N,N-diethylaniline

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

The reproduction toxicity endpoint is considered as a waiver since, data from a robust pre-natal developmental toxicity screening study is present.

Link to relevant study records

Reference

Endpoint:

extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)

Data waiving:

other justification

Justification for data waiving:

the study does not need to be conducted because a pre-natal developmental toxicity study is available

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

The reproduction toxicity endpoint is considered as a waiver since, data from a robust pre-natal developmental toxicity screening study is present.

Effects on developmental toxicity

Description of key information

Developmental Toxicity Study:

A pre-natal developmental toxicity study was conducted to evaluate and assess the effects of the test chemical on the pregnant rats and on the developmental parameters of the fetuses born to the experimental maternal animals. In this study, the test chemical was administered to the Sprague Dawley derived CD rats at the dose levels of 0, 50, 250 and 500 mg/kg bw/day as control, low, mid and high dose groups, respectively. The test chemical was mixed in a vehicle viz. Corn-oil and administered to the animals over the Day 6-15 gestation interval through gastric intubation route.The low and Mid-dose groups contained 24 mated females each. The high-dose group contained a total of 29 mated females. Also, included in the study was a vehicle treated control group (24 mated females). The female rats were about9 weeks at study initiation while the male rats used just for mating purposes were aproximately 26 weeks old. The animals were housed individually, except during the first week of the acclimation period (two females/cage) and mating, in stainless steel suspended cages with wire mesh floors. The animals were fed with Purina Certified Rodent Chow No. 5002 (mesh) fed ad libitum and were provided tap water delivered by automatic watering system, ad libitum. The animals were acclimatized for 21 days to laboratory conditions. The tempaerature was mainatained between 66-74°F. The humidity was maintained between 26-68 % while the photoperiod of 12 hours dark and 12 hours light was maintained. The dosing solutions were prepared fresh daily. Homogeneity of the dosing solutions was determined prior to the initiation of the study. A mock batch of dosing solutions for each dose level was prepared in a volume equivalent to a daily mix and three 10 ml samples weretaken from each dose level from the mixing beaker while it was stirring. One sample was taken from each level of the beaker (top, middle and bottom). A 24-hour stability at room temperature was also determined for each dose level using the dosing solutions prepared for the homogeneitys analyses. During the course of the study, a 10 ml sample of dosing solution was taken at each dose levsl one day of each week of treatment and analyzed for percent recovery. During mating, females selected for mating were placed with male rats in 1:1 ratio. Mating was conducted on 13 days,vaginal smears were taken early in the morning following intervals of cohabitation and females were considered to have mated if sperm and/or a vaginal plug was observed. The day on which evidence of mating was observed was defined as Day 0 of gestation. During observation period, the parental animals were observedtwice daily (morning and afternoon) for signs of pharmacologic or toxicologic effects and mortality. Each female was given a detailed physical examination on Days 0, 6, 10, 12, 15 and 20 of gestation. On Days 6, 10, 12 and 15 of gestation the examinations were given after dosing. Body weights were recorded on Days 0, 6, 10, 12, 15 and 20 of gestation. Food consumption was recorded for the following intervals during gestation day 0 to 6, 6 to 10, 10 to 15 and 15 to 20. In maternal animals, in the low-dose group. three females were noted with excessive salivation during the physical evaluations. No other adverse effects of treatment at this dose level were noted from the physical evaluations. In the mid- and high-dose groups. the following observations were noted with increased frequency during the physical evaluations: excessive salivation; staining of the skin/fur in the ano-genital area and excessive lacrimation. Additionally, in the high-dose group, chromodacryorrhea and/or ned nasal discharge were seen with increased frequency and several females had an unkempt appearance. No mortality occurred in the control. low or mid-dose groups. In the high-dose group. two females died and three females were sacrificed in a moribund condition. An adverse effect of treatment was evident from maternal weight gain data during the treatment interval of gestation. In the low-dose group. Mean weight gains for the Day 6-15 treatment interval of gestation and for the Day 6-20 gestation interval using corrected Day 20 weights were lower than control. However, these differences from control data were not statistically significant. In the mid-dose group, mean weight gains for the Day 6-15 and 6-20 gestation intervals were lower than control however, only for the Day 6-15 gestation interval did these data differ statistically from control data. In the high-dose group. mean weight gain data for the Day 6-15 and 6-20 gestation intervals were markedly lower than control and these data differed statistically from control data. During the Day 15-20 post-treatment interval of gestation, mean weight gain was comparable between the control, low- and hish-dose groups. In the mid-dose group. mean weight gain for the Day 15-20 gestation interval was higher than control and this difference was statistically significant. A number of the treated animals as well as a single untreated control animal showed discolorations of the lungs which were seen most frequently in animals from High Dose Group. All animals with discolored lungs were killed at the end of the study. The lungs of the single untreated control and from severral of the treated animals showed scattered grey foci; one animal from high dose group had lungs which showed both red discoloration and scattered grey foci. The liver of four animals from high dose group only were discolored; one of these animals was killed at the end of the study. two were killed in extremis, and one died spontaneuusly. The pattern of discoloration was variable; for the animal killed at the end of the study the liver was described as being pale, for the two animals killed extremis the liver for one was pale tan and for the other it was red/dark red/tan and for the animal which died spontaneously the liver was mottled tan. These findings were seen either exclusively or most frequently in animals from high dose group. Their toxicological significance with respect to the test chemical, while suggestive, remains equivocal in the absence of a microscopic examination. Other postmortem findings, observed grossly, occurred sporadically in the treated and control animals and were not considered to be related to the test chemical. During the Day 0-6 (pre-treatment) gestation interval, mean food consumption was comparable between the control and treated groups. During the treatment period. mean food consumption was lower than control in each of the treated groups. In the low-dose group, mean food consumption was significantly lower than control during the Day 6-10 gestation interval but was comparable to control for the day 10-15 interval. In the mid-dose group. mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. Similarly, in the high-dose group, mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. During the Day 15-20 (post-treatment) interval of gestation, mean food consumption was comparable between the control. low- and mid-dose groups. In the high-dose group. mean food consumption for the Day 15-20 gestation interval was significantly higher than control and this may represent compensatory feeding in response to the depressed food consumption encountered during the treatment period. the mean number of viable fetuses per pregnant female was comparable between the control and treated groups. The mean number of corpora lutea and uterine implantations was considered comparable between the control and treated groups. Likewise, the mean pre-implantation loss ratio was comparable between these same groups. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. A single female in high dose group, had 10 early resorptions. The incidence of females with at least one resorption site in utero wascomparable between the control and mid-dose group. In the low and high-dose groups, the incidence of females with resorption sites was higher than control; however, these data did not differ statistically from control data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. In fetal parameters, Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. No adverse effect of treatment was evident from-fetal sex distribution data. Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. A considerable spectrum of major malformations was noted among the control and treated groups. In the control group, one fetus had an exencephaly defect. In the low-dose group, one fetus had an omphalocele and a second fetus had a cleft lip/cleft palate defect. In the mid-dose group, one fetus had a cleft palate and a second fetus had a cleft palate and a distortion in shape of the snout i.e a slight ventral protrusion on the upper lip - considered an external variation observation. A third mid-dose fetus had absence of an eye bulge (unilateral-left sided) suggestive of an ocular defect (at visceral evaluation this fetus was noted with anophthalmia of the left eye). Filamentous tail. considered a minor malformation. was seen in two fetuses at the high-dose level. The only external malformation seen with repetition among the treated groups was cleft palate. This was the only external malformation in one mid-dose fetus and was seen in association with other facial irregularities (i.e a cleft lip, distortion in appearance of the snout) in the remaining two fetuses (one fetus each in the low and mid-dose groups).The absence of cleft palate among the other high-dose fetuses suggests that the occurrence of this malformation among several treated group fetuses was not a treatment-related phenomenon. The incidences os skeletal malformations did not differ statistically between the control and treated groups and no adverse effect of treatment was evident from fetal skeletal evaluations. Also, the incidences of ossification variation were comparable between the control. low and high dose groups. In the mid-dose group. the incidence of fetuses with ossification variations was statistically lower than the control data; however. this difference was not considered indicative of an adverse effect of treatment. In the high-dose group. the incidence of fetuses with one or more unossified sternebrae was increased suggestive of a retardation in ossification of the sternebrae. The incidence of fetuses with rudimentary rib structures was increased in the mid and high-dose groups in respect to control; however, these incidences were well within the range and no adverse effect of treatment was indicated. No other effects of treatment were evident from ossification variation data. In the control group. absence of an eye structure (i.e anophthalmia: unilateral - left sided), a major malformation, was seen in one fetus. In the mid-dose group, cleft palate was confirmed for the two fetuses noted externally with this malformation and anophthalmia (unilateral left sided) was seen for the one mid-dose fetus noted during the external examination with absence of eye bulge. The only other visceral malfonmation noted for the mid-dose group was a defect of the aortic arch. This defect involving an abnormal course of the arch passing dorsal, not ventral, to the esophagus/trachea, was seen in one fetus. During the evaluation of head sections of mid-dose fetus, along with confirmation of the externally noted cleft palate Defect, several malformations were noted involving the distortion of the snout. This fetus also had anophthalmia (unilateral left sided); presence of additional nasal folds; and presence of additional brain tissue which distorted the normal brain shape. In the high-dose group, extreme distortion of the lateral ventricles of the brain (i.e internal hydrocephalus), a major malformation, was seen in a fetus. In second fetus, a small eye structure (i.e., microphthalmia unilateral left sided), a major malformation, was observed. No adverse effect of treatment was indicated from visceral malformation data due to the dissimilarity in type of defects noted among the treated groups and the low incidence of fetuses affected. Tortuous and/or distended ureter with or without association of distended renal pelvis with papilla present was seen in two control fetuses, 12 low-dose fetuses, five mid-dose fetuses and. one high dose fetus. The visceral variations as noted among the treated group fetuses occurred at low incidence and did not indicate an adverse effect of treatment. Thus, no adverse effect of treatment was evident from visceral variation data. Thus, based on all the observations and results, it was concluded that, the NOAEL for the maternal toxicity was found to be 50 mg/kg bw/day while NOAEL and NOEL for fetotoxicity was found to be 250 and 50 mg/kg bw/day.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Approx. 120 days

