Formic acid excretion in rats exposed to trichloroethylene: a possible explanation for renal toxicity in long-term studies
Introduction
Trichloroethylene has been shown to be nephrotoxic in animals, particularly the rat, following long-term exposure either by gavage or inhalation. In a number of studies, the nephrotoxicity in male rats has been accompanied by small increases in the incidences of kidney tubular cell adenomas and adenocarcinomas (NTP, 1983, NTP, 1988, NTP, 1990, Maltoni et al., 1986, Maltoni et al., 1988). These tumours have not been seen in the absence of kidney toxicity suggesting that toxicity is a pre-requisite for the development of these tumours.
A number of studies have investigated the mechanism(s) involved in the development of both the nephrotoxicity and the low incidences of kidney tumours seen in male rats exposed to trichloroethylene. These tumours were not seen in female rats or mice exposed under the same conditions. A minor metabolic pathway involving conjugation of trichloroethylene with glutathione was identified several years ago and has been linked with the adverse renal toxicology seen in the long-term studies (Dekant et al., 1986, Dekant et al., 1990, Commandeur and Vermeulen, 1990, Birner et al., 1993, Goeptar et al., 1995, Henschler et al., 1980, Henschler et al., 1995, Birnauer et al., 1996, Green et al., 1997). In vivo, this pathway occurs at very low levels in all species; typically the major urinary metabolite, N-acetyl-S-(1,2-dichlorovinyl)-l-cysteine (N-acetyl DCVC), accounts for approximately 0.005% of the administered dose of trichloroethylene (Commandeur and Vermeulen, 1990, Birner et al., 1993, Birnauer et al., 1996, Green et al., 1997). The precursor to the major urinary metabolite, S-(1,2-dichlorovinyl)-l-cysteine (DCVC), is known to be activated by the renal enzyme β-lyase to reactive species which are nephrotoxic in rodents and mutagenic in bacteria (Green and Odum, 1985), and consequently its formation is suggested to be the basis of the effects seen in the long-term studies.
Quantitation of the pathway leading to DCVC in rats and mice failed to find the expected correlation between metabolism by this pathway and the species difference in renal cancer seen in the bioassays. The levels of N-acetyl DCVC in mouse urine were several fold higher than those in the rat (Birner et al., 1993) and the mouse was significantly more susceptible to the acute nephrotoxic effects of DCVC than the rat (Eyre et al., 1995a, Eyre et al., 1995b, Green et al., 1997). The potency of DCVC as a renal carcinogen has also been questioned. The levels of this metabolite formed from trichloroethylene are approximately three orders of magnitude lower than the acute renal NOEL for DCVC in the rat (Green et al., 1997), and in the long-term, DCVC itself failed to cause renal cancer in rats, again at dose levels which were several orders of magnitude higher than the levels of DCVC found in rats dosed with trichloroethylene (Terracini and Parker, 1965). Thus, in addition to the lack of species specificity, there is a question about the toxicological significance of the very low levels of DCVC formed from trichloroethylene.
The lack of mechanistic data to support the assumption that the nephrotoxicity and low incidence of rat kidney tumours are a result of the metabolism of trichloroethylene to DCVC prompted the present search for an alternative explanation. In this paper, the acute renal toxicity of trichloroethylene has been assessed over a 28-day period, and the novel finding that rats exposed to this chemical excrete large amounts of formic acid is reported.
Section snippets
Chemicals
Trichloroethylene with a minimum purity of >99% was supplied by BDH (Poole, Dorset, UK). All other chemicals were of analytical grade or the highest grade available. [1,2-14C]-1,1,2-Trichloroethylene (14C-trichloroethylene) was supplied in sealed glass ampoules by Cambridge Research Biochemicals (Northwich, Cheshire) (19.3 mCi/mmol; 97%).
Animals
Male Fischer 344 rats (180–220 g bodyweight) were supplied by Harlan Olac UK. The animals were group housed in stainless steel cages in temperature controlled
Acute studies
Rats dosed with trichloroethylene, either by gavage or by inhalation, were found to have exceptionally high levels of formic acid in their urine. In contrast, the urine samples collected from rats prior to dosing or from control animals had little or no formic acid (Fig. 1). The excretion of formic acid following a single 1000 mg/kg gavage dose was higher on day 2 (5.8±2.1 mg/24 h) than day 1 (4.1±1.3 mg/24 h). Similarly, following a single exposure to 500 ppm trichloroethylene, excretion of
Discussion
Trichloroethylene is an extremely weak acute nephrotoxin as exemplified by the present 28-day study. There were no morphological changes in the kidneys at any of the time points, nor were there any changes in the sensitive biochemical markers normally used to detect kidney damage. Similar findings have been reported following gavage administration of high dose levels of trichloroethylene (Stott et al., 1982, Goldsworthy et al., 1988, Green et al., 1997). It would appear therefore that the
Acknowledgements
These studies were sponsored by members of the European Chlorinated Solvent Association (Brussels, Belgium), the Halogenated Solvents Industry Alliance (Washington, DC) and the Japan Association for Hygiene of Chlorinated Solvents (Tokyo, Japan).
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