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As we have seen, economic entomologists celebrated DDT for its high toxicity against a wide range of insect pests. Few questioned the toxicity of DDT when employed in the fight against insects, yet scientists wondered if a chemical that was highly toxic to insects might also be to some degree toxic to plants, animals, and even humans. To explore this possibility, they conducted extensive pharmacologic studies on laboratory animals, including rats, rabbits, chicks, mice, guinea pigs, dogs, and monkeys. The results suggested the potential risks DDT posed to humans. Both the FDA and the PHS undertook ambitious research programs in this direction.
In 1944, Maurice I. Smith (chief pharmacologist at the PHS) and Edward F. Stohlman (associate pharmacologist) published the results of one of the earliest studies of the pharmacologic action of DDT on tissues and body fluids. Recall that Smith also conducted the toxicological analysis on Jamaica ginger in 1930. To test DDT, they devised a method to estimate the quantity of the chemical in its pure form. Noting that DDT given in aqueous suspension was irregularly and poorly absorbed in the stomach and intestines, the scientists administered DDT to rats and rabbits in olive oil, which produced superior results. For rats, the LD50 for DDT given intragastrically in 1 percent to 5 percent solution was 150 mg/kg (of body weight) and for rabbits 300 mg/kg, but death could be delayed for several days. DDT proved to be three times as toxic as phenol in rats and possibly twice as toxic to rabbits. Clearly, its acute toxicity was fairly modest, but scientists also considered the possibility that chronic toxicity might also be a serious problem.
Smith and Stohlman noted: “The effects of DDT in experimental animals are cumulative, and small single doses given repeatedly lead to chronic poisoning. In a group of 10 rats of about 80 grams weight, DDT fed at a level of 0.1 percent in a semisynthetic adequate diet containing 18 percent protein as casein was uniformly fatal in from 18 to 80 days.”32 Chronic toxicity was similar for rabbits and cats, although they were tested differently.33 Two cats that received 50 mg/kg every day or every second or third day developed all the characteristic symptoms of poisoning and died in one case after twelve days with a cumulative dose of 500 mg/kg and in the other after fifteen days (cumulative dose of 300 mg/kg).
Having established levels for acute and chronic toxicity of DDT, Smith and Stohlman examined whether DDT could be absorbed through the skin. They found little dermal absorption of DDT from impregnated clothing, but DDT applied to the skin in a dimethylphthalate solution was definitely toxic (dimethylphthalate was a nonirritant with a low toxicity to rabbits when given orally). Yet the scientists failed to rule out skin adsorption of DDT from impregnated cloths as approximately half of the animals exhibited systemic effects on the central nervous system.34 Thus one of the earliest pharmacologic investigations of the toxic effects of DDT reached a striking conclusion: “The toxicity of DDT combined with its cumulative action and absorbability from the skin places a definite health hazard upon its use. Symptomatically the effects on the central nervous system are the most obvious, damage to the liver is less obvious and for this reason perhaps more serious.”35
M. I. Smith also collaborated with R. D. Lillie (senior surgeon of the PHS) to determine the pathology of the poisoning experiments described above. This study clarified some of Smith’s findings. Despite Smith’s observations of pronounced neurologic symptoms, pathological examination revealed only slight histologic alterations in the central nervous system. As Smith suspected, the liver contained the most striking pathologic alterations. Hyaline degeneration was similar to that described in poisoning by azo-benzene (Lillie, Smith, and Stohlman had previously published their study of azo-benzene). Lillie and Smith also observed a variable amount of fatty degeneration of liver cells (often centrolobular) in cats, rats, and rabbits (all of the animals considered in this study).36
In 1943, a team of researchers from the NIH that included Neal (senior surgeon) and Oettingen (who had become principal industrial toxicologist) conducted experiments to determine the toxicity and other potential dangers of aerosols containing DDT. The research was divided between the Industrial Hygiene Research Laboratory and the Pathology Laboratory. They exposed animals for forty-five minutes to approximately 1 pound of DDT aerosol in a glass chamber. Guinea pigs, rats, and puppies showed no signs of discomfort or poisoning during or following exposure to initial concentrations of 54.4, 12.44, and 6.22 mg of DDT per liter of air over the period described. Mice, however, became “jumpy” shortly afterward and subsequently developed tremors, became hyperexcitable (as in strychnine poisoning), and developed clonic-tonic convulsions shortly before death. Eleven out of twenty mice exposed to an initial concentration of 12.44 mg of DDT per liter of air died during the four days after exposure. Yet the remaining nine mice showed no toxic symptoms and gained weight during the following three weeks. When the researchers dropped the initial concentration of DDT slightly (from 6.2 to 6.1 mg/liter), none of the animals (including mice) exhibited toxic symptoms. Raising the concentration of sesame oil in the aerosol to 9.5 percent induced toxic effects in the mice, prompting the researchers to conclude: “It is, therefore, apparent that the toxicity of DDT for mice may be increased by increasing the percentage of sesame oil in the aerosol mixture.”37 The researchers attributed some of the effect of sesame oil to contamination of the mouse fur (ingested when the mouse licked its fur) and the toxicity of DDT was increased by the presence of fatty material.