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 NTRL report.

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

The above experiment was performed to evaluate and assess the effects of the test chemical on the pregnant rats and on the developmental parameters of the fetuses born to the experimental maternal animals.

GLP compliance:

not specified

Specific details on test material used for the study:

- Molecular formula (if other than submission substance): C10-H15-N
- Molecular weight (if other than submission substance): 149.236 g/mol
- Substance type: Organic
- Physical state: Dark Amber Liquid
- Impurities (identity and concentrations): No Data Available

Species:

rat

Strain:

other: CD (Sprague Dawley Derived)

Details on test animals or test system and environmental conditions:

Details on test animals and env. conditions
TEST ANIMALS
- Source:
Females: Charles River Breeding Laboratories. Inc. Shaver Road, Portage. Michigan 49081.
Males: Proven breeders used solely for mating purposes (Bio/dynamics' in-house breeding colony).
- Age at study initiation: (P) Males: Approx. 26 weeks and Females: 9 weeks
- Weight at study initiation: Females: 167.6-311.1 g
- Fasting period before study: No Data Available
- Housing: Individually. except during the first week of the acclimation period (two females/cage) and mating, in stainless steel suspended cages with wire mesh floors.
- Use of restrainers for preventing ingestion (if dermal): no
- Diet (e.g. ad libitum): Purina. Certified Rodent Chow No. 5002 (mesh) fed ad libitum.
- Water (e.g. ad libitum): Tap water delivered by automatic watering system, ad libitum (Elizabethtown Water Company).
- Acclimation period: 21 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 66-74°F
- Humidity (%): 26-68 %
- Air changes (per hr): No Data Available
- Photoperiod (hrs dark / hrs light): 12 hour light/dark cycle (7 AM to 7 PM)

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

Details on exposure
PREPARATION OF DOSING SOLUTIONS: Dosing solutions were prepared fresh daily. Homogeneity of the dosing solutions was determined prior to the initiation of the study. A mock batch of dosing solutions for each dose level was prepared in a volume equivalent to a daily mix and three 10 ml samples were taken from each dose level from the mixing beaker while it was stirring. One sample was taken from each level of the beaker (top, middle and bottom). A 24-hour stability at room temperature was also determined for each dose level using the dosing solutions prepared for the homogeneitys analyses. During the course of the study, a 10 ml sample of dosing solution was taken at
each dose levsl one day of each week of treatment and analyzed for percent recovery.

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): No Data Available
- Concentration in vehicle: No Data Available
- Amount of vehicle (if gavage): 5 ml/kg of boqy weight/day; dose volumes were derived from Day 6 body weights and were adjusted during the remaining treatment period to the most recent body weights.
- Lot/batch no. (if required): No Data Available
- Purity: No Data Available

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

STANDARD PREPARATION: Dilutions for injection were prepared as follows:
a. 10.0 ml of a 1000 ug/ml std. diluted to 100 ml for a 100 ug/ml std.
b. 5.0 ml of a 1000 ug/ml std. diluted to 100 ml for a 50.0 ug/ml std.
c. 1.0 ml of a 1000 ug/ml std. diluted to 100 ml for a 10.0 ug/ml std.
d. 0.5 ml of a 1000 ug/ml std. diluted to 100 ml for a 5.00 ug/ml std.
Additional standards were not used. Standards were stored in the freezer between analyses. Standards were prepared fresh on a monthly basis.

SAMPLE PREPARATION: One ml of each sample was diluted to 100 ml before the following dilutions were prepared for the appropriate dose levels:
a. 0.00 ug/ml Diluted the same as the lowest level on test
b. 100 - 499 ug/ml 5.0 ml diluted to 10.0 ml
c. 500 - 999 ug/ml 1.0 ml di luted to 10.0 ml
d. 1000 ug/ml 0.5 ml diluted to 10.0 ml
All standards were diluted in hexane. They were stored refrigerated as corn oil solutions and in the freezer as hexane dilutions between analyses.

