United States Environmental Protection Agency Health Effects Research Laboratory Research Triangle Park NC 27711 Research and Development EPA-600/S1-84-019 Jan 1985 Project Summary Water Distribution System as a Potential Source of Mutagens in Drinking Water Dipak K. Basu, Jitendra Saxena, Frederick W. Stoss, Joseph Santodonato, and Michael W. Neal The primary objectives of this study were to examine the changes in concen- tration of six polycyclic aromatic hydro- carbons (PAHs) and the possibility of changes in mutagenic potential of treated waters as a result of leaching during their passage through commonly used distribution pipes in the United States. With the exception of the fin- ished water from one utility in which the total concentration of the six PAHs was 138.5 parts per trillion (ppt), the total PAH concentration in all other treated water ranged from 0. to 13.4 ppt. The corresponding total PAH concentration in water after their passage through the distribution pipes varied from 0 to 61.6 ppt. This demonstrates that PAH con- centration in waters can increase as a result of their passage through the asphalt-line distribution pipes. M utagenic activity was also detected in many of the treated water samples, however, the levels of this activity did not correlate with either the transit of water through the distribution system or the levels of PAH in the water. There was some evidence to indicate that the water treatment process itself may have contributed to the mutagenicity ob- served in the finished water and that compounds responsible for the ob- served activity were different from the low molecular weight chlorinated com- pounds produced during chlorination. This Project Summary was developed by EPA's Health Effects Research Laboratory, Research Triangle Park, NC, to announce key findings of the research project that is fully documented in a separate report of the same title (see Project Report ordering information at back). Introduction Finished waters from the treatment sites are transported to the consumers through a variety of pipelines In the United States, three kinds of pipes are commonly used as transmission/distri- ' bution Imesfor transporting treated water to consumers, cast iron/ductile iron; asbestos/cement, and steel pipes. Other materials occasionally used as water distribution lines are reinforced concrete, a variety of plastic materials, iron, copper, and rarely, lead. With the exception of plastic pipes, coating materials are often used for the interior of the water pipes to prevent corrosion or to aid in curing the pipes. For example, cast iron/ductile iron pipes are lined sometimes with asphalt, cement, or asbestos/cement coatings; concrete pipes with asphalt coating; and steel pipes with cement or coal-tar coat- ings. Depending upon the characteristics, manufacturing conditions, thickness, and age of the pipe or its lining, some leaching of materials can be expected to occur as water passes through the pipes. The leaching of volatile organic compounds from different types of water pipes has also been studied in Europe. Cast iron pipes coated with polyurethane have been shown to add volatile contaminants such as chlorobenzene and aromatic com- pounds in drinking water. Both asphalt and coal tar are known to contain high levels of polycyclic aromatic hydrocarbons (PAHs), some of which are ------- known or suspected carcinogens, al- though the level of PAH in coal tar may be 1000 to 100,000 times higher than in asphalt It is possible that the coating of coal tar-based or asphalt-based paint or sealant on iron, steel, or concrete pipes could cause leach ing of small amounts of PAH into drinking water. The concentra- tion of any given PAH leached into the water is dependent on its solubility, concentration in the coating, contact time, etc. There are reports in the literature that undesirably high levels of PAH can result In a German study, a 10-fold increase in PAH concentration was reported at the end of water supply pipes resulting from the leaching of compounds from the coating used inside the water pipes. In a study in Pascagoula, Mississippi, more than nine PAHs, including fluoranthene at a concentration of 9.7 /ug/L, were reported in water stored in a tank freshly coated with coal-tar pitch. Similarly, potable w-.'er in Portland, Oregon, ob- tained from a terminal point of a water distribution pipe coated with coal tar showed the presence of PAH. Alt hough the leaching of coating mate- rials from the pipes can account for the presence of many compounds including PAH, it may not be the only mechanism which could contribute to the deteriora- tion of the water quality resulting from passage of drinking waters through sup- ply networks. The reaction of residential chlorine with organics leached from pipes, oxidation of leached materials, or microbiological synthesis of compounds on pipe walls and their subsequent release into the water may contaminate the water with compounds not originally present in the finished water. The examination of changes in water quality resulting from distribution pipes can be accomplished by determining changes in the levels of specific chemi- cals Because of their reported presence m some coating materials used inside the distribution lines, and because of the known carcinogenic potential of many of the compounds in this class, PAHs were chosen as a class of chemicals to be monitored in order to determine possible adverse effects of the distribution system on water quality. To monitor the effects of other harmful compounds that may be leached from the pipes or produced in the distribution system, an assay was also employed that was not specific for a particular class of chemicals. This general assay was an in vitro microbiological mutagenicity assay which has the capa- bility of detecting a large variety of compounds