George J. Levinskas

Effects of formaldehyde in the rat and dog following oral exposure

Formaldehyde was administered orally (p.o.) to groups of rats and dogs at dosages up to 150 and 100 mg/kg/day, respectively, for 91 consecutive days. Significant body weight changes were observed at the higher dosages tested in both species. Decreased water consumption was observed in a dose-related pattern in all treated groups of rats. Food consumption and feed efficiency were decreased at higher dosage levels in dogs. Results of other clinical studies and microscopic pathology revealed no specific, treatment-related effects on any organ or tissue examined. These findings suggest that formaldehyde possesses little subacute toxicity following oral exposure.

Evaluation of delayed neurotoxicity and dose-response relationships of phosphate esters in the adult hen.

Abstract An oral multipledose technique in hens was developed to determine the neurotoxic potential of a series of triaryl-, trialkyl-, and alkyl-aryl phosphates. This potential was estimated by observation of abnormal behavioral signs and always verified by histological examination. This procedure permits dose titration of an active material to quantify dose-response relationships. Initial evaluation of a series of phosphates for neurotoxicity yielded three active materials: tricresyl phosphate, cresyl diphenyl phosphate, and o-isopropylphenyl diphenyl phosphate. Subsequent experiments demonstrated that their neurotoxic potential differed markedly. The following conclusions regarding structure-activity relationships may be drawn from this study: (1) Phosphates prepared from o-alkyl-substituted phenols frequently are neurotoxic; however, the o-alkyl group must contain at least one hydrogen atom on the α-carbon for activation. (2) The neurotoxic potency of active o-alkyl-substituted phosphates declines with increased mass and branching of the o-substituent. (3) Neurotoxicity is reduced by further substitution in the ring already containing an o-substituent. (4) A series of unsymmetrical phosphates containing an alkyl group in the p-position of one ring are not neurotoxic even though they contain hydrogen atoms on the α-carbon. (5) p-Tertiary-butylphenyl diphenyl phosphate, with no hydrogen on the α-carbon available for activation, does not produced delayed neurotoxicity.

Subchronic inhalation toxicity of p-nitroaniline and p-nitrochlorobenzene in rats☆

For evaluation of subchronic toxicity of the two single-ring nitroaromatics, p-nitroaniline (PNA) and p-nitrochlorobenzene (PNCB), groups of 10 male and 10 female Sprague-Dawley rats were exposed to an aerosol/vapor of PNA in isopropanol at target concentrations of 0, 10, 30, or 90 mg/m3 or to PNCB vaporized from a solution in ethylene glycol monoethyl ether at target concentrations of 0, 5, 15, or 45 mg/m3 for 6 hr/day, 5 days/week for 4 weeks. Clinical signs of toxicity, body weights, results of ophthalmoscopic exam, hematology and clinical chemistry tests, organ weights, gross and histopathological changes were recorded. Exposure to PNA or PNCB resulted in a dose-related increase in blood methemoglobin levels. Mean red blood cell counts, hematocrit, and hemoglobin were significantly decreased in mid and high level animals exposed to PNCB. Mean spleen weights (absolute and relative to body weight) were significantly increased at the high dose levels in the two studies. A slight increase in spleen weights was also observed at the low concentration level in the PNA study. Absolute and relative liver weights also were increased among animals exposed to 45 mg/m3 PNCB. Microscopic changes were observed mainly in the spleen and included an increase in intensity of extramedullary hematopoiesis and hemosiderosis with both compounds. Spleens of animals exposed to PNCB also exhibited congestion. Neither PNA nor PNCB exhibited significant toxicological effects other than those of methemoglobinemia, anemia, and splenic changes classically associated with nitroaromatics at levels significantly above presently accepted occupational standard. Our data suggest that the current TLV for PNA which is 3 mg/m3 will provide adequate protection to the workers. OSHAs PEL of 1 mg/m3 for PNCB is to be preferred over the current TLV of 3 mg/m3 to provide a comparable margin of safety.

Evaluation of the teratogenic potential of three aliphatic nitriles in the rat

Acetonitrile (ACN), adiponitrile (ADN), and propionitrile (PN), were evaluated for embryotoxic and teratogenic potential in rats. Mated Sprague-Dawley rats were administered one of the three nitriles by gavage on gestation Days 6-19, inclusive. Daily dosage levels (mg/kg body wt) were: ACN at 0, 125, 190, and 275; ADN at 0, 20, 40, and 80; PN at 0, 20, 40, and 80. There was evidence of maternal toxicity in each of the high dose groups treated with ACN, ADN, or PN. Some maternal effects also were seen with ADN at the middle dosage. Embryotoxic effects were observed at the highest dosage tested for ACN and middle and high dose levels for PN. Slight fetotoxicity was observed at the highest dosage for ADN. No teratogenic effects were observed at any dosage level with ACN, ADN, or PN.