LIQUID CHROMATOGRAPHIC ANALYSIS: A standard curve was generated by first injecting at least three different concentrations of standards. Standards were plotted as the peak area of the test chemical versus the test chemical standard concentration. A standard correlation coefficient was then calculated by linear regression. Samples, were injected and their recoveries also calculated by linear regression.

LIQUID CHROMATOGRAPHIC CONDITIONS FOR THE ANALYSIS OF THE TEST CHEMICAL:
Instrument: Waters 510 Pump, Waters 481 LC Spectrophotometer, WISP 710B Autoinjector, HP 3390 A Integrator
Column: Partisil PXS 10/25 PAC 10 urn
Mobile Phase:B 95/5 Hexane/Ethanol or Anhydrous Alcohol
Flow Rate: 1.0 ml minute
Sample Size: 25 ul
Chart Speed: 0.5 cm/minute
AUFS: 0.1 - 0.2
Analytical Wavelength: 280 nm

Details on mating procedure:

Details of mating
- M/F ratio per cage: Females selected for mating were placed with male rats nightly in a 1:1 ratio.
- Length of cohabitation: Mating was conducted on 13 days.
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: Vaginal smears were taken early in the morning following intervals of cohabitation and females were considered to have mated if sperm and/or a vaginal plug was observed. The day on which evidence of mating was observed was defined as Day 0 of gestation.
- 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): Individually
- Any other deviations from standard protocol: No Data Available

Duration of treatment / exposure:

Gestational Days 6 to 15. (Total 10 Days).

Frequency of treatment:

Daily

Duration of test:

Aprroximately 120 days.

Dose / conc.:

0 mg/kg bw/day

Remarks:

Control Group

Dose / conc.:

50 mg/kg bw/day

Remarks:

Low Dose Group

Dose / conc.:

250 mg/kg bw/day

Remarks:

Mid Dose Group

Dose / conc.:

500 mg/kg bw/day

Remarks:

High Dose Group

No. of animals per sex per dose:

The low and mid-dose groups contained 24 mated females and the high-dose group contained 29 mated females.

Control animals:

yes, concurrent vehicle

Details on study design:

No Data Available

Maternal examinations:

Parental animals observation and examinations: Yes,
Twice daily (morning and afternoon) for signs of pharmacologic or toxicologic effects and mortality.

DETAILED CLINICAL OBSERVATIONS: Yes, Each female was given a detailed physical examination on Days 0, 6, 10, 12, 15 and 20 of gestation. On Days 6, 10, 12 and 15 of gestation the examinations were given after dosing.

BODY WEIGHT: Yes Recorded on Days 0, 6, 10, 12, 15 and 20 of gestation.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes, Recorded for the following intervals during gestation: Days 0 to 6, 6 to 10,
10 to 15 and 15 to 20.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data

Ovaries and uterine content:

The intact uterus (ovaries attached) was removed from the abdominal cavity and weighed, and then the number and location of the following were recorded for each horn:
1.) live fetuses
2.) dead fetuses (no evidence of tissuedegeneration).
3.) late resorptions (recognizable dead.
4.) fetus undergoing degeneration regardless of size).
5.) early resorptions (evidence of implantation but no recognizable fetus)
6.) implantation sites
The ovaries were dissected free from the uterus and evaluated for the presence and number of corpora lutea. When no uterine implants were grossly apparent, the uterus was stained with ammonium sulfide. Foci visualized following this staining procedure were counted and females with such foci were included as pregnant in the calculation of pregnancy rate.

Fetal examinations:

All fetuses were gi ven a gross examination for external malformations/variations to include ohservation for palatel defects. Subsequently. Each fetus was weighed and individually identified. The sex of each fetus was noted externally (ano-genital distance) and was confirmed by internal inspection of the gonads at subsequent evaluation. Approximately one-half of the fetuses in each litter (alternating fetuses within the litter) were evaluated for soft tissue malformations/variations using a
microdissection procedures. Evaluations were performed on fresh fetal specimens shortly after removal from the uterus.

Fetuses designated for soft tissue evaluation were decapitated (head placed in appropriately labelled porous paper bags and fixed in Bouin's solution for later evaluation). The fetal specimens were then secured beneath a dissecting microscope and dissected so as to permit evaluation of tissues in the thoracic and abdominal cavities. At the completion of the fetal examination, the fetuses were eviscerated (viscera discarded) and placed in individual plastic cassettes and stored in 70% ethanol solution. Following a period of fixation, the fetal heads were sectioned using a razor blade. The serial, transverse sections generated during this procedure were evaluated for malformations of the palate, eyes and brain under a dissecting microscope (10-20X). Following evaluation, head sections were placed in plastic cassettes for storage (one litter/jar) in a 70% ethanol solution.

The remaining fetuses in each litter were sacrificed via lethal exposure to ether. The intact fetuses were eviscerated (internally sexed by inspection of the gonads) and processed for staining of the ossified skeletal structures using the Alizarin Red S staining procedure. Fetal skeletal specimens were evaluated under a dissecting microscope (10-20X) for ossification variations and malformations. Following evaluation. all fetuses were stored in plastic jars with plastic screw-on lids (1 litter/jar) in a 50/50 solution of 70% ethanol and 100% glycerin.

Late resorptions were examined grossly for external malformations and discarded.

Statistics:

No Data Available

Indices:

Implantation Index, Resorption Index, Pup Viability index.

Historical control data:

No Data Available

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

In the low-dose group. three females were noted with excessive salivation during the physical evaluations. No other adverse effects of treatment at this dose level were noted from the physical evaluations. In the mid- and high-dose groups. the following observations were noted with increased frequency during the physical evaluations: excessive salivation; staining of the skin/fur in the ano-genital area and excessive lacrimation. Additionally, in the high-dose group, chromodacryorrhea and/or ned nasal discharge were seen with increased frequency and several females had an unkempt appearance.

Dermal irritation (if dermal study):

not specified

Mortality:

mortality observed, treatment-related

Description (incidence):

No mortality occurred in the control. low or mid-dose groups. In the high-dose group. two females died and three females were sacrificed in a moribund condition.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

An adverse effect of treatment was evident from maternal weight gain data during the treatment interval of gestation. In the low-dose group. Mean weight gains for the Day 6-15 treatment interval of gestation and for the Day 6-20 gestation interval using corrected Day 20 weights were lower than control. However, these differences from control data were not statistically significant. In the mid-dose group, mean weight gains for the Day 6-15 and 6-20 gestation intervals were lower than control however, only for the Day 6-15 gestation interval did these data differ statistically from control data. In the high-dose group. mean weight gain data for the Day 6-15 and 6-20 gestation intervals were markedly lower than control and these data differed statistically from control data. During the Day 15-20 post-treatment interval of gestation, mean weight gain was comparable between the control, low- and hish-dose groups. In the mid-dose group. mean weight gain for the Day 15-20 gestation interval was higher than control and this difference was statistically significant.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