The NIH team also tested the potential chronic toxicity of DDT aerosols, with experiments similar to those used to test for acute toxicity. Two puppies were exposed to a 1 percent DDT aerosol at a concentration of 12.2 mg/liter for forty-five minutes on two consecutive days one week and four successive days in the following week. Neither puppy exhibited any signs of poisoning, and both gained weight. As in the tests of acute toxicity, mice fared worse than puppies. Thirty mice were exposed to the same conditions as the puppies. Ten mice wrapped in gauze to protect their fur from contamination showed no symptoms of poisoning, but ten unprotected mice died with typical symptoms of DDT poisoning. Finally, ten mice with protective collars (to restrict licking of their fur) manifested typical symptoms that were slightly delayed and less severe, but most of the group died within five days following the exposure. A second series of tests refined the experimental design. In these tests, two monkeys and ten mice were exposed to a concentration of 0.183 mg/liter of DDT daily for a total of five weeks’ duration. This concentration was less than 6 percent of that used for the initial tests of chronic toxicity. Neither the monkeys nor the mice exhibited toxic symptoms. Moreover, the monkeys gained weight (approximately 900 grams) and so did the mice (5 grams each on average).38
In effect, the tests for chronic toxicity amounted to acute toxicity tests stretched out over a longer duration as the researchers made no effort to identify etiology independent of acute symptoms. The PHS also tested humans for the toxicity of DDT (I will discuss these experiments, below, with the other tests conducted on human subjects). Researchers ignored all of these problems when they concluded: “Therefore, it may be concluded that in spite of its inherent toxicity the use of DDT in 1- to 5-percent solutions in 10 percent cyclohexanone with 89 to 95 percent Freon, as aerosol, should offer no serious health hazards if used under conditions required for its use as an insecticide.”39
Neal and the NIH researchers also compared the toxicity of DDT inhaled to that ingested. Only one of three dogs in the inhalation group developed definite signs of poisoning. After closely monitoring this individual’s symptoms, scientists found by autopsy that it had cirrhosis of the liver, prompting them to suggest that liver and kidney dysfunction may precede the onset of nervous symptoms. The three dogs that ingested DDT showed no distinct toxic manifestations, and autopsies of these dogs exhibited no gross pathological changes. After comparing the results of the two experiments, scientists concluded that dogs tolerated comparatively large doses of pure DDT (100 mg per kg) in capsules by mouth and by insufflation (forced inhalation) of the dry powder.40
This set of experiments addressed the crucial factor of body weight, but the techniques used to administer DDT may have affected the results. In the insufflation experiments, researchers blew DDT powder directly into the nostrils of dogs. In the ingestion tests, dogs were fed large quantities of DDT in capsule form. Neither set of experiments was likely to result in an exposure that could be replicated under noncontrolled circumstances. Yet the experiments did raise the possibility that sufflation of large doses of DDT caused definite signs of poisoning, preceded by injury to the liver and kidneys. Such findings broadened the range of possible toxic effects of DDT.