Relationships between Toxicity and Structure of Aliphatic Nitriles

A retrospective structure-activity relationship (SAR) comparison was undertaken of several of the toxicological properties (acute and subchronic toxicity, teratogenicity, and biochemical mechanism studies) of a series of structurally similar aliphatic nitriles based on available published data. Members of this chemical class included several mononitriles (aceto-, propio-, butyro-, and acrylonitrile), dinitriles (succino- and adiponitrile), and cyanohydrin derivatives (lactonitrile and acetone cyanohydrin). Broadly defined, retrospective SAR analysis suggested that the toxicological profiles of this group of aliphatic nitriles were similar; however, unique differences were observed within each of the toxicity categories and nitrile subclasses as the toxicological focus narrowed. Additionally, the toxicological properties associated with acetonitrile, the first of this chemically similar series, were unique and quite different from others within the homologous series. Finally, knowledge regarding the mechanism of toxicological action provided valuable information in relating toxicological properties among the aliphatic nitriles.

Comparative chronic toxicity and carcinogenicity of acrylonitrile by drinking water and oral intubation to Spartan Sprague-Dawley rats.

Groups of 100 male and 100 female Spartan Sprague-Dawley rats were administered lifetime oral doses of Acrylonitrile (AN) by one of two routes of dosing, either at 0.1 or 10 mg/kg per day, 7 day per week by intubation or continually at 1 or 100 ppm AN in their drinking water. The doses selected were designed to approximate the same daily intake of AN in each of two separate studies, whether by a single bolus dose (intubation) or a more continuous dosing regimen in drinking water. Each study had its own untreated control group of 100 rats per sex. In the drinking water study, the equivalent mean dosage of AN administered to males and females were 0, 0.09, and 0.15 mg/kg per day, respectively, at the 1 ppm level, and 0, 8.0 and 10.7 mg/kg per day, respectively, for 100 ppm dose groups. In both studies, groups of ten rats per sex were sacrificed at 6, 12 and 18 months and at study term. Ophthalmoscopic, hematological, clinical biochemistry, urinalysis and full histopathological exams were performed on control and high dose groups of rats in each study. Similar tests were done in lower dose groups, as required, to define dose-responses of observed effects. All animals were necropsied and underwent microscopic examination of target tissues, including brain, ear canal, stomach, spinal cord and any observable tissue masses. High dose male and female rats in both studies exhibited statistically decreased body weights. Food consumption and water intake were reduced only in the drinking water study. Due to increased deaths in groups of high dose rats of both studies receiving AN, all intubation test groups were terminated after 20 months of treatment. Surviving males and females in the drinking water study were terminated after 22 and 19 months, respectively. Small, sometimes statistically significant, reductions in hemoglobin, hematocrit and erythrocyte count were observed in male and female rats in both high dose (10 mg/kg per day intubation and 100 ppm drinking water) groups from both studies. There were increases in absolute or relative organ weight ratios for liver and adrenal in the high dose intubation study groups, but could not be correlated with AN toxicity in the absence of adverse clinical biochemistry or microscopic findings. Similar organ weight findings were not observed in the drinking water study. Again, there were no changes in clinical biochemistry or microscopic findings in these tissues. Absolute kidney weights were increased in high dose male and female rats in the intubation study and high dose female rats only in the drinking water study. Male and female rats from high dose groups in each study had a higher incidence of palpable masses of the head and the nonglandular stomach and, in females only, the mammary region. In both sexes, treatment-related tumors of the central nervous system (brain, spinal cord), ear canal, and gastrointestinal tract, and in females only, the mammary gland (intubation only) were observed in rats administered either 10 mg/kg per day by intubation or 100 ppm in drinking water. Animals from the intubation study had a substantially higher incidence of AN-related site-specific tumors than did their drinking water study counterparts. While a similar spectrum of tumors was produced by both oral dosing regimens, there were some notable differences in organ-specific incidence of tumors. Astrocytomas of the brain and spinal cord were found at a higher incidence in those rats exposed continuously to AN administered in the drinking water versus bolus dosing by intubation. Conversely, a higher incidence of squamous cell carcinomas/papillomas of the forestomach and adenocarcinomas of the intestine and, in females only, carcinomas of the mammary gland were observed in high dose rats receiving AN by intubation. An increase in the degree of severity of forestomach hyperplasia was observed in all high dose groups of animals, irrespective of mode of administration. These effects were more pronounced, were correlated with a much higher incidence of forestomach tumors, and were identified earlier (12 months) in the intubation study in which there was direct tissue contact with a more concentrated AN solution. Elevations in epidermal cysts in high dose males and females in the intubation study and renal hyperplasia in high dose animals of both sexes in both studies may have a treatment relationship. All other clinical and microscopic findings were considered unremarkable. There were no discernable non-neoplastic effects attributable to treatment in groups of low dose male and female rats given AN by intubation at 0.1 mg/kg per day or 1 ppm in drinking water. The results of this study indicate a consistent spectrum of neoplastic and non-neoplastic effects produced by AN in the same rat strain, whether administered orally by bolus or by continuous dosing in the drinking water. While the spectrum of tumors and target organ toxicity produced was similar, bolus dosing clearly increased tumors associated with the gastrointestinal tract. Neoplasms found in several other tissues were most prominently displayed in groups of more continuously dosed rats.

