The Toxicologist - Society of Toxicology

tal neurotoxicity (DNT) test (OECD 426) has been performed on a potential new
generation plasticizer (Grindsted®Soft-N-Safe). This test included measurement
of antiandrogen endpoints. The tested plasticizer is a fully acetylated monoglyceride
based on castor oil. The potential reproductive and developmental neurotoxic effect
was evaluated in Sprague-Dawley rats. Test substance was administered in the diet
continuously for two generations at 1500 to 15000 ppm. The F0 generation animals
and F1 generation animals selected to rear the F2 generation were exposed for
a 10-week premating period, through mating for males and through mating, gestation
and lactation for females. Evaluations included reproductive and postnatal
endpoints, including sperm morphology and motility, estrous cyclicity, offspring
growth and development, and histopathology of reproductive tissues. A potential
endocrine disrupting effect was investigated by sexual maturation (F1offspring),
nipple retention and anogenital distance in the F1 and F2 offspring. Selected F1
offspring were assessed for DNT including a functional observational battery,
motor activity and function, auditory startle habituation, learning and memory and
histopathology of nervous system tissues and, additionally, pre-weaning motor activity
was measured in F2 offspring. Data obtained in this study show no reproductive
or hormone disrupting effects at dose levels up to 15000 ppm by dietary route.
At this stage the plasticizer has shown no significant toxicity to either adults or offspring
which makes it a relevant substitute for existing phthalates.
419 2, 4-DICHLOROPHENOXYACETIC ACID (2, 4-D):
EVALUATION OF SYSTEMIC TOXICITY IN A DIETARY
EXTENDED ONE-GENERATION STUDY IN
CRL:CD(SD) RATS.
J. S. Bus 1 , B. H. Neal 2 , C. L. Zablotny 1 , B. L. Yano 1 , S. Saghir 1 and M. S.
Marty 1 . 1 The Dow Chemical Co., Midland, MI and 2 Exponent, Alexandria, VA.
2,4-D systemic toxicity was assessed in an extended one-generation study that also
evaluated reproductive, developmental neurotoxicity (DNT) and immunotoxicity
(DIT), and endocrine endpoints across multiple life stages. CD rats (27/sex/dose;
P1 generation) were exposed to 0, 100, 300, 600 (female) or 800 (male) ppm 2,4-
D in the diet for 4 weeks pre-mating. P1 males were exposed post mating for 7
weeks and examined for systemic toxicity, including clinical pathology, organ
weights, histopathology and sperm parameters. P1 females were exposed through
gestation and lactation, and examined for systemic toxicity on lactation day 22. F1
offspring were examined for growth, survival, and developmental landmarks. At
weaning (PND 21), F1 offspring were divided into groups: Set 1a (systemic toxicity;
10/sex/dose), Set 1b (DNT; 10/sex/dose), Set 2 (DIT; 20/sex/dose), and Set 3
(reproductive toxicity; 20/sex/dose). Remaining PND 22 rats were used for systemic
toxicity and neuropathology (10-12/sex/dose). Doses were chosen such that
the high dose just exceeded the saturation threshold for renal clearance in adult rats.
High-dose females showed decreased lactation body weight, as did high-dose F1
pups; F1 animals recovered over time. The study confirmed kidney as a target organ
for 2,4-D toxicity. The characteristic renal lesion was very slight to slight degeneration
of the proximal convoluted tubules in the outer zone of the medulla.
Exposure-related kidney findings were in P1 high-dose males only, but extended to
mid- dose F1 adult males and high-dose F1 adult females. No renal lesions were
seen in PND 22 F1 pups. The pattern of kidney findings was likely related to
higher 2,4-D doses in F1 offspring and non-linear toxicokinetics. Across life stages,
the NOAEL for 2,4-D systemic toxicity was 100 ppm in males (5.5 mg/kg/day)
and 300 ppm in females (20.6 mg/kg/day in non-pregnant P1 adults). The male
NOAEL is ~13,000-fold higher than 2,4-D exposures reported in human biomonitoring
studies.
