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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Justification for type of information:
Data is from a National Toxicology Program (NTP) report.

Data source

Reference
Reference Type:
other: Study report
Title:
Unnamed
Year:
1991

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Principles of method if other than guideline:
The above experiment was performed to assess and evaluate the reproductive toxicity of the test chemical in Sprague Dawley rats.
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
not specified
Details on test material:
- Name of test material (as cited in study report): Naphthalene
- Molecular formula (if other than submission substance): C10-H8
- Molecular weight (if other than submission substance): 128.173 g/mol
- Substance type: Organic
- Physical state: No Data Available
- Impurities (identity and concentrations): 1% Impure (99% pure)
Specific details on test material used for the study:
- Molecular formula (if other than submission substance): C10-H8
- Molecular weight (if other than submission substance): 128.173 g/mol
- Substance type: Organic
- Physical state: No Data Available
- Impurities (identity and concentrations):1% Impure (99% pure)

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
Details on test animals and env. conditions
TEST ANIMALS
- Source: Charles River Laboratories, 1 NC, Raleigh, NC
- Age at study initiation: No Data Available
- Weight at study initiation: No Data Available
- Fasting period before study: No Data Available
- Housing: Sperm-positive females were individually housed in solid-bottom polycarbonate cages with stainless steel wire lids (Laboratory Products, Rochelle Park, NJ)
- Use of restrainers for preventing ingestion (if dermal): No
- Diet (e.g. ad libitum): Purina certified Rodent Chow animal diet #5002 was used. Feed was kept available ad libitum.
- Water (e.g. ad libitum): deionized/filtered water was available ad libitum throughout gestation.
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 72°F (range 69-75°F);
- Humidity (%): Average relative humidity was 55% in the first replicate (range 49-60%) and 48% in the second (range 44-63%). A 1-hr drop in relative humidity to 35.4% and a 2-hr drop in relative humidity to 19% were recorded during the second replicate, but these deviations did not affect the integrity of the study.
- Air changes (per hr): No Data Available
- Photoperiod (hrs dark / hrs light): 12 hours photoperiod cycle from 0600 hrs to 1830 hrs.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Timed-mated rats (8-10 weeks of age upon receipt) were administered test chemical (50, 150, or 450 mg/kg) or vehicle (corn oil, by gavage in a volume of 5 mllkg body weight. The corn oil used in the dose formulations contained less than 2 milliequivalents of peroxide/kg corn oil. The actual dose delivered to each animal was adjusted daily, according to a weight taken prior to dosing. For both study replicates, each dose of the test chemical was formulated independently in a sufficient quantity to last the entire dosing period. The chemical/vehicle mixtures had to be refrigerated to insure stability as determined by RTI. To avoid repeated heating and cooling of the test chemical solutions, each dose formulation was divided into aliquotes, such that one aliquot of each formulation was needed for any particular day of doses. Pre- and post-dosing analysis performed by RTI for each replicate were within 96%-108% of their theoretical concentrations. Thus, the test chemical/vehicle formulations here considered to be stable throughout the period of use for each study replicate. Treatment and examination of animals was performed without knowledge of the dose levels.

DIET PREPARATION:
- Rate of preparation of diet (frequency): No Data Available
- Mixing appropriate amounts with (Type of food): No Data Available
- Storage temperature of food: No Data Available