Determining the toxicity of DDT was not the sole purview of the NIH. When Congress cut funding for the FDA, explicitly prohibiting it from conducting research on arsenic and lead, it transferred responsibility for research on the toxicity of pesticides to NIH. Barred from research on lead and arsenic, the FDA pharmacologists shifted their emphasis to the toxicity of Elixir Sulfanilamide and glycols (see chapter 1). Next they analyzed the toxicity of mercury through an army contract. To avoid possible violation of the terms of the congressional appropriation, they undertook the study of the toxicity of DDT under the same contract.41
As early as 1943 the head of the Division of Pharmacy, Herbert O. Calvery, received a sample of DDT for toxicity testing. Calvery and a team of five scientists from the Division of Pharmacology tracked histopathological changes in 117 animals of nine species, including farm animals like chicks, dogs, cows, sheep, and a horse. As in Lillie and Smith’s research, the most characteristic and most frequent lesion produced by the higher dosage levels of DDT was moderate liver damage. No one on the research team had seen the exact counterpart in any other experimental animals that they had ever studied. Because of the severe muscular tremors following large doses of DDT, researchers went to considerable lengths to obtain the brains and spinal cords from affected animals to search for evidence of physical changes to the central nervous system. Although they sectioned the brains of numerous animals and controls, they found no distinct differences between test and control animals. Finally, certain individuals were much more sensitive that others of the same species, but the lesions (i.e., manifestations of injury) were quite consistent throughout the different species.42 In other words, differences between individuals of the same species were greater than differences between different species.
The FDA researchers also studied acute and subacute effects of DDT on small laboratory animals. Rats, mice, guinea pigs, rabbits, and chicks received DDT dissolved in corn oil via a stomach tube. One problem with the design of this experiment was the quantity of corn oil required to dissolve larger doses of DDT. Still, the experiments showed that relatively small doses of DDT intoxicated small animals and that the dosage-mortality curve was flat (that is, survivals and deaths occurred over a wide range of dosage). The researchers also concluded that rats and mice were more sensitive to DDT in single does than guinea pigs and rabbits and that DDT in solution was more readily toxic than DDT in suspension.43
For the tests of subacute toxicity, the FDA researchers fed DDT in different concentrations (ranging from 0.0 percent to 0.10 percent) to four different groups of rats for a period of one year. Several of the rats at the high dosage levels exhibited typical DDT symptoms after a few days and some died. The survivors at this level eventually developed symptoms and perished. The rats exposed to 0.025 percent DDT did not suffer increased mortality over the course of the fifty-two-week experiment. Autopsies of the dead animals most commonly revealed slight to moderate liver damage, occasional testicular atrophy, and some degeneration of the thyroid. Although researchers initially suspected that the rats were developing a tolerance to DDT, when they calculated food intake per kilogram body weight per day and plotted this against age, it became clear that the amount of DDT consumed per kilogram per day gradually declined with age.
The FDA researchers also investigated the impact of DDT on growth rate in rats in a paired experiment. Eight pairs of male rats and eight pairs of female rats were placed on experiment at the beginning of weaning. One member of each pair received 0.05 percent DDT dissolved in corn oil mixed with food, and the other received a control diet containing an equivalent amount of corn oil without DDT. After eleven weeks, the scientists terminated the experiment and concluded that feeding 0.05 percent DDT to rats did not appear to significantly slow the growth rate although the general trend of the average growth rate was downward.44 The conclusions of this report transcended the results of any of the individual experiments and pointed to one of the aspects of DDT that toxicologists found most troubling, namely, the variability in individual susceptibility, which made it difficult to estimate the safely tolerated dose or exposure.45
Calvery’s team of researchers also investigated the dermal or percutaneous absorption of the DDT in anticipation of its eventual use on human body lice. Acute toxicity tests exposed rabbits to 4 g/kg body weight of 5 percent DDT powder (talc). Neither dry nor wetted powder produced toxic symptoms. Having ruled out acute toxicity, the FDA researchers wondered if DDT in other solutions would prove equally “innocuous.” To find out, they conducted a ninety-day subacute experiment.46 They found DDT to be mildly irritating to intact and abraded skin, especially when applied by patch test or on the hands of operators who worked with it on a daily basis for nearly a year. In a carefully worded conclusion, the scientists recommended caution in the use of DDT: “The above data indicated that the unlimited use of DDT solutions on the skin is not free of danger; however, some solutions of DDT have been found safe for restricted use.”47