During the Day 0-6 (pre-treatment) gestation interval, mean food consumption was comparable between the control and treated groups. During the treatment period. mean food consumption was lower than control in each of the treated groups. In the low-dose group, mean food consumption was significantly lower than control during the Day 6-10 gestation interval but was comparable to control for the day 10-15 interval. In the mid-dose group. mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. Similarly, in the high-dose group, mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. During the Day 15-20 (post-treatment) interval of gestation, mean food consumption was comparable between the control. low- and mid-dose groups. In the high-dose group. mean food consumption for the Day 15-20 gestation interval was significantly higher than control and this may represent compensatory feeding in response to the depressed food consumption encountered during the treatment period.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

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:

not specified

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

A number of the treated animals as well as a single untreated control animal showed discolorations of the lungs which were seen most frequently in animals from High Dose Group. All animals with discolored lungs were killed at the end of the study. The lungs of the single untreated control and from severral of the treated animals showed scattered grey foci; one animal from high dose group had lungs which showed both red discoloration and scattered grey foci. The liver of four animals from high dose group only were discolored; one of these animals was killed at the end of the study. two were killed in extremis, and one died spontaneuusly. The pattern of discoloration was variable; for the animal killed at the end of the study the liver was described as being pale, for the two animals killed extremis the liver for one was pale tan and for the other it was red/dark red/tan and for the animal which died spontaneously the liver was mottled tan. These findings were seen either exclusively or most frequently in animals from high dose group. Their toxicological significance with respect to the test chemical, while suggestive, remains equivocal in the absence of a microscopic examination. Other postmortem findings, observed grossly, occurred sporadically in the treated and control animals and were not considered to be related to the test chemical. During the Day 0-6 (pre-treatment) gestation interval, mean food consumption was comparable between the control and treated groups. During the treatment period. mean food consumption was lower than control in each of the treated groups. In the low-dose group, mean food consumption was significantly lower than control during the Day 6-10 gestation interval but was comparable to control for the day 10-15 interval. In the mid-dose group. mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. Similarly, in the high-dose group, mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. During the Day 15-20 (post-treatment) interval of gestation, mean food consumption was comparable between the control. low- and mid-dose groups. In the high-dose group. mean food consumption for the Day 15-20 gestation interval was significantly higher than control and this may represent compensatory feeding in response to the depressed food consumption encountered during the treatment period.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

In the low-dose group. three females were noted with excessive salivation during the physical evaluations. No other adverse effects of treatment at this dose level were noted from the physical evaluations. In the mid- and high-dose groups. the following observations were noted with increased frequency during the physical evaluations: excessive salivation; staining of the skin/fur in the ano-genital area and excessive lacrimation. Additionally, in the high-dose group, chromodacryorrhea and/or ned nasal discharge were seen with increased frequency and several females had an unkempt appearance. No mortality occurred in the control. low or mid-dose groups. In the high-dose group. two females died and three females were sacrificed in a moribund condition. An adverse effect of treatment was evident from maternal weight gain data during the treatment interval of gestation. In the low-dose group. Mean weight gains for the Day 6-15 treatment interval of gestation and for the Day 6-20 gestation interval using corrected Day 20 weights were lower than control. However, these differences from control data were not statistically significant. In the mid-dose group, mean weight gains for the Day 6-15 and 6-20 gestation intervals were lower than control however, only for the Day 6-15 gestation interval did these data differ statistically from control data. In the high-dose group. mean weight gain data for the Day 6-15 and 6-20 gestation intervals were markedly lower than control and these data differed statistically from control data. During the Day 15-20 post-treatment interval of gestation, mean weight gain was comparable between the control, low- and hish-dose groups. In the mid-dose group. mean weight gain for the Day 15-20 gestation interval was higher than control and this difference was statistically significant. A number of the treated animals as well as a single untreated control animal showed discolorations of the lungs which were seen most frequently in animals from High Dose Group. All animals with discolored lungs were killed at the end of the study. The lungs of the single untreated control and from severral of the treated animals showed scattered grey foci; one animal from high dose group had lungs which showed both red discoloration and scattered grey foci. The liver of four animals from high dose group only were discolored; one of these animals was killed at the end of the study. two were killed in extremis, and one died spontaneuusly. The pattern of discoloration was variable; for the animal killed at the end of the study the liver was described as being pale, for the two animals killed extremis the liver for one was pale tan and for the other it was red/dark red/tan and for the animal which died spontaneously the liver was mottled tan. These findings were seen either exclusively or most frequently in animals from high dose group. Their toxicological significance with respect to the test chemical, while suggestive, remains equivocal in the absence of a microscopic examination. Other postmortem findings, observed grossly, occurred sporadically in the treated and control animals and were not considered to be related to the test chemical. During the Day 0-6 (pre-treatment) gestation interval, mean food consumption was comparable between the control and treated groups. During the treatment period. mean food consumption was lower than control in each of the treated groups. In the low-dose group, mean food consumption was significantly lower than control during the Day 6-10 gestation interval but was comparable to control for the day 10-15 interval. In the mid-dose group. mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. Similarly, in the high-dose group, mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. During the Day 15-20 (post-treatment) interval of gestation, mean food consumption was comparable between the control. low- and mid-dose groups. In the high-dose group. mean food consumption for the Day 15-20 gestation interval was significantly higher than control and this may represent compensatory feeding in response to the depressed food consumption encountered during the treatment period.

Number of abortions:

no effects observed

Description (incidence and severity):

The mean number of viable fetuses per pregnant female was comparable between the control and treated groups.

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

The mean number of corpora lutea and uterine implantations was considered comparable between the control and treated groups. Likewise, the mean pre-implantation loss ratio was comparable between these same groups.

Total litter losses by resorption:

no effects observed

Description (incidence and severity):

The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant.

Early or late resorptions:

effects observed, non-treatment-related

Description (incidence and severity):

The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. A single female in high dose group, had 10 early resorptions. The incidence of females with at least one resorption site in utero wascomparable between the control and mid-dose group. In the low and high-dose groups, the incidence of females with resorption sites was higher than control; however, these data did not differ statistically from control data.

Dead fetuses:

no effects observed

Description (incidence and severity):

No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female.

Changes in pregnancy duration:

not specified

Changes in number of pregnant:

not specified

Other effects:

not specified

Details on maternal toxic effects:

The mean number of viable fetuses per pregnant female was comparable between the control and treated groups. The mean number of corpora lutea and uterine implantations was considered comparable between the control and treated groups. Likewise, the mean pre-implantation loss ratio was comparable between these same groups. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. A single female in high dose group, had 10 early resorptions. The incidence of females with at least one resorption site in utero wascomparable between the control and mid-dose group. In the low and high-dose groups, the incidence of females with resorption sites was higher than control; however, these data did not differ statistically from control data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female.

Dose descriptor:

LOAEL

Effect level:

50 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

clinical signs

dead fetuses

early or late resorptions

food consumption and compound intake

gross pathology

mortality

number of abortions

pre and post implantation loss

total litter losses by resorption

Remarks on result:

other: Not Specified

Abnormalities:

not specified

Fetal body weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data.

Reduction in number of live offspring:

no effects observed

Description (incidence and severity):

No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female.

