r C , W a l d m a n n P , F r i m m e l F H . 2 0 0 5 . Washington, DC:American Chemical Society, 36–50. and have not been reported previously in pool Swimming pool water—fractionation and genotoxicologi- Richardson SD. 1998. Drinking Water Disinfection By-products. cal characteri zation of organic constituents. Water Res waters. Bromoform levels were much greater In: The Encyclopedia of Environmental Analysis and 39:4494–4502. in the brominated versus chlorinated pools. Remediation, Vol 3 (Meyers RA, ed). New York:Wiley, Goodman M, Hays S. 2008. Asthma and swimming: a meta- 1398–1421. Compared with previous research on outdoor analysis. J Asthma 45:639–649. Richardson SD, Plewa MJ, Wagner ED, Schoeny R, DeMarini DM. Granville CA, Ross MK, Tornero-Velez R, Hanley NM, Grindstaff pools, we found a much greater number of 2007. Occurrence, genotoxicity, and carcino genicity of RD, Gold A, et al. 2005. Genotoxicity and metabolism of the DBPs in these indoor pools. regulated and emerging disinfection by-products in drink- source-water contaminant 1,1,dichloropropene: activation ing water: a review and roadmap for research. Mutat Res The mutagenicity of these pool waters was by GSTT1-1 and structure-activity considerations. Mutat 636:178–242. Res 572:98–112. similar to that of drinking water, indicating Richardson SD, Thruston AD Jr, Krasner SW, Weinberg HS, Haddad S, Tardif GC, Tardif R. 2006. Development of physiologi- that the levels of mutagenic DBPs are simi- Miltner RJ, Schenck KM, et al. 2008. Integrated dis infection cally based toxicokinetic models for improving the human by-products mixtures research: comprehensive characteri- lar in both waters. Subjects who swam in the indoor exposure assessment to water contaminants: zation of water concentrates prepared from chlorinated trichloroethylene and trihalomethanes. J Toxicol Environ mutagenic, chlorinated pool water evaluated and ozonated/postchlorinated drinking water. J Toxicol Health A 69:2095–2136. in this study had increases in geno toxicity bio- Environ Health A 71:1165–1186. Héry M, Hecht G, Gendre JC, Hubert G, Rebuffaud I. 1995. markers that were associated with the con- S a n t a M a r i n a L , I b a r l u z e a J , B a s t e r r e c h e a M , G o ñ i F , Exposure to chloramines in the atmosphere of indoor Ulibarrena E, Artieda J, et al. 2009. Indoor air and bathing swimming pools. Ann Occup Hyg 39:427–439. centrations of brominated THMs, but not water pollution in indoor swimming pools in Guipuzcoa Honer WG, Ashwood-Smith MJ, Warby C. 1980. Mutagenic chloroform, in their exhaled breath (Kogevinas (Spain). Gac Sanit 23:115–120. activity of swimming-pool water. Mutat Res 78:137–144. | | Environmental Health Perspectives • volume 118 number 11 November 2010 1529 Richardson et al. Shang C, Blatchley ER III. 1999. Differentiation and quantifica - Ventura F, Rivera J. 1985. Factors influencing the high content environmental exposures at swimming pools: state of the tion of free chlorine and inorganic chloramines in aqueous of brominated trihalomethanes in Barcelona’s water supply science and research recommendations. Environ Health solution by MIMS. Environ Sci Technol 33:2218–2223. (Spain). Bull Environ Contam Toxicol 35:73–81. Perspect 117:500–507. Stav D, Stav M. 2005. Asthma and whirlpool baths. N Engl J Villanueva CM, Cantor KP, Cordier S, Jaakkola JJK, King WD, W o r l d H e a l t h O r g a n i z a t i o n . 2 0 0 0 . G u i d e l i n e s f o r S a f e Med 353:1635–1636. Lynch CF, et al. 2004. Disinfection byproducts and bladder Recreational-Water Environments, Vol. 2, Swimming Takanashi H, Kishida M, Nakajima T, Ohki A, Akiba M, Aizawa T. cancer. A pooled analysis. Epidemiology 15:357–367. Pools, Spas and Similar Recreational-Water Environments. 2009. Surveying the mutagenicity of tap water to elicit the Villanueva CM, Cantor KP, Grimalt JO, Malats N, Silverman D, Geneva:World Health Organization. effects of purification processes on Japanese tap water. Tardon A, et al. 2007. Bladder cancer and exposure to Xu X, Weisel CP. 2005. Dermal uptake of chloroform and halo ketones Chemosphere 77:434–439. water disinfection by-products through ingestion, bathing, during bathing. J Exp Anal Environ Epidemiol 15:289–296. Thier R, Müller M, Taylor JB, Pemble SE, Ketterer B, Guengerich showering and swimming pool attendance. Am J Epidemiol Xu W, Weisel CP. 2004. Human respiratory uptake of chloro form FP. 1995. Enhancement of bacterial mutagenicity of bifunc- 165:148–156. and haloketones during showering. J Exp Anal Environ tional alkylating agents by expression of mammalian gluta- Walse SS, Mitch WA. 2008. Nitrosamine carcinogens also swim Epidemiol 15:6–16. thione S-transferase. Chem Res Toxicol 8:465–472. in chlorinated pools. Environ Sci Technol 42:1032–1037. Zwiener C, Richardson SD, DeMarini DM, Grummt T, Glauner T, Thier R, Taylor JB, Pemble SE, Humphreys WG, Persmark M, Weaver WA, Li J, Wen Y, Johnston J, Blatchley MR, Blatchley ER Frimmel FH. 2007. Drowning in disinfection byproducts? Ketterer B, et al. 1993. Expression of mammalian glutathi- III. 2009. Volatile disinfection by-product analysis from chlo- Assessing swimming pool water. Environ Sci Technol one S-transferase 5-5 in Salmonella typhimurium TA1535 rinated indoor swimming pools. Water Res 43:3308–3318. 41:363–372. leads to base-pair mutations upon exposure to dihalo- Weisel CP, Richardson SD, Nemery B, Aggazzotti G, Baraldi E, methanes. Proc Natl Acad Sci USA 90:8576–8580. Blatchley ER III, et al. 2009. Childhood asthma and | | 1530 volume 118 number 11 November 2010 • Environmental Health Perspectives

Journal

Environmental Health Perspectives – Pubmed Central

Published: Sep 12, 2010

There are no references for this article.