with genotoxic potential. Therefore, the present investigation was primarily oriented to provide information on two aspects of water quality: PAH content and the presence of mutagens in source waters, and the effect of treatment and passage of those waters through the distribution pipes on these measures of water quality. The sampling locations for this study represent the largest water supplies within 24 selected counties in the United States. These counties were selected by the U.S. Environmental Protection Agency for an epidemiological study of associa- tion between cancer mortality rates and drinking water quality. Twelve pairs of counties were selected with contrasting (high and low) total mortality rates for colon, rectum, bladder, and liver cancer in females during the years 1 968 to 1974. Each pairof counties was matched on the basis of five demographic characteristics (geographic region, total population, per- cent urbanization, median school years completed, and percent of work force in manufacturing industry). Only counties that were at least 50% urban were included. The studies reported here, were conducted in these counties to determine if the occurrence of PAH or mutagenic activity correlated with the occurrence of any of the types of cancer. The monitoring of the expected low levels of PAH in drinking water required a sample collection method that was cap- able of concentrating PAH from large volumes of water. A method developed in a previous study was found to be suitable and was used; the organic contaminants are adsorbed onto polyurethane foam plugs and subsequently desorbed by elution with an organic solvent. This method also includes a final chemical separation procedure (thin layer chroma- tography) capable of eliminating interfer- ing impurities and a quantification method (fluorescence spectrometry) sen- sitive enough to detect small amounts of PAH. The PAH detection limit for PAHs by this method was determined to be better than with capillary-gas chromatography with flame ionization detection. The overall mutagenic potential was assessed by utilizing the Ames Salmonella assay system. This assay employs a mammalian metabolic activation system, thus allowing the detection of compounds that are biologically active only after having been metabolized. The system was chosen primarily for two reasons. First, it provides a quick screening method for testing mutagenic potential of com- pounds. Second, a high correlation be- tween positive results in the Ames assay and positive results in in vivo animal carcinogenicity assays have been estab- lished for a large number of compounds. The scoring of samples for positive or negative mutagenic activity was inde- pendently conducted by three individuals. Objective evaluation criteria (i.e., doubling of spontaneous reversion rate, and demonstration of dose-related increase in revertants) were employed in the interpretation of all Ames assay results. However, in certain assays where the results were not definitive (e.g., more than a doubling but lack of dose-related increase, or positive dose-response curve, but number of revertants slightly less than twice the spontaneous) a subjective evaluation was also applied to the data. In these cases, a consensus of the three reviewers was obtained before declaring the result to the positive or negative. Results and Conclusions The results indicate that in 67 percent of the total water supplies studied, a measurable increase in concentration of PAH in water does occur as a conse- quence of treated water travelling through petroleum asphalt-lined distribution lines. The increases in PAH concentration, however, were small and amounted to a maximum of 61 ppt in water. It was also determined that the maximum leaching effect occurred with distribution pipes that had recently been coated with asphalt linings. The recently installed asphalt- coated pipes predominantly added the more water-soluble PAH, whereas leach- ates from the older pipes showed higher ratios of less water-soluble to more water-soluble PAHs. In older pipes this is probably due to depletion of the more water-soluble PAH from the surface of the coating. Waters from a few distribu- tion lines showed reductions in PAH concentration instead of the expected enhancements of PAH concentration. This reversing trend can be explained by assuming sorption of PAH on the surface of the distribution pipes or chemical interaction with oxidants in water. Distri- bution pipes that are depleted of the leachable PAH may act as sorbents for these compounds. Thus, depending on the age of the pipes, it is possible to get either desorption or sorption effects. In addition to the pre-selected six PAHs identified and quantified in this study, many other compounds were detected in the water samples. No attempt was made to identify these compounds. The results of bacterial mutagenicity assays indicated the presence of low ------- levels of mutagenic activity in many of the waters sampled. In general, the raw, finished, and distributed drinking waters tested in this investigation demonstrated no significant or consistent mutagenic activity in the Ames test system. Thus, it is not clear whether the leachate from certain distribution pipes actually con- tributes to the presence of mutagens or, on the other hand, whether some pipes adsorb mutagens which had not previous- ly been removed during treatment. Of the 24 water supplies tested, two showed no detectable mutagenic activity in either the raw water or water taken from any of the collection points. Five other water supplies had a number of sampling sites in which an individual assay point demonstrated a mutation ratio (number of revertants in experimental plate divided by number of revertants in control plates) of between 1.5 and 2.0 The random appearance of these weak mutagenic responses and the