420 2, 4-DICHLOROPHENOXYACETIC ACID (2, 4-D):
EVALUATION OF REPRODUCTIVE/ENDOCRINE
ENDPOINTS IN A DIETARY EXTENDED ONE-
GENERATION STUDY IN CRL:CD(SD)RATS.
B. H. Neal 2 , J. S. Bus 1 , C. L. Zablotny 1 , B. L. Yano 1 , S. Saghir 1 and M. S.
Marty 1 . 1 The Dow Chemical Co., Midland, MI and 2 Exponent, Alexandria, VA.
An advantage of the extended one-generation study is that it examines multiple life
stages, and offers flexibility to verify age-related sensitivity and/or reproducibility of
results. Dietary doses to CD rats were 0, 100, 300, 600 (female) or 800 (male) ppm
2,4-D. P1 males exposed for ~ 11 weeks showed decreased seminal vesicle and
prostate weights (>300 ppm), but there were no associated effects on reproductive
function, sperm parameters or pathology. Control relative organ weights exceeded
historical control. Decreased testes weights were seen in PND 22 F1 weanlings; 800
ppm F1 males had slightly delayed preputial separation. In both cases, affected offspring
had decreased body weights; PND 22 pups showed no associated pathology.
Evaluation of groups of F1 males at PND 70 and PND 139 showed no effects on
male reproductive organ weights, pathology, and/or sperm parameters. The F1
males, exposed in utero, via lactation and in the diet, did not reproduce P1 organ
weight effects, despite longer exposures at higher doses (mg/kg/day). Thyroid hormones
(TH), weights and/or pathology were assessed in P1 adults, GD 17 dams,
F1 PND 4 and 22 pups, and F1 PND 70 and 139 adults. High-dose GD 17 dams
had non-significant TH changes and altered pathology (3 of 12), indicating an
adaptive change during the demanding period of gestation. In contrast, TH
changes in high- and mid-dose PND 22 pups had no corresponding thyroid weight
changes or pathology and were deemed not biologically significant. There were no
other effects on endocrine-sensitive endpoints, including anogenital distance or
nipple retention, vaginal opening, estrous cyclicity, reproductive indices or litter parameters
and mating of a second generation was not triggered. Thus, this integrated
study design supports the conclusion that 2,4-D did not induce reproductive toxicity;
endocrine toxicity, seen as adaptive thyroid changes on GD 17, only occurred
at a nonlinear toxicokinetic dose.
421 2, 4-DICHLOROPHENOXYACETIC ACID (2, 4-D):
EVALUATION OF DEVELOPMENTAL NEUROTOXICITY
(DNT) AND DEVELOPMENTAL IMMUNOTOXICITY
(DIT) IN A DIETARY EXTENDED ONE-GENERATION
STUDY IN CRL:CD(SD)RATS.
A. K. Andrus 1 , C. L. Zablotny 1 , B. L. Yano 1 , J. S. Bus 1 , B. H. Neal 2 and M. S.
Marty 1 . 1 The Dow Chemical Co., Midland, MI and 2 Exponent, Alexandria, VA.
The extended one-generation study design allows the flexibility to investigate a variety
of endpoints of interest in animals exposed during critical windows of development.
As part of a 2,4-D extended one-generation study which also characterized
reproductive and endocrine toxicity, DNT and DIT were examined in F1 offspring.
An advantage of this design is that stand-alone studies individually examining
these endpoints would have used > 1500 more animals. CD rats were exposed
in utero, during lactation and directly in the diet starting at weaning to 0, 100, 300,
600 (female) or 800 (male) ppm 2,4-D. For DNT, there were no exposure-related
effects on functional observational battery parameters (hand-held and open-field
observations, grip performance, landing foot splay, rectal temperature), motor activity
or auditory startle response (PND 54-59). Perfused PND 22 and 60 offspring
showed no exposure-related neuropathology in the central or peripheral nervous
system, effects on brain myelin (assessed by special staining) or PND 60 morphometry.
For DIT, 2,4-D had no effect on humoral immune function in males (all dose
levels) or females exposed to