Polyfunctional alkylating agents have been studied for evidence of individual specificity of action on normal tissues and on tumors by using biological systems in which effects on multiple factors can be observed simultaneously. In these systems apparently related agents may reveal different patterns of biological activity.The principal system used in the studies to be reported here was the pregnant rat and its fetuses. There are several reports on the normal development of the rat fetus,'-* and many drugs and modified diets have been shown to produce developmental abnormalities.6-8 Furthermore, nitrogen mustard (HN2) will produce consistent teratogenic effects on the fetal l o a system that thus suggested itself for the study of the comparative activity of other polyfunctional alkylating agents.Three important variables involved in this system are: the fetus, the mother, and the pharmacological properties of the test drug. While these variables cannot be evaluated adequately from the limited data available, they must be considered in the interpretation of our results. The developing fetus presents a complex and evolving organism, and brief exposure to a noxious agent may produce a transient and specific action on active processes that subsequently may be magnified into extensive and specific developmental abnormalities in fetuses surviving to the end of gestation. While a specific teratogenic effect is a function, in part, of the stage of development when a noxious agent is applied, the effect is also related to the type of agent used.Another factor is the close relationship between the mother and fetus. This involves a consideration of the toxic effects of the drug on the mother; its metabolism, detoxification, and excretion; the possible presence of protective substances in the mother; its effects on her endocrine system; and the efficiency with which the drug or its metabolites are transmitted to the fetus. Furthermore, drugs with the same type of biochemical action may, because of prosthetic groups of unique chemical configuration, have different properties, such as solubilities, metabolic fates, or rates of transport into cells, that may influence their pattern of biological effects.Preliminary results from the comparison of several polyfunctional alkylating agents were available also on two systems in the chick embryo." In one system the drugs were tested for teratogenic activity by injection into