Environ. Sci. Technol. 1991, 25, 93-98
Subchronic Toxicity Study of Ozonated and OzonatedKhlorinated Humic Acids in Sprague-Dawley Rats: A Model System for Drinking Water Disinfection F. Bernard Danlel," Merrel Robinson, H. Paul Rlnghand, Judy A. Stober, Norbert P. Page,+ and Greg R. Olson' Health Effects Research Laboratory, US. Environmental Protection Agency, Cincinnati, Ohio 45268
Male and female Sprague-Dawley rats were administered drinking water containing humic acids either nondisinfected or following ozonation (0,) or ozonation/ chlorination (03/C12)for 90 consecutive days. Test animals drank either of two concentrations of humic acids, 0.25 and 1.0 g/L total organic carbon (TOC), while controls received phosphate-buffered, distilled water. No consistent significant treatment-related effects were observed in body weight gain, organ weights, food or water consumption, or hematological and clinical chemistry parameters. No target organs were identified from the histopathological examination of the tissues. The most significant observation, an increase in liver to body weight ratio for the male animals in the 1.0 g/L 03/Cl, humic acid group, was not observed in any other group, nor was it corroborated via any biochemical measurements or histopathological analysis. Kidney lesions, primarily chronic progressive nephropathy, were a common observation in both controls and treated groups with no apparent relationship to either humic acid concentration or the disinfection process.
Introduction A search for alternative processes to the disinfection of drinking water by chlorination is underway based on the concern over hazardous byproducts formed by chlorination. Ozonation represents an attractive alternative and has been used to disinfect drinking water in France since 1905. It is currently in widespread use there and to a limited extent elsewhere in Europe. In contrast, ozonation has seen only minor usage in the United States, mainly for the disinfection of waste streams and for specialized disinfection of drinking water supplies e.g., bottled water. As discussed in the reviews by Anderson et al. (1) and Carmichael et al. ( 2 ) ,ozone has several advantages over alternative disinfectants. It is a strong oxidizing agent reacting with a wide variety of organic compounds and it is highly effective in controlling algal growth and in the elimination of microbes, including bacteria, amoebae, and viruses. In addition, ozone is effective in controlling and imparting desirable color, taste,and odor to finished water. It does not produce trihalomethanes (THM) in water as does chlorination and has, in fact, been shown to lower the levels of precursor organics that lead to THM formation. However, there are at least two characteristics about ozonation that are drawbacks to its use as a nationwide replacement for chlorination as the predisinfectant for drinking water: (1)with current technology for application, it is more costly, and (2) ozone has a rather short half-life (usually -20 min) in water. While this latter feature may
* Address correspondence to: Biochemical and Molecular Toxicology Branch, Genetic Toxicology Division, Health Effects Research Laboratory, U S . Environmental Protection Agency, Andrew W. Breidenbach Environmental Research Center, 26 West Martin Luther King Drive, Cincinnati, OH 45268. 'Page Associates, 17601 Stoneridge Court, Gaithersburg, MD 20878. Pathology Associates, 6217 Centre Park Drive, West Chester, OH 45069. 0013-936X/91/0925-0093$02.50/0
be desirable from the viewpoint of human consumption, it is an undesirable feature, overall, since there is no residual protection of the water supply to prevent microorganism growth in the drinking water distribution system. To compensate for the lack of a residual effect, it has been proposed that small amounts of chlorine or chloramine be added to ozone-purified water as postdisinfectants to provide residual protection for the distribution system. A major concern of the United States Environmental Protection Agency centers on the relative lack of knowledge regarding the chemical nature of organic reaction products of the ozonation process and the potential toxicity of these reaction byproducts. Still less is known about the health effects associated with byproducts of combined ozonation and chlorination treatments. This toxicity study is intended to be a preliminary evaluation of the toxic effects that might result from a protracted (90-day) ingestion of a very high concentration of disinfectant byproducts generated via the reaction of ozone or ozone followed by chlorine with aqueous humic acids. Thus, these humic acid solutions represent one approach to the generation of drinking water disinfection byproducts (DBP) at levels suitable for toxicological testing. The rationale for this approach centers about the observation that a majority of soluble organic materials found in surface waters are in the form of humic acid or fulvic acids. These are a heterogeneous and complex group of compounds, primarily organic acids, probably produced by oxidative coupling of phenolic and aliphatic organics. While there have been no studies comparable to those reported herein, there have been two related studies: one evaluated the toxic properties of organic concentrates extracted from drinking water treated with various disinfectants in a series of bioassays (3),and the other examined the 90-day subchronic toxicity of water containing gram per liter concentrations of humic acids identical with those reported here which had been previously treated with chlorine ( 4 ) . In like manner, this study evaluates the comparative toxicity of a 90-day exposure to drinking water containing gram per liter levels of either nondisinfected humic acids or humic acids that had been ozonated or ozonated and subsequently chlorinated (ozonated/chlorinated).
Methods Preparation of Test Materials. Three materials were prepared for administration to the test animals: (a) untreated (control) humic acid solutions of 1.0 g/L total organic carbon (TOG), (b) humic acid solutions of 0.25 and 1.0 g/L TOC that were ozonated, and finally, (c) humic acid solutions of 0.25 and 1.0 g/L TOC that were first ozonated and subsequently chlorinated. Phosphate buffered (0.05 M) distilled water (pH 8.02) was used as the aqueous vehicle and was administered to the control groups. The distilled water (conductivity 51 ppm, TOC