Although most of the early DDT experiments addressed its acute toxicity, a team of researchers from the Pharmacology Section of the Chemical Warfare Service explored the chronic toxicity of DDT in dogs. Daily doses of 100 mg/kg initiated “moderate, coarse tremor,” which disappeared after DDT was withdrawn. Raising the dose from 150 to 250 mg/kg of DDT produced more severe neurological disturbances (intense tremors involving all muscle groups, aberrations in gait, exaggeration of the stretch reflexes), but symptoms diminished and disappeared a few days after the treatment ended. At higher doses (greater than 250 mg/kg), persistent neurological signs developed: “severe hypermetria [high stepping each step], rigid, hyper-extended and abnormally abducted legs, and aberrations in gait” (dogs could not walk in a straight line but proceeded in a zigzag fashion). Other symptoms included inability to eat, leading to severe weight loss and dehydration. Researchers were most concerned, however, that the tolerance of dogs to DDT declined markedly: a 40 mg/kg dose (which yielded few or no symptoms in a normal dog) produced severe symptoms in dogs on which they repeated the experiment. They concluded: “It was apparent from these observations that irreversible symptoms had been produced by the prolonged administration of DDT. Their clinical nature indicated that injury to the cerebellum might have played a major rôle in their production.”48 The researchers also found that DDT damaged the liver, sometimes only moderately but in some cases so severely that the animals died. DDT had no detectable effect, however, on renal function in dogs.
For the most part, these DDT investigations did not require methodological innovations. Most of the experiments involved the calculation of LD50s and pathological examinations. One exception was the research of Edwin P. Laug, one of the pharmacologists in the FDA Division of Pharmacology. Laug developed a biological assay for the determination of DDT in animal tissues and excreta based on the toxic response of the housefly (Musca domestica). The great sensitivity of the housefly to DDT (and other chemicals) provided an excellent indicator. According to Laug, under proper conditions, the LD50 was on the order of 2.5 mg/kg bodyweight for houseflies.49 He extracted the fat from the tissue of a DDT-poisoned animal, placed the extract in a flask, and introduced 100 flies, which picked up residues of the extract while walking around the flask. After a specified amount of time, Laug counted the number of living and dead flies and plotted a curve to determine the point at which half of the flies had
died. Using this bioassay, which he referred to as his “flyo-assay,” Laug could determine quantities of DDT on the order of 2.5 ppm. In time, Laug’s flyo-assay would be superseded by more refined chemical methods, but it was a useful technique during the early analysis of the toxicity of DDT and other novel chemicals.50
Most of the early tests of the acute or subacute toxicity of DDT on laboratory animals lasted for a few months at most. Even the tests for chronic toxicity continued less than six months. Edwin Laug and his colleague at the FDA, O. Garth Fitzhugh, changed that state of affairs when they conducted experiments on rats lasting for at least six months and up to two years. In one paired feeding experiment, rats received diets containing either 800 or 1,200 parts per million (ppm) for six months, after which all the animals showed characteristic symptoms of DDT poisoning. After six months, tissue analysis revealed measurable amounts of DDT in all tissues (save two kidneys). DDT seemed to be particularly concentrated in the perirenal fat (i.e., surrounding the kidney), at 50 to 100 times as great as in any of the other tissues. The most compelling finding of this study came from a two-year exposure to DDT, after which there was a definite correlation between tissue level, most clearly seen in perirenal fat and the level of DDT consumed. Yet at 800 ppm, the DDT concentration in perirenal fat was of roughly the same order of magnitude after six months’ exposure as after two years of exposure. In contrast, DDT levels in the kidneys continued to rise during the two years, reaching levels four to five times as high as that recorded after six months.51 DDT was the first insecticide subjected to long-term studies lasting up to two years despite the fact that heavy metal insecticides like lead arsenate were arguably more toxic. Studies of the toxicity of lead arsenate to dogs and rats preceded the DDT studies by a few years even though the insecticide had been in use for decades. Not surprisingly, FDA pharmacologists conducted this research.52