IDENTIFICATION OF ENVIRONMENTAL 
GENETIC TOXICANTS 
WITH CULTURED MAMMALIAN CELLS 
Alexander R. Malcolm, Robert R. Young, 
and Carolyn J. Wilcox 
U.S. Environmental Protection Agency 
Environmental Research Laboratory 
Narragansett, R.l. 02892 
ABSTRACT 
Experiments designed to detect small-scale mutations leading to auxotrophy 
were carried out in vitro with the Chinese hamster ovarian (CHO) cell system 
(5-bromodeoxyuridine/visible light selection) initially described by Puck and 
Kao (43). The system was standardized with ethylmethanesulfonate (EMS), a 
known mutagen previously demonstrated to be active in CHO cells (27), and 
5-bromodeoxyuridine (BrdU), another known mutagen (7) utilized in the 
selection procedure, but not previously evaluated for mutagenic activity in the 
CHO Cell/BrdU-VL assay. Both EMS and BrdU routinely yielded glycine, 
hypoxanthine or triple-requiring (glycine/hypoxanthine/thymidine) 
auxotrophs and showed dose response. For a series of inorganic compounds 
known to be or suspected of being genetic toxicants, statistically significant 
numbers of auxotrophs were obtained only with the chloride salts of cadmium 
and manganese. Neither cadmium nor maganese were consistently mutagenic, 
cadmium showing activity in about 20 percent of experiments, manganese in 
50 percent of experiments. It was not possible to demonstrate dose response 
with these compounds. A water extract of JP-5 jet fuel was also found to be 
mutagenic in a single test. Variant cell types other than auxotrophs were 
isolated from cell populations treated with three different carcinogenic agents 
(EMS, CrO^, PbAc 0 .3H->0) but not from control experiments. These cells, 
exhibiting either a rounded cell morphology or potential contact inhibition, 
may reflect mutation in additional loci of possible value as genetic markers. 
Other data are presented to illustrate special problems associated with the 
application of in vitro cell systems. 
INTRODUCTION 
Serious concern for the possible effects of genetic toxicants in the 
environment developed approximately a decade ago with the discovery of 
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