O’Hara et al. (1983) conducted a published, peer-reviewed subchronic-duration oral study in which male Sprague-Dawley (COBS-CD(S-D)BR) rats were administered nitrofen (95.7% purity, impurities unknown) for 95 days before mating with untreated females. The study also evaluated male reproductive effects. Compliance with good laboratory practice (GLP) is unknown. Males received 0-, 100-, 500-, or 2500-ppm nitrofen in their food for 95 days prior to cohabitation and then 0-, 6-, 30-, or 155-mg/kg nitrofen in corn oil in 10-mL/kg volumes via gavage during cohabitation with females, up to 10 days until mating. Control rats were fed normal diet or a gavage of corn oil only. The study authors reported the averaged daily doses of nitrofen were 0, 7, 37, and 186 mg/kg-day before mating and during cohabitation. Both sexes were of equal age and were obtained from Charles River Breeding Farms (Kingston, NY). Unmated male and female rats were housed in stainless steel cages with wire mesh floors and fronts. After mating, suspected pregnant female rats were placed in larger cages (17"W × 10"D × 7.5"H) lined with bedding. Cages were inspected daily for births after 20 days of gestation. Environmental conditions were described by a temperature of 23 ± 2°C, relative humidity of 50 ± 15%, and a 12-hour light/12-hour dark cycle. Animals were fed Purina Rodent Laboratory Chow (No. 5001-Meal) and water ad libitum.

Researchers observed male rats twice daily to monitor for any ill health, morbidity, mortality, or reactions to nitrofen treatment (O’Hara et al., 1983). Male rats were weighed 1 week before administering the first dietary nitrofen dose and then weighed every week for the first 13 weeks (91 days) of the 95-day premating period. Mean body weights and food consumption were averaged weekly within dosed male study groups. After a maximum of 10 days of cohabitation with females, male rats were fasted overnight, and blood was removed from the orbital sinus for hematological and chemical analyses. Male rats were then sacrificed by exsanguination, necropsied, and examined thoroughly for gross abnormalities. Male kidneys, liver, and testes were weighed and then reported as percentage of total body weight. These organs were then fixed in 10%-buffered formalin, sectioned in 5-μm slices, and stained with hematoxylin and eosin for histopathological examination. Pregnant females were observed at different days of gestation, and offspring were observed twice daily for 13 weeks after birth. Pregnant dams were weighed on gestation days (GDs) 0, 6, 10, 16, and 21 and on lactation days (LDs) 0, 4, 10, 15, and 21. Three days after their expected delivery date, females were sacrificed and then grossly examined for lesions in their abdominal region and for the number of implantation sites present in their uterine horns. Subchronic data were analyzed using Sheffe’s Multiple Range Test.

No effect on food consumption was observed in males from any dose group (O’Hara et al., 1983). At the highest dose, a decrease in body-weight gain in males was seen throughout the first 13 weeks of exposure to nitrofen. Males exposed to 2500-ppm (186-mg/kg-day) nitrofen for 15 weeks exhibited statistically significant differences in blood glucose (86% of control value), total protein (113% of control value), albumin (111% of control value), globulin (116% of control value), and cholesterol levels (215% of control value) (see Table B.1). Male organ weights were increased in the liver (absolute—2500 ppm [186 mg/kg-day]; relative—≥500 ppm [37 mg/kg-day]), kidneys (relative—≥500 ppm [37 mg/kg-day]), and the testes (absolute and relative—≥500 ppm [37 mg/kg-day]) (see Table B.2). Gross necropsy revealed enlarged (6/25) and darkened (3/25) livers at 2500 ppm (186 mg/kg-day). The same dose resulted in slight-to-marked hydrotrophy of the liver and increased cytoplasmic basophilia of the male centribular hepatocytes (no further details reported). At 500 ppm (37 mg/kg-day), slight-to-very slight effects were seen in the hepatocytes of males (no further details reported). While histopathological data are discussed and pictures provided to illustrate the effects, no effort was made by the study authors to quantify the histopathological effects of nitrofen to male livers. No effects on gestation, fertility, litter size, weight, or sex ratio were seen in any dose group. Based on increased relative weights of the male liver, kidney, and testes, a LOAEL of 500 ppm (37 mg/kg-day) and a NOAEL of 100 ppm (7 mg/kg-day) are assigned for subchronic-duration oral exposure to nitrofen.

O'Hara et al. 1983

Risk of bias

Animal bioassay experiments

HAWC