Changes in sex ratio:

no effects observed

Description (incidence and severity):

No adverse effect of treatment was evident from-fetal sex distribution data.

Changes in litter size and weights:

no effects observed

Description (incidence and severity):

Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data.

Changes in postnatal survival:

not specified

External malformations:

effects observed, non-treatment-related

Description (incidence and severity):

A considerable spectrum of major malformations was noted among the control and treated groups. In the control group, one fetus had an exencephaly defect. In the low-dose group, one fetus had an omphalocele and a second fetus had a cleft lip/cleft palate defect. In the mid-dose group, one fetus had a cleft palate and a second fetus had a cleft palate and a distortion in shape of the snout i.e a slight ventral protrusion on the upper lip - considered an external variation observation. A third mid-dose fetus had absence of an eye bulge (unilateral-left sided) suggestive of an ocular defect (at visceral evaluation this fetus was noted with anophthalmia of the left eye). Filamentous tail. considered a minor malformation. was seen in two fetuses at the high-dose level. The only external malformation seen with repetition among the treated groups was cleft palate. This was the only external malformation in one mid-dose fetus and was seen in association with other facial irregularities (i.e a cleft lip, distortion in appearance of the snout) in the remaining two fetuses (one fetus each in the low and mid-dose groups).The absence of cleft
palate among the other high-dose fetuses suggests that the occurrence of this malformation among several treated group fetuses was not a treatment-related
phenomenon.

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

The incidences os skeletal malformations did not differ statistically between the control and treated groups and no adverse effect of treatment was evident from fetal
skeletal evaluations. Also, the incidences of ossification variation were comparable between the control. low and high dose groups. In the mid-dose group. the incidence of fetuses with ossification variations was statistically lower than the control data; however. this difference was not considered indicative of an adverse effect of treatment. In the high-dose group. the incidence of fetuses with one or more unossified sternebrae was increased suggestive of a retardation in ossification of the sternebrae. The incidence of fetuses with rudimentary rib structures was increased in the mid and high-dose groups in respect to control; however, these incidences were well within the range and no adverse effect of treatment was indicated. No other effects of treatment were evident from ossification variation data.

Visceral malformations:

effects observed, non-treatment-related

Description (incidence and severity):

In the control group. absence of an eye structure (i.e anophthalmia: unilateral - left sided), a major malformation, was seen in one fetus. In the mid-dose group, cleft palate was confirmed for the two fetuses noted externally with this malformation and anophthalmia (unilateral left sided) was seen for the one mid-dose fetus noted during the external examination with absence of eye bulge. The only other visceral malfonmation noted for the mid-dose group was a defect of the aortic arch. This defect involving an abnormal course of the arch passing dorsal, not ventral, to the esophagus/trachea, was seen in one fetus. During the evaluation of head sections of mid-dose fetus, along with confirmation of the externally noted cleft palate Defect, several malformations were noted involving the distortion of the snout. This fetus also had anophthalmia (unilateral left sided); presence of additional nasal folds; and presence of additional brain tissue which distorted the normal brain shape. In the high-dose group, extreme distortion of the lateral ventricles of the brain (i.e internal hydrocephalus), a major malformation, was seen in a fetus. In second fetus, a small eye structure (i.e., microphthalmia unilateral left sided), a major malformation, was observed. No adverse effect of treatment was indicated from visceral malformation data due to the dissimilarity in type of defects noted among the treated groups and the low incidence of fetuses affected. Tortuous and/or
distended ureter with or without association of distended renal pelvis with papilla present was seen in two control fetuses, 12 low-dose fetuses, five mid-dose fetuses and. one high dose fetus. The visceral variations as noted among the treated group fetuses occurred at low incidence and did not indicate an adverse effect
of treatment. Thus, no adverse effect of treatment was evident from visceral variation data.

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. No adverse effect of treatment was evident from-fetal sex distribution data. Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. A considerable spectrum of major malformations was noted among the control and treated groups. In the control group, one fetus had an exencephaly defect. In the low-dose group, one fetus had an omphalocele and a second fetus had a cleft lip/cleft palate defect. In the mid-dose group, one fetus had a cleft palate and a second fetus had a cleft palate and a distortion in shape of the snout i.e a slight ventral protrusion on the upper lip - considered an external variation observation. A third mid-dose fetus had absence of an eye bulge (unilateral-left sided) suggestive of an ocular defect (at visceral evaluation this fetus was noted with anophthalmia of the left eye). Filamentous tail. considered a minor malformation. was seen in two fetuses at the high-dose level. The only external malformation seen with repetition among the treated groups was cleft palate. This was the only external malformation in one mid-dose fetus and was seen in association with other facial irregularities (i.e a cleft lip, distortion in appearance of the snout) in the remaining two fetuses (one fetus each in the low and mid-dose groups).The absence of cleft
palate among the other high-dose fetuses suggests that the occurrence of this malformation among several treated group fetuses was not a treatment-related
phenomenon. The incidences os skeletal malformations did not differ statistically between the control and treated groups and no adverse effect of treatment was evident from fetal skeletal evaluations. Also, the incidences of ossification variation were comparable between the control. low and high dose groups. In the mid-dose group. the incidence of fetuses with ossification variations was statistically lower than the control data; however. this difference was not considered indicative of an adverse effect of treatment. In the high-dose group. the incidence of fetuses with one or more unossified sternebrae was increased suggestive of a retardation in ossification of the sternebrae. The incidence of fetuses with rudimentary rib structures was increased in the mid and high-dose groups in respect to control; however, these incidences were well within the range and no adverse effect of treatment was indicated. No other effects of treatment were evident from ossification variation data. In the control group. absence of an eye structure (i.e anophthalmia: unilateral - left sided), a major malformation, was seen in one fetus. In the mid-dose group, cleft palate was confirmed for the two fetuses noted externally with this malformation and anophthalmia (unilateral left sided) was seen for the one mid-dose fetus noted during the external examination with absence of eye bulge. The only other visceral malfonmation noted for the mid-dose group was a defect of the aortic arch. This defect involving an abnormal course of the arch passing dorsal, not ventral, to the esophagus/trachea, was seen in one fetus. During the evaluation of head sections of mid-dose fetus, along with confirmation of the externally noted cleft palate Defect, several malformations were noted involving the distortion of the snout. This fetus also had anophthalmia (unilateral left sided); presence of additional nasal folds; and presence of additional brain tissue which distorted the normal brain shape. In the high-dose group, extreme distortion of the lateral ventricles of the brain (i.e internal hydrocephalus), a major malformation, was seen in a fetus. In second fetus, a small eye structure (i.e., microphthalmia unilateral left sided), a major malformation, was observed. No adverse effect of treatment was indicated from visceral malformation data due to the dissimilarity in type of defects noted among the treated groups and the low incidence of fetuses affected. Tortuous and/or distended ureter with or without association of distended renal pelvis with papilla present was seen in two control fetuses, 12 low-dose fetuses, five mid-dose fetuses and. one high dose fetus. The visceral variations as noted among the treated group fetuses occurred at low incidence and did not indicate an adverse effect of treatment. Thus, no adverse effect of treatment was evident from visceral variation data.