VEHICLE:
- Justification for use and choice of vehicle (if other than water): The test chemical was best miscible in corn oil.
- Concentration in vehicle: 0, 50, 150, or 450 mg/kg
- Amount of vehicle (if gavage): No Data Available
- Lot/batch no. (if required): No Data Available
- Purity: No Data Available
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
No Data Available
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: Overnight
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: Females were examined for the presence of sperm by vaginal smear. The day of sperm detection was designated as gestational day (GD 0).
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility.No Data Available
- Further matings after two unsuccessful attempts: [no / yes (explain)]: No Data Available
- After successful mating each pregnant female was caged (how): On GD 0, sperm-positive females were assigned to dose groups by stratified randomization so that body weights did not differ among groups.
Duration of treatment / exposure:
Gestation Day 6 through Gestation 15.
Frequency of treatment:
No Data Available
Duration of test:
No Data Available
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
Control Group
Dose / conc.:
50 mg/kg bw/day (actual dose received)
Remarks:
Low Dose Group (G1)
Dose / conc.:
150 mg/kg bw/day (actual dose received)
Remarks:
Mid Dose Group (G2)
Dose / conc.:
450 mg/kg bw/day (actual dose received)
Remarks:
High Dose Group (G3)
No. of animals per sex per dose:
28 animals per dose were used in the study.
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose range selected for this study was based on preliminary data. The effects of the test chemical (0, 100, 400, 500, 600 or 800 mg/kg/day administered by gavage on gestational days 6 through 15) on a limited number of maternal and fetal endpoints.
- Rationale for animal assignment (if not random): The animals were assigned randomly to the groups on the basis of the body weights
- Other: No Data Available

Examinations

Maternal examinations:
Maternal examinations were performed and included observations on the clinical signs, mortality, body weight analysis, food and water consumption, increase in organ weights and reproductive performance.
Ovaries and uterine content:
Relative and absolute uterine weights of maternal parameters, Number of pre-implantation and post implantation losses, Number of Resorptions were analyzed and contents were examined.
Fetal examinations:
Fetal Body weights, Changes in litter weights, postnatal survival, viability and mortality of the pups, External Malformations, Skeletal and Visceral Malformaions were observed in the pups.
Statistics:
General Linear Models (GLM) procedures ,were applied for the analyses of variance (ANOVA) o f maternal and fetal parameters. Prior to GLM analysis, an arcsine-square root transformation was performed on all litter derived percentage data (Snedecor and Cochran, 1967) and Bartlett's test for homogeneity of variance was performed on all data to be analyzed by ANOVA. GLM analysis determined the significance o f dose-response relationships and the significance of dose effects , replicate effects and dose x replicate interactions . When ANOVA revealed a significant dose effects, William and Dunnet’s Multiple Comparison Tests compared the test chemical-exposed to control groups. One-tailed test were used for all pairwise comparisons except maternal body and organ weights, maternal food and water consumption, fet al body weight and percent males per litre. Nonsignificant dose x replicate effects on selected fetal parametric measures were considered justification for pooling data across replicates for nonparametric analysis on related measures. When a significant dose x replicate interaction occurred, the data for that endpoint and for any related nominal scale data were analyzed separately for dose effects within each replicate in the study, as well as for all replicates combined. Nominal scale measures were analyzed by a test for independence and by a test for linear trend on proportions. When a test differences, a one-tailed Fisher's probability test was used for pairwise comparisons of the test chemical and control groups.
Indices:
Fetal weights, Pups survival index
Historical control data:
No Data Available

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Test chemical treated animals exhibited clinical signs of toxicity which were observed in all treatment groups on the first day of dosing. Lethargy, slow breathing, and prone body posture were the most comon clinical signs during the first five days of dosing, with the incidence of these effects showing a marked dose-dependence.
Dermal irritation (if dermal study):
not specified
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Two dams died during dosing and both were in the 50 mg/kg/day group. Necropsy data indicated that the deaths were unrelated to dosing errors.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The test chemical also caused a significant dose-dependent decrease i n maternal body weight and body weight gain; the effect was confined to the 150 mg/kg and 450 mg/kg groups, the same groups which exhibited suppressed food and water consumption.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
The test chemical also reduced maternal food and water consumption in the two highest dose groups. On GD 6 t o 9, relative food intake was suppressed 25% by 150 mg/kg of the test chemical and 37% by the higher dose. However, by GD 9 to 12, food consumption in the two groups had returned to control levels. A significant elevation in food consumption was noted on gd 18 to 20 in the 450 mg/kg/day group.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
Relative water intake in both groups displayed a pattern similar to food consumption. From GD 6 to 9, a 19% decrease was observed in the 150
mg/kg/day group (significant) and a 13% reduction i n the 450 mg/kg/day group ( non significant ); water consumption in both groups showed significant elevations of 15%-25% on GD 9 to 12 through GD 15 t o 18, before returning to control levels by GD 18 to 20. Absolute food and water consumption were similarly affected.
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Maternal liver weights (absolute or relative) were comparable across groups. There was a trend toward decreased gravid uterine weight with
increasing dose of the test chemical. Gravid uterine weights of the 50, 150, and 450 mg/kg/day groups were 105%, 95%, and 89% of control values respectively, but there is no significant difference among groups by ANOVA.
Gross pathological findings:
not specified
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Details on results:
No Data Available