concomitant lack of a dose-related increase in mutation rate indicate that these values fall within the normal variation of the Ames assay When samples produced no mutagenic response the reason was probably that there were insufficient quantities of mutagens present. There was also the possibility, though, that some of these samples contained mutagenic compounds but the biological activity of these com- pounds was masked by the presence of toxic substances in the samples. Since toxicity tests were not performed on any of the water samples or their concen- trates, the only detectable evidence of severe toxicity in our investigation would be the absence of a bacterial lawn (the limited growth of histidine-dependent bacteria allowed by the trace amount of histidine supplied by the media in the minimal plate). The absence of a bacterial lawn did not result from the testing of any samples in this study. However, slight bacterial toxicity was occasionally encoun- tered as indicated by a dose-dependent decrease in the number of revertants with some samples. An example of this toxic effect was observed with the raw water samplefrom Fremont, Ohio, where average number of revertants for strain TA1535 declined from 79.5 for control to 62.0, 50.0, 56.5, and 8.4 as the dose increased. It is apparent from the variable effect of metabolic activation on bacterial toxicity (in some samples S-9 increased activity while in others it decreased toxicity) that the chemical composition and/or amounts of the toxic agents in individual samples varied extensively among the different collection locations. In most samples, the lowest dose tested appeared to be nontoxic to the bacteria. The types of pipe associated with the two water samples that demonstrated no mutagenic activity were cast iron and asbestos/cement. Although many vari- ables are unknown about the pipes such as age, condition, and the physical prop- erties (pH, types of solutes) of the water, it is apparent that this study shows no consistent relationship between the type of pipe and the absence of observed mutagenic activity in the water which has been transported by these pipes. Fifteen of the water supplies provided at least one sampling point which either produced a mutagenic response or a weak mutagenic response in at least one of the five strains of bacteria tested. In this study a response was considered to be weakly mutagenic if the reversion ratio fell between 1.5 and 2.0 and was accom- panied by a positive dose-response trend. A clear mutagenic response in this study was considered to occur when the rever- sion ratio equaled or exceeded 2.0. It should be noted that it is not unexpected to observe mutagenic responses with environmental samples where the high dose point has a lower ratio than the intermediate and low dose point. Such data are generally indicative of the toxicity to the tester strain of bacteria used in the Ames assay. Even in samples demonstra- ting clear mutagenic potential in this study, the response was not great, with most ratios being only slightly greater than 2.0. Recommendations On the basis of the results of the present project, the investigators recom- mend the following areas for further investigations: 1. Samples that showed mutagenicity should be fractionated to isolate and identify the component(s) re- sponsible for the mutagenic effect. 2. Since no dramatic PAH leaching was observed with the petroleum asphalt-lined pipes, further study should be directed towards the leaching effects from distribution pipes lined with coal tar-based products. 3. In the present investigation, the age of the installed pipes was found to be one of the most critical factors in the leachability of PAH from the pipe linings. In future studies, the age of pipes, condition of coating, and contact time should be given primary considerations in the selec- tion of sampling sites. Recently installed pipes are expected to provide more dramatic enhance- ment of PAH concentration than the older pipes. 4. Factors, such as pH, residual chlo- rine, aggressive index of water, and conditioning of the pipes after ap- plication of coat ings (all factors that may effect the leaching of PAHs), should be morethoroughly studied. 5. The effect of chlorination of drinking water on some of the leached compounds should be investigated. A few recent studies suggest that the leached compounds are con- verted to chlorinated or oxidized products in the presence of chlorine. 6. The apparent increase in mutagenic activity resulting from conventional water treatment processes should be studied in greater detail This should include an investigation of the production of high molecular weight compounds having biologi- cal activity as a result of chlorina"- tion. 7. The mutagenic activity of trace contaminants in treatment chemi- cals used for the production of finished water should be investi- gated as a possible source of muta- genic activity in drinking water. ------- D. K. Basu, J. Saxena, F. W. Stoss, J. Santodonato, and M. W. Neal are with Syracuse Research Corporation, Syracuse, NY 13210-4080. Frederick P. Kopfler is the EPA Project Officer (see below). The complete report, entitled "Water Distribution System as a Potential Source of Mutagens in Drinking Water,"(Order No. PB 85-125 474; Cost: $19.00, subject to change) will be available only from: National Technical Information Service 5285 Port Royal Road Springfield, VA 22161 Telephone: 703-487-4650 The EPA Project Officer can be contacted at: Health Effects Research Laboratory U.S. Environmental Protection Agency Research Triangle Park, NC 27711 * U S GOVERNMENT PRINTING OFFICE, 1985 — 559-016/7875 United States Environmental Protection Agency Center for Environmental Research Information Cincinnati OH 45268 BULK RATE POSTAGE & FEES EPA PERMIT No G-3 Official Business Penalty for Private Use $300 ------- |