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

changes in sex ratio

fetal/pup body weight changes

changes in litter size and weights

external malformations

skeletal malformations

visceral malformations

Remarks on result:

other: Not Specified

Dose descriptor:

NOEL

Effect level:

50 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

changes in sex ratio

fetal/pup body weight changes

changes in litter size and weights

external malformations

skeletal malformations

visceral malformations

Remarks on result:

other: Not Specified

Abnormalities:

not specified

Developmental effects observed:

not specified

Treatment related:

not specified

Conclusions:

Based on all the observations and results, it was concluded that, the NOAEL for the maternal toxicity was found to be 50 mg/kg bw/day while NOAEL and NOEL for fetotoxicity was found to be 250 and 50 mg/kg bw/day.

Executive summary:

A pre-natal developmental toxicity study was conducted to evaluate and assess the effects of the test chemical on the pregnant rats and on the developmental parameters of the fetuses born to the experimental maternal animals. In this study, the test chemical was administered to the Sprague Dawley derived CD rats at the dose levels of 0, 50, 250 and 500 mg/kg bw/day as control, low, mid and high dose groups, respectively. The test chemical was mixed in a vehicle viz. Corn-oil and administered to the animals over the Day 6-15 gestation interval through gastric intubation route. The low and Mid-dose groups contained 24 mated females each. The high-dose group contained a total of 29 mated females. Also, included in the study was a vehicle treated control group (24 mated females). The female rats were about9 weeks at study initiation while the male rats used just for mating purposes were aproximately 26 weeks old. The animals were housed individually, except during the first week of the acclimation period (two females/cage) and mating, in stainless steel suspended cages with wire mesh floors. The animals were fed with Purina Certified Rodent Chow No. 5002 (mesh) fed ad libitum and were provided tap water delivered by automatic watering system, ad libitum. The animals were acclimatized for 21 days to laboratory conditions. The tempaerature was mainatained between 66-74°F. The humidity was maintained between 26-68 % while the photoperiod of 12 hours dark and 12 hours light was maintained. The dosing solutions were prepared fresh daily. Homogeneity of the dosing solutions was determined prior to the initiation of the study. A mock batch of dosing solutions for each dose level was prepared in a volume equivalent to a daily mix and three 10 ml samples weretaken from each dose level from the mixing beaker while it was stirring. One sample was taken from each level of the beaker (top, middle and bottom). A 24-hour stability at room temperature was also determined for each dose level using the dosing solutions prepared for the homogeneitys analyses. During the course of the study, a 10 ml sample of dosing solution was taken at each dose levsl one day of each week of treatment and analyzed for percent recovery. During mating, females selected for mating were placed with male rats in 1:1 ratio. Mating was conducted on 13 days, vaginal smears were taken early in the morning following intervals of cohabitation and females were considered to have mated if sperm and/or a vaginal plug was observed. The day on which evidence of mating was observed was defined as Day 0 of gestation. During observation period, the parental animals were observedtwice daily (morning and afternoon) for signs of pharmacologic or toxicologic effects and mortality. Each female was given a detailed physical examination on Days 0, 6, 10, 12, 15 and 20 of gestation. On Days 6, 10, 12 and 15 of gestation the examinations were given after dosing. Body weights were recorded on Days 0, 6, 10, 12, 15 and 20 of gestation. Food consumption was recorded for the following intervals during gestation day 0 to 6, 6 to 10, 10 to 15 and 15 to 20. In maternal animals, in the low-dose group. three females were noted with excessive salivation during the physical evaluations. No other adverse effects of treatment at this dose level were noted from the physical evaluations. In the mid- and high-dose groups. the following observations were noted with increased frequency during the physical evaluations: excessive salivation; staining of the skin/fur in the ano-genital area and excessive lacrimation. Additionally, in the high-dose group, chromodacryorrhea and/or ned nasal discharge were seen with increased frequency and several females had an unkempt appearance. No mortality occurred in the control. low or mid-dose groups. In the high-dose group. two females died and three females were sacrificed in a moribund condition. An adverse effect of treatment was evident from maternal weight gain data during the treatment interval of gestation. In the low-dose group. Mean weight gains for the Day 6-15 treatment interval of gestation and for the Day 6-20 gestation interval using corrected Day 20 weights were lower than control. However, these differences from control data were not statistically significant. In the mid-dose group, mean weight gains for the Day 6-15 and 6-20 gestation intervals were lower than control however, only for the Day 6-15 gestation interval did these data differ statistically from control data. In the high-dose group. mean weight gain data for the Day 6-15 and 6-20 gestation intervals were markedly lower than control and these data differed statistically from control data. During the Day 15-20 post-treatment interval of gestation, mean weight gain was comparable between the control, low- and hish-dose groups. In the mid-dose group. mean weight gain for the Day 15-20 gestation interval was higher than control and this difference was statistically significant. A number of the treated animals as well as a single untreated control animal showed discolorations of the lungs which were seen most frequently in animals from High Dose Group. All animals with discolored lungs were killed at the end of the study. The lungs of the single untreated control and from severral of the treated animals showed scattered grey foci; one animal from high dose group had lungs which showed both red discoloration and scattered grey foci. The liver of four animals from high dose group only were discolored; one of these animals was killed at the end of the study. two were killed in extremis, and one died spontaneuusly. The pattern of discoloration was variable; for the animal killed at the end of the study the liver was described as being pale, for the two animals killed extremis the liver for one was pale tan and for the other it was red/dark red/tan and for the animal which died spontaneously the liver was mottled tan. These findings were seen either exclusively or most frequently in animals from high dose group. Their toxicological significance with respect to the test chemical, while suggestive, remains equivocal in the absence of a microscopic examination. Other postmortem findings, observed grossly, occurred sporadically in the treated and control animals and were not considered to be related to the test chemical. During the Day 0-6 (pre-treatment) gestation interval, mean food consumption was comparable between the control and treated groups. During the treatment period. mean food consumption was lower than control in each of the treated groups. In the low-dose group, mean food consumption was significantly lower than control during the Day 6-10 gestation interval but was comparable to control for the day 10-15 interval. In the mid-dose group. mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. Similarly, in the high-dose group, mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. During the Day 15-20 (post-treatment) interval of gestation, mean food consumption was comparable between the control. low- and mid-dose groups. In the high-dose group. mean food consumption for the Day 15-20 gestation interval was significantly higher than control and this may represent compensatory feeding in response to the depressed food consumption encountered during the treatment period. the mean number of viable fetuses per pregnant female was comparable between the control and treated groups. The mean number of corpora lutea and uterine implantations was considered comparable between the control and treated groups. Likewise, the mean pre-implantation loss ratio was comparable between these same groups. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. A single female in high dose group, had 10 early resorptions. The incidence of females with at least one resorption site in utero wascomparable between the control and mid-dose group. In the low and high-dose groups, the incidence of females with resorption sites was higher than control; however, these data did not differ statistically from control data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. In fetal parameters, Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. No adverse effect of treatment was evident from-fetal sex distribution data. Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. A considerable spectrum of major malformations was noted among the control and treated groups. In the control group, one fetus had an exencephaly defect. In the low-dose group, one fetus had an omphalocele and a second fetus had a cleft lip/cleft palate defect. In the mid-dose group, one fetus had a cleft palate and a second fetus had a cleft palate and a distortion in shape of the snout i.e a slight ventral protrusion on the upper lip - considered an external variation observation. A third mid-dose fetus had absence of an eye bulge (unilateral-left sided) suggestive of an ocular defect (at visceral evaluation this fetus was noted with anophthalmia of the left eye). Filamentous tail. considered a minor malformation. was seen in two fetuses at the high-dose level. The only external malformation seen with repetition among the treated groups was cleft palate. This was the only external malformation in one mid-dose fetus and was seen in association with other facial irregularities (i.e a cleft lip, distortion in appearance of the snout) in the remaining two fetuses (one fetus each in the low and mid-dose groups).The absence of cleft palate among the other high-dose fetuses suggests that the occurrence of this malformation among several treated group fetuses was not a treatment-related phenomenon. The incidences os skeletal malformations did not differ statistically between the control and treated groups and no adverse effect of treatment was evident from fetal skeletal evaluations. Also, the incidences of ossification variation were comparable between the control. low and high dose groups. In the mid-dose group. the incidence of fetuses with ossification variations was statistically lower than the control data; however. this difference was not considered indicative of an adverse effect of treatment. In the high-dose group. the incidence of fetuses with one or more unossified sternebrae was increased suggestive of a retardation in ossification of the sternebrae. The incidence of fetuses with rudimentary rib structures was increased in the mid and high-dose groups in respect to control; however, these incidences were well within the range and no adverse effect of treatment was indicated. No other effects of treatment were evident from ossification variation data. In the control group. absence of an eye structure (i.e anophthalmia: unilateral - left sided), a major malformation, was seen in one fetus. In the mid-dose group, cleft palate was confirmed for the two fetuses noted externally with this malformation and anophthalmia (unilateral left sided) was seen for the one mid-dose fetus noted during the external examination with absence of eye bulge. The only other visceral malfonmation noted for the mid-dose group was a defect of the aortic arch. This defect involving an abnormal course of the arch passing dorsal, not ventral, to the esophagus/trachea, was seen in one fetus. During the evaluation of head sections of mid-dose fetus, along with confirmation of the externally noted cleft palate Defect, several malformations were noted involving the distortion of the snout. This fetus also had anophthalmia (unilateral left sided); presence of additional nasal folds; and presence of additional brain tissue which distorted the normal brain shape. In the high-dose group, extreme distortion of the lateral ventricles of the brain (i.e internal hydrocephalus), a major malformation, was seen in a fetus. In second fetus, a small eye structure (i.e., microphthalmia unilateral left sided), a major malformation, was observed. No adverse effect of treatment was indicated from visceral malformation data due to the dissimilarity in type of defects noted among the treated groups and the low incidence of fetuses affected. Tortuous and/or distended ureter with or without association of distended renal pelvis with papilla present was seen in two control fetuses, 12 low-dose fetuses, five mid-dose fetuses and. one high dose fetus. The visceral variations as noted among the treated group fetuses occurred at low incidence and did not indicate an adverse effect of treatment. Thus, no adverse effect of treatment was evident from visceral variation data. Thus, based on all the observations and results, it was concluded that, the NOAEL for the maternal toxicity was found to be 50 mg/kg bw/day while NOAEL and NOEL for fetotoxicity was found to be 250 and 50 mg/kg bw/day.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