A Review of the Subchronic Toxicity of Butyl Benzyl Phthalate

This review compares the subchronic toxicity of butyl benzyl phthalate (BBP) across several species. Data from the published literature as well as previously unpublished studies sponsored by Monsanto are presented. BBP-induced toxicity occurs only at relatively high levels of exposure and is dependent on the species, age and strain of test animals used. These factors should be considered in extrapolating findings from animal toxicology studies to humans when assessing the safety of BBP.

Chronic toxicity and oncogenic dose-response effects of lifetime oral acrylonitrile exposure to Fischer 344 rats

Acrylonitrile (AN) was administered in the drinking water for approximately 2 years to groups of 100 male and 100 female Fischer 344 rats at nominal concentrations of 1, 3, 10, 30, and 100 ppm. Two groups, each of 100 males and 100 females, were used as untreated controls. Average daily intake was 0.1, 0.3, 0.8, 2.5 or 8.4 mg AN per kg body weight per day, respectively, for treated male rats and 0.1, 0.4, 1.3, 3.7, or 10.9 mg AN per kg per body weight per day, respectively, for dosed females. Clinical biochemistry, interim necropsies, organ weights and microscopic evaluation of tissues and organs were performed on groups of ten rats per sex per group at months 6, 12, and 18 and at study termination. Females were sacrificed in the 24th month and males were terminated after 26 months of dosing. A consistent decrease in survival, lower body weight and reduced water intake, as well as small reductions in hematological parameters, were observed in both sexes of the 100 ppm group. Elevated numbers of early deaths were observed in groups of males receiving 10 ppm AN and females receiving 30 ppm AN. Organ:body weight ratios at various study intervals were consistently elevated in the high dose group and likely were related to lower body weights. At these same intervals, mean absolute weights were either comparable to controls or only slightly elevated and few changes in weight ratios were seen when organ weights were compared with brain weights. No biochemical changes suggested a treatment-related effect. An increase in urine specific gravity in 100 ppm male rats was reflective of a decrease in liquid intake at this level. The only significant non-neoplastic finding observed histologically was a dose-related increase in hyperplasia/hyperkeratosis in squamous cells of the forestomach in male and female rats given 3 ppm and higher AN. This observation correlated with the induction of treatment-related squamous cell tumors (papillomas and carcinomas) of the forestomach seen primarily in rats at 3 ppm AN and higher. Mammary gland carcinomas were observed only in female groups. Both sexes given 10 ppm AN or more in their drinking water for their lifetime had astrocytomas of the brain/spinal cord and adenomas/carcinomas of the Zymbals gland.

Teratogenic Response of Dimethylacetamide in Rats

Pregnant CD rats (25/group) were used to determine the teratogenic potential of dimethylacetamide (DMAC). DMAC was administered in deionized water once a day by gavage on Days 6 through 19 of gestation at dosages of 0, 65, 160, and 400 mg/kg/day. Cesarean sections were performed on all females on Gestation Day 20. No treatment-related effects were observed in survival, appearance, or behavior at necropsy. Mean maternal body weight gain was reduced significantly only at the 400 mg/kg/day level. Fetotoxicity manifested by increased postimplantation loss was seen at the 400 mg/kg/day level while reduction in mean fetal body weights was noted at the 160 and 400 mg/kg/day test levels. Developmental variations (reduced ossification and unossified skeletal variations) were increased at the 400 mg/kg/day test level and corresponded to the reduced fetal body weights which were observed. Treatment-related malformations of the heart, major vessels and oral cavity, and anasarca were seen at the 400 mg/kg/day DMAC level. No teratogenic effect of DMAC treatment was observed at or below dosage levels of 160 mg/kg/day.

Teratology and multigeneration reproduction studies with maleic anhydride in rats

These studies were initiated to evaluate the effects of maleic anhydride on development and reproduction in CD rats. In the teratology study, pregnant rats (19-23/group) received 0, 30, 90, or 140 mg/kg/day maleic anhydride in corn oil orally from Days 6-15 of gestation and fetuses were examined for gross soft tissue and skeletal defects. A reduced weight gain or weight loss was observed in all maleic anhydride-treated groups between Days 6 and 9; however, mean weights of all groups were within 5% of control on Days 15 and 20. No treatment-related effects on fetal development were observed. In the multigeneration study, rats (10 males and 20 females/group) received 0, 20, 55, or 150 mg/kg/day maleic anhydride in corn oil orally and were mated to produce two generations, each with two litters. Groups of the same size from the second litter were used for subsequent generations and were given the same dose of maleic anhydride as were their parents. The high-dose group was terminated during the second generation due to treatment-related mortality in adults. Renal cortical necrosis occurred in high-dose Fo males and females. Increased kidney weights were observed in low- and mid-dose adult F1 females. No treatment-related effects on reproduction were observed with maleic anhydride at doses up to 55 mg/kg/day over two generations.