Maternal developmental toxicity

Number of abortions:
effects observed, treatment-related
Description (incidence and severity):
The incidence of conceptuses adversely affected ( i e non live or malformed) was 8%, 7%, 13% and 20% in the control through high-dose groups, respectively.
Pre- and post-implantation loss:
effects observed, treatment-related
Description (incidence and severity):
The number of implantation sites per litter in the the test chemical-treated dams were within 95-102% of control values. However, there was a significant trend test for the percent adversely affected implants per litter.
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
There were also no statistically significant effects of treatment upon the incidence of resorptions.
Early or late resorptions:
no effects observed
Description (incidence and severity):
There were also no statistically significant effects of treatment upon the incidence of resorptions.
Dead fetuses:
no effects observed
Description (incidence and severity):
There were also no statistically significant effects of treatment upon late fetal deaths.
Changes in pregnancy duration:
not specified
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not specified
Changes in number of pregnant:
not specified
Other effects:
not specified

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
mortality
food consumption and compound intake
total litter losses by resorption
early or late resorptions
dead fetuses
Dose descriptor:
LOAEL
Effect level:
150 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
clinical signs
body weight and weight gain
water consumption and compound intake
organ weights and organ / body weight ratios
number of abortions
pre and post implantation loss

Maternal abnormalities

Abnormalities:
not specified

Results (fetuses)

Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
Average fetal body weight per litter also exhibited a significant trend test. Fetal body weights in the 50, 150, and 450 mg/kg/day groups were 105%, 99% and 95%) of control values respectively. Thus, a reduction trend was observed in the weights of the fetuses.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): effects observed, treatment-related
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): Average fetal body weight per litter also exhibited a significant trend test. Fetal body weights in the 50, 150, and 450 mg/kg/day groups were 105%, 99% and 95%) of control values respectively. Thus, a reduction trend was observed in the weights of the fetuses.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
There were also no statistically significant effects of treatment upon late fetal deaths.
Changes in sex ratio:
not specified
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
Statistical analysis did not detect a significant overall effect of dose for either the percent adversely affected fetuses or the average fetal body weight per litter.
Changes in postnatal survival:
no effects observed
Description (incidence and severity):
There were also no statistically significant effects of treatment on postnatal pup survival.
External malformations:
effects observed, treatment-related
Description (incidence and severity):
Analysis of anatomical defects indicated that the test chemical treatment tended to increase the incidence of malformations. Both the percent fetuses malormed per litter (4%, 4%, 7% and 10%) and the percent litters with malformed fetuses (23%. 274, 33% and 50%) showed a significant trend test. The largest difference was seen in the 450 mg/kg/day group where the percent fetuses malformed per litter were 2.5 times greater than controls (ie., 10% vs. 4%).
Skeletal malformations:
no effects observed
Visceral malformations:
effects observed, treatment-related
Description (incidence and severity):
The individual malformations suggested an effect of treatment on the incidence of enlarged ventricles of the brain. A separate statistical analysis of this malformation indicated that there was a significant dose-dependent increase in the percent fetuses per litter and the percent litters with enlarged ventricles.
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
No Data Available

Effect levels (fetuses)

Dose descriptor:
LOAEL
Effect level:
<= 50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
external malformations
visceral malformations

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified
Treatment related:
not specified
Relation to maternal toxicity:
not specified
Dose response relationship:
not specified
Relevant for humans:
not specified

Applicant's summary and conclusion

Conclusions:
Based on all the above observations and results, it was concluded that the LOAEL for the test chemical based on the fetus observation was found out to be <= 50 mg/kg bw/day.
Executive summary:

The above study was performed to evaluate and assess the reproductive and the developmental toxicity of the test chemical by dosing the Sprague-Dawley rats. In this study, the test chemical was dissolved in the vehicle (corn-oil) and the doses used in this study were 0, 50, 150, and 450 mg/kg bw/day as the control group, Low dose group, Mid Dose group and High dose group respectively. The animals were obtained fromCharles River Laboratories, 1 NC, Raleigh, NC annd were quarantined for 7 days before the study. The males were also obtained from a different dose and were used for mating. The males and female animals were co-housed together for mating kept together overnight. The next morning the female rats were checked foror the presence of sperm by vaginal smear. The day of sperm detection was designated as gestational day (GD 0). The animals were then dosed with the test chemicals from Gestational Day 6 through Gestational Day 15. The animals were then observed the different parameters. Test chemical treated animals exhibited clinical signs of toxicity which were observed in all treatment groups on the first day of dosing. Lethargy, slow breathing, and prone body posture were the most comon clinical signs during the first five days of dosing, with the incidence of these effects showing a marked dose-dependence. Two dams died during dosing and both were in the 50 mg/kg/day group. Necropsy data indicated that the deaths were unrelated to dosing errors. Further, the test chemical also caused a significant dose-dependent decrease in maternal body weight and body weight gain; the effect was confined to the 150 mg/kg and 450 mg/kg groups. The test chemical also reduced maternal food and water consumption in the two highest dose groups. On GD 6 to 9, relative food intake was suppressed 25% by 150 mg/kg of the test chemical and 37% by the higher dose. However, by GD 9 to 12, food consumption in the two groups had returned to control levels. A significant elevation in food consumption was noted on gd 18 to 20 in the 450 mg/kg/day group. Relative water intake in both groups displayed a pattern similar to food consumption. From GD 6 to 9, a 19% decrease was observed in the 150 mg/kg/day group (significant) and a 13% reduction i n the 450 mg/kg/day group ( non significant ); water consumption in both groups showed significant elevations of 15%-25% on GD 9 to 12 through GD 15 t o 18, before returning  to control levels by GD 18 to 20. Absolute food and water consumption were similarly affected. Maternal liver weights (absolute or relative) were comparable across groups. There was a trend toward decreased gravid uterine weight with increasing dose of the test chemical. Gravid uterine weights of the 50, 150, and 450 mg/kg/day groups were 105%, 95%, and 89% of control values respectively, but there is no significant difference among groups by ANOVA. In reproductive parameters, the incidence of conceptuses was adversely affected ( i.e. nonlive or malformed) was 8%, 7%, 13% and 20% in the control through high-dose groups, respectively. In fetal parameters, the incidence of conceptuses adversely affected ( i e nonlive or malformed) was 8%, 7%, 13% and 20% in the control through high-dose groups, respectively. Thus, the an upward trend was observed in mortality/non-viability of the pups. Average fetal body weight per litter also exhibited a significant trend test. Fetal body weights in the 50, 150, and 450 mg/kg/day groups were 105%, 99% and 95%) of control values respectively. Thus, a reduction trend was observed in the weights of the fetuses. A treatment related change, on the incidence of enlarged ventricles of the brain was observed in fetuses. Analysis of anatomical defects indicated that the test chemical treatment tended to increase the incidence of malformations. Both the percent fetuses malormed per litter (4%, 4%, 7% and 10%) and the percent litters with malformed fetuses (23%. 274, 33% and 50%) showed a significant trend test. The largest difference was seen in the 450 mg/kg/day group where the percent fetuses malformed per litter were 2.5 times greater than controls (ie., 10% vs. 4%).A separate statistical analysis of this malformation indicated that there was a significant dose-dependent increase in the percent fetuses per litter and the percent litters with enlarged ventricles (significant trend test for both). Thus, based on all the above observations and results, it was concluded that NOAEL for the test chemical for reproductive toxicity was found to be 50 mg/kg bw/day while the LOAEL for the test chemical based on the fetus observation was found out to be <= 50 mg/kg bw/day.