250 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

The data is from a Klimisch 2 database and gives a robust study summary.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Developmental Toxicity Study:

A pre-natal developmental toxicity study was conducted to evaluate and assess the effects of the test chemical on the pregnant rats and on the developmental parameters of the fetuses born to the experimental maternal animals. In this study, the test chemical was administered to the Sprague Dawley derived CD rats at the dose levels of 0, 50, 250 and 500 mg/kg bw/day as control, low, mid and high dose groups, respectively. The test chemical was mixed in a vehicle viz. Corn-oil and administered to the animals over the Day 6-15 gestation interval through gastric intubation route.The low and Mid-dose groups contained 24 mated females each. The high-dose group contained a total of 29 mated females. Also, included in the study was a vehicle treated control group (24 mated females). The female rats were about9 weeks at study initiation while the male rats used just for mating purposes were aproximately 26 weeks old. The animals were housed individually, except during the first week of the acclimation period (two females/cage) and mating, in stainless steel suspended cages with wire mesh floors. The animals were fed with Purina Certified Rodent Chow No. 5002 (mesh) fed ad libitum and were provided tap water delivered by automatic watering system, ad libitum. The animals were acclimatized for 21 days to laboratory conditions. The tempaerature was mainatained between 66-74°F. The humidity was maintained between 26-68 % while the photoperiod of 12 hours dark and 12 hours light was maintained. The dosing solutions were prepared fresh daily. Homogeneity of the dosing solutions was determined prior to the initiation of the study. A mock batch of dosing solutions for each dose level was prepared in a volume equivalent to a daily mix and three 10 ml samples weretaken from each dose level from the mixing beaker while it was stirring. One sample was taken from each level of the beaker (top, middle and bottom). A 24-hour stability at room temperature was also determined for each dose level using the dosing solutions prepared for the homogeneitys analyses. During the course of the study, a 10 ml sample of dosing solution was taken at each dose levsl one day of each week of treatment and analyzed for percent recovery. During mating, females selected for mating were placed with male rats in 1:1 ratio. Mating was conducted on 13 days,vaginal smears were taken early in the morning following intervals of cohabitation and females were considered to have mated if sperm and/or a vaginal plug was observed. The day on which evidence of mating was observed was defined as Day 0 of gestation. During observation period, the parental animals were observedtwice daily (morning and afternoon) for signs of pharmacologic or toxicologic effects and mortality. Each female was given a detailed physical examination on Days 0, 6, 10, 12, 15 and 20 of gestation. On Days 6, 10, 12 and 15 of gestation the examinations were given after dosing. Body weights were recorded on Days 0, 6, 10, 12, 15 and 20 of gestation. Food consumption was recorded for the following intervals during gestation day 0 to 6, 6 to 10, 10 to 15 and 15 to 20. In maternal animals, in the low-dose group. three females were noted with excessive salivation during the physical evaluations. No other adverse effects of treatment at this dose level were noted from the physical evaluations. In the mid- and high-dose groups. the following observations were noted with increased frequency during the physical evaluations: excessive salivation; staining of the skin/fur in the ano-genital area and excessive lacrimation. Additionally, in the high-dose group, chromodacryorrhea and/or ned nasal discharge were seen with increased frequency and several females had an unkempt appearance. No mortality occurred in the control. low or mid-dose groups. In the high-dose group. two females died and three females were sacrificed in a moribund condition. An adverse effect of treatment was evident from maternal weight gain data during the treatment interval of gestation. In the low-dose group. Mean weight gains for the Day 6-15 treatment interval of gestation and for the Day 6-20 gestation interval using corrected Day 20 weights were lower than control. However, these differences from control data were not statistically significant. In the mid-dose group, mean weight gains for the Day 6-15 and 6-20 gestation intervals were lower than control however, only for the Day 6-15 gestation interval did these data differ statistically from control data. In the high-dose group. mean weight gain data for the Day 6-15 and 6-20 gestation intervals were markedly lower than control and these data differed statistically from control data. During the Day 15-20 post-treatment interval of gestation, mean weight gain was comparable between the control, low- and hish-dose groups. In the mid-dose group. mean weight gain for the Day 15-20 gestation interval was higher than control and this difference was statistically significant. A number of the treated animals as well as a single untreated control animal showed discolorations of the lungs which were seen most frequently in animals from High Dose Group. All animals with discolored lungs were killed at the end of the study. The lungs of the single untreated control and from severral of the treated animals showed scattered grey foci; one animal from high dose group had lungs which showed both red discoloration and scattered grey foci. The liver of four animals from high dose group only were discolored; one of these animals was killed at the end of the study. two were killed in extremis, and one died spontaneuusly. The pattern of discoloration was variable; for the animal killed at the end of the study the liver was described as being pale, for the two animals killed extremis the liver for one was pale tan and for the other it was red/dark red/tan and for the animal which died spontaneously the liver was mottled tan. These findings were seen either exclusively or most frequently in animals from high dose group. Their toxicological significance with respect to the test chemical, while suggestive, remains equivocal in the absence of a microscopic examination. Other postmortem findings, observed grossly, occurred sporadically in the treated and control animals and were not considered to be related to the test chemical. During the Day 0-6 (pre-treatment) gestation interval, mean food consumption was comparable between the control and treated groups. During the treatment period. mean food consumption was lower than control in each of the treated groups. In the low-dose group, mean food consumption was significantly lower than control during the Day 6-10 gestation interval but was comparable to control for the day 10-15 interval. In the mid-dose group. mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. Similarly, in the high-dose group, mean food consumption was significantly lower than control during both the Day 6-10 and 10-15 gestation intervals. During the Day 15-20 (post-treatment) interval of gestation, mean food consumption was comparable between the control. low- and mid-dose groups. In the high-dose group. mean food consumption for the Day 15-20 gestation interval was significantly higher than control and this may represent compensatory feeding in response to the depressed food consumption encountered during the treatment period. the mean number of viable fetuses per pregnant female was comparable between the control and treated groups. The mean number of corpora lutea and uterine implantations was considered comparable between the control and treated groups. Likewise, the mean pre-implantation loss ratio was comparable between these same groups. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. The mean number of resorption sites per pregnant female was comparable between the control. low and mid-dose groups. In the high-dose group, the mean number of resorption sites was higher than control; however, this difference was not statistically significant. A single female in high dose group, had 10 early resorptions. The incidence of females with at least one resorption site in utero wascomparable between the control and mid-dose group. In the low and high-dose groups, the incidence of females with resorption sites was higher than control; however, these data did not differ statistically from control data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. In fetal parameters, Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. No dead fetuses were recovered from the control, low or mid-dose groups. In the high-dose group, one dead fetus was recovered from female. No adverse effect of treatment was evident from-fetal sex distribution data. Mean fetal weight data as a composite for both sexes and when distinguished by sex, were comparable between the control, low and mid-dose groups. At the high-dose level, mean fetal weight data were lower than control and these differences were statistically significant. Thus. only at the high-dose level was an adverse effect of treatment evident from fetal weight data. A considerable spectrum of major malformations was noted among the control and treated groups. In the control group, one fetus had an exencephaly defect. In the low-dose group, one fetus had an omphalocele and a second fetus had a cleft lip/cleft palate defect. In the mid-dose group, one fetus had a cleft palate and a second fetus had a cleft palate and a distortion in shape of the snout i.e a slight ventral protrusion on the upper lip - considered an external variation observation. A third mid-dose fetus had absence of an eye bulge (unilateral-left sided) suggestive of an ocular defect (at visceral evaluation this fetus was noted with anophthalmia of the left eye). Filamentous tail. considered a minor malformation. was seen in two fetuses at the high-dose level. The only external malformation seen with repetition among the treated groups was cleft palate. This was the only external malformation in one mid-dose fetus and was seen in association with other facial irregularities (i.e a cleft lip, distortion in appearance of the snout) in the remaining two fetuses (one fetus each in the low and mid-dose groups).The absence of cleft palate among the other high-dose fetuses suggests that the occurrence of this malformation among several treated group fetuses was not a treatment-related phenomenon. The incidences os skeletal malformations did not differ statistically between the control and treated groups and no adverse effect of treatment was evident from fetal skeletal evaluations. Also, the incidences of ossification variation were comparable between the control. low and high dose groups. In the mid-dose group. the incidence of fetuses with ossification variations was statistically lower than the control data; however. this difference was not considered indicative of an adverse effect of treatment. In the high-dose group. the incidence of fetuses with one or more unossified sternebrae was increased suggestive of a retardation in ossification of the sternebrae. The incidence of fetuses with rudimentary rib structures was increased in the mid and high-dose groups in respect to control; however, these incidences were well within the range and no adverse effect of treatment was indicated. No other effects of treatment were evident from ossification variation data. In the control group. absence of an eye structure (i.e anophthalmia: unilateral - left sided), a major malformation, was seen in one fetus. In the mid-dose group, cleft palate was confirmed for the two fetuses noted externally with this malformation and anophthalmia (unilateral left sided) was seen for the one mid-dose fetus noted during the external examination with absence of eye bulge. The only other visceral malfonmation noted for the mid-dose group was a defect of the aortic arch. This defect involving an abnormal course of the arch passing dorsal, not ventral, to the esophagus/trachea, was seen in one fetus. During the evaluation of head sections of mid-dose fetus, along with confirmation of the externally noted cleft palate Defect, several malformations were noted involving the distortion of the snout. This fetus also had anophthalmia (unilateral left sided); presence of additional nasal folds; and presence of additional brain tissue which distorted the normal brain shape. In the high-dose group, extreme distortion of the lateral ventricles of the brain (i.e internal hydrocephalus), a major malformation, was seen in a fetus. In second fetus, a small eye structure (i.e., microphthalmia unilateral left sided), a major malformation, was observed. No adverse effect of treatment was indicated from visceral malformation data due to the dissimilarity in type of defects noted among the treated groups and the low incidence of fetuses affected. Tortuous and/or distended ureter with or without association of distended renal pelvis with papilla present was seen in two control fetuses, 12 low-dose fetuses, five mid-dose fetuses and. one high dose fetus. The visceral variations as noted among the treated group fetuses occurred at low incidence and did not indicate an adverse effect of treatment. Thus, no adverse effect of treatment was evident from visceral variation data. Thus, based on all the observations and results, it was concluded that, the NOAEL for the maternal toxicity was found to be 50 mg/kg bw/day while NOAEL and NOEL for fetotoxicity was found to be 250 and 50 mg/kg bw/day.

Justification for classification or non-classification

Based on the available information, the test chemical does not show effects on reproductive and developmental parameters of the animals. Thus, the test chemical cannot be classified as the reproductive and developmental toxicant according to CLP regulation.